WO2023193789A1 - Wee1 degrading compounds - Google Patents
Wee1 degrading compounds Download PDFInfo
- Publication number
- WO2023193789A1 WO2023193789A1 PCT/CN2023/086879 CN2023086879W WO2023193789A1 WO 2023193789 A1 WO2023193789 A1 WO 2023193789A1 CN 2023086879 W CN2023086879 W CN 2023086879W WO 2023193789 A1 WO2023193789 A1 WO 2023193789A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- phenyl
- methyl
- amino
- pyrazolo
- pyrimidin
- Prior art date
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- 150000001875 compounds Chemical class 0.000 title claims abstract description 149
- 101150040313 Wee1 gene Proteins 0.000 title description 6
- 230000000593 degrading effect Effects 0.000 title description 2
- 238000000034 method Methods 0.000 claims abstract description 32
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 22
- 201000010099 disease Diseases 0.000 claims abstract description 17
- 238000011282 treatment Methods 0.000 claims abstract description 15
- -1 amino, hydroxyl Chemical group 0.000 claims description 531
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 114
- 125000004194 piperazin-1-yl group Chemical group [H]N1C([H])([H])C([H])([H])N(*)C([H])([H])C1([H])[H] 0.000 claims description 114
- 229910052736 halogen Inorganic materials 0.000 claims description 112
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 108
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims description 105
- 150000003839 salts Chemical class 0.000 claims description 94
- 229910052739 hydrogen Inorganic materials 0.000 claims description 79
- 239000001257 hydrogen Substances 0.000 claims description 79
- 239000003446 ligand Substances 0.000 claims description 73
- 239000012453 solvate Substances 0.000 claims description 73
- 239000000651 prodrug Substances 0.000 claims description 59
- 229940002612 prodrug Drugs 0.000 claims description 59
- 125000000738 acetamido group Chemical group [H]C([H])([H])C(=O)N([H])[*] 0.000 claims description 51
- 239000012634 fragment Substances 0.000 claims description 48
- 125000000217 alkyl group Chemical group 0.000 claims description 45
- 125000000623 heterocyclic group Chemical group 0.000 claims description 36
- 125000000753 cycloalkyl group Chemical group 0.000 claims description 35
- 125000004435 hydrogen atom Chemical class [H]* 0.000 claims description 34
- 125000001301 ethoxy group Chemical group [H]C([H])([H])C([H])([H])O* 0.000 claims description 29
- 125000006273 (C1-C3) alkyl group Chemical group 0.000 claims description 28
- 125000000113 cyclohexyl group Chemical group [H]C1([H])C([H])([H])C([H])([H])C([H])(*)C([H])([H])C1([H])[H] 0.000 claims description 25
- 125000000547 substituted alkyl group Chemical group 0.000 claims description 22
- 150000002367 halogens Chemical class 0.000 claims description 21
- 125000004429 atom Chemical group 0.000 claims description 20
- 125000004093 cyano group Chemical group *C#N 0.000 claims description 18
- 125000001424 substituent group Chemical group 0.000 claims description 18
- 206010028980 Neoplasm Diseases 0.000 claims description 17
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 16
- 201000011510 cancer Diseases 0.000 claims description 16
- 230000007115 recruitment Effects 0.000 claims description 16
- 125000004214 1-pyrrolidinyl group Chemical group [H]C1([H])N(*)C([H])([H])C([H])([H])C1([H])[H] 0.000 claims description 15
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 claims description 15
- 125000004169 (C1-C6) alkyl group Chemical group 0.000 claims description 14
- 125000002572 propoxy group Chemical group [*]OC([H])([H])C(C([H])([H])[H])([H])[H] 0.000 claims description 14
- 239000008194 pharmaceutical composition Substances 0.000 claims description 12
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 10
- 125000004482 piperidin-4-yl group Chemical group N1CCC(CC1)* 0.000 claims description 9
- 239000002202 Polyethylene glycol Substances 0.000 claims description 8
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 8
- 229920001223 polyethylene glycol Polymers 0.000 claims description 8
- 125000002993 cycloalkylene group Chemical group 0.000 claims description 7
- 125000001147 pentyl group Chemical group C(CCCC)* 0.000 claims description 7
- 241001465754 Metazoa Species 0.000 claims description 6
- 230000001613 neoplastic effect Effects 0.000 claims description 6
- 125000006274 (C1-C3)alkoxy group Chemical group 0.000 claims description 5
- 239000003814 drug Substances 0.000 claims description 5
- 125000004943 pyrimidin-6-yl group Chemical group N1=CN=CC=C1* 0.000 claims description 5
- 125000004051 hexyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])* 0.000 claims description 4
- 125000001995 cyclobutyl group Chemical group [H]C1([H])C([H])([H])C([H])(*)C1([H])[H] 0.000 claims description 3
- 238000004519 manufacturing process Methods 0.000 claims description 3
- 125000004115 pentoxy group Chemical group [*]OC([H])([H])C([H])([H])C([H])([H])C(C([H])([H])[H])([H])[H] 0.000 claims description 3
- IVXQBCUBSIPQGU-UHFFFAOYSA-N piperazine-1-carboxamide Chemical compound NC(=O)N1CCNCC1 IVXQBCUBSIPQGU-UHFFFAOYSA-N 0.000 claims description 3
- 125000000592 heterocycloalkyl group Chemical group 0.000 claims description 2
- 125000004566 azetidin-1-yl group Chemical group N1(CCC1)* 0.000 claims 3
- DPBWFNDFMCCGGJ-UHFFFAOYSA-N 4-Piperidine carboxamide Chemical compound NC(=O)C1CCNCC1 DPBWFNDFMCCGGJ-UHFFFAOYSA-N 0.000 claims 1
- 125000004567 azetidin-3-yl group Chemical group N1CC(C1)* 0.000 claims 1
- 125000000484 butyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims 1
- QXEIHYDRWFWNLE-UHFFFAOYSA-N n-[[4-(4-methyl-1,3-thiazol-5-yl)phenyl]methyl]pyrrolidine-2-carboxamide Chemical compound N1=CSC(C=2C=CC(CNC(=O)C3NCCC3)=CC=2)=C1C QXEIHYDRWFWNLE-UHFFFAOYSA-N 0.000 claims 1
- 101000621390 Homo sapiens Wee1-like protein kinase Proteins 0.000 abstract description 37
- 102100023037 Wee1-like protein kinase Human genes 0.000 abstract description 35
- 230000004063 proteosomal degradation Effects 0.000 abstract description 4
- 230000034512 ubiquitination Effects 0.000 abstract description 3
- 238000010798 ubiquitination Methods 0.000 abstract description 3
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- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 358
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- 239000000243 solution Substances 0.000 description 211
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 199
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- 238000006243 chemical reaction Methods 0.000 description 162
- 239000012044 organic layer Substances 0.000 description 143
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 141
- 125000000999 tert-butyl group Chemical group [H]C([H])([H])C(*)(C([H])([H])[H])C([H])([H])[H] 0.000 description 139
- 239000012071 phase Substances 0.000 description 113
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- 125000005843 halogen group Chemical group 0.000 description 101
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- 125000002023 trifluoromethyl group Chemical group FC(F)(F)* 0.000 description 91
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 90
- IAZDPXIOMUYVGZ-WFGJKAKNSA-N Dimethyl sulfoxide Chemical compound [2H]C([2H])([2H])S(=O)C([2H])([2H])[2H] IAZDPXIOMUYVGZ-WFGJKAKNSA-N 0.000 description 84
- 235000019439 ethyl acetate Nutrition 0.000 description 82
- 239000012267 brine Substances 0.000 description 76
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical compound O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 76
- 238000010898 silica gel chromatography Methods 0.000 description 72
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 65
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 65
- 125000002947 alkylene group Chemical group 0.000 description 53
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 52
- 125000004432 carbon atom Chemical group C* 0.000 description 49
- 230000002829 reductive effect Effects 0.000 description 49
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 44
- 102000053200 Von Hippel-Lindau Tumor Suppressor Human genes 0.000 description 44
- 108700031765 Von Hippel-Lindau Tumor Suppressor Proteins 0.000 description 44
- 238000005160 1H NMR spectroscopy Methods 0.000 description 43
- 238000002953 preparative HPLC Methods 0.000 description 42
- 150000002431 hydrogen Chemical class 0.000 description 41
- MPKYJUACTUTJRY-UHFFFAOYSA-N 1-[6-(2-hydroxypropan-2-yl)pyridin-2-yl]-6-(4-piperazin-1-ylanilino)-2-prop-2-enylpyrazolo[3,4-d]pyrimidin-3-one Chemical compound CC(C)(O)C1=CC=CC(N2C3=NC(NC=4C=CC(=CC=4)N4CCNCC4)=NC=C3C(=O)N2CC=C)=N1 MPKYJUACTUTJRY-UHFFFAOYSA-N 0.000 description 40
- 239000007821 HATU Substances 0.000 description 40
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 40
- RAXXELZNTBOGNW-UHFFFAOYSA-N imidazole Natural products C1=CNC=N1 RAXXELZNTBOGNW-UHFFFAOYSA-N 0.000 description 39
- 125000001072 heteroaryl group Chemical group 0.000 description 37
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- 150000003254 radicals Chemical class 0.000 description 35
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- 229910000027 potassium carbonate Inorganic materials 0.000 description 26
- 239000007832 Na2SO4 Substances 0.000 description 24
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- RIOQSEWOXXDEQQ-UHFFFAOYSA-N triphenylphosphine Chemical compound C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1 RIOQSEWOXXDEQQ-UHFFFAOYSA-N 0.000 description 24
- 125000004419 alkynylene group Chemical group 0.000 description 23
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 23
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- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 19
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- PLDXRPSSERMPSV-UHFFFAOYSA-N spiro[3.6]decane Chemical compound C1CCC21CCCCCC2 PLDXRPSSERMPSV-UHFFFAOYSA-N 0.000 description 1
- PHICBFWUYUCFKS-UHFFFAOYSA-N spiro[4.4]nonane Chemical compound C1CCCC21CCCC2 PHICBFWUYUCFKS-UHFFFAOYSA-N 0.000 description 1
- NWTSRIZEBKLUFM-UHFFFAOYSA-N spiro[4.6]undecane Chemical compound C1CCCC21CCCCCC2 NWTSRIZEBKLUFM-UHFFFAOYSA-N 0.000 description 1
- NECLQTPQJZSWOE-UHFFFAOYSA-N spiro[5.5]undecane Chemical compound C1CCCCC21CCCCC2 NECLQTPQJZSWOE-UHFFFAOYSA-N 0.000 description 1
- ZDNANLAHKKNCTC-UHFFFAOYSA-N spiro[5.6]dodecane Chemical compound C1CCCCC21CCCCCC2 ZDNANLAHKKNCTC-UHFFFAOYSA-N 0.000 description 1
- NGGUAMSYAYRZGK-UHFFFAOYSA-N spiro[6.6]tridecane Chemical compound C1CCCCCC21CCCCCC2 NGGUAMSYAYRZGK-UHFFFAOYSA-N 0.000 description 1
- CTDQAGUNKPRERK-UHFFFAOYSA-N spirodecane Chemical compound C1CCCC21CCCCC2 CTDQAGUNKPRERK-UHFFFAOYSA-N 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- LSNNMFCWUKXFEE-UHFFFAOYSA-L sulfite Chemical class [O-]S([O-])=O LSNNMFCWUKXFEE-UHFFFAOYSA-L 0.000 description 1
- 150000003467 sulfuric acid derivatives Chemical class 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000001308 synthesis method Methods 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000002906 tartaric acid Nutrition 0.000 description 1
- 239000011975 tartaric acid Substances 0.000 description 1
- 150000003892 tartrate salts Chemical class 0.000 description 1
- 238000003419 tautomerization reaction Methods 0.000 description 1
- QQLXPUHVMNANLN-UHFFFAOYSA-N tert-butyl 10-bromodecanoate Chemical compound CC(C)(C)OC(=O)CCCCCCCCCBr QQLXPUHVMNANLN-UHFFFAOYSA-N 0.000 description 1
- ISIJQEHRDSCQIU-UHFFFAOYSA-N tert-butyl 2,7-diazaspiro[4.5]decane-7-carboxylate Chemical compound C1N(C(=O)OC(C)(C)C)CCCC11CNCC1 ISIJQEHRDSCQIU-UHFFFAOYSA-N 0.000 description 1
- RLUMXXFNHHSSIF-UHFFFAOYSA-N tert-butyl 2-(4-iodobutoxy)acetate Chemical compound CC(C)(C)OC(=O)COCCCCI RLUMXXFNHHSSIF-UHFFFAOYSA-N 0.000 description 1
- LEOVLGAAQMGPBM-UHFFFAOYSA-N tert-butyl 2-[2-(2-bromoethoxy)ethoxy]acetate Chemical compound CC(C)(C)OC(=O)COCCOCCBr LEOVLGAAQMGPBM-UHFFFAOYSA-N 0.000 description 1
- NFYYXORESKNQKM-UHFFFAOYSA-N tert-butyl 3-(3-hydroxypropoxy)propanoate Chemical compound CC(C)(C)OC(=O)CCOCCCO NFYYXORESKNQKM-UHFFFAOYSA-N 0.000 description 1
- UOYAYHGUAATLSG-UHFFFAOYSA-N tert-butyl 3-[2-(2-bromoethoxy)ethoxy]propanoate Chemical compound CC(C)(C)OC(=O)CCOCCOCCBr UOYAYHGUAATLSG-UHFFFAOYSA-N 0.000 description 1
- RXFHRKPNLPBDGE-UHFFFAOYSA-N tert-butyl 4-(4-aminophenyl)piperazine-1-carboxylate Chemical compound C1CN(C(=O)OC(C)(C)C)CCN1C1=CC=C(N)C=C1 RXFHRKPNLPBDGE-UHFFFAOYSA-N 0.000 description 1
- UYDIIUHJHUZDME-UHFFFAOYSA-N tert-butyl 5-bromopentanoate Chemical compound CC(C)(C)OC(=O)CCCCBr UYDIIUHJHUZDME-UHFFFAOYSA-N 0.000 description 1
- ZTWGIZOCAXOUOE-UHFFFAOYSA-N tert-butyl n-amino-n-prop-2-enylcarbamate Chemical compound CC(C)(C)OC(=O)N(N)CC=C ZTWGIZOCAXOUOE-UHFFFAOYSA-N 0.000 description 1
- 150000003512 tertiary amines Chemical class 0.000 description 1
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 1
- WHRNULOCNSKMGB-UHFFFAOYSA-N tetrahydrofuran thf Chemical compound C1CCOC1.C1CCOC1 WHRNULOCNSKMGB-UHFFFAOYSA-N 0.000 description 1
- 125000003718 tetrahydrofuranyl group Chemical group 0.000 description 1
- 125000001412 tetrahydropyranyl group Chemical group 0.000 description 1
- 125000000147 tetrahydroquinolinyl group Chemical group N1(CCCC2=CC=CC=C12)* 0.000 description 1
- CZDYPVPMEAXLPK-UHFFFAOYSA-N tetramethylsilane Chemical compound C[Si](C)(C)C CZDYPVPMEAXLPK-UHFFFAOYSA-N 0.000 description 1
- 125000003831 tetrazolyl group Chemical group 0.000 description 1
- WROMPOXWARCANT-UHFFFAOYSA-N tfa trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F.OC(=O)C(F)(F)F WROMPOXWARCANT-UHFFFAOYSA-N 0.000 description 1
- 229960004559 theobromine Drugs 0.000 description 1
- 125000001113 thiadiazolyl group Chemical group 0.000 description 1
- 125000001984 thiazolidinyl group Chemical group 0.000 description 1
- 125000000335 thiazolyl group Chemical group 0.000 description 1
- 125000005985 thienyl[1,3]dithianyl group Chemical group 0.000 description 1
- LBLYYCQCTBFVLH-UHFFFAOYSA-M toluenesulfonate group Chemical group C=1(C(=CC=CC1)S(=O)(=O)[O-])C LBLYYCQCTBFVLH-UHFFFAOYSA-M 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 238000000844 transformation Methods 0.000 description 1
- 230000001052 transient effect Effects 0.000 description 1
- 230000014616 translation Effects 0.000 description 1
- 238000011269 treatment regimen Methods 0.000 description 1
- 125000004306 triazinyl group Chemical group 0.000 description 1
- 125000001425 triazolyl group Chemical group 0.000 description 1
- INQOMBQAUSQDDS-FIBGUPNXSA-N trideuterio(iodo)methane Chemical compound [2H]C([2H])([2H])I INQOMBQAUSQDDS-FIBGUPNXSA-N 0.000 description 1
- YFTHZRPMJXBUME-UHFFFAOYSA-N tripropylamine Chemical compound CCCN(CCC)CCC YFTHZRPMJXBUME-UHFFFAOYSA-N 0.000 description 1
- 125000005455 trithianyl group Chemical group 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 229950009811 ubenimex Drugs 0.000 description 1
- 229920002554 vinyl polymer Polymers 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
- C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
- C07D487/04—Ortho-condensed systems
Definitions
- WEE1 is a kinase belonging to the Ser/Thr family of protein kinases and is a key regulator of cell cycle progression. Pharmacological intervention in the cell cycle is strategy for the treatment of cancer and other proliferative disease.
- the present disclosure provides compounds, compositions and methods for the reduction or elimination of WEE1 activity via the targeting of WEE1 protein in a cell through ubiquitination and proteasomal degradation.
- One embodiment provides a compound, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I) :
- R 1 , R 2 , R 3 , and R 4 are each independently selected from hydrogen, optionally substituted straight or branched C 1-6 alkyl, optionally substituted C 1-3 alkoxy, halogen, amino, hydroxyl, and cyano; or
- R 1 and R 2 , R 2 and R 3 , or R 3 and R 4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocycloalkyl ring;
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*;
- Ak 1 is selected from - (CR 5 R 6 ) k -;
- Ak 2 is selected from - (CR 5 R 6 ) m -;
- Ak 3 is selected from - (CR 5 R 6 ) n -;
- each of k, m, and n is selected from 0 to 10
- R 5 and R 6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl;
- Z 1 and Z 2 are each independently selected from bond, -O-, -NR 7 -, -S-, heterocyclene, and cycloalkylene;
- R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced;
- L is a polyethylene glycol containing moiety
- U is a VHL ligand.
- One embodiment provides a pharmaceutical composition
- a pharmaceutical composition comprising a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, and at least one pharmaceutically acceptable excipient.
- One embodiment provides a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof for use in a method of treatment of a human or animal.
- One embodiment provides a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, for use in a method of treatment of cancer or neoplastic disease.
- One embodiment provides the use of a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, in the manufacture of a medicament for the treatment of cancer or neoplastic disease.
- One embodiment provides a method of treating a disease or disorder in a patient in need thereof comprising administering to the patient a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof. Another embodiment provides the method wherein the disease or disorder is cancer.
- One embodiment provides a method of treating cancer in a patient in need thereof comprising administering to the patient a pharmaceutical composition comprising a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, and a pharmaceutically acceptable excipient.
- Amino refers to the —NH 2 radical.
- Niro refers to the -NO 2 radical.
- Oxa refers to the -O-radical.
- Alkyl refers to a straight or branched hydrocarbon chain radical consisting solely of carbon and hydrogen atoms, containing no unsaturation, having from one to fifteen carbon atoms (e.g., C 1 -C 15 alkyl) .
- an alkyl comprises one to thirteen carbon atoms (e.g., C 1 -C 13 alkyl) .
- an alkyl comprises one to eight carbon atoms (e.g., C 1 -C 8 alkyl) .
- an alkyl comprises one to five carbon atoms (e.g., C 1 -C 5 alkyl) .
- an alkyl comprises one to four carbon atoms (e.g., C 1 -C 4 alkyl) . In other embodiments, an alkyl comprises one to three carbon atoms (e.g., C 1 -C 3 alkyl) . In other embodiments, an alkyl comprises one to two carbon atoms (e.g., C 1 -C 2 alkyl) . In other embodiments, an alkyl comprises one carbon atom (e.g., C 1 alkyl) . In other embodiments, an alkyl comprises five to fifteen carbon atoms (e.g., C 5 -C 15 alkyl) .
- an alkyl comprises five to eight carbon atoms (e.g., C 5 -C 8 alkyl) . In other embodiments, an alkyl comprises two to five carbon atoms (e.g., C 2 -C 5 alkyl) . In other embodiments, an alkyl comprises three to five carbon atoms (e.g., C 3 -C 5 alkyl) .
- the alkyl group is selected from methyl, ethyl, 1-propyl (n-propyl) , 1-methylethyl (iso-propyl) , 1-butyl (n-butyl) , 1-methylpropyl (sec-butyl) , 2-methylpropyl (iso-butyl) , 1, 1-dimethylethyl (tert-butyl) , and 1-pentyl (n-pentyl) .
- the alkyl is attached to the rest of the molecule by a single bond.
- an alkyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where t
- Alkoxy refers to a radical bonded through an oxygen atom of the formula –O-alkyl, where alkyl is an alkyl chain as defined above.
- Alkenyl refers to a straight or branched hydrocarbon chain radical group consisting solely of carbon and hydrogen atoms, containing at least one carbon-carbon double bond, and having from two to twelve carbon atoms. In certain embodiments, an alkenyl comprises two to eight carbon atoms. In other embodiments, an alkenyl comprises two to four carbon atoms. The alkenyl is attached to the rest of the molecule by a single bond, for example, ethenyl (i.e., vinyl) , prop-1-enyl (i.e., allyl) , but-1-enyl, pent-1-enyl, penta-1, 4-dienyl, and the like.
- ethenyl i.e., vinyl
- prop-1-enyl i.e., allyl
- pent-1-enyl penta-1, 4-dienyl, and the like.
- an alkenyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where
- Alkynyl refers to a straight or branched hydrocarbon chain radical group consisting solely of carbon and hydrogen atoms, containing at least one carbon-carbon triple bond, having from two to twelve carbon atoms.
- an alkynyl comprises two to eight carbon atoms.
- an alkynyl comprises two to six carbon atoms.
- an alkynyl comprises two to four carbon atoms.
- the alkynyl is attached to the rest of the molecule by a single bond, for example, ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.
- an alkynyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where
- Alkylene or "alkylene chain” refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing no unsaturation and having from one to twelve carbon atoms, for example, methylene, ethylene, propylene, n-butylene, and the like.
- the alkylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond.
- the points of attachment of the alkylene chain to the rest of the molecule and to the radical group are through one carbon in the alkylene chain or through any two carbons within the chain.
- an alkylene comprises one to eight carbon atoms (e.g., C 1 -C 8 alkylene) . In other embodiments, an alkylene comprises one to five carbon atoms (e.g., C 1 -C 5 alkylene) . In other embodiments, an alkylene comprises one to four carbon atoms (e.g., C 1 -C 4 alkylene) . In other embodiments, an alkylene comprises one to three carbon atoms (e.g., C 1 -C 3 alkylene) . In other embodiments, an alkylene comprises one to two carbon atoms (e.g., C 1 -C 2 alkylene) .
- an alkylene comprises one carbon atom (e.g., C 1 alkylene) . In other embodiments, an alkylene comprises five to eight carbon atoms (e.g., C 5 -C 8 alkylene) . In other embodiments, an alkylene comprises two to five carbon atoms (e.g., C 2 -C 5 alkylene) . In other embodiments, an alkylene comprises three to five carbon atoms (e.g., C 3 -C 5 alkylene) .
- an alkylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where t
- Alkenylene or “alkenylene chain” refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing at least one carbon-carbon double bond, and having from two to twelve carbon atoms.
- the alkenylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond.
- an alkenylene comprises two to eight carbon atoms (e.g., C 2 -C 8 alkenylene) .
- an alkenylene comprises two to five carbon atoms (e.g., C 2 -C 5 alkenylene) .
- an alkenylene comprises two to four carbon atoms (e.g., C 2 -C 4 alkenylene) . In other embodiments, an alkenylene comprises two to three carbon atoms (e.g., C 2 -C 3 alkenylene) . In other embodiments, an alkenylene comprises two carbon atoms (e.g., C 2 alkenylene) . In other embodiments, an alkenylene comprises five to eight carbon atoms (e.g., C 5 -C 8 alkenylene) . In other embodiments, an alkenylene comprises three to five carbon atoms (e.g., C 3 -C 5 alkenylene) .
- an alkenylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where
- Alkynylene or “alkynylene chain” refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing at least one carbon-carbon triple bond, and having from two to twelve carbon atoms.
- the alkynylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond.
- an alkynylene comprises two to eight carbon atoms (e.g., C 2 -C 8 alkynylene) .
- an alkynylene comprises two to five carbon atoms (e.g., C 2 -C 5 alkynylene) .
- an alkynylene comprises two to four carbon atoms (e.g., C 2 -C 4 alkynylene) . In other embodiments, an alkynylene comprises two to three carbon atoms (e.g., C 2 -C 3 alkynylene) . In other embodiments, an alkynylene comprises two carbon atoms (e.g., C 2 alkynylene) . In other embodiments, an alkynylene comprises five to eight carbon atoms (e.g., C 5 -C 8 alkynylene) . In other embodiments, an alkynylene comprises three to five carbon atoms (e.g., C 3 -C 5 alkynylene) .
- an alkynylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -OR a , -SR a , -OC (O) -R a , -N (R a ) 2 , -C (O) R a , -C (O) OR a , -C (O) N (R a ) 2 , -N (R a ) C (O) OR a , -OC (O) -N (R a ) 2 , -N (R a ) C (O) R a , -N (R a ) S (O) t R a (where t is 1 or 2) , -S (O) t OR a (where t is 1 or 2) , -S (O) t OR a (where
- Aryl refers to a radical derived from an aromatic monocyclic or multicyclic hydrocarbon ring system by removing a hydrogen atom from a ring carbon atom.
- the aromatic monocyclic or multicyclic hydrocarbon ring system contains only hydrogen and carbon from five to eighteen carbon atoms, where at least one of the rings in the ring system is fully unsaturated, i.e., it contains a cyclic, delocalized (4n+2) ⁇ –electron system in accordance with the Hückel theory.
- the ring system from which aryl groups are derived include, but are not limited to, groups such as benzene, fluorene, indane, indene, tetralin and naphthalene.
- aryl or the prefix “ar-” (such as in “aralkyl” ) is meant to include aryl radicals optionally substituted by one or more substituents independently selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -R b -OR a , -R b -OC (O) -R a , -R b -OC (O) -OR a , -R b -OC (O)
- Alkyl refers to a radical of the formula -R c -aryl where R c is an alkylene chain as defined above, for example, methylene, ethylene, and the like.
- the alkylene chain part of the aralkyl radical is optionally substituted as described above for an alkylene chain.
- the aryl part of the aralkyl radical is optionally substituted as described above for an aryl group.
- alkenyl refers to a radical of the formula –R d -aryl where R d is an alkenylene chain as defined above.
- the aryl part of the aralkenyl radical is optionally substituted as described above for an aryl group.
- the alkenylene chain part of the aralkenyl radical is optionally substituted as defined above for an alkenylene group.
- Alkynyl refers to a radical of the formula -R e -aryl, where R e is an alkynylene chain as defined above.
- the aryl part of the aralkynyl radical is optionally substituted as described above for an aryl group.
- the alkynylene chain part of the aralkynyl radical is optionally substituted as defined above for an alkynylene chain.
- Alkoxy refers to a radical bonded through an oxygen atom of the formula -O-R c -aryl where R c is an alkylene chain as defined above, for example, methylene, ethylene, and the like.
- R c is an alkylene chain as defined above, for example, methylene, ethylene, and the like.
- the alkylene chain part of the aralkyl radical is optionally substituted as described above for an alkylene chain.
- the aryl part of the aralkyl radical is optionally substituted as described above for an aryl group.
- Carbocyclyl refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms, which includes fused, bridged, or spirocyclic ring systems, having from three to fifteen carbon atoms.
- a carbocyclyl comprises three to ten carbon atoms.
- a carbocyclyl comprises five to seven carbon atoms.
- the carbocyclyl is attached to the rest of the molecule by a single bond. Carbocyclyl is saturated (i.e., containing single C-C bonds only) or unsaturated (i.e., containing one or more double bonds or triple bonds) .
- a fully saturated carbocyclyl radical is also referred to as "cycloalkyl.
- monocyclic cycloalkyls include, e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
- An unsaturated carbocyclyl is also referred to as “cycloalkenyl.
- Examples of monocyclic cycloalkenyls include, e.g., cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl.
- Polycyclic carbocyclyl radicals include, for example, adamantyl, norbornyl (i.e., bicyclo [2.2.1] heptanyl) , norbornenyl, decalinyl, 7, 7-dimethyl-bicyclo [2.2.1] heptanyl, and the like.
- Spirocyclic carbocyclyl or cycloalkyl radicals include, for example, spiro [2.2] pentane, spiro [2.3] hexane, spiro [2.4] heptane, spiro [2.5] octane, spiro [2.6] nonane, spiro [3.3] heptane, spiro [3.4] octane, spiro [3.5] nonane, spiro [3.6] decane, spiro [4.4] nonane, spiro [4.5] decane, spiro [4.6] undecane, spiro [5.5] undecane, spiro [5.6] dodecane, spiro [6.6] tridecane, and the like.
- carbocyclyl is meant to include carbocyclyl radicals that are optionally substituted by one or more substituents independently selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -R b -OR a , -R b -OC (O) -R a , -R b -OC (O) -OR a , -R b -OC (O) -OR a , -R b -OC
- Cycloalkylene refers to a divalent carbocyclyl or cycloalkyl linking the rest of the molecule to a radical group.
- Carbocyclylalkyl refers to a radical of the formula –R c -carbocyclyl where R c is an alkylene chain as defined above. The alkylene chain and the carbocyclyl radical is optionally substituted as defined above.
- Carbocyclylalkynyl refers to a radical of the formula –R c -carbocyclyl where R c is an alkynylene chain as defined above. The alkynylene chain and the carbocyclyl radical is optionally substituted as defined above.
- Carbocyclylalkoxy refers to a radical bonded through an oxygen atom of the formula –O-R c -carbocyclyl where R c is an alkylene chain as defined above.
- R c is an alkylene chain as defined above.
- the alkylene chain and the carbocyclyl radical is optionally substituted as defined above.
- carboxylic acid bioisostere refers to a functional group or moiety that exhibits similar physical, biological and/or chemical properties as a carboxylic acid moiety.
- Examples of carboxylic acid bioisosteres include, but are not limited to, and the like.
- Halo or halogen refers to bromo, chloro, fluoro or iodo substituents.
- Fluoroalkyl refers to an alkyl radical, as defined above, that is substituted by one or more fluoro radicals, as defined above, for example, trifluoromethyl, difluoromethyl, fluoromethyl, 2, 2, 2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, and the like.
- the alkyl part of the fluoroalkyl radical is optionally substituted as defined above for an alkyl group.
- Heterocyclyl refers to a stable 3-to 18-membered non-aromatic ring radical that comprises two to twelve carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur. Unless stated otherwise specifically in the specification, the heterocyclyl radical is a monocyclic, bicyclic, tricyclic, tetracyclic, or spirocyclic ring system, which optionally includes fused or bridged ring systems. The heteroatoms in the heterocyclyl radical are optionally oxidized. One or more nitrogen atoms, if present, are optionally quaternized. The heterocyclyl radical is partially or fully saturated. The heterocyclyl is attached to the rest of the molecule through any atom of the ring (s) .
- heterocyclyl radicals include, but are not limited to, dioxolanyl, thienyl [1, 3] dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorph
- heterocyclyl is meant to include heterocyclyl radicals as defined above that are optionally substituted by one or more substituents selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -R b -OR a , -R b -OC (O) -R a , -R b -OC (O) -OR a , -R b -OC (O)
- Heterocyclene refers to a divalent heterocyclyl linking the rest of the molecule to a radical group.
- N-heterocyclyl or “N-attached heterocyclyl” refers to a heterocyclyl radical as defined above containing at least one nitrogen and where the point of attachment of the heterocyclyl radical to the rest of the molecule is through a nitrogen atom in the heterocyclyl radical.
- An N-heterocyclyl radical is optionally substituted as described above for heterocyclyl radicals. Examples of such N-heterocyclyl radicals include, but are not limited to, 1-morpholinyl, 1-piperidinyl, 1-piperazinyl, 1-pyrrolidinyl, pyrazolidinyl, imidazolinyl, and imidazolidinyl.
- C-heterocyclyl or “C-attached heterocyclyl” refers to a heterocyclyl radical as defined above containing at least one heteroatom and where the point of attachment of the heterocyclyl radical to the rest of the molecule is through a carbon atom in the heterocyclyl radical.
- a C-heterocyclyl radical is optionally substituted as described above for heterocyclyl radicals. Examples of such C-heterocyclyl radicals include, but are not limited to, 2-morpholinyl, 2-or 3-or 4-piperidinyl, 2-piperazinyl, 2-or 3-pyrrolidinyl, and the like.
- Heterocyclylalkyl refers to a radical of the formula –R c -heterocyclyl where R c is an alkylene chain as defined above. If the heterocyclyl is a nitrogen-containing heterocyclyl, the heterocyclyl is optionally attached to the alkyl radical at the nitrogen atom.
- the alkylene chain of the heterocyclylalkyl radical is optionally substituted as defined above for an alkylene chain.
- the heterocyclyl part of the heterocyclylalkyl radical is optionally substituted as defined above for a heterocyclyl group.
- Heterocyclylalkoxy refers to a radical bonded through an oxygen atom of the formula –O-R c -heterocyclyl where R c is an alkylene chain as defined above. If the heterocyclyl is a nitrogen-containing heterocyclyl, the heterocyclyl is optionally attached to the alkyl radical at the nitrogen atom.
- the alkylene chain of the heterocyclylalkoxy radical is optionally substituted as defined above for an alkylene chain.
- the heterocyclyl part of the heterocyclylalkoxy radical is optionally substituted as defined above for a heterocyclyl group.
- Heteroaryl refers to a radical derived from a 3-to 18-membered aromatic ring radical that comprises two to seventeen carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur.
- the heteroaryl radical is a monocyclic, bicyclic, tricyclic or tetracyclic ring system, wherein at least one of the rings in the ring system is fully unsaturated, i.e., it contains a cyclic, delocalized (4n+2) ⁇ –electron system in accordance with the Hückel theory.
- Heteroaryl includes fused or bridged ring systems.
- the heteroatom (s) in the heteroaryl radical is optionally oxidized.
- heteroaryl is attached to the rest of the molecule through any atom of the ring (s) .
- heteroaryls include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzindolyl, 1, 3-benzodioxolyl, benzofuranyl, benzooxazolyl, benzo [d] thiazolyl, benzothiadiazolyl, benzo [b] [1, 4] dioxepinyl, benzo [b] [1, 4] oxazinyl, 1, 4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzo
- heteroaryl is meant to include heteroaryl radicals as defined above which are optionally substituted by one or more substituents selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, haloalkenyl, haloalkynyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -R b -OR a , -R b -OC (O) -R a , -R b -
- N-heteroaryl refers to a heteroaryl radical as defined above containing at least one nitrogen and where the point of attachment of the heteroaryl radical to the rest of the molecule is through a nitrogen atom in the heteroaryl radical.
- An N-heteroaryl radical is optionally substituted as described above for heteroaryl radicals.
- C-heteroaryl refers to a heteroaryl radical as defined above and where the point of attachment of the heteroaryl radical to the rest of the molecule is through a carbon atom in the heteroaryl radical.
- a C-heteroaryl radical is optionally substituted as described above for heteroaryl radicals.
- Heteroarylalkyl refers to a radical of the formula –R c -heteroaryl, where R c is an alkylene chain as defined above. If the heteroaryl is a nitrogen-containing heteroaryl, the heteroaryl is optionally attached to the alkyl radical at the nitrogen atom.
- the alkylene chain of the heteroarylalkyl radical is optionally substituted as defined above for an alkylene chain.
- the heteroaryl part of the heteroarylalkyl radical is optionally substituted as defined above for a heteroaryl group.
- Heteroarylalkoxy refers to a radical bonded through an oxygen atom of the formula –O-R c -heteroaryl, where R c is an alkylene chain as defined above. If the heteroaryl is a nitrogen-containing heteroaryl, the heteroaryl is optionally attached to the alkyl radical at the nitrogen atom.
- the alkylene chain of the heteroarylalkoxy radical is optionally substituted as defined above for an alkylene chain.
- the heteroaryl part of the heteroarylalkoxy radical is optionally substituted as defined above for a heteroaryl group.
- the compounds disclosed herein in some embodiments, contain one or more asymmetric centers and thus give rise to enantiomers, diastereomers, and other stereoisomeric forms that are defined, in terms of absolute stereochemistry, as (R) -or (S) -. Unless stated otherwise, it is intended that all stereoisomeric forms of the compounds disclosed herein are contemplated by this disclosure. When the compounds described herein contain alkene double bonds, and unless specified otherwise, it is intended that this disclosure includes both E and Z geometric isomers (e.g., cis or trans. ) Likewise, all possible isomers, as well as their racemic and optically pure forms, and all tautomeric forms are also intended to be included.
- geometric isomer refers to E or Z geometric isomers (e.g., cis or trans) of an alkene double bond.
- positional isomer refers to structural isomers around a central ring, such as ortho-, meta-, and para-isomers around a benzene ring.
- a "tautomer” refers to a molecule wherein a proton shift from one atom of a molecule to another atom of the same molecule is possible.
- the compounds disclosed herein are used in different enriched isotopic forms, e.g., enriched in the content of 2 H, 3 H, 11 C, 13 C and/or 14 C.
- the compound is deuterated in at least one position.
- deuterated forms can be made by the procedure described in U.S. Patent Nos. 5,846,514 and 6,334,997. As described in U.S. Patent Nos. 5,846,514 and 6,334,997, deuteration can improve the metabolic stability and or efficacy, thus increasing the duration of action of drugs.
- structures depicted herein are intended to include compounds which differ only in the presence of one or more isotopically enriched atoms.
- compounds having the present structures except for the replacement of a hydrogen by a deuterium or tritium, or the replacement of a carbon by 13 C-or 14 C-enriched carbon are within the scope of the present disclosure.
- the compounds of the present disclosure optionally contain unnatural proportions of atomic isotopes at one or more atoms that constitute such compounds.
- the compounds may be labeled with isotopes, such as for example, deuterium ( 2 H) , tritium ( 3 H) , iodine-125 ( 125 I) or carbon-14 ( 14 C) .
- isotopes such as for example, deuterium ( 2 H) , tritium ( 3 H) , iodine-125 ( 125 I) or carbon-14 ( 14 C) .
- Isotopic substitution with 2 H, 11 C, 13 C, 14 C, 15 C, 12 N, 13 N, 15 N, 16 N, 16 O, 17 O, 14 F, 15 F, 16 F, 17 F, 18 F, 33 S, 34 S, 35 S, 36 S, 35 Cl, 37 Cl, 79 Br, 81 Br, 125 I are all contemplated.
- isotopic substitution with 18 F is contemplated. All isotopic variations
- the compounds disclosed herein have some or all of the 1 H atoms replaced with 2 H atoms.
- the methods of synthesis for deuterium-containing compounds are known in the art and include, by way of non-limiting example only, the following synthetic methods.
- Deuterium substituted compounds are synthesized using various methods such as described in: Dean, Dennis C.; Editor. Recent Advances in the Synthesis and Applications of Radiolabeled Compounds for Drug Discovery and Development. [Curr., Pharm. Des., 2000; 6 (10) ] 2000, 110 pp; George W.; Varma, Rajender S. The Synthesis of Radiolabeled Compounds via Organometallic Intermediates, Tetrahedron, 1989, 45 (21) , 6601-21; and Evans, E. Anthony. Synthesis of radiolabeled compounds, J. Radioanal. Chem., 1981, 64 (1-2) , 9-32.
- Deuterated starting materials are readily available and are subjected to the synthetic methods described herein to provide for the synthesis of deuterium-containing compounds.
- Large numbers of deuterium-containing reagents and building blocks are available commercially from chemical vendors, such as Aldrich Chemical Co.
- CD 3 I iodomethane-d 3
- LiAlD 4 lithium aluminum deuteride
- Deuterium gas and palladium catalyst are employed to reduce unsaturated carbon-carbon linkages and to perform a reductive substitution of aryl carbon-halogen bonds as illustrated, by way of example only, in the reaction schemes below.
- the compounds disclosed herein contain one deuterium atom. In another embodiment, the compounds disclosed herein contain two deuterium atoms. In another embodiment, the compounds disclosed herein contain three deuterium atoms. In another embodiment, the compounds disclosed herein contain four deuterium atoms. In another embodiment, the compounds disclosed herein contain five deuterium atoms. In another embodiment, the compounds disclosed herein contain six deuterium atoms. In another embodiment, the compounds disclosed herein contain more than six deuterium atoms. In another embodiment, the compound disclosed herein is fully substituted with deuterium atoms and contains no non-exchangeable 1 H hydrogen atoms. In one embodiment, the level of deuterium incorporation is determined by synthetic methods in which a deuterated synthetic building block is used as a starting material.
- “Pharmaceutically acceptable salt” includes both acid and base addition salts.
- a pharmaceutically acceptable salt of any one of the heteroaromatic WEE1 inhibitory compounds described herein is intended to encompass any and all pharmaceutically suitable salt forms.
- Preferred pharmaceutically acceptable salts of the compounds described herein are pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.
- “Pharmaceutically acceptable acid addition salt” refers to those salts which retain the biological effectiveness and properties of the free bases, which are not biologically or otherwise undesirable, and which are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid, hydrofluoric acid, phosphorous acid, and the like. Also included are salts that are formed with organic acids such as aliphatic mono-and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, alkanedioic acids, aromatic acids, aliphatic and. aromatic sulfonic acids, etc.
- acetic acid trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.
- Exemplary salts thus include sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, nitrates, phosphates, monohydrogenphosphates, dihydrogenphosphates, metaphosphates, pyrophosphates, chlorides, bromides, iodides, acetates, trifluoroacetates, propionates, caprylates, isobutyrates, oxalates, malonates, succinate suberates, sebacates, fumarates, maleates, mandelates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, phthalates, benzenesulfonates, toluenesulfonates, phenylacetates, citrates, lactates, malates, tartrates, methanesulfonates, and the like.
- salts of amino acids such as arginates, gluconates, and galacturonates (see, for example, Berge S.M. et al., “Pharmaceutical Salts, " Journal of Pharmaceutical Science, 66: 1-19 (1997) ) .
- Acid addition salts of basic compounds are, in some embodiments, prepared by contacting the free base forms with a sufficient amount of the desired acid to produce the salt according to methods and techniques with which a skilled artisan is familiar.
- “Pharmaceutically acceptable base addition salt” refers to those salts that retain the biological effectiveness and properties of the free acids, which are not biologically or otherwise undesirable. These salts are prepared from addition of an inorganic base or an organic base to the free acid. Pharmaceutically acceptable base addition salts are, in some embodiments, formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum salts and the like.
- Salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, for example, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, N, N-dibenzylethylenediamine, chloroprocaine, hydrabamine, choline, betaine, ethylenediamine, ethylenedianiline, N-methylglucamine, glucosamine, methylglucamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins and the like. See Berge et al
- solvates refers to a composition of matter that is the solvent addition form.
- solvates contain either stoichiometric or non-stoichiometric amounts of a solvent, and are formed during the process of making with pharmaceutically acceptable solvents such as water, ethanol, and the like. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of compounds described herein are conveniently prepared or formed during the processes described herein. The compounds provided herein optionally exist in either unsolvated as well as solvated forms.
- subject or “patient” encompasses mammals.
- mammals include, but are not limited to, any member of the Mammalian class: humans, non-human primates such as chimpanzees, and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, swine; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents, such as rats, mice and guinea pigs, and the like.
- the mammal is a human.
- Prodrug refers to a compound that undergoes biotransformation before exhibiting its pharmacological effects. Prodrugs can thus be viewed as drugs containing specialized protective groups used in a transient manner to alter pharmacological properties in the parent molecule.
- treatment or “treating, ” or “palliating” or “ameliorating” are used interchangeably. These terms refer to an approach for obtaining beneficial or desired results including but not limited to therapeutic benefit and/or a prophylactic benefit.
- therapeutic benefit is meant eradication or amelioration of the underlying disorder being treated.
- a therapeutic benefit is achieved with the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disorder such that an improvement is observed in the patient, notwithstanding that the patient is still afflicted with the underlying disorder.
- the compositions are, in some embodiments, administered to a patient at risk of developing a particular disease, or to a patient reporting one or more of the physiological symptoms of a disease, even though a diagnosis of this disease has not been made.
- WEE1 is a kinase belonging to the Ser/Thr family of protein kinases and is a key regulator of cell cycle progression.
- the WEE1 kinase is encoded by the WEE1 gene.
- WEE1 is an important regulator of the G2/M cell cycle checkpoint.
- WEE1 mediates cell-cycle arrest by regulating the phosphorylation of cyclin-dependent kinase 1 (Cdk1) .
- Cdk1 cyclin-dependent kinase 1
- WEE1 inhibits Cdk1 by phosphorylating it on two different sites, Tyr15 and Thr14. Phosphorylated Cdk1 has reduced kinase activity and therefore prevents entry into mitosis.
- WEE1 As a cell progress through the cell cycle, WEE1 is inactivated and degraded, and the cell is allowed to enter mitosis.
- Cancer cells are often deficient in p53 signaling, display genomic instability, and rely on the WEE1 and G2/M cell cycle checkpoint to avoid mitotic catastrophe due to DNA damage. Inhibition or degradation of WEE1 sensitizes cancers to DNA-damaging therapies. Additional information may be found in a) LL Parker, and H Piwnica-Worms.
- the levels of proteins within a cell are determined by both the rate of protein synthesis and the rate of protein degradation.
- two pathways exist for selective protein degradation the ubiquitin-proteasome pathway and the lysosomal proteolysis pathway.
- selective protein degradation is mediated by the presence of a recruitment motif which promotes binding of degradation proteins, such as proteasomal degradation proteins, or proteins associated with ubiquitin-proteasome pathway.
- Recruitment motifs include E3 ligase recognition agents and proteasome recognition agents. Conjugation of recruitment motifs with high affinity ligands for WEE1 kinase provides compounds capable of selectively directing pathways for protein degradation to the WEE1 kinase protein itself. Such an outcome will reduce levels of WEE1 kinase activity.
- heteroaromatic WEE1 degradation compound having the general formula provided below:
- the WEE1 kinase affinity motif is a molecular construct having high affinity for the WEE1 kinase protein independent of the linking motif and/or the recruitment motif
- the linking motif is a molecular construct providing a covalent bond to both the WEE1 kinase affinity motif and the recruitment motif
- the recruitment motif is a molecular construct having the ability to selectively target and recruit protein degradation.
- Recruitment motifs include E3 ligase recognition agents and proteasome recognition agents.
- the recruitment motif is derived from, VHL ligand, nutlin, bestatin, HIF-1 ⁇ –VHL binding peptide, hydroxy proline-HIF-1 ⁇ -VHL binding peptide, SCFb-TRCP targeting peptide or an inhibitor of apoptosis protein ligand.
- the recruitment motif is selected from a molecular construct related to VHL ligand as illustrated below:
- the recruitment motif is selected from a molecular construct related to VHL ligand as illustrated below:
- the recruitment motif is In some embodiments, the recruitment motif is In some embodiments, the recruitment motif is
- VHL ligand VHL binder
- VHL E3 ubiquitin ligase binder VHL E3 ubiquitin ligase binder
- the linking motif is a molecular construct providing a covalent bond to both the WEE1 kinase affinity motif and the recruitment motif.
- the linking motif comprises a cyclic moiety.
- the linking motif comprises an acyclic moiety.
- the linking motif comprises an unsaturated moiety.
- the linking motif comprises between 4 and 50 non-hydrogen atoms in a linear sequence.
- the linking motif comprises between 4 and 20 non-hydrogen atoms in a linear sequence.
- the linking motif comprises between 4 and 25 non-hydrogen atoms in a linear sequence.
- the linking motif comprises between 4 and 30 non-hydrogen atoms in a linear sequence.
- the linking motif comprises between 6 and 18 atoms. In some embodiments, the linking motif comprises between 5 and 10 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 non-hydrogen atoms in a linear sequence.
- the linking motif is represented by the following formula:
- Ak 1 is selected from - (CR 5 R 6 ) k -;
- Ak 2 is selected from - (CR 5 R 6 ) m -;
- Ak 3 is selected from - (CR 5 R 6 ) n -;
- each of k, m, and n is selected from 0 to 10;
- R 5 and R 6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl;
- Z 1 and Z 2 are each independently selected from bond, -O-, -NR 7 -, -S-, heterocyclene, and cycloalkylene;
- R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced;
- the linking motif is a polyethylene glycol containing moiety.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5)
- said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) .
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from - (CR 5 R 6 ) n -NR 7 – (CR 5 R 6 ) m -*, wherein n is selected from 0 to 10, m is selected from 0 to 10, R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced; and the point of bonding to the VHL ligand indicated by the asterisk. In some embodiments, n is selected from 1 to 10, m is selected from 1 to 10.
- said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5)
- said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5)
- said n is selected from 1 to 7
- said m is selected from 0-3.
- R 7 is hydrogen or methyl.
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*wherein k is selected from 1 to 10, n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- said k is selected from 1 to 5
- said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5)
- said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) .
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- X, X1, and X2 at each instance are independently CH or N and wherein the point of bonding to the VHL ligand is indicated by the asterisk.
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif the -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- the linking motif -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is selected from a molecular fragment illustrated below:
- each R 7 is independently hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced , wherein the point of bonding to the VHL ligand is indicated by the asterisk.
- R 7 is hydrogen or methyl.
- k is1, 2, 3, 4, 5, 6, or 7, and m is 0, 1, 2, 3, 4, or 5.
- k is1, 2, 3, 4, 5, 6, or 7, and m is 0.
- k is1, 2, 3, 4, or 5 and m is 1, 2, 3, 4, or 5.
- the linking motif the linking motif is a polyethylene glycol selected from– (O-CH 2 -CH 2 ) n -O-and – (O-CH 2 -CH 2 ) n -N (R 23 ) -*, wherein n is selected from 1 to 20, and R 23 is optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced; and wherein the point of bonding to the VHL ligand is indicated by the asterisk.
- n is 1, 2, 3, 4, or 5
- Adavosertib (MK-1775, AZD1775) is a potent and selective Wee1 inhibitor with IC 50 of 5.2 nM in a cell-free assay as described in Hirai et al., “Small-molecule inhibition of Wee1 kinase by MK-1775 selectively sensitizes p53-deficient tumor cells to DNA-damaging agents” Mol. Can. Therap. 2009. DOI: 10.1158/1535-7163. MCT-09-0463 which is incorporated herein by reference.
- ZN-c3 also is a potent and selective Wee1 inhibitor with IC 50 of 3.8 nM in a cell-free assay as described in Huang et al., “Discovery of ZN-c3, a Highly Potent and Selective Wee1 inhibitor Undergoing Evaluation in Clinical Trials for the Treatment of Cancer” J. Med. Chem. 2021. DOI: 10.1021/acs. jmedchem. 1c01121which is incorporated herein by reference.
- R 1 , R 2 , R 3 , and R 4 are each independently selected from hydrogen, optionally substituted straight or branched C 1-6 alkyl, optionally substituted C 1-3 alkoxy, halogen, amino, hydroxyl, and cyano; or
- R 1 and R 2 , R 2 and R 3 , or R 3 and R 4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring;
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*;
- Ak 1 is selected from - (CR 5 R 6 ) k -;
- Ak 2 is selected from - (CR 5 R 6 ) m -;
- Ak 3 is selected from - (CR 5 R 6 ) n -;
- each of k, m, and n is selected from 0 to 10
- R 5 and R 6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl;
- Z 1 and Z 2 are each independently selected from bond, -O-, -NR 7 -, -S-, heterocyclene, and cycloalkylene;
- R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced;
- L is a polyethylene glycol containing moiety
- U is a VHL ligand.
- provided herein is a compound of Formula (I) , or a pharmaceutically acceptable salt or solvate thereof.
- R 1 , R 2 , R 3 , and R 4 are each independently selected from hydrogen, optionally substituted straight or branched C 1-6 alkyl, optionally substituted C 1-3 alkoxy, halogen, amino, hydroxyl, and cyano, wherein the alkyl is optionally substituted with hydroxy.
- R 1 , R 2 , R 3 , and R 4 are each independently selected from hydrogen, optionally substituted straight or branched C 1-6 alkyl, optionally substituted C 1-3 alkoxy, and halogen, wherein the alkyl is optionally substituted with hydroxyl.
- R 1 , R 2 , R 3 , and R 4 are each independently selected from hydrogen and optionally substituted straight or branched C 1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy.
- R 1 is optionally substituted branched C 1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy.
- R 1 is optionally substituted branched C 1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy and R 2 , R 3 , and R 4 are each hydrogen.
- R 1 is
- R 1 and R 2 , R 2 and R 3 , or R 3 and R 4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring, wherein the cycloalkyl and heterocyclyl are optionally substituted with one or more substituents selected from C 1-6 alkyl and hydroxyl.
- R 1 and R 2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring, wherein the cycloalkyl and heterocyclyl are optionally substituted with one or more substituents selected from C 1-6 alkyl and hydroxyl, and R 3 and R 4 are hydrogen.
- R 1 and R 2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl.
- R 1 and R 2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl, wherein the cycloalkyl is optionally substituted with one or more substituents selected from C 1-6 alkyl and hydroxyl.
- R 1 and R 2 taken together with the intervening atoms to which they are attached form a substituted 5-6 membered cycloalkyl, wherein the cycloalkyl is substituted with C 1-6 alkyl and hydroxyl.
- R 1 and R 2 taken together with the intervening atoms to which they are attached form
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*;
- Ak 1 is selected from - (CR 5 R 6 ) k -;
- Ak 2 is selected from - (CR 5 R 6 ) m -;
- Ak 3 is selected from - (CR 5 R 6 ) n -;
- each of k, m, and n is selected from 0 to 10
- R 5 and R 6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl;
- Z 1 and Z 2 are each independently selected from bond, -O-, -NR 7 -, -S-, heterocyclene, and cycloalkylene;
- R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced;
- L is a polyethylene glycol containing moiety
- U is a VHL ligand.
- provided herein is a compound of Formula (I-a) , or a pharmaceutically acceptable salt or solvate thereof.
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*;
- Ak 1 is selected from - (CR 5 R 6 ) k -;
- Ak 2 is selected from - (CR 5 R 6 ) m -;
- Ak 3 is selected from - (CR 5 R 6 ) n -;
- each of k, m, and n is selected from 0 to 10
- R 5 and R 6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl;
- Z 1 and Z 2 are each independently selected from bond, -O-, -NR 7 -, -S-, heterocyclene, and cycloalkylene;
- R 7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH 2 -units are replaced with O provided that two adjacent -CH 2 -units are not both replaced;
- L is a polyethylene glycol containing moiety
- U is a VHL ligand.
- provided herein is a compound of Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4, and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is - (CR 5 R 6 ) n -NR 7 – (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O– – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-7, m is 0-5, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0-3, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0, and R 7 is hydrogen or methyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is, wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2.
- R 5 and R 6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, hydroxyl, C 1-3 alkyl, and C 1-3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R 5 and R 6 are both hydrogen.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is .
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is .
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is,
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4, and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is - (CR 5 R 6 ) n -NR 7 – (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-7, m is 0-5, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0-3, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0, and R 7 is hydrogen or methyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is , wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2.
- R 5 and R 6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, hydroxyl, C 1-3 alkyl, and C 1-3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R 5 and R 6 are both hydrogen.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*
- provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I-b)
- L is -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 1, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 2, and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 3 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4, and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 4 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 5 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 3.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 4.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein n is 6 and m is 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is - (CR 5 R 6 ) n -NR 7 – (CR 5 R 6 ) m -*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 1.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 1, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 2, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 1, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 2, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 3, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is – (CR 5 R 6 ) k -O–– (CR 5 R 6 ) n -O– (CR 5 R 6 ) m -*, wherein k is 4, n is 3, and m is 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is and each R 7 is independently hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-7, m is 0-5, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0-3, and R 7 is hydrogen or C 1-3 alkyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein k is 1-6, m is 0, and R 7 is hydrogen or methyl.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is, wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2.
- R 5 and R 6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, hydroxyl, C 1-3 alkyl, and C 1-3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, halogen, C 1-3 alkyl, and C 3 cycloalkyl. In some embodiments, R 5 and R 6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R 5 and R 6 are both hydrogen.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2.
- -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -* is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*is, In some embodiments, -Ak 1 -Z 1 -Ak 2 -Z 2 -Ak 3 -*
- the compounds as described herein have a structure provided in Table 1, Table 2, or Table 3. In some embodiments, the compounds as described herein have a structure provided in Table 1.
- compounds as described herein has a structure provided in Table 2.
- compounds as described herein has a structure provided in Table 3.
- the heteroaromatic WEE1 degradation compound described e.g., a compound of Formula (I) , Formula (I-a) , or Formula (I-b) ) herein is administered as a pure chemical.
- the heteroaromatic WEE1 degradation compound described herein is combined with a pharmaceutically suitable or acceptable carrier (also referred to herein as a pharmaceutically suitable (or acceptable) excipient, physiologically suitable (or acceptable) excipient, or physiologically suitable (or acceptable) carrier) selected on the basis of a chosen route of administration and standard pharmaceutical practice as described, for example, in Remington: The Science and Practice of Pharmacy (Gennaro, 21 st Ed. Mack Pub. Co., Easton, PA (2005) ) .
- a pharmaceutical composition comprising at least one heteroaromatic WEE1 degradation compound as described herein (e.g., a compound of Formula (I) , Formula (I-a) , or Formula (I-b) ) , or a stereoisomer, pharmaceutically acceptable salt, hydrate, or solvate thereof, together with one or more pharmaceutically acceptable carriers.
- the carrier (s) or excipient (s) ) is acceptable or suitable if the carrier is compatible with the other ingredients of the composition and not deleterious to the recipient (i.e., the subject or the patient) of the composition.
- One embodiment provides a pharmaceutical composition
- a pharmaceutical composition comprising a pharmaceutically acceptable excipient and a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof.
- One embodiment provides a method of preparing a pharmaceutical composition comprising mixing a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier.
- the heteroaromatic WEE1 degradation compound as described by Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof is substantially pure, in that it contains less than about 5%, or less than about 1%, or less than about 0.1%, of other organic small molecules, such as unreacted intermediates or synthesis by-products that are created, for example, in one or more of the steps of a synthesis method.
- Suitable oral dosage forms include, for example, tablets, pills, sachets, or capsules of hard or soft gelatin, methylcellulose or of another suitable material easily dissolved in the digestive tract.
- suitable nontoxic solid carriers include, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, talcum, cellulose, glucose, sucrose, magnesium carbonate, and the like. (See, e.g., Remington: The Science and Practice of Pharmacy (Gennaro, 21 st Ed. Mack Pub. Co., Easton, PA (2005) ) .
- the heteroaromatic WEE1 degradation compound as described by Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof is formulated for administration by injection.
- the injection formulation is an aqueous formulation.
- the injection formulation is a non-aqueous formulation.
- the injection formulation is an oil-based formulation, such as sesame oil, or the like.
- the dose of the composition comprising at least one heteroaromatic WEE1 degradation compound as described herein differs depending upon the subject or patient's (e.g., human) condition. In some embodiments, such factors include general health status, age, and other factors.
- compositions are administered in a manner appropriate to the disease to be treated (or prevented) .
- An appropriate dose and a suitable duration and frequency of administration will be determined by such factors as the condition of the patient, the type and severity of the patient's disease, the particular form of the active ingredient, and the method of administration.
- an appropriate dose and treatment regimen provides the composition (s) in an amount sufficient to provide therapeutic and/or prophylactic benefit (e.g., an improved clinical outcome, such as more frequent complete or partial remissions, or longer disease-free and/or overall survival, or a lessening of symptom severity.
- Optimal doses are generally determined using experimental models and/or clinical trials. The optimal dose depends upon the body mass, weight, or blood volume of the patient.
- One embodiment provides a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, for use in a method of treatment of the human or animal body.
- One embodiment provides a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, for use in a method of treatment of cancer or neoplastic disease.
- One embodiment provides a use of a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, in the manufacture of a medicament for the treatment of cancer or neoplastic disease.
- described herein is a method of treating cancer in a patient in need thereof comprising administering to the patient a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof.
- described herein is a method of treating cancer in a patient in need thereof comprising administering to the patient a pharmaceutical composition comprising a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable excipient.
- Provided herein is the method wherein the pharmaceutical composition is administered orally. Provided herein is the method wherein the pharmaceutical composition is administered by injection.
- the compounds disclosed herein are synthesized according to the following examples.
- the following abbreviations unless otherwise indicated, shall be understood to have the following meanings: °C degrees Celsius ⁇ H chemical shift in parts per million downfield from tetramethylsilane DCM dichloromethane (CH 2 Cl 2 ) DMF dimethylformamide DMSO dimethylsulfoxide EA ethyl acetate ESI electrospray ionization Et ethyl g gram (s) h hour (s) HPLC high performance liquid chromatography Hz hertz J coupling constant (in NMR spectrometry) LCMS liquid chromatography mass spectrometry ⁇ micro m multiplet (spectral) ; meter (s) ; milli M molar M + parent molecular ion Me methyl MHz megahertz min minute (s) mol mole (s) ; molecular (as in mol wt
- Step 1 methyl 4- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ butanoate
- Step 2 4- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ butanoic acid
- Step 1 tert-butyl 6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanoate
- Step 2 6- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ hexanoic acid
- Step 2 7- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ heptanoic acid
- Step 1 ethyl 4- (2-allyl-2- (tert-butoxycarbonyl) hydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate
- Step 8 ethyl 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoate
- Step 9 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoic acid
- Step 1 ethyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoate
- Step 2 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoic acid
- Step 1 tert-butyl 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoate
- Step 2 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoic acid
- Step 1 methyl 11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanoate
- Step 4 methyl 4- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) butanoate
- Step 1 methyl 4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanoate
- Step 2 4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanoic acid
- Step 3 tert-butyl 2- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) oxy) acetate
- Step 1 tert-butyl 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butoxy) acetate
- Step 3 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 12)
- Step 1 tert-butyl 2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetate
- reaction was purified by reverse column to afford [ (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) methoxy] acetic acid.
- Step 1 tert-butyl 2- [ (5- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ pentyl) oxy] acetate
- Step 2 [ (5- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ pentyl) oxy] acetic acid
- Step 1 tert-butyl 5- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) pentanoate
- Step 2 5- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) pentanoic acid
- Step 1 tert-butyl 2- [2- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) ethoxy] acetate
- Step 2 methyl dodec-11-ynoate [2- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) ethoxy] acetic acid
- Step 1 tert-butyl 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetate
- Step 4 tert-butyl 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoate
- the resulting mixture was stirred at 50 °C overnight.
- the reaction solution was quenched with water (20 mL) .
- the resulting mixture was extracted with EtOAc (3 x 50 mL) .
- the combined organic layers were washed with brine (4 x 100 mL) and then concentrated.
- Step 3 4- (4- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ butoxy) butanoic acid
- Step 3 tert-butyl 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetate
- Step 4 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetic acid
- Step 1 10, 10, 11, 11-tetramethyl-1-phenyl-2, 6, 9-trioxa-10-siladodecane
- Step 7 [2- (3- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ propoxy) ethoxy] acetic acid
- Step 4 methyl 6- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) hexanoate
- Step 5 6- (2- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ ethoxy) hexanoic acid
- Step 1 tert-butyl 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoate
- Step 2 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoic acid
- the resulting mixture was stirred for 1 h at 60 °C in air atmosphere.
- NaBH 3 CN 51.66 mg, 0.822 mmol, 1.6 equiv.
- the resulting mixture was stirred for additional for 5 h at room temperature. Desired product could be detected by LCMS.
- the reaction was quenched with sat. NaHCO 3 (aq. ) at room temperature.
- the resulting mixture was extracted with EA (3 x 10 mL) . The combined organic layers were dried over anhydrous Na 2 SO 4 .
- Step 6 4'- ⁇ 4- [4- ( ⁇ 1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ - [1, 1'-bi (cyclohexane) ] -4-carboxylic acid
- reaction mixture was further purified by Prep-HPLC (Column: Xselect CSH C18 OBD Column 30*150mm 5 ⁇ m, n; Mobile Phase A: Water (10 mmol/L NH 4 HCO 3 +0.1%NH 3 .
- Step 1 methyl 4- (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) cyclohexane-1-carboxylate
- Step 3 methyl 4- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) cyclohexane-1-carboxylate
- Step 4 4- (4- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) cyclohexane-1-carboxylic acid
- Step 1 tert-butyl 4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidine-1-carboxylate
- the resulting mixture was stirred at 25 °C for 3 h.
- the reaction was quenched with H 2 O (10 mL) .
- the mixture was basified to pH ⁇ 9 with NaHCO 3 aqueous solution and extracted with EA (3 x 50 mL) , washed with brine (3 x 70 mL) , the organic layers were combined and concentrated under vacuum.
- Step 4 6- ( (4- (4- ( [1, 4'-bipiperidin] -4-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one
- Step 6 4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) - [1, 4'-bipiperidine] -1'-carboxamide (Compound 29)
- Step 1 ethyl 1- ⁇ 1, 4-dioxaspiro [4.5] decan-8-yl ⁇ piperidine-4-carboxylate
- Step 4 1- (4- ⁇ 4- [4- ( ⁇ 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl ⁇ amino) phenyl] piperazin-1-yl ⁇ cyclohexyl) piperidine-4-carboxylic acid
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Abstract
Provided herein are compounds which target WEE1 kinase proteins for ubiquitination and proteasomal degradation. Also provided herein are methods for using said compounds for the treatment of diseases.
Description
CROSS-REFERENCE
This application claims the benefit of International Application PCT/CN2022/085833 filed April 8, 2022, which is incorporated herein by reference in its entirety.
WEE1 is a kinase belonging to the Ser/Thr family of protein kinases and is a key regulator of cell cycle progression. Pharmacological intervention in the cell cycle is strategy for the treatment of cancer and other proliferative disease. The present disclosure provides compounds, compositions and methods for the reduction or elimination of WEE1 activity via the targeting of WEE1 protein in a cell through ubiquitination and proteasomal degradation.
BRIEF SUMMARY OF THE INVENTION
Provided herein are compounds which target WEE1 proteins for ubiquitination and proteasomal degradation. Also provided herein are methods for using said compounds for the treatment of diseases.
One embodiment provides a compound, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I) :
wherein,
R1, R2, R3, and R4 are each independently selected from hydrogen, optionally substituted straight or branched C1-6 alkyl, optionally substituted C1-3 alkoxy, halogen, amino, hydroxyl, and cyano; or
R1 and R2, R2 and R3, or R3 and R4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocycloalkyl ring;
L is -Ak1-Z1-Ak2-Z2-Ak3-*;
Ak1 is selected from - (CR5R6) k-;
Ak2 is selected from - (CR5R6) m-;
Ak3 is selected from - (CR5R6) n-;
each of k, m, and n is selected from 0 to 10
R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;
Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;
R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced;
wherein at least one of Z1 and Z2 is not bond; or
L is a polyethylene glycol containing moiety;
the point of bonding to the VHL ligand is indicated by the asterisk; and
U is a VHL ligand.
One embodiment provides a pharmaceutical composition comprising a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, and at least one pharmaceutically acceptable excipient.
One embodiment provides a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof for use in a method of treatment of a human or animal.
One embodiment provides a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, for use in a method of treatment of cancer or neoplastic disease.
One embodiment provides the use of a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, in the manufacture of a medicament for the treatment of cancer or neoplastic disease.
One embodiment provides a method of treating a disease or disorder in a patient in need thereof comprising administering to the patient a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof. Another embodiment provides the method wherein the disease or disorder is cancer.
One embodiment provides a method of treating cancer in a patient in need thereof comprising administering to the patient a pharmaceutical composition comprising a compound of Formula (I) , or pharmaceutically acceptable salt solvate, prodrug, stereoisomer, or tautomer thereof, and a pharmaceutically acceptable excipient.
INCORPORATION BY REFERENCE
All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference for the specific purposes identified herein.
As used herein and in the appended claims, the singular forms "a, " "and, " and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "an agent" includes a plurality of such agents, and reference to "the cell" includes reference to one or more cells (or to a plurality of cells) and equivalents thereof known to those skilled in the art, and so forth. When ranges are used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formulae, all combinations and subcombinations of ranges and specific embodiments therein are intended to be included. The term "about" when referring to a number or a numerical range means that the number or numerical range referred to is an approximation within experimental variability (or within statistical experimental error) , and thus the number or numerical range, in some instances, will vary between 1%and 15%of the stated number or numerical range. The term "comprising" (and related terms such as "comprise" or "comprises" or "having" or "including" ) is not intended to exclude that in other certain embodiments, for example, an embodiment of any composition of matter, composition, method, or process, or the like, described herein, "consist of" or "consist essentially of" the described features.
Definitions
As used in the specification and appended claims, unless specified to the contrary, the following terms have the meaning indicated below.
"Amino" refers to the –NH2 radical.
"Cyano" refers to the -CN radical.
"Nitro" refers to the -NO2 radical.
"Oxa" refers to the -O-radical.
"Oxo" refers to the =O radical.
"Thioxo" refers to the =S radical.
"Imino" refers to the =N-H radical.
"Oximo" refers to the =N-OH radical.
"Hydrazino" refers to the =N-NH2 radical.
"Alkyl" refers to a straight or branched hydrocarbon chain radical consisting solely of carbon and hydrogen atoms, containing no unsaturation, having from one to fifteen carbon atoms (e.g., C1-C15 alkyl) . In certain embodiments, an alkyl comprises one to thirteen carbon atoms (e.g., C1-C13 alkyl) . In certain embodiments, an alkyl comprises one to eight carbon atoms (e.g., C1-C8 alkyl) . In other embodiments, an alkyl comprises one to five carbon atoms (e.g., C1-C5 alkyl) . In other embodiments, an alkyl comprises one to four carbon atoms (e.g., C1-C4 alkyl) . In other embodiments, an alkyl comprises one to three carbon atoms (e.g., C1-C3 alkyl) . In other
embodiments, an alkyl comprises one to two carbon atoms (e.g., C1-C2 alkyl) . In other embodiments, an alkyl comprises one carbon atom (e.g., C1 alkyl) . In other embodiments, an alkyl comprises five to fifteen carbon atoms (e.g., C5-C15 alkyl) . In other embodiments, an alkyl comprises five to eight carbon atoms (e.g., C5-C8 alkyl) . In other embodiments, an alkyl comprises two to five carbon atoms (e.g., C2-C5 alkyl) . In other embodiments, an alkyl comprises three to five carbon atoms (e.g., C3-C5 alkyl) . In other embodiments, the alkyl group is selected from methyl, ethyl, 1-propyl (n-propyl) , 1-methylethyl (iso-propyl) , 1-butyl (n-butyl) , 1-methylpropyl (sec-butyl) , 2-methylpropyl (iso-butyl) , 1, 1-dimethylethyl (tert-butyl) , and 1-pentyl (n-pentyl) . The alkyl is attached to the rest of the molecule by a single bond. Unless stated otherwise specifically in the specification, an alkyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa (where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Alkoxy" refers to a radical bonded through an oxygen atom of the formula –O-alkyl, where alkyl is an alkyl chain as defined above.
"Alkenyl" refers to a straight or branched hydrocarbon chain radical group consisting solely of carbon and hydrogen atoms, containing at least one carbon-carbon double bond, and having from two to twelve carbon atoms. In certain embodiments, an alkenyl comprises two to eight carbon atoms. In other embodiments, an alkenyl comprises two to four carbon atoms. The alkenyl is attached to the rest of the molecule by a single bond, for example, ethenyl (i.e., vinyl) , prop-1-enyl (i.e., allyl) , but-1-enyl, pent-1-enyl, penta-1, 4-dienyl, and the like. Unless stated otherwise specifically in the specification, an alkenyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa
(where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Alkynyl" refers to a straight or branched hydrocarbon chain radical group consisting solely of carbon and hydrogen atoms, containing at least one carbon-carbon triple bond, having from two to twelve carbon atoms. In certain embodiments, an alkynyl comprises two to eight carbon atoms. In other embodiments, an alkynyl comprises two to six carbon atoms. In other embodiments, an alkynyl comprises two to four carbon atoms. The alkynyl is attached to the rest of the molecule by a single bond, for example, ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like. Unless stated otherwise specifically in the specification, an alkynyl group is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa (where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Alkylene" or "alkylene chain" refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing no unsaturation and having from one to twelve carbon atoms, for example, methylene, ethylene, propylene, n-butylene, and the like. The alkylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. The points of
attachment of the alkylene chain to the rest of the molecule and to the radical group are through one carbon in the alkylene chain or through any two carbons within the chain. In certain embodiments, an alkylene comprises one to eight carbon atoms (e.g., C1-C8 alkylene) . In other embodiments, an alkylene comprises one to five carbon atoms (e.g., C1-C5 alkylene) . In other embodiments, an alkylene comprises one to four carbon atoms (e.g., C1-C4 alkylene) . In other embodiments, an alkylene comprises one to three carbon atoms (e.g., C1-C3 alkylene) . In other embodiments, an alkylene comprises one to two carbon atoms (e.g., C1-C2 alkylene) . In other embodiments, an alkylene comprises one carbon atom (e.g., C1 alkylene) . In other embodiments, an alkylene comprises five to eight carbon atoms (e.g., C5-C8 alkylene) . In other embodiments, an alkylene comprises two to five carbon atoms (e.g., C2-C5 alkylene) . In other embodiments, an alkylene comprises three to five carbon atoms (e.g., C3-C5 alkylene) . Unless stated otherwise specifically in the specification, an alkylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa (where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Alkenylene" or "alkenylene chain" refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing at least one carbon-carbon double bond, and having from two to twelve carbon atoms. The alkenylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. In certain embodiments, an alkenylene comprises two to eight carbon atoms (e.g., C2-C8 alkenylene) . In other embodiments, an alkenylene comprises two to five carbon atoms (e.g., C2-C5 alkenylene) . In other embodiments, an alkenylene comprises two to four carbon atoms (e.g., C2-C4 alkenylene) . In other embodiments, an alkenylene comprises two to three carbon atoms (e.g., C2-C3 alkenylene) . In other embodiments, an alkenylene comprises two carbon atoms (e.g., C2 alkenylene) . In other embodiments, an alkenylene comprises five to eight carbon
atoms (e.g., C5-C8 alkenylene) . In other embodiments, an alkenylene comprises three to five carbon atoms (e.g., C3-C5 alkenylene) . Unless stated otherwise specifically in the specification, an alkenylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa (where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Alkynylene" or "alkynylene chain" refers to a straight or branched divalent hydrocarbon chain linking the rest of the molecule to a radical group, consisting solely of carbon and hydrogen, containing at least one carbon-carbon triple bond, and having from two to twelve carbon atoms. The alkynylene chain is attached to the rest of the molecule through a single bond and to the radical group through a single bond. In certain embodiments, an alkynylene comprises two to eight carbon atoms (e.g., C2-C8 alkynylene) . In other embodiments, an alkynylene comprises two to five carbon atoms (e.g., C2-C5 alkynylene) . In other embodiments, an alkynylene comprises two to four carbon atoms (e.g., C2-C4 alkynylene) . In other embodiments, an alkynylene comprises two to three carbon atoms (e.g., C2-C3 alkynylene) . In other embodiments, an alkynylene comprises two carbon atoms (e.g., C2 alkynylene) . In other embodiments, an alkynylene comprises five to eight carbon atoms (e.g., C5-C8 alkynylene) . In other embodiments, an alkynylene comprises three to five carbon atoms (e.g., C3-C5 alkynylene) . Unless stated otherwise specifically in the specification, an alkynylene chain is optionally substituted by one or more of the following substituents: halo, cyano, nitro, oxo, thioxo, imino, oximo, trimethylsilanyl, -ORa, -SRa, -OC (O) -Ra, -N (Ra) 2, -C (O) Ra, -C (O) ORa, -C (O) N (Ra) 2, -N (Ra) C (O) ORa, -OC (O) -N (Ra) 2, -N (Ra) C (O) Ra, -N (Ra) S (O) tRa (where t is 1 or 2) , -S (O) tORa (where t is 1 or 2) , -S (O) tRa (where t is 1 or 2) and -S (O) tN (Ra) 2 (where t is 1 or 2) where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, carbocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , carbocyclylalkyl
(optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) .
"Aryl" refers to a radical derived from an aromatic monocyclic or multicyclic hydrocarbon ring system by removing a hydrogen atom from a ring carbon atom. The aromatic monocyclic or multicyclic hydrocarbon ring system contains only hydrogen and carbon from five to eighteen carbon atoms, where at least one of the rings in the ring system is fully unsaturated, i.e., it contains a cyclic, delocalized (4n+2) π–electron system in accordance with the Hückel theory. The ring system from which aryl groups are derived include, but are not limited to, groups such as benzene, fluorene, indane, indene, tetralin and naphthalene. Unless stated otherwise specifically in the specification, the term "aryl" or the prefix "ar-" (such as in "aralkyl" ) is meant to include aryl radicals optionally substituted by one or more substituents independently selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -Rb-ORa, -Rb-OC (O) -Ra, -Rb-OC (O) -ORa, -Rb-OC (O) -N (Ra) 2, -Rb-N (Ra) 2, -Rb-C (O) Ra, -Rb-C (O) ORa, -Rb-C (O) N (Ra) 2, -Rb-O-Rc-C (O) N (Ra) 2, -Rb-N (Ra) C (O) ORa, -Rb-N (Ra) C (O) Ra, -Rb-N (Ra) S (O) tRa (where t is 1 or 2) , -Rb-S (O) tRa (where t is 1 or 2) , -Rb-S (O) tORa (where t is 1 or 2) and -Rb-S (O) tN (Ra) 2 (where t is 1 or 2) , where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, cycloalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , cycloalkylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , each Rb is independently a direct bond or a straight or branched alkylene or alkenylene chain, and Rc is a
straight or branched alkylene or alkenylene chain, and where each of the above substituents is unsubstituted unless otherwise indicated.
"Aralkyl" refers to a radical of the formula -Rc-aryl where Rc is an alkylene chain as defined above, for example, methylene, ethylene, and the like. The alkylene chain part of the aralkyl radical is optionally substituted as described above for an alkylene chain. The aryl part of the aralkyl radical is optionally substituted as described above for an aryl group.
"Aralkenyl" refers to a radical of the formula –Rd-aryl where Rd is an alkenylene chain as defined above. The aryl part of the aralkenyl radical is optionally substituted as described above for an aryl group. The alkenylene chain part of the aralkenyl radical is optionally substituted as defined above for an alkenylene group.
"Aralkynyl" refers to a radical of the formula -Re-aryl, where Re is an alkynylene chain as defined above. The aryl part of the aralkynyl radical is optionally substituted as described above for an aryl group. The alkynylene chain part of the aralkynyl radical is optionally substituted as defined above for an alkynylene chain.
"Aralkoxy" refers to a radical bonded through an oxygen atom of the formula -O-Rc-aryl where Rc is an alkylene chain as defined above, for example, methylene, ethylene, and the like. The alkylene chain part of the aralkyl radical is optionally substituted as described above for an alkylene chain. The aryl part of the aralkyl radical is optionally substituted as described above for an aryl group.
"Carbocyclyl" refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms, which includes fused, bridged, or spirocyclic ring systems, having from three to fifteen carbon atoms. In certain embodiments, a carbocyclyl comprises three to ten carbon atoms. In other embodiments, a carbocyclyl comprises five to seven carbon atoms. The carbocyclyl is attached to the rest of the molecule by a single bond. Carbocyclyl is saturated (i.e., containing single C-C bonds only) or unsaturated (i.e., containing one or more double bonds or triple bonds) . A fully saturated carbocyclyl radical is also referred to as "cycloalkyl. " Examples of monocyclic cycloalkyls include, e.g., cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. An unsaturated carbocyclyl is also referred to as "cycloalkenyl. " Examples of monocyclic cycloalkenyls include, e.g., cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl. Polycyclic carbocyclyl radicals include, for example, adamantyl, norbornyl (i.e., bicyclo [2.2.1] heptanyl) , norbornenyl, decalinyl, 7, 7-dimethyl-bicyclo [2.2.1] heptanyl, and the like. Spirocyclic carbocyclyl or cycloalkyl radicals include, for example, spiro [2.2] pentane, spiro [2.3] hexane, spiro [2.4] heptane, spiro [2.5] octane, spiro [2.6] nonane, spiro [3.3] heptane, spiro [3.4] octane, spiro [3.5] nonane, spiro [3.6] decane, spiro [4.4] nonane, spiro [4.5] decane, spiro [4.6] undecane, spiro [5.5] undecane, spiro [5.6] dodecane,
spiro [6.6] tridecane, and the like. Unless otherwise stated specifically in the specification, the term "carbocyclyl" is meant to include carbocyclyl radicals that are optionally substituted by one or more substituents independently selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -Rb-ORa, -Rb-OC (O) -Ra, -Rb-OC (O) -ORa, -Rb-OC (O) -N (Ra) 2, -Rb-N (Ra) 2, -Rb-C (O) Ra, -Rb-C (O) ORa, -Rb-C (O) N (Ra) 2, -Rb-O-Rc-C (O) N (Ra) 2, -Rb-N (Ra) C (O) ORa, -Rb-N (Ra) C (O) Ra, -Rb-N (Ra) S (O) tRa (where t is 1 or 2) , -Rb-S (O) tRa (where t is 1 or 2) , -Rb-S (O) tORa (where t is 1 or 2) and -Rb-S (O) tN (Ra) 2 (where t is 1 or 2) , where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, cycloalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , cycloalkylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , each Rb is independently a direct bond or a straight or branched alkylene or alkenylene chain, and Rc is a straight or branched alkylene or alkenylene chain, and where each of the above substituents is unsubstituted unless otherwise indicated.
“Cycloalkylene” refers to a divalent carbocyclyl or cycloalkyl linking the rest of the molecule to a radical group.
"Carbocyclylalkyl" refers to a radical of the formula –Rc-carbocyclyl where Rc is an alkylene chain as defined above. The alkylene chain and the carbocyclyl radical is optionally substituted as defined above.
"Carbocyclylalkynyl" refers to a radical of the formula –Rc-carbocyclyl where Rc is an alkynylene chain as defined above. The alkynylene chain and the carbocyclyl radical is optionally substituted as defined above.
"Carbocyclylalkoxy" refers to a radical bonded through an oxygen atom of the formula –O-Rc-carbocyclyl where Rc is an alkylene chain as defined above. The alkylene chain and the carbocyclyl radical is optionally substituted as defined above.
As used herein, “carboxylic acid bioisostere” refers to a functional group or moiety that exhibits similar physical, biological and/or chemical properties as a carboxylic acid moiety. Examples of carboxylic acid bioisosteres include, but are not limited to,
and the like.
"Halo" or "halogen" refers to bromo, chloro, fluoro or iodo substituents.
"Fluoroalkyl" refers to an alkyl radical, as defined above, that is substituted by one or more fluoro radicals, as defined above, for example, trifluoromethyl, difluoromethyl, fluoromethyl, 2, 2, 2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, and the like. In some embodiments, the alkyl part of the fluoroalkyl radical is optionally substituted as defined above for an alkyl group.
"Heterocyclyl" refers to a stable 3-to 18-membered non-aromatic ring radical that comprises two to twelve carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur. Unless stated otherwise specifically in the specification, the heterocyclyl radical is a monocyclic, bicyclic, tricyclic, tetracyclic, or spirocyclic ring system, which optionally includes fused or bridged ring systems. The heteroatoms in the heterocyclyl radical are optionally oxidized. One or more nitrogen atoms, if present, are optionally quaternized. The heterocyclyl radical is partially or fully saturated. The heterocyclyl is attached to the rest of the molecule through any atom of the ring (s) . Examples of such heterocyclyl radicals include, but are not limited to, dioxolanyl, thienyl [1, 3] dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, and 1, 1-dioxo-thiomorpholinyl, 1-azaspiro [2.2] pentane, 1-azaspiro [2.3] hexane, 1-azaspiro [2.4] heptane, 1-azaspiro [2.5] octane, 1-azaspiro [2.6] nonane, 2-azaspiro [3.3] heptane, 2-azaspiro [3.4] octane, 2-azaspiro [3.5] nonane, 2-azaspiro [3.6] decane, 2-azaspiro [4.4] nonane, 2-azaspiro [4.5] decane, 2-azaspiro [4.6] undecane, 3-azaspiro [5.5] undecane, 3-azaspiro [5.6] dodecane, 3-azaspiro [6.6] tridecane, 5-azaspiro [2.3] hexane, 5-azaspiro [2.4] heptane, 6-azaspiro [2.5] octane, 6-azaspiro [2.6] nonane, 6-azaspiro [3.4] octane, 7-azaspiro [3.5] nonane, 7-azaspiro [3.6] decane, 8-azaspiro [4.5] decane, 8-azaspiro [4.6] undecane, 9-azaspiro [5.6] dodecane, 1, 4-diazaspiro [2.2] pentane, 1, 5-diazaspiro [2.3] hexane, 1, 5-diazaspiro [2.4] heptane, 1, 6-
diazaspiro [2.5] octane, 1, 6-diazaspiro [2.6] nonane, 2, 6-diazaspiro [3.3] heptane, 2, 6-diazaspiro [3.4] octane, 2, 7-diazaspiro [3.5] nonane, 2, 7-diazaspiro [3.6] decane, 2, 7-diazaspiro [4.4] nonane, 2, 8-diazaspiro [4.5] decane, 2, 8-diazaspiro [4.6] undecane, 3, 9-diazaspiro [5.5] undecane, 3, 9-diazaspiro [5.6] dodecane, and 3, 10-diazaspiro [6.6] tridecane. Unless stated otherwise specifically in the specification, the term "heterocyclyl" is meant to include heterocyclyl radicals as defined above that are optionally substituted by one or more substituents selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -Rb-ORa, -Rb-OC (O) -Ra, -Rb-OC (O) -ORa, -Rb-OC (O) -N (Ra) 2, -Rb-N (Ra) 2, -Rb-C (O) Ra, -Rb-C (O) ORa, -Rb-C (O) N (Ra) 2, -Rb-O-Rc-C (O) N (Ra) 2, -Rb-N (Ra) C (O) ORa, -Rb-N (Ra) C (O) Ra, -Rb-N (Ra) S (O) tRa (where t is 1 or 2) , -Rb-S (O) tRa (where t is 1 or 2) , -Rb-S (O) tORa (where t is 1 or 2) and -Rb-S (O) tN (Ra) 2 (where t is 1 or 2) , where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, cycloalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , cycloalkylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , each Rb is independently a direct bond or a straight or branched alkylene or alkenylene chain, and Rc is a straight or branched alkylene or alkenylene chain, and where each of the above substituents is unsubstituted unless otherwise indicated.
“Heterocyclene” refers to a divalent heterocyclyl linking the rest of the molecule to a radical group.
"N-heterocyclyl" or “N-attached heterocyclyl” refers to a heterocyclyl radical as defined above containing at least one nitrogen and where the point of attachment of the heterocyclyl radical to the rest of the molecule is through a nitrogen atom in the heterocyclyl radical. An N-heterocyclyl radical is optionally substituted as described above for heterocyclyl radicals. Examples of such N-heterocyclyl radicals include, but are not limited to, 1-morpholinyl, 1-piperidinyl, 1-piperazinyl, 1-pyrrolidinyl, pyrazolidinyl, imidazolinyl, and imidazolidinyl.
"C-heterocyclyl" or “C-attached heterocyclyl” refers to a heterocyclyl radical as defined above containing at least one heteroatom and where the point of attachment of the heterocyclyl radical to the rest of the molecule is through a carbon atom in the heterocyclyl radical. A C-heterocyclyl radical is optionally substituted as described above for heterocyclyl radicals. Examples of such C-heterocyclyl radicals include, but are not limited to, 2-morpholinyl, 2-or 3-or 4-piperidinyl, 2-piperazinyl, 2-or 3-pyrrolidinyl, and the like.
"Heterocyclylalkyl" refers to a radical of the formula –Rc-heterocyclyl where Rc is an alkylene chain as defined above. If the heterocyclyl is a nitrogen-containing heterocyclyl, the heterocyclyl is optionally attached to the alkyl radical at the nitrogen atom. The alkylene chain of the heterocyclylalkyl radical is optionally substituted as defined above for an alkylene chain. The heterocyclyl part of the heterocyclylalkyl radical is optionally substituted as defined above for a heterocyclyl group.
"Heterocyclylalkoxy" refers to a radical bonded through an oxygen atom of the formula –O-Rc-heterocyclyl where Rc is an alkylene chain as defined above. If the heterocyclyl is a nitrogen-containing heterocyclyl, the heterocyclyl is optionally attached to the alkyl radical at the nitrogen atom. The alkylene chain of the heterocyclylalkoxy radical is optionally substituted as defined above for an alkylene chain. The heterocyclyl part of the heterocyclylalkoxy radical is optionally substituted as defined above for a heterocyclyl group.
"Heteroaryl" refers to a radical derived from a 3-to 18-membered aromatic ring radical that comprises two to seventeen carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur. As used herein, the heteroaryl radical is a monocyclic, bicyclic, tricyclic or tetracyclic ring system, wherein at least one of the rings in the ring system is fully unsaturated, i.e., it contains a cyclic, delocalized (4n+2) π–electron system in accordance with the Hückel theory. Heteroaryl includes fused or bridged ring systems. The heteroatom (s) in the heteroaryl radical is optionally oxidized. One or more nitrogen atoms, if present, are optionally quaternized. The heteroaryl is attached to the rest of the molecule through any atom of the ring (s) . Examples of heteroaryls include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzindolyl, 1, 3-benzodioxolyl, benzofuranyl, benzooxazolyl, benzo [d] thiazolyl, benzothiadiazolyl, benzo [b] [1, 4] dioxepinyl, benzo [b] [1, 4] oxazinyl, 1, 4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl) , benzothieno [3, 2-d] pyrimidinyl, benzotriazolyl, benzo [4, 6] imidazo [1, 2-a] pyridinyl, carbazolyl, cinnolinyl, cyclopenta [d] pyrimidinyl, 6, 7-dihydro-5H-cyclopenta [4, 5] thieno [2, 3-d] pyrimidinyl, 5, 6-dihydrobenzo [h] quinazolinyl, 5, 6-dihydrobenzo [h] cinnolinyl, 6, 7-dihydro-5H-benzo [6, 7] cyclohepta [1, 2-c] pyridazinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanonyl,
furo [3, 2-c] pyridinyl, 5, 6, 7, 8, 9, 10-hexahydrocycloocta [d] pyrimidinyl, 5, 6, 7, 8, 9, 10-hexahydrocycloocta [d] pyridazinyl, 5, 6, 7, 8, 9, 10-hexahydrocycloocta [d] pyridinyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl, isoindolyl, indolinyl, isoindolinyl, isoquinolyl, indolizinyl, isoxazolyl, 5, 8-methano-5, 6, 7, 8-tetrahydroquinazolinyl, naphthyridinyl, 1, 6-naphthyridinonyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl, oxiranyl, 5, 6, 6a, 7, 8, 9, 10, 10a-octahydrobenzo [h] quinazolinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl, pyrazolo [3, 4-d] pyrimidinyl, pyridinyl, pyrido [3, 2-d] pyrimidinyl, pyrido [3, 4-d] pyrimidinyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl, quinazolinyl, quinoxalinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, 5, 6, 7, 8-tetrahydroquinazolinyl, 5, 6, 7, 8-tetrahydrobenzo [4, 5] thieno [2, 3-d] pyrimidinyl, 6, 7, 8, 9-tetrahydro-5H-cyclohepta [4, 5] thieno [2, 3-d] pyrimidinyl, 5, 6, 7, 8-tetrahydropyrido [4, 5-c] pyridazinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, thieno [2, 3-d] pyrimidinyl, thieno [3, 2-d] pyrimidinyl, thieno [2, 3-c] pridinyl, and thiophenyl (i.e. thienyl) . Unless stated otherwise specifically in the specification, the term "heteroaryl" is meant to include heteroaryl radicals as defined above which are optionally substituted by one or more substituents selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, haloalkenyl, haloalkynyl, oxo, thioxo, cyano, nitro, optionally substituted aryl, optionally substituted aralkyl, optionally substituted aralkenyl, optionally substituted aralkynyl, optionally substituted carbocyclyl, optionally substituted carbocyclylalkyl, optionally substituted heterocyclyl, optionally substituted heterocyclylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -Rb-ORa, -Rb-OC (O) -Ra, -Rb-OC (O) -ORa, -Rb-OC (O) -N (Ra) 2, -Rb-N (Ra) 2, -Rb-C (O) Ra, -Rb-C (O) ORa, -Rb-C (O) N (Ra) 2, -Rb-O-Rc-C (O) N (Ra) 2, -Rb-N (Ra) C (O) ORa, -Rb-N (Ra) C (O) Ra, -Rb-N (Ra) S (O) tRa (where t is 1 or 2) , -Rb-S (O) tRa (where t is 1 or 2) , -Rb-S (O) tORa (where t is 1 or 2) and -Rb-S (O) tN (Ra) 2 (where t is 1 or 2) , where each Ra is independently hydrogen, alkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , fluoroalkyl, cycloalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , cycloalkylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , aralkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heterocyclylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , heteroaryl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , or heteroarylalkyl (optionally substituted with halogen, hydroxy, methoxy, or trifluoromethyl) , each Rb is independently a direct bond or a straight or
branched alkylene or alkenylene chain, and Rc is a straight or branched alkylene or alkenylene chain, and where each of the above substituents is unsubstituted unless otherwise indicated.
"N-heteroaryl" refers to a heteroaryl radical as defined above containing at least one nitrogen and where the point of attachment of the heteroaryl radical to the rest of the molecule is through a nitrogen atom in the heteroaryl radical. An N-heteroaryl radical is optionally substituted as described above for heteroaryl radicals.
"C-heteroaryl" refers to a heteroaryl radical as defined above and where the point of attachment of the heteroaryl radical to the rest of the molecule is through a carbon atom in the heteroaryl radical. A C-heteroaryl radical is optionally substituted as described above for heteroaryl radicals.
"Heteroarylalkyl" refers to a radical of the formula –Rc-heteroaryl, where Rc is an alkylene chain as defined above. If the heteroaryl is a nitrogen-containing heteroaryl, the heteroaryl is optionally attached to the alkyl radical at the nitrogen atom. The alkylene chain of the heteroarylalkyl radical is optionally substituted as defined above for an alkylene chain. The heteroaryl part of the heteroarylalkyl radical is optionally substituted as defined above for a heteroaryl group.
"Heteroarylalkoxy" refers to a radical bonded through an oxygen atom of the formula –O-Rc-heteroaryl, where Rc is an alkylene chain as defined above. If the heteroaryl is a nitrogen-containing heteroaryl, the heteroaryl is optionally attached to the alkyl radical at the nitrogen atom. The alkylene chain of the heteroarylalkoxy radical is optionally substituted as defined above for an alkylene chain. The heteroaryl part of the heteroarylalkoxy radical is optionally substituted as defined above for a heteroaryl group.
The compounds disclosed herein, in some embodiments, contain one or more asymmetric centers and thus give rise to enantiomers, diastereomers, and other stereoisomeric forms that are defined, in terms of absolute stereochemistry, as (R) -or (S) -. Unless stated otherwise, it is intended that all stereoisomeric forms of the compounds disclosed herein are contemplated by this disclosure. When the compounds described herein contain alkene double bonds, and unless specified otherwise, it is intended that this disclosure includes both E and Z geometric isomers (e.g., cis or trans. ) Likewise, all possible isomers, as well as their racemic and optically pure forms, and all tautomeric forms are also intended to be included. The term “geometric isomer” refers to E or Z geometric isomers (e.g., cis or trans) of an alkene double bond. The term “positional isomer” refers to structural isomers around a central ring, such as ortho-, meta-, and para-isomers around a benzene ring.
A "tautomer" refers to a molecule wherein a proton shift from one atom of a molecule to another atom of the same molecule is possible. The compounds presented herein, in certain
embodiments, exist as tautomers. In circumstances where tautomerization is possible, a chemical equilibrium of the tautomers will exist. The exact ratio of the tautomers depends on several factors, including physical state, temperature, solvent, and pH. Some examples of tautomeric equilibrium include:
The compounds disclosed herein, in some embodiments, are used in different enriched isotopic forms, e.g., enriched in the content of 2H, 3H, 11C, 13C and/or 14C. In one particular embodiment, the compound is deuterated in at least one position. Such deuterated forms can be made by the procedure described in U.S. Patent Nos. 5,846,514 and 6,334,997. As described in U.S. Patent Nos. 5,846,514 and 6,334,997, deuteration can improve the metabolic stability and or efficacy, thus increasing the duration of action of drugs.
Unless otherwise stated, structures depicted herein are intended to include compounds which differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures except for the replacement of a hydrogen by a deuterium or tritium, or the replacement of a carbon by 13C-or 14C-enriched carbon are within the scope of the present disclosure.
The compounds of the present disclosure optionally contain unnatural proportions of atomic isotopes at one or more atoms that constitute such compounds. For example, the compounds may be labeled with isotopes, such as for example, deuterium (2H) , tritium (3H) , iodine-125 (125I) or carbon-14 (14C) . Isotopic substitution with 2H, 11C, 13C, 14C, 15C, 12N, 13N, 15N, 16N, 16O, 17O, 14F, 15F, 16F, 17F, 18F, 33S, 34S, 35S, 36S, 35Cl, 37Cl, 79Br, 81Br, 125I are all contemplated. In some embodiments, isotopic substitution with 18F is contemplated. All isotopic variations of the compounds of the present invention, whether radioactive or not, are encompassed within the scope of the present invention.
In certain embodiments, the compounds disclosed herein have some or all of the 1H atoms replaced with 2H atoms. The methods of synthesis for deuterium-containing compounds are known in the art and include, by way of non-limiting example only, the following synthetic methods.
Deuterium substituted compounds are synthesized using various methods such as described in: Dean, Dennis C.; Editor. Recent Advances in the Synthesis and Applications of Radiolabeled Compounds for Drug Discovery and Development. [Curr., Pharm. Des., 2000; 6 (10) ] 2000, 110 pp; George W.; Varma, Rajender S. The Synthesis of Radiolabeled Compounds via Organometallic Intermediates, Tetrahedron, 1989, 45 (21) , 6601-21; and Evans, E. Anthony. Synthesis of radiolabeled compounds, J. Radioanal. Chem., 1981, 64 (1-2) , 9-32.
Deuterated starting materials are readily available and are subjected to the synthetic methods described herein to provide for the synthesis of deuterium-containing compounds. Large numbers of deuterium-containing reagents and building blocks are available commercially from chemical vendors, such as Aldrich Chemical Co.
Deuterium-transfer reagents suitable for use in nucleophilic substitution reactions, such as iodomethane-d3 (CD3I) , are readily available and may be employed to transfer a deuterium-substituted carbon atom under nucleophilic substitution reaction conditions to the reaction substrate. The use of CD3I is illustrated, by way of example only, in the reaction schemes below.
Deuterium-transfer reagents, such as lithium aluminum deuteride (LiAlD4) , are employed to transfer deuterium under reducing conditions to the reaction substrate. The use of LiAlD4 is illustrated, by way of example only, in the reaction schemes below.
Deuterium gas and palladium catalyst are employed to reduce unsaturated carbon-carbon linkages and to perform a reductive substitution of aryl carbon-halogen bonds as illustrated, by way of example only, in the reaction schemes below.
In one embodiment, the compounds disclosed herein contain one deuterium atom. In another embodiment, the compounds disclosed herein contain two deuterium atoms. In another embodiment, the compounds disclosed herein contain three deuterium atoms. In another embodiment, the compounds disclosed herein contain four deuterium atoms. In another embodiment, the compounds disclosed herein contain five deuterium atoms. In another embodiment, the compounds disclosed herein contain six deuterium atoms. In another embodiment, the compounds disclosed herein contain more than six deuterium atoms. In another embodiment, the compound disclosed herein is fully substituted with deuterium atoms and contains no non-exchangeable 1H hydrogen atoms. In one embodiment, the level of deuterium incorporation is determined by synthetic methods in which a deuterated synthetic building block is used as a starting material.
"Pharmaceutically acceptable salt" includes both acid and base addition salts. A pharmaceutically acceptable salt of any one of the heteroaromatic WEE1 inhibitory compounds described herein is intended to encompass any and all pharmaceutically suitable salt forms. Preferred pharmaceutically acceptable salts of the compounds described herein are pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.
"Pharmaceutically acceptable acid addition salt" refers to those salts which retain the biological effectiveness and properties of the free bases, which are not biologically or otherwise undesirable, and which are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid, hydrofluoric acid, phosphorous acid, and the like. Also included are salts that are formed with organic acids such as aliphatic mono-and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, alkanedioic acids, aromatic acids, aliphatic and. aromatic sulfonic acids, etc. and include, for example, acetic acid, trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Exemplary salts thus include sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, nitrates, phosphates, monohydrogenphosphates, dihydrogenphosphates, metaphosphates, pyrophosphates, chlorides, bromides, iodides, acetates, trifluoroacetates, propionates, caprylates, isobutyrates, oxalates,
malonates, succinate suberates, sebacates, fumarates, maleates, mandelates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, phthalates, benzenesulfonates, toluenesulfonates, phenylacetates, citrates, lactates, malates, tartrates, methanesulfonates, and the like. Also contemplated are salts of amino acids, such as arginates, gluconates, and galacturonates (see, for example, Berge S.M. et al., "Pharmaceutical Salts, " Journal of Pharmaceutical Science, 66: 1-19 (1997) ) . Acid addition salts of basic compounds are, in some embodiments, prepared by contacting the free base forms with a sufficient amount of the desired acid to produce the salt according to methods and techniques with which a skilled artisan is familiar.
"Pharmaceutically acceptable base addition salt" refers to those salts that retain the biological effectiveness and properties of the free acids, which are not biologically or otherwise undesirable. These salts are prepared from addition of an inorganic base or an organic base to the free acid. Pharmaceutically acceptable base addition salts are, in some embodiments, formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum salts and the like. Salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, for example, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, N, N-dibenzylethylenediamine, chloroprocaine, hydrabamine, choline, betaine, ethylenediamine, ethylenedianiline, N-methylglucamine, glucosamine, methylglucamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins and the like. See Berge et al., supra.
"Pharmaceutically acceptable solvate" refers to a composition of matter that is the solvent addition form. In some embodiments, solvates contain either stoichiometric or non-stoichiometric amounts of a solvent, and are formed during the process of making with pharmaceutically acceptable solvents such as water, ethanol, and the like. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of compounds described herein are conveniently prepared or formed during the processes described herein. The compounds provided herein optionally exist in either unsolvated as well as solvated forms.
The term “subject” or “patient” encompasses mammals. Examples of mammals include, but are not limited to, any member of the Mammalian class: humans, non-human primates such as chimpanzees, and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, swine; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents, such as rats, mice and guinea pigs, and the like. In one aspect, the mammal is a human.
"Prodrug" refers to a compound that undergoes biotransformation before exhibiting its pharmacological effects. Prodrugs can thus be viewed as drugs containing specialized protective groups used in a transient manner to alter pharmacological properties in the parent molecule.
As used herein, “treatment” or “treating, ” or “palliating” or “ameliorating” are used interchangeably. These terms refer to an approach for obtaining beneficial or desired results including but not limited to therapeutic benefit and/or a prophylactic benefit. By “therapeutic benefit” is meant eradication or amelioration of the underlying disorder being treated. Also, a therapeutic benefit is achieved with the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disorder such that an improvement is observed in the patient, notwithstanding that the patient is still afflicted with the underlying disorder. For prophylactic benefit, the compositions are, in some embodiments, administered to a patient at risk of developing a particular disease, or to a patient reporting one or more of the physiological symptoms of a disease, even though a diagnosis of this disease has not been made.
WEE1 Kinase
WEE1 is a kinase belonging to the Ser/Thr family of protein kinases and is a key regulator of cell cycle progression. The WEE1 kinase is encoded by the WEE1 gene. WEE1 is an important regulator of the G2/M cell cycle checkpoint. WEE1 mediates cell-cycle arrest by regulating the phosphorylation of cyclin-dependent kinase 1 (Cdk1) . WEE1 inhibits Cdk1 by phosphorylating it on two different sites, Tyr15 and Thr14. Phosphorylated Cdk1 has reduced kinase activity and therefore prevents entry into mitosis. As a cell progress through the cell cycle, WEE1 is inactivated and degraded, and the cell is allowed to enter mitosis. DNA damage caused by, for example, genomic instability, errors in DNA synthesis, or DNA damaging agents, activates p53 and reduces WEE1 degradation thereby arresting the cell cycle and preventing progression into mitosis. Cancer cells are often deficient in p53 signaling, display genomic instability, and rely on the WEE1 and G2/M cell cycle checkpoint to avoid mitotic catastrophe due to DNA damage. Inhibition or degradation of WEE1 sensitizes cancers to DNA-damaging therapies. Additional information may be found in a) LL Parker, and H Piwnica-Worms. Inactivation of the p34cdc2-cyclin B complex by the human WEE1 tyrosine kinase. Science. 1992 Sep 25; 257 (5078) : 1955-1957, b) N Watanabe, M Broome, and T Hunter. Regulation of the human WEE1Hu CDK tyrosine 15-kinase during the cell cycle. EMBO J. 1995 May 1; 14 (9) : 1878-1891, c) CH McGowan, and P Russell. Cell cycle regulation of human WEE1. EMBO J. 1995 May 15; 14 (10) : 2166-2175, and d) Philip C De Witt Hamer, Shahryar E Mir, David Noske, Cornelis J F Van Noorden, Tom Wurdinger. WEE1 kinase targeting combined with DNA-damaging cancer therapy catalyzes mitotic catastrophe. Clin Cancer Res. 1022 Jul 1; 17 (13) , 4200–4207, each of which is incorporated herein by reference.
Selective Protein Degradation
The levels of proteins within a cell are determined by both the rate of protein synthesis and the rate of protein degradation. In eukaryotic cells two pathways exist for selective protein degradation, the ubiquitin-proteasome pathway and the lysosomal proteolysis pathway. In general, selective protein degradation is mediated by the presence of a recruitment motif which promotes binding of degradation proteins, such as proteasomal degradation proteins, or proteins associated with ubiquitin-proteasome pathway. Recruitment motifs include E3 ligase recognition agents and proteasome recognition agents. Conjugation of recruitment motifs with high affinity ligands for WEE1 kinase provides compounds capable of selectively directing pathways for protein degradation to the WEE1 kinase protein itself. Such an outcome will reduce levels of WEE1 kinase activity.
Heteroaromatic WEE1 degradation compounds
In one aspect, provided herein is a heteroaromatic WEE1 degradation compound having the general formula provided below:
wherein the WEE1 kinase affinity motif is a molecular construct having high affinity for the WEE1 kinase protein independent of the linking motif and/or the recruitment motif, the linking motif is a molecular construct providing a covalent bond to both the WEE1 kinase affinity motif and the recruitment motif, and the recruitment motif is a molecular construct having the ability to selectively target and recruit protein degradation.
Recruitment motif
Recruitment motifs include E3 ligase recognition agents and proteasome recognition agents. In some embodiments the recruitment motif is derived from, VHL ligand, nutlin, bestatin, HIF-1α –VHL binding peptide, hydroxy proline-HIF-1α-VHL binding peptide, SCFb-TRCP targeting peptide or an inhibitor of apoptosis protein ligand.
In some embodiments, the recruitment motif is selected from a molecular construct related to VHL ligand as illustrated below:
In some embodiments, the recruitment motif is selected from a molecular construct related to VHL ligand as illustrated below:
with the point of bonding to the linking motif as indicated by the wavy bond. In some embodiments, the recruitment motif is
In some embodiments, the recruitment motif is In some embodiments, the recruitment motif is
The terms “VHL ligand” , “VHL binder” , and “VHL E3 ubiquitin ligase binder” are used herein interchangeably to refer to a compound or motif that binds Von Hippel–Lindau tumor suppressor (VHL) .
Linking Motif
The linking motif is a molecular construct providing a covalent bond to both the WEE1 kinase affinity motif and the recruitment motif. In some embodiments, the linking motif comprises a cyclic moiety. In some embodiments, the linking motif comprises an acyclic moiety. In some embodiments, the linking motif comprises an unsaturated moiety. In some embodiments, the linking motif comprises between 4 and 50 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises between 4 and 20 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises between 4 and 25 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises between 4 and 30 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises between 6 and 18 atoms. In some embodiments, the linking motif comprises between 5 and 10 non-hydrogen atoms in a linear sequence. In some embodiments, the linking motif comprises 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 non-hydrogen atoms in a linear sequence.
In some embodiments, the linking motif is represented by the following formula:
-Ak1-Z1-Ak2-Z2-Ak3-*
wherein,
Ak1 is selected from - (CR5R6) k-;
Ak2 is selected from - (CR5R6) m-;
Ak3 is selected from - (CR5R6) n-;
each of k, m, and n is selected from 0 to 10;
R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;
Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;
R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced; and
wherein at least one of Z1 and Z2 is not bond and the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif is a polyethylene glycol containing moiety.
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is selected from – (CR5R6) n-O– (CR5R6) m-*, wherein n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk. In some further embodiments, said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) , and said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) .
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from
- (CR5R6) n-NR7– (CR5R6) m-*, wherein n is selected from 0 to 10, m is selected from 0 to 10, R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced; and the point of bonding to the VHL ligand indicated by the asterisk. In some embodiments, n is selected from 1 to 10, m is selected from 1 to 10. In some further embodiments, said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) , and said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) . In some embodiments, said n is selected from 1 to 7, and said m is selected from 0-3. In some embodiments, R7 is hydrogen or methyl.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*wherein k is selected from 1 to 10, n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk. In some further embodiments, said k is selected from 1 to 5, said n is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) , and said m is selected from 1 to 5 (e.g. 1, 2, 3, 4 or 5) .
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
, wherein X, X1, and X2 at each instance are independently CH or N and wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif the -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk.
In some embodiments, the linking motif -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein each R7 is independently hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced , wherein the point of bonding to the VHL ligand is indicated by the asterisk. In some embodiments, R7 is hydrogen or methyl. In some embodiments, k is1, 2, 3, 4, 5, 6, or 7,
and m is 0, 1, 2, 3, 4, or 5. In some embodiments, k is1, 2, 3, 4, 5, 6, or 7, and m is 0. In some embodiments, k is1, 2, 3, 4, or 5 and m is 1, 2, 3, 4, or 5. In some embodiments, is
In some embodiments, the linking motif the linking motif is a polyethylene glycol selected from– (O-CH2-CH2) n-O-and – (O-CH2-CH2) n-N (R23) -*, wherein n is selected from 1 to 20, and R23 is optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced; and wherein the point of bonding to the VHL ligand is indicated by the asterisk. In some embodiments, n is 1, 2, 3, 4, or 5
WEE1 Kinase Affinity Motif
Adavosertib (MK-1775, AZD1775) is a potent and selective Wee1 inhibitor with IC50 of 5.2 nM in a cell-free assay as described in Hirai et al., “Small-molecule inhibition of Wee1 kinase by MK-1775 selectively sensitizes p53-deficient tumor cells to DNA-damaging agents” Mol. Can. Therap. 2009. DOI: 10.1158/1535-7163. MCT-09-0463 which is incorporated herein by reference.
ZN-c3 also is a potent and selective Wee1 inhibitor with IC50 of 3.8 nM in a cell-free assay as described in Huang et al., “Discovery of ZN-c3, a Highly Potent and Selective Wee1 inhibitor Undergoing Evaluation in Clinical Trials for the Treatment of Cancer” J. Med. Chem. 2021. DOI: 10.1021/acs. jmedchem. 1c01121which is incorporated herein by reference.
Heteroaromatic WEE1 degradation compounds
In some embodiments, provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I) :
wherein,
R1, R2, R3, and R4 are each independently selected from hydrogen, optionally substituted straight or branched C1-6 alkyl, optionally substituted C1-3 alkoxy, halogen, amino, hydroxyl, and cyano; or
R1 and R2, R2 and R3, or R3 and R4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring;
L is -Ak1-Z1-Ak2-Z2-Ak3-*;
Ak1 is selected from - (CR5R6) k-;
Ak2 is selected from - (CR5R6) m-;
Ak3 is selected from - (CR5R6) n-;
each of k, m, and n is selected from 0 to 10
R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;
Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;
R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced;
wherein at least one of Z1 and Z2 is not bond; or
L is a polyethylene glycol containing moiety; and
wherein the point of bonding to the VHL ligand is indicated by the asterisk; and
U is a VHL ligand.
In some embodiments, provided herein is a compound of Formula (I) , or a pharmaceutically acceptable salt or solvate thereof.
In some embodiments of Formula (I) , R1, R2, R3, and R4 are each independently selected from hydrogen, optionally substituted straight or branched C1-6 alkyl, optionally substituted C1-3 alkoxy, halogen, amino, hydroxyl, and cyano, wherein the alkyl is optionally substituted with
hydroxy. In some embodiments, R1, R2, R3, and R4 are each independently selected from hydrogen, optionally substituted straight or branched C1-6 alkyl, optionally substituted C1-3 alkoxy, and halogen, wherein the alkyl is optionally substituted with hydroxyl. In some embodiments, R1, R2, R3, and R4 are each independently selected from hydrogen and optionally substituted straight or branched C1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy. In some embodiments, R1, is optionally substituted branched C1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy. In some embodiments, R1 is optionally substituted branched C1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy and R2, R3, and R4 are each hydrogen. In some embodiments, R1 is
In some embodiments of Formula (I) , R1 and R2, R2 and R3, or R3 and R4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring, wherein the cycloalkyl and heterocyclyl are optionally substituted with one or more substituents selected from C1-6 alkyl and hydroxyl. In some embodiments, R1 and R2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocyclyl ring, wherein the cycloalkyl and heterocyclyl are optionally substituted with one or more substituents selected from C1-6 alkyl and hydroxyl, and R3 and R4 are hydrogen. In some embodiments, R1 and R2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl. In some embodiments, R1 and R2 taken together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl, wherein the cycloalkyl is optionally substituted with one or more substituents selected from C1-6 alkyl and hydroxyl. In some embodiments, R1 and R2 taken together with the intervening atoms to which they are attached form a substituted 5-6 membered cycloalkyl, wherein the cycloalkyl is substituted with C1-6 alkyl and hydroxyl. In some embodiments, R1 and R2 taken together with the intervening atoms to which they are attached form
In some embodiments, provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I-a) :
wherein,
L is -Ak1-Z1-Ak2-Z2-Ak3-*;
Ak1 is selected from - (CR5R6) k-;
Ak2 is selected from - (CR5R6) m-;
Ak3 is selected from - (CR5R6) n-;
each of k, m, and n is selected from 0 to 10
R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;
Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;
R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced;
wherein at least one of Z1 and Z2 is not bond; or
L is a polyethylene glycol containing moiety; and
wherein the point of bonding to the VHL ligand is indicated by the asterisk; and
U is a VHL ligand.
In some embodiments, provided herein is a compound of Formula (I-a) , or a pharmaceutically acceptable salt or solvate thereof.
In some embodiments, provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I-b) :
wherein,
L is -Ak1-Z1-Ak2-Z2-Ak3-*;
Ak1 is selected from - (CR5R6) k-;
Ak2 is selected from - (CR5R6) m-;
Ak3 is selected from - (CR5R6) n-;
each of k, m, and n is selected from 0 to 10
R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;
Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;
R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced;
wherein at least one of Z1 and Z2 is not bond; or
L is a polyethylene glycol containing moiety; and
wherein the point of bonding to the VHL ligand is indicated by the asterisk; and
U is a VHL ligand.
In some embodiments, provided herein is a compound of Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof.
In some embodiments of Formula (I) , Formula (I-a) , and Formula (I-b) , -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O–
(CR5R6) m-*, wherein n is 4, and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is - (CR5R6) n-NR7– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–
– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isand each R7 is independently hydrogen or C1-3 alkyl. -In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isand each R7 is independently hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein k is 1-7, m is 0-5, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*iswherein k is 1-6, m is 0-3, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein k is 1-6, m is 0, and R7 is hydrogen or methyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, hydroxyl, C1-3 alkyl, and C1-3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R5 and R6 are both hydrogen. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I-a)
wherein,
U is
L is -Ak1-Z1-Ak2-Z2-Ak3-*. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n
is 2 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4, and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is - (CR5R6) n-NR7– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is –
(CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is and each R7 is independently hydrogen or C1-3 alkyl. -In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isand each R7 is independently hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein k is 1-7, m is 0-5, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*iswherein k is 1-6, m is 0-3, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein k is 1-6, m is 0, and R7 is hydrogen or methyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is , wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, hydroxyl, C1-3 alkyl, and C1-3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R5 and R6 are both hydrogen. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some
embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is, In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some
embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, provided herein is a compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I-b)
wherein,
U is
L is -Ak1-Z1-Ak2-Z2-Ak3-*. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5, or 6 and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 1, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2
and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 2, and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 3 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4, and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 4 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 5 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 3. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 4. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is 6 and m is 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is - (CR5R6) n-NR7– (CR5R6) m-*, wherein n is 1, 2, 3, 4, 5 or 6, and m is 1, 2, 3, 4, or 5. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, 2, 3, or 4, n is 1, 2, or 3, and m is 1 or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O–
(CR5R6) m-*, wherein k is 1, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 1. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 1, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 2, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 1, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 2, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 3, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is 4, n is 3, and m is 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is and each R7 is independently hydrogen or C1-3 alkyl. -In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isand each R7 is independently
hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein k is 1-7, m is 0-5, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*iswherein k is 1-6, m is 0-3, and R7 is hydrogen or C1-3 alkyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein k is 1-6, m is 0, and R7 is hydrogen or methyl. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is, wherein X, X1, and X2 are each independently CH or N and wherein each k is independently 0, 1 or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is wherein X1, X2, X3, and X4 are each independently CH or N and wherein each k is independently 0, 1 or 2, each m is independently 0, 1, or 2, and each n is independently 0, 1, or 2. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, hydroxyl, C1-3 alkyl, and C1-3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, halogen, C1-3 alkyl, and C3 cycloalkyl. In some embodiments, R5 and R6 are independently selected from hydrogen, fluoro, and methyl. In some embodiments, R5 and R6 are both hydrogen. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
wherein X1 and X2 are each independently CH or N and wherein each k is independently 0, 1, or 2, and each m is independently 0, 1, or 2. In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is, In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*isIn some embodiments, -Ak1-Z1-Ak2-Z2-Ak3-*is,
In some embodiments, the compounds as described herein have a structure provided in Table 1, Table 2, or Table 3. In some embodiments, the compounds as described herein have a structure provided in Table 1.
Table 1
In some embodiments, compounds as described herein has a structure provided in Table 2.
Table 2
In some embodiments, compounds as described herein has a structure provided in Table 3.
Table 3
Preparation of Compounds
The compounds used in the reactions described herein are made according to organic synthesis techniques known to those skilled in this art, starting from commercially available chemicals and/or from compounds described in the chemical literature. "Commercially available chemicals" are obtained from standard commercial sources including Acros Organics (Pittsburgh, PA) , Aldrich Chemical (Milwaukee, WI, including Sigma Chemical and Fluka) , Apin Chemicals Ltd. (Milton Park, UK) , Avocado Research (Lancashire, U.K. ) , BDH Inc. (Toronto, Canada) , Bionet (Cornwall, U.K. ) , Chemservice Inc. (West Chester, PA) , Crescent Chemical Co. (Hauppauge, NY) , Eastman Organic Chemicals, Eastman Kodak Company (Rochester, NY) , Fisher Scientific Co. (Pittsburgh, PA) , Fisons Chemicals (Leicestershire, UK) , Frontier Scientific (Logan, UT) , ICN Biomedicals, Inc. (Costa Mesa, CA) , Key Organics (Cornwall, U.K. ) , Lancaster Synthesis (Windham, NH) , Maybridge Chemical Co. Ltd. (Cornwall, U.K. ) , Parish Chemical Co. (Orem, UT) , Pfaltz &Bauer, Inc. (Waterbury, CN) , Polyorganix (Houston, TX) , Pierce Chemical Co. (Rockford, IL) , Riedel de Haen AG (Hanover, Germany) , Spectrum Quality Product, Inc. (New Brunswick, NJ) , TCI America (Portland, OR) , Trans World Chemicals, Inc. (Rockville, MD) , and Wako Chemicals USA, Inc. (Richmond, VA) .
Suitable reference books and treatise that detail the synthesis of reactants useful in the preparation of compounds described herein, or provide references to articles that describe the preparation, include for example, "Synthetic Organic Chemistry" , John Wiley &Sons, Inc., New York; S.R. Sandler et al., "Organic Functional Group Preparations, " 2nd Ed., Academic Press, New York, 1983; H.O. House, "Modern Synthetic Reactions" , 2nd Ed., W.A. Benjamin, Inc. Menlo Park, Calif. 1972; T.L. Gilchrist, "Heterocyclic Chemistry" , 2nd Ed., John Wiley &Sons, New York, 1992; J. March, "Advanced Organic Chemistry: Reactions, Mechanisms and Structure" , 4th Ed., Wiley-Interscience, New York, 1992. Additional suitable reference books and treatise that detail the synthesis of reactants useful in the preparation of compounds described
herein, or provide references to articles that describe the preparation, include for example, Fuhrhop, J. and Penzlin G. "Organic Synthesis: Concepts, Methods, Starting Materials" , Second, Revised and Enlarged Edition (1994) John Wiley &Sons ISBN: 3-527-29074-5; Hoffman, R. V. "Organic Chemistry, An Intermediate Text" (1996) Oxford University Press, ISBN 0-19-509618-5; Larock, R.C. "Comprehensive Organic Transformations: A Guide to Functional Group Preparations" 2nd Edition (1999) Wiley-VCH, ISBN: 0-471-19031-4; March, J. "Advanced Organic Chemistry: Reactions, Mechanisms, and Structure" 4th Edition (1992) John Wiley &Sons, ISBN: 0-471-60180-2; Otera, J. (editor) "Modern Carbonyl Chemistry" (2000) Wiley-VCH, ISBN: 3-527-29871-1; Patai, S. "Patai's 1992 Guide to the Chemistry of Functional Groups" (1992) Interscience ISBN: 0-471-93022-9; Solomons, T.W.G. "Organic Chemistry" 7th Edition (2000) John Wiley &Sons, ISBN: 0-471-19095-0; Stowell, J.C., "Intermediate Organic Chemistry" 2nd Edition (1993) Wiley-Interscience, ISBN: 0-471-57456-2; "Industrial Organic Chemicals: Starting Materials and Intermediates: An Ullmann's Encyclopedia" (1999) John Wiley &Sons, ISBN: 3-527-29645-X, in 8 volumes; "Organic Reactions" (1942-2000) John Wiley &Sons, in over 55 volumes; and "Chemistry of Functional Groups" John Wiley &Sons, in 73 volumes.
Specific and analogous reactants are optionally identified through the indices of known chemicals prepared by the Chemical Abstract Service of the American Chemical Society, which are available in most public and university libraries, as well as through on-line databases (contact the American Chemical Society, Washington, D.C. for more details) . Chemicals that are known but not commercially available in catalogs are optionally prepared by custom chemical synthesis houses, where many of the standard chemical supply houses (e.g., those listed above) provide custom synthesis services. A reference useful for the preparation and selection of pharmaceutical salts of the compounds described herein is P.H. Stahl &C.G. Wermuth "Handbook of Pharmaceutical Salts" , Verlag Helvetica Chimica Acta, Zurich, 2002.
Pharmaceutical Compositions
In certain embodiments, the heteroaromatic WEE1 degradation compound described (e.g., a compound of Formula (I) , Formula (I-a) , or Formula (I-b) ) herein is administered as a pure chemical. In other embodiments, the heteroaromatic WEE1 degradation compound described herein is combined with a pharmaceutically suitable or acceptable carrier (also referred to herein as a pharmaceutically suitable (or acceptable) excipient, physiologically suitable (or acceptable) excipient, or physiologically suitable (or acceptable) carrier) selected on the basis of a chosen route of administration and standard pharmaceutical practice as described, for example, in Remington: The Science and Practice of Pharmacy (Gennaro, 21st Ed. Mack Pub. Co., Easton, PA (2005) ) .
Provided herein is a pharmaceutical composition comprising at least one heteroaromatic WEE1 degradation compound as described herein (e.g., a compound of Formula (I) , Formula (I-a) , or Formula (I-b) ) , or a stereoisomer, pharmaceutically acceptable salt, hydrate, or solvate thereof, together with one or more pharmaceutically acceptable carriers. The carrier (s) (or excipient (s) ) is acceptable or suitable if the carrier is compatible with the other ingredients of the composition and not deleterious to the recipient (i.e., the subject or the patient) of the composition.
One embodiment provides a pharmaceutical composition comprising a pharmaceutically acceptable excipient and a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof.
One embodiment provides a method of preparing a pharmaceutical composition comprising mixing a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier.
In certain embodiments, the heteroaromatic WEE1 degradation compound as described by Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, is substantially pure, in that it contains less than about 5%, or less than about 1%, or less than about 0.1%, of other organic small molecules, such as unreacted intermediates or synthesis by-products that are created, for example, in one or more of the steps of a synthesis method.
Suitable oral dosage forms include, for example, tablets, pills, sachets, or capsules of hard or soft gelatin, methylcellulose or of another suitable material easily dissolved in the digestive tract. In some embodiments, suitable nontoxic solid carriers are used which include, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, talcum, cellulose, glucose, sucrose, magnesium carbonate, and the like. (See, e.g., Remington: The Science and Practice of Pharmacy (Gennaro, 21st Ed. Mack Pub. Co., Easton, PA (2005) ) .
In some embodiments, the heteroaromatic WEE1 degradation compound as described by Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, is formulated for administration by injection. In some instances, the injection formulation is an aqueous formulation. In some instances, the injection formulation is a non-aqueous formulation. In some instances, the injection formulation is an oil-based formulation, such as sesame oil, or the like.
The dose of the composition comprising at least one heteroaromatic WEE1 degradation compound as described herein (e.g., a compound of Formula (I) , Formula (I-a) , or Formula (I-b) ) differs depending upon the subject or patient's (e.g., human) condition. In some embodiments, such factors include general health status, age, and other factors.
Pharmaceutical compositions are administered in a manner appropriate to the disease to be treated (or prevented) . An appropriate dose and a suitable duration and frequency of
administration will be determined by such factors as the condition of the patient, the type and severity of the patient's disease, the particular form of the active ingredient, and the method of administration. In general, an appropriate dose and treatment regimen provides the composition (s) in an amount sufficient to provide therapeutic and/or prophylactic benefit (e.g., an improved clinical outcome, such as more frequent complete or partial remissions, or longer disease-free and/or overall survival, or a lessening of symptom severity. Optimal doses are generally determined using experimental models and/or clinical trials. The optimal dose depends upon the body mass, weight, or blood volume of the patient.
Methods of Treatment
One embodiment provides a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, for use in a method of treatment of the human or animal body.
One embodiment provides a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, for use in a method of treatment of cancer or neoplastic disease.
One embodiment provides a use of a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, in the manufacture of a medicament for the treatment of cancer or neoplastic disease.
In some embodiments, described herein is a method of treating cancer in a patient in need thereof comprising administering to the patient a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof. In some embodiments, described herein is a method of treating cancer in a patient in need thereof comprising administering to the patient a pharmaceutical composition comprising a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable excipient.
Provided herein is the method wherein the pharmaceutical composition is administered orally. Provided herein is the method wherein the pharmaceutical composition is administered by injection.
Other embodiments and uses will be apparent to one skilled in the art in light of the present disclosures. The following examples are provided merely as illustrative of various embodiments and shall not be construed to limit the invention in any way.
EXAMPLES
I. Chemical Synthesis
In some embodiments, the compounds disclosed herein are synthesized according to the
following examples. As used below, and throughout the description of the invention, the following abbreviations, unless otherwise indicated, shall be understood to have the following meanings:
℃ degrees Celsius
δH chemical shift in parts per million downfield from tetramethylsilane
DCM dichloromethane (CH2Cl2)
DMF dimethylformamide
DMSO dimethylsulfoxide
EA ethyl acetate
ESI electrospray ionization
Et ethyl
g gram (s)
h hour (s)
HPLC high performance liquid chromatography
Hz hertz
J coupling constant (in NMR spectrometry)
LCMS liquid chromatography mass spectrometry
μ micro
m multiplet (spectral) ; meter (s) ; milli
M molar
M+ parent molecular ion
Me methyl
MHz megahertz
min minute (s)
mol mole (s) ; molecular (as in mol wt)
mL milliliter
MS mass spectrometry
nm nanometer (s)
NMR nuclear magnetic resonance
pH potential of hydrogen; a measure of the acidity or basicity of an aqueous
solution
PE petroleum ether
RT room temperature
s singlet (spectral)
t triplet (spectral)
T temperature
TFA trifluoroacetic acid
THF tetrahydrofuran
DIEA diisopropylethyl amine
HATU O- (7-Azabenzotriazol-1-yl) -N, N, N, N-tetramethyl uranium
hexafluorophosphate
℃ degrees Celsius
δH chemical shift in parts per million downfield from tetramethylsilane
DCM dichloromethane (CH2Cl2)
DMF dimethylformamide
DMSO dimethylsulfoxide
EA ethyl acetate
ESI electrospray ionization
Et ethyl
g gram (s)
h hour (s)
HPLC high performance liquid chromatography
Hz hertz
J coupling constant (in NMR spectrometry)
LCMS liquid chromatography mass spectrometry
μ micro
m multiplet (spectral) ; meter (s) ; milli
M molar
M+ parent molecular ion
Me methyl
MHz megahertz
min minute (s)
mol mole (s) ; molecular (as in mol wt)
mL milliliter
MS mass spectrometry
nm nanometer (s)
NMR nuclear magnetic resonance
pH potential of hydrogen; a measure of the acidity or basicity of an aqueous
solution
PE petroleum ether
RT room temperature
s singlet (spectral)
t triplet (spectral)
T temperature
TFA trifluoroacetic acid
THF tetrahydrofuran
DIEA diisopropylethyl amine
HATU O- (7-Azabenzotriazol-1-yl) -N, N, N, N-tetramethyl uranium
hexafluorophosphate
Compound 1
Step 1. methyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) and methyl 4-bromobutanoate (119.05 mg, 0.658 mmol, 1.6 equiv. ) . The resulting mixture was stirred at 50 ℃ for 3 h. After cooling down to rt., the reaction was quenched with H2O (10 mL) , then extracted with DCM (3*10 mL) , washed with brine (4*30 mL) . The organic layers were combined and concentrated. The residue obtained was purified by TLC (1/14 MeOH/DCM) to afford methyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoate.
LC/MS: 587.50 [M+H] +.
Step 2. 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoic acid
To a solution of methyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoate (116 mg, 0.198 mmol, 1.00 equiv. ) in THF (3 mL) was added LiOH. H2O (41.48 mg, 0.990 mmol, 5 equiv. ) and H2O (3 mL) . The resulting mixture was stirred at 25 ℃ for 1 h. The pH value of the resulting mixture was adjusted to 6 by using 2N HCl solution. The resulting mixture was filtered to afford 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoic acid.
LC/MS: 573.35 [M+H] +.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 1)
To a solution of 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butanoic acid (98 mg, 0.171 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (155.85 mg, 1.539 mmol, 9 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (88.42 mg, 0.205 mmol, 1.2 equiv. ) (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (88.42 mg, 0.205 mmol, 1.2 equiv. ) , HATU (84.59 mg, 0.222 mmol, 1.3 equiv. ) . The mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3*15 mL) . The organic layers were combined, washed with brine (4*15 mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 30*150 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3+0.1%NH3. H2O, Mobile Phase B: CH3CN (20 %to 47%in 11 min, 47 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 20 %in 0.1 min, 20 %to 20 %in 1 min; Detector: UV 254 nm to afford Compound 1 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-
yl} amino) phenyl] piperazin-1-yl} butanamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 8.98 (s, 1H) , 8.83 (s, 1H) , 8.57 (t, J = 6.0 Hz, 1H) , 8.02 -8.07 (m, 1H) , 7.85 -7.91 (m, 1H) , 7.72 -7.78 (m, 1H) , 7.55 -7.59 (m, 3H) , 7.35 -7.47 (m, 4 H) , 6.92 (d, J = 9.2 Hz, 2H) , 5.60 -5.74 (m, 1H) , 5.33 (s, 1H) , 5.11 -5.17 (m, 1H) , 4.96 -5.03 (m, 1H) , 4.78 -4.88 (m, 1H) , 4.66 -4.72 (m, 2H) , 4.53 -4.60 (m, 1H) , 4.39 -4.49 (m, 2H) , 4.30 -4.40 (m, 1H) , 4.17 -4.30 (m, 1H) , 3.62 -3.73 (m, 2H) , 3.08 -3.14 (m, 4H) , 2.47 (s, 4H) , 2.45 (s, 3H) , 2.24 -2.33 (m, 3H) , 2.13 -2.24 (m, 1H) , 2.00 -2.09 (m, 1H) , 1.86 -1.97 (m, 1H) , 1.64 -1.76 (m, 2H) , 1.47 (s, 6H) , 0.95 (s, 9H) .
LC/MS: 985.50 [M+H] +.
Compound 2
Step 1. tert-butyl 6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (280 mg, 0.575 mmol) in DMF (5 mL) was added tert-butyl 6-bromohexanoate (158.98 mg, 0.633mmol) . The resulting mixture was stirred at 50 ℃ for 6 h. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted
with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (4 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-5%MeOH/DCM) to afford tert-butyl 6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanoate.
LCMS: 657.4 [M+H] +.
Step 2. 6- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} hexanoic acid
To a solution of tert-butyl 6- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} hexanoate (260 mg, 0.396 mmol) in DCM (5mL) was added TFA (1.5mL) . The resulting mixture was stirred at 25 ℃ for 1.5 hour. The reaction was concentrated to afford the 6- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} hexanoic acid.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (Compound 2)
To a solution of 6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanoic acid (240 mg, 0.400 mmol) in DMF (5 mL) was added TEA (121.28 mg, 1.200 mmol) , HATU (197.48 mg, 0.520 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (189.22 mg, 0.440 mmol) , The resulting mixture was stirred at rt 4 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /10: 1) then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3; Phase B: CH3CN (28%CH3CN up to 58%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 58%CH3CN in 0.1min, hold 28%in 0.9min) ; Detector, UV220 &254nm to afford 58.0 mg of Compound 2 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-
yl] hexanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.51 -8.57 (m, 1H) , 8.03 -8.18 (m, 1H) , 7.84 (d, J = 9.2 Hz, 1H) , 7.76 -7.77 (m, 1H) , 7.53 -7.63 (m, 3H) , 7.39 -7.44 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.60 -5.73 (m, 1H) , 5.31 (s, 1H) , 5.11 -5.12 (m, 1H) , 5.00 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 -4.73 (m, 2H) , 4.55 (d, J = 9.6 Hz, 1H) , 4.40 -4.48 (m, 2H) , 4.34 -4.38 (m, 1H) , 4.20 -4.26 (m, 1H) , 3.62 -3.71 (m, 2H) , 3.30 (s, 2H) , 3.10 (s, 4H) , 2.50 (s, 2H) , 2.47 (s, 3H) , 2.22 -2.32 (m, 3H) , 2.10 -2.20 (m, 1H) , 2.02 -2.04 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.39 -1.61 (m, 10H) , 1.23 -1.33 (m, 2H) , 0.93 (s, 9H) .
LCMS: 1013.5, [M+H] +.
Compound 3
Step 1. tert-butyl 7- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] heptanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (280 mg, 0.575 mmol) in DMF (5 mL) was added tert-butyl 6-bromohexanoate (167.86 mg, 0.633mmol) . The resulting mixture was stirred at
50 ℃ for 6 h. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (4 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-6%MeOH/DCM) to afford tert-butyl 6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexanoate.
LCMS: 671.4 [M+H] +.
Step 2. 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} heptanoic acid
To a solution of tert-butyl 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} heptanoate (280 mg, 0.418 mmol) in DCM (5mL) was added TFA (1.5mL) . The resulting mixture was stirred at 25℃ for 1.5 hour. The reaction was concentrated to afford the 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} heptanoic acid.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (7- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] heptanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (Compound 3)
To a solution of 7- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] heptanoic acid (240mg, 0.39mmol) in DMF (5mL) was added TEA (118.51mg, 1.170mmol) , HATU (192.98mg, 0.507mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (184.90 mg, 0.429 mmol) , The resulting mixture was stirred at rt 4h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /10: 1) then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3; Phase B: CH3CN (31%CH3CN up to 57%in 9 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 57%CH3CN in 0.1min, hold 31%in 0.9min) ; Detector, UV220 &254nm to afford 66.9 mg of Compound 3 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (7- [4- [4- ( [1- [6- (2-hydroxypropan-2-
yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] heptanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.51 -8.57 (m, 1H) , 8.03 -8.18 (m, 1H) , 7.84 (d, J = 9.2 Hz, 1H) , 7.76 -7.77 (m, 1H) , 7.53 -7.63 (m, 3H) , 7.39 -7.44 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.60 -5.73 (m, 1H) , 5.31 (s, 1H) , 5.11 -5.12 (m, 1H) , 5.00 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 -4.73 (m, 2H) , 4.55 (d, J = 9.6 Hz, 1H) , 4.40 -4.48 (m, 2H) , 4.34 -4.38 (m, 1H) , 4.20 -4.26 (m, 1H) , 3.62 -3.71 (m, 2H) , 3.30 (s, 2H) , 3.10 (s, 4H) , 2.50 (s, 2H) , 2.47 (s, 3H) , 2.22 -2.32 (m, 3H) , 2.10 -2.20 (m, 1H) , 2.02 -2.04 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.39 -1.61 (m, 10H) , 1.23 -1.33 (m, 4H) , 0.93 (s, 9H) .
LCMS: 1027.5 [M+H] +.
Compound 4
Step 1. ethyl 4- (2-allyl-2- (tert-butoxycarbonyl) hydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate
To a solution of ethyl 4-chloro-2- (methylthio) pyrimidine-5-carboxylate (12.5 g, 53.722 mmol) , tert-butyl 1-allylhydrazine-1-carboxylate (9.25 g, 53.722 mmol) and DIEA (23.39 mL, 134.305 mmol) in THF (120 mL) was added. The resulting mixture was stirred at 70 ℃ 18 h. After cooling
down to rt. The reaction was concentrated and purified by silica gel chromatography (0-10%EtOAc/petroleum ether) to afford ethyl 4- (2-allyl-2- (tert-butoxycarbonyl) hydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate .
LC/MS: 369.2 [M+H] +.
Step 2. ethyl 4- (2-allylhydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate
To a solution of ethyl 4- (2-allyl-2- (tert-butoxycarbonyl) hydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate (8.51 g, 23.097 mmol) in DCM (20 mL) was added TFA (13 mL, 175.019 mmol) . The resulting mixture was stirred at ℃ for 1 h. The reaction was concentrated to afford ethyl 4- (2-allylhydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate.
LC/MS: 269.1 [M+H] +.
Step 3. 2-allyl-6- (methylthio) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of ethyl 4- (2-allylhydrazineyl) -2- (methylthio) pyrimidine-5-carboxylate (6.123 g, 22.818 mmol) in EtOH (60 mL) , the mixture was stirred at 0℃, then 6M NaOH (120 mL) was added. The resulting mixture was stirred at 25 ℃ 30 min. The pH value of the resulting mixture was adjusted to 1 used concentrated hydrochloric acid. After removing ethanol by concentration and filtration, the solid obtained were washed with ethyl ether and afford 2-allyl-6- (methylthio) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 223.1 [M+H] +.
Step 4. 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- (methylsulfanyl) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of dodec-To a solution of 6- (methylsulfanyl) -2- (prop-2-en-1-yl) -1H-pyrazolo [3, 4-d] pyrimidin-3-one (4.20 g, 18.896 mmol) in 1, 4-dioxane (50 mL) was added 2- (6-bromopyridin-2-yl) propan-2-ol (5.31 g, 24.565 mmol) , then added K2CO3 (3.66 g, 26.454 mmol) and CuI (3.60 g, 18.903 mmol) finally added methyl [2- (methylamino) ethyl] amine (3.33 g, 37.775 mmol) . The resulting mixture was stirred at 100 ℃ under N2 protection overnight. After cooling down to rt, the reaction was quenched with H2O (100 mL) , then filtered. The resulting mixture was extracted with EtOAc (3 x 100 mL) . The combined organic layers were washed with brine (2 x 100 mL) , then concentrated under reduced pressure. The residue obtained was purified by silica gel
chromatography (35-45%EtOAc/petroleum ether) to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- (methylsulfanyl) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 358.1 [M+H] +.
Step 5. 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6-methanesulfinyl-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- (methylsulfanyl) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (4.48 g, 12.534 mmol) in DCM (100 mL) was added mCPBA (2.16 g, 12.517 mmol) at 0℃. The resulting mixture was stirred at rt 2 h. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with DCM (3 x 100 mL) , concentrated. The residue obtained was purified by silica gel chromatography (0-5%MeOH/DCM) to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6-methanesulfinyl-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 374.1 [M+H] +.
Step 6. tert-butyl 4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazine-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6-methanesulfinyl-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (4.5 g, 12.050 mmol) in toluene (80 mL) was added DIEA (4.67 g, 36.151 mmol) , tert-butyl 4- (4-aminophenyl) piperazine-1-carboxylate (3.34 g, 12.050 mmol) . The resulting mixture was stirred at 80 ℃ for 4 h. After cooling down to rt, the reaction was concentrated, dissolved in DCM then concentrated. The residue obtained was purified by silica gel chromatography (0-5%MeOH/DCM) to afford tert-butyl 4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazine-1-carboxylate.
LC/MS: 587.30 [M+H] +.
Step 7. tert-butyl 4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazine-1-carboxylate
To a solution of tert-butyl 4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazine-1-carboxylate (4.80 g, 8.181 mmol) in DCM (8 mL) was added TFA (6 mL) . The resulting mixture was stirred at rt 1.5 h. The resulting
mixture was concentrated, dissolved in DMF (6mL) . The residue was purified by reverse flash chromatography with the following conditions: column: C18 silica gel; mobile phase: MeCN in water, 0%to 50%gradient in 30 min; detector: UV254, 220 nm to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 487.25 [M+H] +.
Step 8. ethyl 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (450 mg, 0.925 mmol) in DMF (5 mL) was added ethyl 8-bromooctanoate (255.51 mg, 1.018 mmol) . The resulting mixture was stirred at rt overnight. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted with EtOAc (3 x 100 mL) . The combined organic layers were washed with brine (5 x 100 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-5%MeOH/DCM) to afford ethyl 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoate.
LC/MS: 657.38 [M+H] +.
Step 9. 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoic acid
To a solution of ethyl 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoate (250 mg, 0.381 mmol) in DCM (5 mL) was added TFA (1.5 mL) . The resulting mixture was stirred at rt 1.5 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) . The organic layers were washed with brine (5 x 30 mL) then concentrated. The residue obtained was purified by silica gel chromatography (10-20%MeOH/DCM) to afford 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoic acid.
LC/MS: 629.4 [M+H] +.
Step 10. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (Compound 4)
To a solution of 8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanoic acid (250 mg, 0.398 mmol) in DMF (5 mL) was added TEA (120.70 mg, 1.193 mmol) , HATU (196.53 mg, 0.517 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (171.19 mg, 0.398 mmol) , The resulting mixture was stirred at rt 4 h. The reaction was quenched with H2O (10 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by silica gel chromatography (5-15%MeOH/) then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (42%CH3CN up to 62%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 62%CH3CN in 0.1min, hold 42%in 0.9min) ; Detector, UV220 &254nm to afford 110.3 mg of Compound 4 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (8- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] octanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 8.98 (s, 1H) , 8.83 (s, 1H) , 8.53 -8.61 (m, 1H) , 7.93 -8.16 (m, 1H) , 7.85 (d, J = 6.9 Hz, 1H) , 7.71 -7.81 (m, 1H) , 7.53 -7.68 (m, 3H) , 7.31 -7.48 (m, 4H) , 6.92 (d, J = 6.6 Hz, 2H) , 5.59 -5.84 (m, 1H) , 5.33 (s, 1H) , 5.13 (s, 1H) , 5.00 (d, J = 6.0 Hz, 1H) , 4.82 (d, J = 7.2 Hz, 1H) , 4.68 (s, 2H) , 4.55 (d, J = 6.0 Hz, 1H) , 4.39 -4.49 (m, 2H) , 4.35 (brs, 1H) , 4.17 -4.28 (m, 1H) , 3.61 -3.74 (m, 2H) , 3.30 (s, 2H) , 3.08 (brs, 4H) , 2.41-2.49 (m, 5H) , 2.22 -2.36 (m, 3H) , 1.99 -2.17 (m, 2H) , 1.84 -1.96 (m, 1H) , 1.46 (s, 10H) , 1.27 (s, 6H) , 0.93 (s, 9H) .
LC/MS: 1041.55 [M+H] +.
Compound 5
Step 1. ethyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoate
To a solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (450 mg, 0.925 mmol) in DMF (10 mL) was added K2CO3 (638 mg, 4.629 mmol) , and ethyl 9-bromononanoate (268 mg, 1.017 mmol) . The resulting mixture was stirred for 6 h at 50℃. The reaction was monitored by LCMS. Upon completion, the reaction was quenched by H2O (15 ml) , extracted by EA (15 ml *3) and washed by brine (4*20) . Then the organic layer was concentrated under vacuum, the residue was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford ethyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoate.
LCMS: 671.6 [M+H] +.
Step 2. 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoic acid
To a solution of ethyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoate (300 mg, 0.447 mmol) in THF (3 mL) and H2O (3 ml) was added LiOH (54 mg, 2.238 mmol) . The resulting mixture was stirred for 16 h at 25℃. The reaction was monitored by LCMS. Upon completion, the reaction was adjusted to pH 6 with 2N HCl, extracted with EA (15 ml *3) and dried with Na2SO4. Then the organic layer was concentrated under vacuum to afford 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoic acid.
LCMS: 643.6 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 5)
9- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] nonanoic acid (250mg, 0.389mmol, 1.0equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (167.46 mg, 0.389 mmol, 1.0 equiv. ) , HATU (192.24 mg, 0.506 mmol, 1.3 equiv. ) and TEA (196.77 mg, 1.945 mmol, 5.0 equiv. ) were dissolved in DMF (10 ml) . The mixture was stirred at room temperature overnight. 30 ml of Water was added. The solution was extracted with EA (100 ml*3) , washed with (80 ml*3) and concentrated. The crude was purified by TLC with DCM/MeOH (10/1) , then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (50%CH3CN up to 66%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 66%CH3CN in 0.1min, hold 50%in 0.9min) ; Detector, UV220 &254nm. This resulted in 124.9 mg of Compound 5 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (9- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] nonanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 8.98 (s, 1H) , 8.83 (s, 1H) , 8.53 -8.61 (m, 1H) , 7.93 -8.16 (m, 1H) , 7.85 (d, J = 6.9 Hz, 1H) , 7.71 -7.81 (m, 1H) , 7.53 -7.68 (m, 3H) , 7.31 -7.48 (m, 4H) , 6.92 (d, J = 6.6 Hz, 2H) , 5.59 -5.84 (m, 1H) , 5.33 (s, 1H) , 5.13 (s, 1H) , 5.00 (d, J = 6.0 Hz, 1H) , 4.82 (d, J = 7.2 Hz, 1H) , 4.68 (s, 2H) , 4.55 (d, J = 6.0 Hz, 1H) , 4.39 -4.49 (m, 2H) , 4.35 (brs, 1 H) , 4.17 -4.28 (m, 1H) , 3.61 -3.74 (m, 2H) , 3.31 (s, 2H) , 3.09 (brs, 4H) , 2.41-2.49 (m, 5H) , 2.21 -2.37 (m, 3H) , 1.99 -2.17 (m, 2H) , 1.84 -1.96 (m, 1H) , 1.46 (s, 10H) , 1.27 (s, 8H) , 0.94 (s, 9H) .
LCMS: 1055.6 [M+H] +.
Compound 6
Step 1. tert-butyl 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoate
To a solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (450 mg, 0.925 mmol) in DMF (10 mL) was added K2CO3 (638 mg, 4.629 mmol) , and tert-butyl 10-bromodecanoate (283 mg, 1.017 mmol) . The resulting mixture was stirred for 6 h at 50℃. The reaction was monitored by LCMS. Upon completion, the reaction was quenched by H2O (15 ml) , extracted by EA (15 ml *3) and washed by brine (4*20) . Then the organic layer was concentrated under vacuum, the residue was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford tert-butyl 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoate.
LCMS: 713.6 [M+H] +.
Step 2. 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoic acid
To a solution of e tert-butyl 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoate (300 mg, 0.421 mmol) in DCM (5 ml) was added TFA (1.5 ml) . The resulting mixture was stirred for 2 h at 25℃. The reaction was monitored by LCMS. Upon completion, the reaction was concentrated under vacuum to afford 10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanoic acid.
LCMS: 657.6 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (10- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) decanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 6)
10- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] decanoic acid (250 mg, 0.381 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (163.88 mg, 0.381 mmol, 1.0 equiv. ) , HATU (188.14 mg, 0.495 mmol, 1.3 equiv. ) and TEA (192.57 mg, 1.905 mmol, 5.0 equiv. ) were dissolved in DMF (10 ml) . The mixture was stirred at room temperature for overnight. 30 ml Water was added. The solution was extracted with EA (100 ml*3) , washed with (80 ml*3) and concentrated. The crude product was purified by TLC with DCM/MeOH (10/1) , then purified by
Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (50%CH3CN up to 66%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 66%CH3CN in 0.1min, hold 50%in 0.9min) ; Detector, UV220 &254nm. This resulted in 144.3 mg of Compound 6 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (10- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] decanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 8.98 (s, 1H) , 8.83 (s, 1H) , 8.53 -8.61 (m, 1H) , 7.93 -8.16 (m, 1H) , 7.85 (d, J = 6.9 Hz, 1H) , 7.71 -7.81 (m, 1H) , 7.53 -7.68 (m, 3H) , 7.31 -7.48 (m, 4H) , 6.92 (d, J = 6.6 Hz, 2H) , 5.59 -5.84 (m, 1H) , 5.33 (s, 1H) , 5.13 (s, 1H) , 5.00 (d, J = 6.0 Hz, 1H) , 4.82 (d, J = 7.2 Hz, 1H) , 4.68 (s, 2H) , 4.55 (d, J = 6.0 Hz, 1H) , 4.39 -4.49 (m, 2H) , 4.35 (brs, 1 H) , 4.17 -4.28 (m, 1H) , 3.61 -3.76 (m, 2H) , 3.33 (s, 2H) , 3.09 (brs, 4H) , 2.41-2.47 (m, 5H) , 2.21 -2.37 (m, 3H) , 1.99 -2.17 (m, 2H) , 1.84 -1.96 (m, 1H) , 1.46 (s, 10H) , 1.26 (s, 10H) , 0.94 (s, 9H) .
LCMS: 1069.6 [M+H] +.
Compound 7
Step 1. methyl 11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanoate
To a solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (450 mg, 0.925 mmol) in DMF (10 mL) was added K2CO3 (638 mg, 4.629 mmol) , and methyl 11-bromoundecanoate (282 mg, 1.017 mmol) . The resulting mixture was stirred for 6 h at 50℃. The reaction was monitored by LCMS. On completion, the reaction was quenched with H2O (15 ml) , extracted with EA (15 ml *3) and washed with brine (4*20) . Then the organic layer was concentrated under vacuum, the residue was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford methyl 11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanoate.
LCMS: 685.6 [M+H] +.
Step 2. 11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanoic acid
To a solution of ethyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) nonanoate (300 mg, 0.438 mmol) in THF (3 mL) and H2O (3 ml) was added LiOH (52.63 mg, 2.19 mmol) . The resulting mixture was stirred for 16 h at 25℃. The reaction was monitored by LCMS. Upon completion, the reaction was adjusted to pH 6 with 2N HCl , extracted by EA (15 ml *3) and dried by Na2SO4. Then the organic layer was concentrated under vacuum to afford 11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanoic acid.
LCMS: 671.6 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (11- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) undecanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 7)
11- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] undecanoic acid (250mg, 0.373mmol, 1.0equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (160.45 mg, 0.373 mmol, 1.0 equiv. ) , HATU (184.2 mg, 0.485 mmol, 1.3 equiv. ) and TEA (188.55 mg, 1.865 mmol, 5.0 equiv. ) were dissolved in DMF (10 ml) . The mixture was stirred at room temperature for overnight. 30 ml of water was added. The solution was extracted with EA (100 ml*3) , washed with (80 ml*3) and concentrated. The crude product was purified by TLC with DCM/MeOH (10/1) , then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (54%CH3CN up to 74%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 74%CH3CN in 0.1min, hold 54%in 0.9min) ; Detector, UV220 &254nm. This resulted in 127.9 mg of Compound 7 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (11- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-
yl] undecanamido) -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 8.98 (s, 1H) , 8.82 (s, 1H) , 8.46 -8.63 (m, 1H) , 7.95 -8.18 (m, 1H) , 7.85 (d, J = 6.9 Hz, 1H) , 7.71 -7.81 (m, 1H) , 7.53 -7.68 (m, 3H) , 7.31 -7.48 (m, 4H) , 6.92 (d, J = 6.6 Hz, 2H) , 5.59 -5.84 (m, 1H) , 5.33 (s, 1H) , 5.13 (s, 1H) , 5.00 (d, J = 6.0 Hz, 1H) , 4.82 (d, J = 7.2 Hz, 1H) , 4.68 (s, 2H) , 4.55 (d, J = 6.0 Hz, 1H) , 4.39 -4.49 (m, 2H) , 4.35 (brs, 1 H) , 4.17 -4.28 (m, 1H) , 3.61 -3.76 (m, 2H) , 3.32 (s, 2H) , 3.09 (brs, 4H) , 2.41-2.48 (m, 5H) , 2.21 -2.38 (m, 3H) , 1.99 -2.17 (m, 2H) , 1.87 -1.96 (m, 1H) , 1.46 (s, 10H) , 1.26 (s, 12H) , 0.94 (s, 9H) .
LCMS: 1083.6 [M+H] +.
Compound 8
Step 1. methyl (E) -4- (2- (benzyloxy) ethoxy) but-2-enoate
To a solution of DMSO (800 mg, 10.2 mmol) in DCM (15 mL) was slowly added oxalyl dichloride (970 mg, 7.65 mmol) at -78 ℃ under N2 atmosphere. The resulting mixture was stirred for 0.5 h at this temperature. Then 2- (2- (benzyloxy) ethoxy) ethan-1-ol (1 g, 5.1 mmol) was added and stirred for 1 h at this temperature. Subsequently, TEA (2.58 g 25.5 mmol) was added. Then the reaction was allowed to warm to r.t. slowly and stirred for 1 h. The reaction was monitored by TLC. Upon completion, methyl 2- (triphenyl-l5-phosphaneylidene) acetate (2.05 g, 6.12 mmol) was added and the reaction was stirred for 2 h at r.t. . The reaction was monitored by LCMS. The reaction was quenched with H2O (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) and the organic layer was concentrated under vacuum. The residue obtained was purified by silica gel chromatography (0-15%EtOAc/petroleum ether) to afford methyl (E) -4- (2- (benzyloxy) ethoxy) but-2-enoate.
LCMS: 251.1 [M+H] +.
Step 2. methyl 4- (2-hydroxyethoxy) butanoate
To a solution of methyl (E) -4- (2- (benzyloxy) ethoxy) but-2-enoate (900 mg, 3.6 mmol) in MeOH (15 mL) was added 10%Pd/C (800 mg) , . Under 10 atm of H2 atmosphere the resulting mixture was stirred at r.t. for 16 h. The reaction was monitored by LCMS. On completion the mixture was filtrated and concentrated under vacuum. 550 mg of methyl 4- (2-hydroxyethoxy) butanoate was obtained.
LCMS: 163.1 [M+H] +.
Step 3. methyl 4- (2-iodoethoxy) butanoate
To a solution of methyl 4- (2-hydroxyethoxy) butanoate (300 mg, 1.850 mmol) in DCM (5 mL) was added Imidazole (151.11 mg, 2.220 mmol) , I2 (516.43 mg, 2.035 mmol) , PPh3 (533.68 mg, 2.035 mmol) , The resulting mixture was stirred at r.t. overnight, the reaction was quenched with Na2S2O3 (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) , washed with brine (2 x 30 mL) . The organic layers were combined then concentrated. The residue obtained was purified by silica gel chromatography (10-20%EtOAc/petroleum ether) to afford methyl 4- (2-iodoethoxy) butanoate.
Step 4. methyl 4- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) butanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (300 mg, 0.617 mmol) in DMF (5 mL) was added K2CO3 (426.05 mg, 3.085 mmol) , methyl 4- (2-hydroxyethoxy) butanoate (138.53 mg, 0.678 mmol) , The resulting mixture was stirred at r.t. overnight. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0 -6%MeOH/DCM) to afford 200 mg of methyl 4- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) butanoate.
LC/MS: 631.3 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (dodec-11-ynamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide
To a solution of methyl 4- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) butanoate (200 mg, 0.317 mmol) in TFA (3 mL) , H2O (3 mL) was added LiOH (37.97 mg, 1.585 mmol) . The resulting mixture was stirred at r.t. overnight. The reaction was acidified to pH 2-4 with 2N HCl. The resulting mixture was filtered and concentrated. to afford 4- (2- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] ethoxy) butanoic acid.
LC/MS: 617.3 [M+H] +.
Step 6. (2S, 4R) -1- ( (S) -2- (4- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) butanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (thiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 8)
To a solution of 4- (2- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] ethoxy) butanoic acid (240 mg, 0.389 mmol) in DMF (5 mL) was added TEA (118.14 mg, 1.167 mmol) , HATU (192.36 mg, 0.506 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-
thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (184.31 mg, 0.428 mmol) , The resulting mixture was stirred at rt for 2 hour. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /6: 1) then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 50mmol/L NH4HCO3; Phase B: CH3CN (21%CH3CN up to 51%in 9 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 51%CH3CN in 0.1min, hold 21%in 0.9min) ; Detector, UV220 &254nm to afford 128.3 mg of Compound 8 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (2- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] ethoxy) butanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.51 -8.57 (m, 1H) , 8.03 -8.18 (m, 1H) , 7.88 (d, J = 9.6 Hz, 1H) , 7.76 -7.77 (m, 1H) , 7.53 -7.63 (m, 3H) , 7.39 -7.44 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.60 -5.73 (m, 1H) , 5.31 (s, 1H) , 5.11 -5.12 (m, 1H) , 5.00 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 -4.73 (m, 2H) , 4.54 (d, J = 2.4 Hz, 1H) , 4.40 -4.48 (m, 2H) , 4.34 -4.38 (m, 1H) , 4.20 -4.26 (m, 1H) , 3.62 -3.71 (m, 2H) , 3.45 -3.56 (m, 2H) , 3.35 -3.40 (m, 2H) , 3.10 (s, 4H) , 2.51 -2.60 (m, 6H) , 2.39 (s, 3H) 2.23 -2.30 (m, 1H) , 2.16 -2.23 (m, 1H) , 2.10 -2.20 (m, 1H) , 2.02 -2.04 (m, 1H) , 1.63 -1.78 (m, 2H) , 1.45 (s, 6H) , 0.81 -1.00 (m, 9H) .
LC/MS: 1029.5 [M+H] +.
Compound 9
Step 1. tert-butyl 3- (4-hydroxybutoxy) propanoate
To a solution of 1, 4-butanediol (500 mg, 5.548 mmol) in DMF (5 mL) was added NaH (199.71 mg, 8.322 mmol) at 0 ℃. The resulting mixture was stirred at rt for 0.5 h. Then add tert-butyl 3-bromopropanoate reaction was stirred at rt for 1 h. The reaction was quenched with H2O (15 mL) . Then resulting mixture was extracted with EtOAc (3 x 20 mL) . The combined organic layers were washed with brine (4 x 30 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-30%EtOAc/petroleum ether) to afford tert-butyl 3- (4-hydroxybutoxy) propanoate, as a colorless oil.
Step 2. tert-butyl 3- (4-iodobutoxy) propanoate
To a solution of tert-butyl 3- (4-hydroxybutoxy) propanoate (450 mg, 2.061 mmol) in DCM (5 mL) was added imidazole (168.41 mg, 2.474 mmol) , I2 (575.54 mg, 2.268 mmol) and PPh3 (594.76
mg, 2.268 mmol) . The resulting mixture was stirred at r.t. overnight, the reaction was quenched with Na2S2O3 (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) , washed with brine (2 x 30 mL) The organic layers were combined then concentrated. The residue obtained was purified by silica gel chromatography (10-20%EtOAc/petroleum ether) to afford tert-butyl 3- (4-iodobutoxy) propanoate.
Step 3. tert-butyl 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250 mg, 0.514 mmol) in DCM (5 mL) was added K2CO3 (355.04 mg, 2.569 mmol) , tert-butyl 3- (4-iodobutoxy) propanoate (185.48 mg, 0.565 mmol) , The resulting mixture was stirred at rt overnight. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0 -5%MeOH/DCM) to afford 240 mg of tert-butyl 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoate.
LCMS: 687.4 [M+H] +.
Step 4. 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoic acid
To a solution of tert-butyl 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoate (280 mg, 0.408 mmol) , DCM (5 mL) , TFA (2 mL) The resulting mixture was stirred at rt for 1 h and concentrated, to afford 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoic acid.
LC/MS: 631.3 [M+H] +
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (Compound 9)
To a solution of 3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanoic acid (270 mg, 0.428 mmol) in DMF (5 mL) was added TEA (129.95 mg, 1.284 mmol) , HATU (211.59 mg, 0.556 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (202.74 mg, 0.471 mmol) , The resulting mixture was stirred at rt 4 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /10: 1) then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 50mmol/L NH4HCO3; Phase B: CH3CN (36%CH3CN up to 56%in 9 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 56%CH3CN in 0.1min, hold 36%in 0.9min) ; Detector, UV220 &254nm to afford 182.2 mg of Compound 9 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (s, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.51 –8.57 (m, 1H) , 8.03 –8.18 (m, 1H) , 7.84 (d, J = 9.2 Hz, 1H) , 7.76 –7.77 (m, 1H) , 7.53 –7.63 (m, 3H) , 7.39 –7.44 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.60 –5.73 (m, 1H) , 5.31 (s, 1H) , 5.11 –5.12 (m, 1H) , 5.00 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 –4.73 (m, 2H) , 4.57 (d, J = 9.6 Hz, 1H) , 4.36 –4.59 (m, 3H) , 4.19 –4.28 (m, 1H) , 3.52 –3.71 (m, 5H) , 3.30 –3.41 (m, 2H) , 3.04 –3.12 (m, 4H) , 2.48 –2.60 (m, 3H) , 2.45 (s, 3H) , 2.29 –2.38 (m, 3H) , 2.00 –2.10 (m, 1H) , 1.90 –1.98 (m, 1H) , 1.49 –1.53 (m, 4H) , 1.48 (s, 7H) , 0.94 (s, 9H) .
LCMS: 1043.5 [M+H] +.
Compound 10
Step 1. methyl 4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250.00 mg, 0.514 mmol) in DMF (4 mL) was added methyl 4- (3-iodopropoxy) butanoate (161.70 mg, 0.565 mmol) and K2CO3 (355.04 mg, 2.569 mmol) . The resulting mixture was stirred at 25 ℃ for 5 h. The reaction was quenched with water (5mL) . The resulting mixture was extracted with ethyl acetate (3*15mL) . The organic layers were combined, washed with brine (4*15mL) and concentrated. The residue obtained was purified by silica gel chromatography (0-10%DCM/MeOH) to afford the methyl 4- (3- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] propoxy) butanoate.
LC/MS: 645.3 [M+H] +.
Step 2. 4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanoic acid
To a solution of methyl 4- (3- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] propoxy) butanoate (146.00 mg, 0.226 mmol) in THF (2 mL) and H2O (2 mL) was added LiOH. H2O (47.51 mg, 1.132 mmol) . The resulting mixture was stirred at 25 ℃ for 16 h. The reaction was quenched with water (5 mL) . The pH value of the aqueous phase was adjusted to 5 used HCl solution (2 M) , then filtered and concentrated to afford the 4- (3- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] propoxy) butanoic acid.
LC/MS: 631.25 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 10)
Added (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (82.60 mg, 0.192 mmol) , HATU (86.20 mg, 0.227 mmol) and DIEA (67.62 mg, 0.523 mmol) . The resulting mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, washed with brine (4x15mL) and concentrated. The residue obtained was purified by TLC (1/10 MeOH/DCM) , then purified by Prep-HPLC with the following conditions: Column, Xselect CSH OBD Column 30*150mm 5um; Mobile Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (25%CH3CN up to 55%in 9 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 25%CH3CN in 0.1min, hold 25%in 0.9min) ; Detector, UV220 &254 nm. This resulted in 77.2 mg of Compound 10 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (3- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] propoxy) butanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 9.00 (s, 1H) , 8.82 (s, 1H) , 8.55 (t, J = 4 Hz, 1H) , 7.99 -8.18 (m, 1H) , 7.87 (d, J = 12 Hz, 1H) , 7.68-7.82 (m, 1H) , 7.51 -7.67 (m, 3H) , 7.37 -7.50 (m, 4H) , 6.82 -6.92 (m, 2H) , 5.55 -5.75 (m, 1H) , 5.31 (s, 1H) , 5.10 -5.13 (m, 1H) , 4.97 -5.05 (m, 1H) , 4.80 -4.90 (m, 1H) , 4.61 -4.75 (m, 2H) , 4.51 -4.62 (m, 1H) , 4.40 -4.50 (m, 2H) , 4.35 -4.39 (m, 1H) , 4.12 -4.30 (m, 1H) , 3.66 -3.80 (m, 2H) , 3.37 -3.42 (m, 2H) , 3.32 (s, 2H) , 3.02 -3.20 (m, 4H) , 2.44 (s, 6H) , 2.28 -2.40 (m, 4H) , 2.18 -2.22 (m, 1H) , 1.99 -2.11 (m, 1H) , 1.81 -1.95 (m, 1H) , 1.60 -1.80 (m, 4H) , 1.46 (s, 6H) , 0.94 (s, 9H) . LC/MS: 1043.65 [M+H] +.
Compound 11
Step 1. tert-butyl 2- ( (6-hydroxyhexyl) oxy) acetate
To a solution of hexane-1, 6-diol (1.00 g, 8.462 mmol) in DMF (20 mL) was added NaH (0.51 g, 12.693 mmol) , at 0℃. the resulting mixture was stirred at r.t. 0.5 hour. Then add butyl 2-bromoacetate (1.65 mL, 8.462 mmol) , the resulting mixture was stirred at r.t. 1 hour. The reaction was quenched with H2O (20 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (4 x 100 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%EtOAc/petroleum ether) to afford tert-butyl 2- ( (6-hydroxyhexyl) oxy) acetate.
LC/MS: 233.2 [M+H] +.
Step 2. tert-butyl 2- ( (6-iodohexyl) oxy) acetate
To a solution of tert-butyl 2- [ (6-hydroxyhexyl) oxy] acetate (500 mg, 2.152 mmol) in DCM (5 mL) was added imidazole (175.82 mg, 2.583 mmol) , I2 (600.87, 2.367 mmol) and PPh3 (620.94 mg, 2.367 mmol) . The resulting mixture was stirred at r.t. overnight. The reaction was quenched with Na2S2O3 (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) , washed with brine (2 x 30 mL) . The organic layers were combined then concentrated. The residue obtained was purified by silica gel chromatography (10-20%EtOAc/petroleum ether) to afford tert-butyl 2- [ (6-iodohexyl) oxy] acetate.
Step 3. tert-butyl 2- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) oxy) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ [4- (piperazin-1-yl) phenyl] amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (280 mg, 0.575 mmol) in DMF (5 mL) was added K2CO3 (397.65mg, 2.877 mmol) , methyl 4- (2-hydroxyethoxy) butanoate (216.62 mg, 0.633 mmol) , The resulting mixture was stirred at rt overnight. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0 -6%MeOH/DCM) to afford tert-butyl 2- [ (6- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} hexyl) oxy] acetate.
LC/MS: 702.4 [M+H] +.
Step 4.2- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) oxy) acetic acid
To a solution of tert-butyl 2- [ (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexyl) oxy] acetate (220 mg, 0.314 mmol) , DCM (5 mL) , TFA (2 mL) The resulting mixture was stirred at rt for 1.5 h and concentrated, to afford [ (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexyl) oxy] acetic acid as a yellow solid.
LCMS: 645.3 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (2- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) oxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 11)
To a solution of [ (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexyl) oxy] acetic acid (200 mg, 0.310 mmol) in DMF (5 mL) was added TEA (94.16 mg, 0.930 mmol) , HATU (153.32 mg, 0.403 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide (146.91 mg, 0.341 mmol) . The resulting mixture was stirred at rt 2 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /10: 1) then purified by Prep-HPLC under the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) 4; mobile phase, Phase A: water with 50mmol/L NH4HCO3; Phase B: CH3CN (38%CH3CN up to 68%in 9 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 68%CH3CN in 0.1min, hold 38%in 0.9min) ; Detector, UV220 &254nm to afford 136.4 mg of Compound 11 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- [ (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] hexyl) oxy] acetamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.50 -8.70 (m, 1H) , 7.90 -8.10 (m, 1H) , 7.71 -7.80 (m, 1H) , 7.52 -7.63 (m, 3H) , 7.31 -7.43 (m, 5H) , 6.81 -6.97 (m, 2H) , 5.51 -5.79 (m, 1H) , 5.29 (s, 1H) , 5.15 (s, 1H) , 4.98 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.63 -4.72 (m, 2H) , 4.55 (d, J = 1.2 Hz, 1H) , 4.32 -4.50 (m, 3H) , 4.22 -4.30 (m, 1H) , 3.87 -3.98 (m, 2H) , 3.60 -3.70 (m, 2H) , 3.47 -3.52 (m, 2H) , 3.00 -3.12 (m, 4H) , 2.40 -2.50 (m, 6H) , 2.13 -2.40 (m, 3H) , 2.00 -2.10 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.52 -1.62 (m, 2H) , 1.46 (s, 8H) , 1.30 -1.40 (m, 4H) , 0.95 (s, 9H) .
LC/MS: 1057.33 [M+H] +.
Compound 12
Step 1. tert-butyl 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butoxy) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 2- (4-iodobutoxy) acetate (154.96 mg, 0.493 mmol, 1.2 equiv. ) . The resulting mixture was stirred at 50 ℃ for 5 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford tert-butyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetate (240 mg, 79.32%) .
LC/MS: 673.3 [M+H] +.
Step 2. 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butoxy) acetic acid
To a solution of tert-butyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetate (240 mg, 0.357 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 220.49 equiv. ) was added TFA (3 mL, 40.389 mmol, 113.23 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was purified by reverse column to afford (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetic acid.
LC/MS: 617.3 [M+H] +.
Step 3. 2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 12)
To a solution of (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetic acid (220 mg, 0.357 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added TEA (180.48 mg, 1.785 mmol, 5 equiv. ) , HATU (176.33 mg, 0.464 mmol, 1.3 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (168.95 mg, 0.393 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 40 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the crude and the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 35%B to 45%B in 11 min, then 45%to 100%in 1 min, hold 100%in 1 min, down to 35%in 1 min, hold 35%in 1 min; Wave Length: 254 nm to afford Compound 12 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ: 10.12 (brs, 1H) , 9.00 (s, 1H) , 8.82 (s, 1H) , 8.58 -8.68 (m, 1H) , 8.00 -8.10 (m, 1H) , 7.75 (s, 1H) , 7.56 -7.60 (m, 3H) , 7.35 -7.41 (m, 5H) , 6.90 (d, J = 8.8 Hz, 2H) , 5.65 -5.67 (m, 1H) , 5.32 (s, 1H) , 5.18 (s, 1H) , 4.96 -5.00 (m, 1H) , 4.75 -4.90 (m, 1H) , 4.60 -4.72 (m, 2H) , 4.57 (d, J = 9.6 Hz, 1H) , 4.42 -4.52 (m, 1H) , 4.30 -4.41 (m, 2H) , 4.20 -4.29 (m, 1H) , 3.90 -4.03 (m, 2H) , 3.58 -3.72 (m, 2H) , 3.44 -3.55 (m, 2H) , 3.00 -3.15 (m, 4H) , 2.50 (s, 3H) , 2.28 -2.45 (m, 6H) , 2.00 -2.12 (m, 1H) , 1.85 -1.95 (m, 1H) , 1.58 -1.60 (m, 4H) , 1.46 (s, 6H) , 0.96 (s, 9H) .
LCMS: 1029.5 [M+H] +.
Compound 13
Step 1. tert-butyl 3- [3- (benzyloxy) propoxy] propanoate
To a solution of 3- (benzyloxy) propan-1-ol (1.5 g, 9.024 mmol, 1.00 equiv. ) in DCM (10 mL, 157.300 mmol, 17.43 equiv. ) was added tert-butyl 3-bromopropanoate (2.83 g, 13.536 mmol, 1.5 equiv. ) , NaOH (10 mL, 250.018 mmol, 27.71 equiv. ) and tetrabutylazanium chloride (2.51 g, 9.024 mmol, 1 equiv. ) . The mixture was stirred at 25 ℃ overnight. The reaction was quenched
with H2O (15 mL) . The resulting mixture was extracted with DCM (3 x 15 mL) . The organic layers were combined concentrated. The residue obtained was purified by silica gel column chromatography, eluted with (50%PE/EA) to afford the tert-butyl 3- [3- (benzyloxy) propoxy] propanoate.
Step 2. tert-butyl 3- (3-iodopropoxy) propanoate
To a solution of tert-butyl 3- [3- (benzyloxy) propoxy] propanoate (810 mg, 2.751 mmol, 1.00 equiv. ) in MeOH (10 mL, 246.989 mmol, 89.77 equiv. ) was added Pd/C (600 mg, 5.638 mmol, 2.05 equiv. ) . The mixture was maintained under hydrogen atmosphere and stirred at 25 ℃ overnight, filtered and concentrated to afford the tert-butyl 3- (3-iodopropoxy) propanoate (549 mg, 63.51%) as a colorless oil.
Step 3. tert-butyl 3- (3-iodopropoxy) propanoate
To a solution of tert-butyl 3- (3-hydroxypropoxy) propanoate (549 mg, 2.688 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 29.26 equiv. ) was added PPh3 (775.43 mg, 2.957 mmol, 1.1 equiv. ) , Imidazole (219.56 mg, 3.226 mmol, 1.2 equiv. ) , I2 (750.37 mg, 2.957 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 4 h. The reaction was quenched with Na2S2O3 (50 mL) . The resulting mixture was extracted with DCM (3 x 50 mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (0-50%ethyl acetate/petroleum ether) to afford the tert-butyl 3- (3-iodopropoxy) propanoate.
Step 4. tert-butyl 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) . The resulting mixture was stirred at 50 ℃ for 3h. After cooling down to rt, the reaction was quenched with water (10 mL) . The resulting mixture was extracted with EA (3 x 10mL) , washed with brine (6*10 mL) . The organic layers were combined and concentrated. The residue was purified by Prep-TLC (DCM/MeOH 14/1) to afford tert-butyl 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoate.
LC/MS: 673.40 [M+H] +.
Step 5. 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoic acid
To a solution of tert-butyl 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoate (145 mg, 0.216 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 364.95 equiv. ) was added TFA (2 mL, 26.926 mmol, 124.94 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated to afford the 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoic acid.
LC/MS: 617.30M+H] +.
Step 6. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 13)
To a solution of 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanoic acid (130 mg, 0.211 mmol, 1.00 equiv. ) in DMF (4 mL) was added TEA (106.65 mg, 1.055 mmol, 5 equiv. ) , HATU (104.19 mg, 0.274 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (108.91 mg, 0.253 mmol, 1.2 equiv. ) . The mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, then washed with brine (4 x 15 mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then purified by Prep-HPLC with the following conditions: Column, XBridge Prep C18 OBD Column, 30*100 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3, Mobile Phase B: CH3CN (21 %to 41%in 10 min, 41 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 21 %in 0.1 min, 21 %to 21 %in 1 min; Detector, UV 254 nm to afford 16.7 mg of Compound 13 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ10.10 (brs, 1 H) , 8.95 (s, 1 H) , 8.81 (s, 1 H) , 8.54 -8.60 (m, 1 H) , 7.99 -8.10 (m, 1 H) , 7.71 -7.98 (m, 1 H) , 7.70 -7.80 (m, 1 H) , 7.50 -7.68 (m, 3 H) , 7.30 -7.41 (m, 4 H) , 6.89 (d, J = 9.3 Hz, 2 H) , 5.49 -5.80 (m, 1 H) , 5.29 (s, 1 H) , 5.10 -5.19 (m, 1 H) , 4.90 -5.00 (m, 1 H) , 4.75 -4.89 (m, 1 H) , 4.61 -4.70 (m, 2 H) , 4.53 -4.57 (m, 1 H) , 4.30 -4.50 (m, 3 H) , 4.19 -4.29 (m, 1 H) , 3.48 -3.79 (m, 4 H) , 3.35 -3.42 (m, 2 H) , 3.06 (brs, 4 H) , 2.45 -2.47 (m, 5 H) , 2.42 (s, 3 H) , 2.30 -2.39 (m, 3 H) , 1.99 -2.10 (m, 1 H) , 1.82 -1.98 (m, 1 H) , 1.60 -1.73 (m, 2 H) , 1.44 (s, 6 H) , 0.93 (s, 9 H) . LCMS: 1029.70 [M+H] +.
Compound 14
Step 1. tert-butyl 2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 0.103 mmol, 1.00 equiv. ) in DMF
(5.00 mL) was added K2CO3 (71.01 mg, 0.515 mmol, 5 equiv. ) , tert-butyl 2- [ (2-iodoethoxy) methoxy] acetate (35.73 mg, 0.113 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 50 ℃ for 5 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the tert-butyl 2- [ (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) methoxy] acetate.
LC/MS: 675.3 [M+H] +.
Step 2. 2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetic acid
To a solution of tert-butyl 2- [ (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) methoxy] acetate (45 mg, 0.067 mmol, 1.00 equiv. ) in DCM (3 mL) was added TFA (1 mL) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was purified by reverse column to afford [ (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) methoxy] acetic acid.
LC/MS: 619.3 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 14)
To a solution of [ (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) methoxy] acetic acid (135 mg, 0.218 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added TEA (110.40 mg, 1.090 mmol, 5 equiv. ) , HATU (107.86 mg, 0.283 mmol, 1.3 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (112.74 mg, 0.262 mmol, 1.2 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 40 mL) , washed with brine (3 x 20 mL) . The organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the
crude and the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 53%B to 60%B in 9 min, then 60%to 100%in 1 min, hold 100%in 1 min, down to 53%in 1 min, hold 53%in 1 min; Wave Length: 254 nm to afford Compound 14 (2S, 4R) -1- ( (S) -2- (2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ: 10.18 (brs, 1H) , 9.00 (s, 1H) , 8.83 (s, 1H) , 8.61 -8.63 (m, 1H) , 8.00 -8.10 (m, 1H) , 7.74 -7.76 (m, 1H) , 7.48 -7.62 (m, 4H) , 7.40 (s, 4H) , 6.91 (d, J = 9.2 Hz, 2H) , 5.66 -5.67 (m, 1H) , 5.33 (s, 1H) , 5.18 (s, 1H) , 4.92 -5.02 (m, 1H) , 4.60 -4.85 (m, 6H) , 4.69 (d, J = 5.2 Hz, 1H) , 4.30 -4.52 (m, 3H) , 4.18 -4.29 (m, 1H) , 3.95 -4.15 (m, 2H) , 3.55 -3.72 (m, 4H) , 3.08 -3.10 (m, 4H) , 2.51 -2.58 (m, 5H) , 2.44 (s, 3H) , 2.00 -2.15 (m, 1H) , 1.82 -1.92 (m, 1H) , 1.46 (s, 6H) , 0.96 (s, 9H) .
LC/MS: 1031.5 [M+H] +.
Compound 15
Step 1. tert-butyl 2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 2- [ (5-iodopentyl) oxy] acetate (148.39 mg, 0.452 mmol, 1.1 equiv. ) The resulting mixture was d stirred at 50 ℃ for 3 h. After cooling down to rt, the reaction was quenched with water (10 mL) . The resulting mixture was extracted with EA (3 x 10mL) , washed with brine (6*10 mL) . The organic layers were combined and concentrated. The residue was purified by Prep-TLC (DCM/MeOH 14/1) to afford tert-butyl 2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetate.
LC/MS: 687.35 [M+H] +.
Step 2. [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetic acid
To a solution of tert-butyl 2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetate (152 mg, 0.221 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 355.40 equiv. ) was added TFA (2 mL, 26.926 mmol, 121.67 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated to afford the [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetic acid.
LC/MS: 631.25 M+H] +.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 15)
To a solution of [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetic acid (140 mg, 0.222 mmol, 1.00 equiv. ) in DMF (4 mL, 51.687 mmol, 232.87 equiv. ) was added TEA (202.14 mg, 1.998 mmol, 9 equiv. ) , HATU (109.71 mg, 0.289 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (114.68 mg, 0.266 mmol, 1.2 equiv. ) . The mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, washed with brine (4x15mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then purified by Prep-HPLC with the following conditions: Column, XBridge Prep C18 OBD Column, 30*100 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3, Mobile Phase B: CH3CN (30 %to 42%in 10 min, 42 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 30 %in 0.1 min, 30 %to 30 %in 1 min; Detector, UV 254 nm to afford 74.8 mg of Compound 15 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ10.10 (brs, 1 H) , 8.95 (s, 1 H) , 8.81 (s, 1 H) , 8.59 (t, J = 6.0 Hz, 1 H) , 7.98 -8.02 (m, 1 H) , 7.71 -7.74 (m, 1 H) , 7.50 -7.60 (m, 3 H) , 7.30 -7.42 (m, 5 H) , 6.90 (d, J = 9.0 Hz, 2 H) , 5.56 -5.72 (m, 1 H) , 5.29 (s, 1 H) , 5.13 -5.14 (m, 1 H) , 4.97 (d, J = 12 Hz, 1 H) , 4.77 -4.90 (m, 1 H) , 4.60 -4.72 (m, 2 H) , 4.50 -4.59 (m, 1 H) , 4.13 -4.49 (m, 4 H) , 3.90 (s, 2 H) , 3.64 -3.88 (m, 2 H) , 3.48 (t, J = 6.3 Hz, 2 H) , 3.04 (brs, 4 H) , 2.42 -2.49 (m, 7 H) , 2.20 -2.32 (m, 2 H) , 1.99 -2.10 (m, 1 H) , 1.82 -1.98 (m, 1 H) , 1.51 -1.61 (m, 2 H) , 1.30 -1.50 (m, 11 H) , 0.91 (s, 9 H) .
LC/MS: 1044.10 [M+H] +.
Compound 16
Step 1. tert-butyl 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in
DMF) was added tert-butyl 5- (2-iodoethoxy) pentanoate (148.39 mg, 0.452 mmol, 1.1 equiv. ) , The resulting mixture was stirred at 50℃ for 4 h . The reaction was quenched with H2O (mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) . The combined organic layers were washed with brine (4 x 50mL) then concentrated. The residue obtained was purified by prep-TLC (MeOH: DCM=1: 8) to afford tert-butyl 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoate.
LC/MS: 687.39 [M+H] +.
Step 2. 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoic acid
To a solution of tert-butyl 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoate (175 mg, 0.255 mmol, 1.00 equiv. ) DCM (5 mL) , was added TFA (1.5 mL) The resulting mixture was stirred at r.t. for 1.5 h and concentrated, to afford 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoic acid.
LC/MS: 631.3 [M+H] +.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 16)
To a solution of 5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanoic acid (150 mg, 0.238 mmol) in DMF (5 mL) was added TEA (120.32 mg, 1.19 mmol) , HATU (117.55 mg, 0.309 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (102.39 mg, 0.238 mmol) , The resulting mixture was stirred at r.t. 4 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /8: 1) then further purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L
NH4HCO3+0.1NH3. H2O; Phase B: CH3CN (35%CH3CN up to 55%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 55%CH3CN in 0.1min, hold 35%in 0.9min) ; Detector, UV220 &254nm to afford 90.5 mg of Compound 16 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.15 (brs, 1H) , 9.00 (s, 1H) , 8.83 (s, 1H) , 8.50 -8.60 (m, 1H) , 8.00 -8.16 (m, 1H) , 7.89 (d, J = 12.00 Hz, 1H) , 7.69 -7.82 (m, 1H) , 7.50 -7.64 (m, 3H) , 7.32 -7.48 (m, 4H) , 6.92 (d, J = 8.0 Hz, 2H) , 5.59 -5.74 (m, 1H) , 5.42 (s, 1H) , 5.09 -5.16 (m, 1H) , 4.97 (d, J = 8.0 Hz, 1H) , 4.83 (d, J = 12 Hz, 1H) , 4.60 -4.73 (m, 2H) , 4.56 (d, J = 8.0 Hz, 1H) , 4.40 -4.50 (m, 2H) , 4.30 -4.38 (m, 1H) , 4.17 -4.28 (m, 1H) , 3.59 -3.86 (m, 2H) , 3.48 -3.52 (m, 2H) , 3.36 -3.44 (m, 2H) , 3.02 -3.16 (m, 4H) , 2.53 -2.61 (m, 6H) , 2.43 (s, 3H) , 2.26 -2.35 (m, 1H) , 2.09 -2.19 (m, 1H) , 1.98 -2.07 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.41 -1.57 (m, 10H) , 0.95 (s, 9H) .
LC/MS: 1043.52 [M+H] +.
Compound 17
Step 1. tert-butyl 2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (4 mL, 51.687 mmol, 125.75 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 2- [2- (2-bromoethoxy) ethoxy] acetate (128.03 mg, 0.452 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 50 ℃ for 8 h. After cooling down to r.t., the reaction was quenched with water (10 mL) . The resulting mixture was extracted with EA (3 *10mL) , washed with brine (6 *10 mL) . The organic layers were combined and concentrated. The residue was purified by Prep-TLC (DCM/MeOH 14/1) to afford the tert-butyl 2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetate.
LC/MS: 689.55 [M+H] +.
Step 2. methyl dodec-11-ynoate [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetic acid
To a solution of tert-butyl 2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetate (110 mg, 0.160 mmol, 1.00 equiv. ) in DCM (4 mL, 62.920 mmol, 394.01 equiv. ) was added TFA (2 mL, 26.926 mmol, 168.61 equiv. ) . The resulting mixture was d stirred at 25 ℃ for 1 h. The reaction was concentrated to afford [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetic acid.
LC/MS: 633.20 [M+H] +.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 17)
To a solution of [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetic acid (100 mg, 0.158 mmol, 1.00 equiv. ) in DMF (4 mL, 51.687 mmol, 327.04 equiv. ) was added TEA (143.94 mg, 1.422 mmol, 9 equiv. ) , HATU (78.12 mg, 0.205 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (81.66 mg, 0.190 mmol, 1.2 equiv. ) . The mixture was stirred at 25 ℃ for 5 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, washed with brine (4 x 15mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then further purified by Prep-HPLC with the following conditions: Column, XBridge Shield RP18 OBD Column, 30*150 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3, Mobile Phase B: CH3CN (36 %to 43%in 10 min, 43 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 43 %in 0.1 min, 43 %to 43 %in 1 min; Detector, UV 254 nm to afford 70.6 mg of Compound 17 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ10.15 (brs, 1 H) , 8.96 (s, 1 H) , 8.82 (s, 1 H) , 8.59 (t, J = 6.0 Hz, 1 H) , 8.03 (t, J = 7.6 Hz, 1 H) , 7.74 -7.75 (m, 1 H) , 7.50 -7.61 (m, 3 H) , 7.41 -7.59 (m, 5 H) , 6.88 -6.91 (m, 2 H) , 5.60 -5.75 (m, 1 H) , 5.32 (s, 1 H) , 5.16 (d, J = 2.8 Hz, 1 H) , 4.97 -5.05 (m, 1 H) , 4.80 -4.84 (m, 1 H) , 4.65 -4.72 (m, 2 H) , 4.51 -4.62 (m, 1 H) , 4.31 -4.49 (m, 3 H) , 4.20 -4.30 (m, 1 H) , 3.99 (s, 2 H) , 3.55 -3.70 (m, 8 H) , 3.06 (brs, 4 H) , 2.50 -2.56 (m, 6 H) , 2.43 (s, 3 H) , 2.04 -2.11 (m, 1 H) , 1.85 -1.93 (m, 1 H) , 1.46 (s, 6 H) , 0.94 (s, 9 H) .
LC/MS: 1045.65 [M+H] +.
Compound 18
Step 1. tert-butyl 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) . The resulting mixture was stirred at 50 ℃ for 5 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography to afford tert-butyl 2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetate.
LC/MS: 689.3 [M+H] +.
Step 2. 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetic acid
To a solution of tert-butyl 2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetate (200 mg, 0.290 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 270.88 equiv. ) was added TFA (3 mL, 40.389 mmol, 139.11 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was purified by reverse column chromatography to afford the [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetic acid.
LC/MS: 633.3 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 18)
To a solution of [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetic acid (180 mg, 0.284 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added TEA (143.94 mg, 1.420 mmol, 5 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (146.99 mg, 0.341 mmol, 1.2 equiv. ) , HATU (140.62 mg, 0.369 mmol, 1.3 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the crude and the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 34%to 40%in 15 min, then 40%to 100%in 1 min, hold 100%in 1 min, down to 34%in 1 min, hold 34%in 1 min; Wave Length: 254 nm to afford Compound 18 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-
d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ: 10.10 (brs, 1H) , 9.00 (s, 1H) , 8.85 (s, 1H) , 8.58 -8.60 (m, 1H) , 8.00 -8.10 (m, 1H) , 7.60 (d, J = 7.6 Hz, 1H) , 7.59 -7.64 (m, 3H) , 7.45 -7.47 (m, 1H) , 7.40 (s, 4H) , 6.90 (d, J = 9.2 Hz, 2H) , 5.56 -5.78 (m, 1H) , 5.30 (s, 1H) , 5.15 (d, J = 3.2 Hz, 1H) , 4.97 -5.02 (m, 1H) , 4.79 -4.87 (m, 1H) , 4.72 (s, 2H) , 4.62 -4.71 (m, 2H) , 4.58 (d, J = 9.6 Hz, 1H) , 4.35 -4.52 (m, 3H) , 4.20 -4.32 (m, 1H) , 3.90 -4.15 (m, 2H) , 3.50 -3.75 (m, 4H) , 3.06 -3.10 (m, 4H) , 2.51 -2.52 (m, 4H) , 2.49 (s, 3H) , 2.30 -2.49 (m, 2H) , 2.00 -2.12 (m, 1H) , 1.85 -1.96 (m, 1H) , 1.65 -1.76 (m, 2H) , 1.46 (s, 6H) , 0.95 (s, 9H) . LC/MS: 1045.5 [M+H] +.
Compound 19
Step 1. tert-butyl 3- ( (5- (benzyloxy) pentyl) oxy) propanoate
To a solution of 5- (benzyloxy) pentan-1-ol (1 g, 5.15 mmol) in DCM (10 mL) was added tert-butyl 3-bromopropanoate (1.6 g 7.73mmol) , 35%NaOH (10 ml) and Bu4NCl (1.4 g 5.15 mmol) . The resulting mixture was stirred for 16 h at 25℃. The reaction was monitored by LCMS. Upon completion, the reaction was quenched by H2O and extracted by DCM (20 ml *3) . Then the organic layer was concentrated under vacuum the resulted crude was purified by silica gel column (0 –15%PE/EA) to afford the tert-butyl 3- ( (5- (benzyloxy) pentyl) oxy) propanoate.
LC/MS: 267 [M+H-tBu] +.
Step 2. tert-butyl 3- ( (5-hydroxypentyl) oxy) propanoate
To a solution of tert-butyl 3- ( (5- (benzyloxy) pentyl) oxy) propanoate (520 mg, 1.61 mmol) in MeOH (10 mL) was added 10%Pd/C (300 mg) . The resulting mixture was stirred for 16 h at 25℃ under H2 atmosphere. The reaction was monitored by LCMS. Upon completion, the reaction mixture was filtrated, and the organic phase was concentrated under vacuum to afford tert-butyl 3- ( (5-hydroxypentyl) oxy) propanoate.
LC/MS: 177 [M+H-tBu] +.
Step 3. tert-butyl 3- ( (5-iodopentyl) oxy) propanoate
To a solution of tert-butyl 3- ( (5-hydroxypentyl) oxy) propanoate (310 mg, 1.33 mmol) in DCM (10ml) was added PPh3 (383 mg 1.46 mmol) , 1H-imidazole (108 mg 1.596 mmol) and I2 (369 mg 1.46 mmol) . The resulting mixture was stirred for 4 h at 25℃. The reaction was monitored by TLC. Upon completion, the reaction was quenched by aqueous of Na2S2O3 and extracted by DCM (15ml*3) . The result solution was concentrated under vacuum and the residue was purified by silica gel column to afford the tert-butyl 3- ( (5-iodopentyl) oxy) propanoate.
Step 4. tert-butyl 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoate
To a stirred solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) and tert-butyl 3- [ (5-iodopentyl) oxy] propanoate (182.86 mg, 0.534 mmol, 1.3 equiv. ) in DCM was added K2CO3 (170.42 mg, 1.233 mmol, 3 equiv. ) . The resulting mixture was stirred at 50 ℃ overnight. The reaction solution was quenched with water (20 mL) . The resulting mixture
was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (4 x 100 mL) and then concentrated. The residue obtained was purified by silica gel chromatography (0-10%DCM/MeOH) to afford tert-butyl 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoate.
LC/MS: 701.40 [M+H] +.
Step 5. 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoic acid
To a stirred solution of tert-butyl 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoate (190 mg, 0.271 mmol, 1.00 equiv. ) in DCM was added TFA (1.5 mL, 20.195 mmol, 74.50 equiv. ) dropwise. The mixture was stirred at room temperature for 1.5 h. Then the solution was dissolved in DCM (30*2 mL) and concentrated under reduced pressure to afford 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoic acid.
LC/MS: 645.35 [M+H] +.
Step 6. (2S, 4R) -1- ( (S) -2- (3- (5- (4- (4- (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-ylamino) phenyl) piperazin-1-yl) pentyloxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 19)
To a solution of 3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) oxy) propanoic acid (180 mg, 0.279 mmol) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (120 mg, 0.279 mmol) in DMF (5 mL) , HATU (138 mg, 0.363 mmol) and TEA (0.5 mL) were added. The mixture was stirred at room temperature overnight. the reaction solution was quenched with water and extracted with ethyl acetate, the combined organic layers was washed with water and brine and dried over Na2SO4, the solid be removed, the filtrate was concentrated under reduced pressure. The residue was purified by flash chromatography to give 100 mg of P1 as yellow solid. The residue was purified by Prep-HPLC with the following conditions: column: XBridge Shield RP18
OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 39%B to 50%B in 11 min, 50%B; Wave Length: 254 nm; RT1 (min) : 9.2; Number Of Runs: 0. This resulted in Compound 19 (2S, 4R) -1- ( (S) -2- (3- (5- (4- (4- (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-ylamino) phenyl) piperazin-1-yl) pentyloxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ10.18 (s, 1H) , 8.98 (s, 1H) , 8.83 (s, 1H) , 8.59 -8.68 (m, 1H) , 8.01 -8.13 (m, 1H) , 7.87 -7.99 (m, 1H) , 7.72 -7.85 (m, 1H) , 7.50 -7.71 (m, 3H) , 7.38 -7.49 (m, 4H) , 6.87 -7.02 (m, 2H) , 5.57 -5.82 (m, 1H) , 5.31 (s, 1H) , 5.10 -5.19 (m, 1H) , 4.91 -5.02 (m, 1H) , 4.73 -4.85 (m, 1H) , 4.62 -4.73 (m, 2H) , 4.51 -4.61 (m, 1H) , 4.38 -4.49 (m, 2H) , 4.32 -4.49 (m, 1H) , 4.18 -4.31 (m, 1H) , 3.50 -3.78 (m, 4H) , 3.18 -3.49 (m, 2H) , 3.10 (s, 4H) , 2.55 -2.65 (m, 5H) , 2.41 -2.43 (m, 3H) , 2.22 -2.37 (m, 3H) , 2.00 -2.11 (m, 1H) , 1.82 -1.99 (m, 1H) , 1.39 -1.58 (m, 10H) , 1.25 -1.38 (m, 2H) , 0.93 -0.95 (m, 9H) .
LC/MS: 1057.70 [M+H] +.
Compound 20
Step 1. tert-butyl 4- (4-iodobutoxy) butanoate
To a solution of tert-butyl 4- (4-hydroxybutoxy) butanoate (590 mg, 2.540 mmol, 1.00 equiv. ) in DCM (5 mL) was added imidazole (207.47 mg, 3.048 mmol, 1.2 equiv. ) , I2 (709.03 mg, 2.794 mmol, 1.1 equiv. ) and PPh3 (732.71 mg, 2.794 mmol, 1.1 equiv. ) . The resulting mixture was stirred at r.t. overnight, then quenched with Na2S2O3 (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) , washed with brine (2 x 30 mL) . The organic layers were combined then concentrated. The residue obtained was purified by silica gel chromatography (10-20%EtOAc/petroleum ether) to afford tert-butyl 4- (4-iodobutoxy) butanoate.
LC/MS: 343.07 [M+H] +.
Step 2. tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 4- (4-iodobutoxy) butanoate (154.73 mg, 0.452 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 50 ℃ for 4 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 25 mL) . The combined organic layers were washed with brine (4 x 50 mL) then concentrated. The residue was purified by Prep-TLC [MeOH/DCM] [1: 8] to afford tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoate.
LC/MS: 701.41 [M+H] +.
Step 3. 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoic acid
To a solution of tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoate (194 mg, 0.277 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 284.15 equiv. ) was added TFA (1.5 mL, 20.195 mmol, 72.96 equiv. ) , The resulting mixture was stirred at r.t. for 1 h. The reaction was concentrated to afford 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoic acid.
LC/MS: 645.35 [M+H] +.
Step 4. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 20)
To a solution of 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanoic acid (160 mg, 0.248 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (125.55 mg, 1.241 mmol) , HATU
(122.66 mg, 0.323 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (106.85 mg, 0.248 mmol) , The resulting mixture was stirred at r.t. for 2h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /8: 1) then further purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1NH3. H2O; Phase B: CH3CN (40%CH3CN up to 70%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 70%CH3CN in 0.1min, hold 40%in 0.9min) ; Detector, UV220 &254 nm to afford 105.3 mg of Compound 20 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.83 (s, 1H) , 8.51 -8.57 (m, 1H) , 7.95 -8.18 (m, 1H) , 7.84 (d, J = 9.2 Hz, 1H) , 7.74 -7.78 (m, 1H) , 7.57 -7.60 (m, 3H) , 7.37 -7.44 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.60 -5.73 (m, 1H) , 5.33 (s, 1H) , 5.13 -5.14 (m, 1H) , 5.01 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 -4.73 (m, 2H) , 4.55 (d, J = 9.6 Hz, 1H) , 4.40 -4.48 (m, 2H) , 4.31 -4.42 (m, 1H) , 4.18 -4.30 (m, 1H) , 3.62 -3.71 (m, 2H) , 3.38 -3.42 (m, 3H) , 3.31 -3.34 (m, 1H) , 3.09 -3.14 (m, 4H) , 2.49 -2.50 (m, 4H) , 2.45 (s, 3H) , 2.29 -2.33 (m, 3H) , 2.18 -2.20 (m, 1H) , 1.98 -2.10 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.71 -1.72 (m, 2H) , 1.45 -1.52 (m, 10H) , 0.95 (s, 9H) .
LC/MS: 1057.54 [M+H] +.
Compound 21
Step 1. tert-butyl 2- ( (4-hydroxybutoxy) methoxy) acetate
To a solution of tert-butyl 2- { [4- (benzyloxy) butoxy] methoxy} acetate (700 mg, 2.158 mmol, 1.00 equiv. ) in MeOH (15 mL, 370.484 mmol, 171.70 equiv. ) was added Pd/C (550 mg, 2.819 mmol, 1.31 equiv. ) . The resulting mixture was stirred at 25 ℃ under H2 for 16 h. The reaction was filtered and concentrated to afford the tert-butyl 2- [ (4-hydroxybutoxy) methoxy] acetate.
Step 2. tert-butyl 2- ( (4-iodobutoxy) methoxy) acetate
To a solution of tert-butyl 2- [ (4-hydroxybutoxy) methoxy] acetate (435 mg, 1.857 mmol, 1.00 equiv. ) in DCM was added PPh3 (535.67 mg, 2.043 mmol, 1.1 equiv. ) , imidazole (151.68 mg, 2.228 mmol, 1.2 equiv. ) and I2 (518.36 mg, 2.043 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 16 h. The reaction was quenched with saturated sodium thiosulfate solution (30 mL) and extracted with DCM (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography to afford tert-butyl 2- [ (4-iodobutoxy) methoxy] acetate.
Step 3. tert-butyl 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 2- [ (3-iodopropoxy) methoxy] acetate (162.85 mg, 0.493 mmol, 1.2 equiv. ) . The resulting mixture was stirred at 50 ℃ for 5 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford tert-butyl 2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetate.
LC/MS: 689.3 [M+H] +.
Step 4. 2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetic acid
To a solution of tert-butyl 2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetate (124 mg, 0.180 mmol, 1.00 equiv. ) in THF (3 mL, 37.029 mmol, 205.70 equiv. ) , H2O (3 mL, 166.525 mmol, 925.06 equiv. ) and MeOH (3 mL, 74.097 mmol, 411.61 equiv. ) was added LiOH (75 mg, 3.132 mmol, 17.40 equiv. ) . The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated and purified by reverse column to afford the [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetic acid.
LC/MS: 633.3 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (2- ( (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) methoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 21)
To a solution of [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) methoxy] acetic acid (110 mg, 0.170 mmol, 1.00 equiv. ) in DMF (5.00 mL) was added TEA (86.05 mg, 0.850 mmol, 5 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (87.88 mg, 0.204 mmol, 1.2 equiv. ) , HATU (77.60 mg, 0.204 mmol, 1.2 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 20 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the crude and the crude product was purified by Prep-HPLC with the following conditions: Column: Kinetex EVO C18 Column, 30*150, 5um; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 39%to 46%in 9 min, then 46%to 100%in 1 min, hold 100%in 1 min, down to 39%in 1 min, hold 39%in 1 min; Wave Length: 254 nm to afford Compound 21 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) methoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ: 10.15 (brs, 1H) , 9.00 (s, 1H) , 8.85 (s, 1H) , 8.58 -8.68 (m, 1H) , 8.00 -8.10 (m, 1H) , 7.70 -7.80 (m, 1H) , 7.45 -7.64 (m, 4H) , 7.40 (s, 4H) , 6.91 (d, J = 9.0 Hz, 2H) , 5.56 -5.78 (m, 1H) , 5.32 (s, 1H) , 5.15 (brs, 1H) , 4.92 -5.02 (m, 1H) , 4.76 -4.87 (m, 1H) , 4.50 -4.75 (m, 5H) , 4.18 -4.48 (m, 4H) , 3.90 -4.10 (m, 2H) , 3.50 -3.72 (m, 4H) , 3.10 (brs, 4H) , 2.40 (s, 6H) , 2.20 -2.39 (m, 3H) , 1.80 -2.15 (m, 2H) , 1.40 -1.65 (m, 10H) , 0.95 (s, 9H) .
LC/MS: 1045.5 [M+H] +.
Compound 22
Step 1. [ (3- { [5- (tert-butoxy) pentyl] oxy} propoxy) methyl] benzene
To a stirred solution of 3- (benzyloxy) propan-1-ol (1 g, 6.016 mmol, 1.00 equiv. ) and tert-butyl 5-bromopentanoate (2.14 g, 9.024 mmol, 1.5 equiv. ) in NaOH (12 mL, 300.022 mmol, 49.87 equiv. ) , DCM (12 mL, 188.761 mmol, 31.38 equiv. ) was added tetra butyl ammonium chloride (1.67 g, 6.016 mmol, 1 equiv. ) in portions at r.t. for overnight. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with water (2 x 100 mL) and brine (2 x 100 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-20%EtOAc /PE) to afford [ (3- { [5- (tert-butoxy) pentyl] oxy} propoxy) methyl] benzene.
LC/MS: 229.18 [M+H] +.
Step 2.3- { [5- (tert-butoxy) pentyl] oxy} propan-1-ol
To a stirred solution of [ (3- { [5- (tert-butoxy) pentyl] oxy} propoxy) methyl] benzene (780 mg, 2.529 mmol, 1.00 equiv. ) and Pd/C (500 mg, 4.698 mmol, 1.86 equiv. ) in MeOH (5 mL, 123.495 mmol, 48.84 equiv. ) were stirred at r.t. under H2 atmosphere overnight, then concentrated to afford 3- { [5- (tert-butoxy) pentyl] oxy} propan-1-ol.
LC/MS: 219.19 [M+H] +.
Step 3. tert-butyl 5- (3-iodopropoxy) pentanoate
To a solution of tert-butyl 5- (3-hydroxypropoxy) pentanoate (550 mg, 2.367 mmol, 1.00 equiv. ) in DCM (5 mL) was added imidazole (193.40 mg, 2.840 mmol, 1.2 equiv. ) , I2 (660.96 mg, 2.604 mmol, 1.1 equiv. ) and PPh3 (683.04 mg, 2.604 mmol, 1.1 equiv. ) . The resulting mixture was stirred at r.t. overnight, the reaction was quenched with Na2S2O3 (20 mL) . The resulting mixture was extracted with EtOAc (3 x 20 mL) , washed with brine (2 x 30 mL) The organic layers were combined then concentrated. The residue obtained was purified by silica gel chromatography (10-20%EtOAc/petroleum ether) to afford tert-butyl 5- (3-iodopropoxy) pentanoate.
LC/MS: 343.07, [M+H] +.
Step 4. tert-butyl 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoat
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (30 mg, 0.062 mmol, 1.00 equiv. ) in DMF (5 mL) , was added K2CO3 (42.61 mg, 0.310 mmol, 5 equiv. ) methane; tert-butyl 5- (3-iodopropoxy) pentanoate (24.30 mg, 0.068 mmol, 1.1 equiv. ) . The resulting mixture was stirred at r.t. for 4 h. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue was purified by Prep-TLC (MeOH/DCM 1: 8) to afford 20 mg of tert-butyl 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoate.
LC/MS: 701.41 [M+H] +.
Step 5. 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoic acid
To a solution of tert-butyl 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoate (180 mg, 1 equiv. ) in DCM (5 mL) , added TFA (1.5 mL) The resulting mixture was stirred at r.t. for 1.5 h. The reaction was concentrated, to afford 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoic acid.
LC/MS: 645.35 [M+H] +.
Step 6. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 22)
To a solution of 5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanoic acid (160 mg, 0.248 mmol) in DMF (5 mL) was added TEA (125.55 mg, 1.240 mmol) , HATU (122.66 mg, 0.322 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (106.85 mg, 0.248 mmol) , The resulting mixture was stirred at r.t. for 2 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /8: 1) then further purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1NH3. H2O; Phase B: CH3CN (40%CH3CN up to 60%in 8 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 60%CH3CN in 0.1min, hold 40%in 0.9min) ; Detector, UV220 &254nm to afford 73.9 mg of Compound 22 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.10 (brs, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.51 -8.57 (m, 1H) , 8.00 -8.18 (m, 1H) , 7.87 (d, J = 8.8 Hz, 1H) , 7.74 -7.76 (m, 1H) , 7.57 -7.62 (m, 3H) , 7.38 -7.40
(m, 4H) , 6.92 (d, J = 8.8 Hz, 2H) , 5.60 -5.73 (m, 1H) , 5.33 (s, 1H) , 5.11 -5.12 (m, 1H) , 5.00 (d, J = 1.2 Hz, 1H) , 4.85 (d, J = 1.2 Hz, 1H) , 4.64 -4.73 (m, 2H) , 4.55 (d, J = 9.6 Hz, 1H) , 4.40 -4.48 (m, 2H) , 4.21 -4.32 (m, 1H) , 4.20 -4.26 (m, 1H) , 3.62 -3.71 (m, 2H) , 3.38 -3.42 (m, 4H) , 3.09 -3.21 (m, 4H) , 2.45 -2.50 (m, 7H) , 2.33 -2.39 (m, 3H) , 2.20 -2.27 (m, 1H) , 2.01 -2.16 (m, 1H) , 1.89 -1.96 (m, 1H) , 1.64 -1.70 (m, 2H) , 1.52 -1.54 (m, 10H) , 0.95 (s, 9H) .
LC/MS: 1057.54 [M+H] +.
Compound 23
Step 1. 10, 10, 11, 11-tetramethyl-1-phenyl-2, 6, 9-trioxa-10-siladodecane
To a solution of 2- [ (tert-butyldimethylsilyl) oxy] ethanol (2.0 g, 11.342 mmol, 1.00 equiv. ) in DMF (30 mL, 387.653 mmol, 34.18 equiv. ) was added 60%NaH (0.33 g, 13.610 mmol, 1.2 equiv. ) , the mixture was stirred for 1 h at 0 ℃ then added [ (3-bromopropoxy) methyl] benzene (2.86 g, 12.476 mmol, 1.1 equiv. ) . The resulting mixture was stirred for 2 h at 25 ℃. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with EA (3*50mL) . The residue was purified by silica gel column chromatography, eluted with (0-30%PE/EA) to afford the 10, 10, 11, 11-tetramethyl-1-phenyl-2, 6, 9-trioxa-10-siladodecane (1.89 g, 51.34%) 10, 10, 11, 11-tetramethyl-1-phenyl-2, 6, 9-trioxa-10-siladodecane.
Step 2. 2- [3- (benzyloxy) propoxy] ethanol
To a solution of 10, 10, 11, 11-tetramethyl-1-phenyl-2, 6, 9-trioxa-10-siladodecane (1.6 g, 4.930 mmol, 1.00 equiv. ) in 1, 4-dioxane (10 mL, 113.501 mmol, 23.02 equiv. ) was added a solution of HCl in 1, 4-dioxane (10 mL, 329.119 mmol, 66.76 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with water (15 mL) . The resulting mixture was extracted with EA (3*15mL) , washed with brine (4*15mL) . The residue obtained was purified by silica gel chromatography (0-50%ethyl acetate/petroleum ether) to afford the 2- [3- (benzyloxy) propoxy] ethanol.
Step 3. tert-butyl 2- {2- [3- (benzyloxy) propoxy] ethoxy} acetate
To a solution of 2- [3- (benzyloxy) propoxy] ethanol (466 mg, 2.216 mmol, 1.00 equiv. ) in DCM (6 mL, 94.380 mmol, 42.59 equiv. ) was added tert-butyl 2-bromoacetate (648.41 mg, 3.324 mmol, 1.5 equiv. ) , 35%NaOH (6 mL, 150.011 mmol, 67.69 equiv. ) and tetrabutylammonium chloride (615.91 mg, 2.216 mmol, 1 equiv. ) . The mixture was stirred at 25 ℃ overnight. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted with DCM (3 x 15 mL) . The organic layers were combined concentrated. The residue obtained was purified by silica gel column chromatography (0-20%ethyl acetate/petroleum ether) to afford the tert-butyl 2- {2- [3- (benzyloxy) propoxy] ethoxy} acetate.
Step 4. tert-butyl 2- [2- (3-hydroxypropoxy) ethoxy] acetate
To a solution of tert-butyl 2- {2- [3- (benzyloxy) propoxy] ethoxy} acetate (353.0 mg, 1.088 mmol, 1.00 equiv. ) in MeOH (5 mL, 123.495 mmol, 113.49 equiv. ) was added Pd/C (300 mg, 2.819 mmol, 2.59 equiv. ) . The resulting mixture was maintained under hydrogen and stirred at 40 ℃ overnight. The reaction was filtered and concentrated to afford the tert-butyl 2- [2- (3-hydroxypropoxy) ethoxy] acetate.
Step 5. tert-butyl 2- [2- (3-iodopropoxy) ethoxy] acetate
To a solution of tert-butyl 2- [2- (3-hydroxypropoxy) ethoxy] acetate (227.0 mg, 0.969 mmol, 1.00 equiv. ) in DCM (4 mL, 62.920 mmol, 64.94 equiv. ) was added Ph3P (279.53 mg, 1.066 mmol, 1.1 equiv. ) , imidazole (79.15 mg, 1.163 mmol, 1.2 equiv. ) and I2 (270.50 mg, 1.066 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1h. The reaction was quenched with Na2S2O3 (15 mL) . The resulting mixture was extracted with DCM (3*15 mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (0-30%ethyl acetate/petroleum ether) to afford the tert-butyl 2- [2- (3-iodopropoxy) ethoxy] acetate.
Step 6. tert-butyl 2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200.00 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , tert-butyl 2- [2- (3-iodopropoxy) ethoxy] acetate (155.62 mg, 0.452 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 50 ℃ for 3 h. After cooling down to rt, the reaction was quenched with water (15 mL) . The resulting mixture was extracted with EA (3*15mL) , washed with brine (6*15 mL) . The organic layers were combined and concentrated. The residue was purified by Prep-TLC (DCM/MeOH 14/1) to afford the tert-butyl 2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetate.
LC/MS: 703.45 [M+H] +.
Step 7. [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetic acid
To a solution of tert-butyl 2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetate (166.00 mg, 0.236 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 333.01 equiv. ) was added TFA (3 mL, 40.389 mmol, 171.01 equiv. ) The resulting mixture was stirred at 25 ℃ for 1 h . The reaction was concentrated to afford the [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetic acid.
LC/MS: 647.25 [M+H] +.
Step 8. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 23)
To a solution of [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetic acid (150 mg, 0.232 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 278.57 equiv. ) was added TEA (211.22 mg, 2.088 mmol, 9 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (119.83 mg, 0.278 mmol, 1.2 equiv. ) and HATU (114.64 mg, 0.302 mmol, 1.3 equiv. ) . The mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, washed with brine (4 x 15 mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then purified by Prep-HPLC with the following conditions: Column, XBridge Shield RP18 OBD column, 19*150 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3+0.1%NH3. H2O, Mobile Phase B: CH3CN (38 %to 45%in 11 min, 45 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 38%in 0.1 min, 38 %to 38%in 1 min; Detector, UV 254 nm to afford the Compound 23 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ10.15 (brs, 1 H) , 8.97 (s, 1 H) , 8.82 (s, 1 H) , 8.60 (t, J = 7.6 Hz, 1 H) , 8.05 (t, J = 7.6 Hz, 1 H) , 7.71 -7.80 (m, 1 H) , 7.55 -7.62 (m, 3 H) , 7.40 -7.43 (m, 5 H) , 6.90 (d, J = 9.6 Hz, 2 H) , 5.60 -5.72 (m, 1 H) , 5.32 (s, 1 H) , 5.16 (d, J = 3.6 Hz, 1 H) , 4.97 -5.01 (m, 1 H) , 4.79 -4.89 (m, 1 H) , 4.61 -4.71 (m, 2 H) , 4.51 -4.60 (m, 1 H) , 4.32 -4.50 (m, 3 H) , 4.19 -4.31 (m, 1 H) , 4.00 (s, 2 H) , 3.60 -3.70 (m, 4 H) , 3.51 -3.59 (m, 2 H) , 3.40 -3.50 (m, 2 H) , 3.00 -3.11 (m, 4 H) , 2.48 -2.50 (m, 4H) , 2.45 (s, 3 H) , 2.31 -2.39 (m, 2 H) , 2.01 -2.11 (m, 1 H) , 1.89 -1.98 (m, 1 H) , 1.71 -1.73 (m, 2 H) , 1.46 (s, 6 H) , 0.96 (s, 9 H) .
LC/MS: 1059.70 [M+H] +.
Compound 24
Step 1. methyl 6- [2- (benzyloxy) ethoxy] hexanoate
To a solution of benzyl glycol (2 g, 13.141 mmol, 1.00 equiv. ) in DMF (15 mL, 193.826 mmol, 14.75 equiv. ) was added 60%NaH (0.38 g, 15.769 mmol, 1.2 equiv. ) , the mixture was stirred for 1 h at 0 ℃ then added [ (3-bromopropoxy) methyl] benzene (2.86 g, 12.476 mmol, 1.1 equiv. ) . The resulting mixture was stirred for 2 h at 25 ℃. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with EA (3*50mL) . The residue was purified by silica gel column chromatography, eluted with (0-30%PE/EA) to afford the methyl 6- [2- (benzyloxy) ethoxy] hexanoate.
Step 2. methyl 6- (2-hydroxyethoxy) hexanoate
To a solution of methyl 6- [2- (benzyloxy) ethoxy] hexanoate (660 mg, 2.354 mmol, 1.00 equiv. ) in MeOH (6 mL, 148.193 mmol, 62.95 equiv. ) was added Pd/C (550 mg, 5.168 mmol, 2.20 equiv. ) . The resulting mixture was maintained under hydrogen and stirred at 40 ℃ overnight. The reaction was filtered and concentrated to afford the methyl 6- (2-hydroxyethoxy) hexanoate.
Step 3. methyl 6- (2-iodoethoxy) hexanoate
To a solution of methyl 6- (2-hydroxyethoxy) hexanoate (398 mg, 2.092 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 37.59 equiv. ) was added PPh3 (603.60 mg, 2.301 mmol, 1.1 equiv. ) , imidazole (170.91 mg, 2.510 mmol, 1.2 equiv. ) and I2 (584.09 mg, 2.301 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1h. The reaction was quenched with Na2S2O3 (15 mL) . The resulting mixture was extracted with DCM (3*15 mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (0-30%ethyl acetate/petroleum ether) to afford the methyl 6- (2-iodoethoxy) hexanoate.
Step 4. methyl 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) , methyl 6- (2-iodoethoxy) hexanoate (135.70 mg, 0.452 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 50 ℃ for 3 h. After cooling down to rt, the reaction was quenched with water (15 mL) . The resulting mixture was extracted with EA (3*15mL) , washed with brine (6*15 mL) . The organic layers were combined and concentrated. The residue was purified by Prep-TLC (DCM/MeOH 14/1) to afford methyl 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoate.
LC/MS: 659.45 [M+H] +.
Step 5. 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoic acid
To a solution of methyl 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoate (139.0 mg, 0.211 mmol, 1.00 equiv. ) in THF (3 mL, 37.029 mmol, 175.50 equiv. ) and H2O (3 mL,
166.525 mmol, 789.26 equiv. ) was added LiOH. H2O (44.27 mg, 1.055 mmol, 5 equiv. ) . The resulting mixture was stirred at 25 ℃ 1 h. The pH value of the resulting mixture was adjusted to 6 using 2N HCl solution. Then extracted with EA (3*15 mL) , washed with brine (4*15mL) . The organic layers were combined, dried over Na2SO4, filtered and concentrated to afford the 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoic acid.
LC/MS: 645.30 [M+H] +.
Step 6. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 24)
To a solution of 6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanoic acid (100 mg, 0.155 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 416.58 equiv. ) was added TEA (141.24 mg, 1.395 mmol, 9 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (80.13 mg, 0.186 mmol, 1.2 equiv. ) and HATU (76.66 mg, 0.202 mmol, 1.3 equiv. ) . The mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The organic layers were combined, washed with brine (4 x 15 mL) . The residue obtained was purified by TLC (1 : 8 MeOH/DCM) then purified by Prep-HPLC with the following conditions: Column, YMC-Actus Triart C18 ExRS, 30*150 mm 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3+0.1%NH3. H2O, Mobile Phase B: CH3CN (36 %to 60%in 8 min, 60 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 36%in 0.1 min, 36 %to 36%in 1 min; Detector, UV 254 nm to afford the Compound 24 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ10.14 (brs, 1 H) , 8.98 (s, 1 H) , 8.82 (s, 1 H) , 8.55 (t, J = 6.7 Hz, 1 H) , 8.00 -8.10 (m, 1 H) , 7.70 -7.89 (m, 2 H) , 7.52 -7.67 (m, 3 H) , 7.37 -7.47 (m, 4 H) , 6.89 -6.99 (m, 2 H) , 5.57 -5.71 (m, 1 H) , 5.30 (s, 1 H) , 5.12 (d, J = 3.2 Hz, 1 H) , 4.98 -5.01 (m, 1 H) , 4.80 -4.90 (m, 1 H) , 4.65 -4.71 (m, 2 H) , 4.51 -4.60 (m, 1 H) , 4.40 -4.50 (m, 2 H) , 4.29 -4.39 (m, 1 H) , 4.20 -4.25 (m, 1 H) , 3.61 -3.70 (m, 2 H) , 3.51 (t, J = 6.0 Hz, 2 H) , 3.32 (s, 2 H) , 3.10
(brs, 4 H) , 2.59 -2.61 (m, 3 H) , 2.51 (s, 3 H) , 2.44 (s, 3 H) , 2.21 -2.31 (m, 1 H) , 2.10 -2.20 (m, 1 H) , 2.00 -2.09 (m, 1 H) , 1.89 -1.99 (m, 1 H) , 1.46 -1.52 (m, 10 H) , 1.21 -1.31 (m, 2 H) , 0.94 (s, 9 H) .
LC/MS: 1057.70 [M+H] +.
Compound 25
Step 1. 3- [2- (benzyloxy) ethoxy] propan-1-ol
To a solution of 3- [ (tert-butyldimethylsilyl) oxy] propan-1-ol (2.0 g, 10.507 mmol, 1.00 equiv. ) in DMF (30 mL, 387.653 mmol, 36.90 equiv. ) was added 60%NaH (0.55 g, 13.659 mmol, 1.3
equiv. ) , the mixture was stirred for 1 h at 0 ℃ then added [ (2-bromoethoxy) methyl] benzene (2.71 g, 12.608 mmol, 1.2 equiv. ) . The resulting mixture was stirred for 4 h at 25 ℃. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with EA (3*50mL) . The residue was purified by silica gel column chromatography, eluted with (0-30%PE/EA) to afford the 3- [2- (benzyloxy) ethoxy] propan-1-ol.
Step 2. tert-butyl 2- {3- [2- (benzyloxy) ethoxy] propoxy} acetate
To a solution of 3- [2- (benzyloxy) ethoxy] propan-1-ol (537.00 mg, 2.554 mmol, 1.00 equiv. ) in DCM (7 mL, 110.110 mmol, 43.12 equiv. ) was added tert-butyl 2-bromoacetate (747.21 mg, 3.831 mmol, 1.5 equiv. ) , 35%NaOH (7 mL, 175.013 mmol, 68.53 equiv. ) and tetrabutylammonium chloride (709.75 mg, 2.554 mmol, 1 equiv. ) . The mixture was stirred at 25 ℃ overnight. The reaction was quenched with H2O (50 mL) . The resulting mixture was extracted with DCM (3*50 mL) . The organic layers were combined concentrated. The residue obtained was purified by silica gel column chromatography (0-30%ethyl acetate/petroleum ether) to afford the tert-butyl 2- {3- [2- (benzyloxy) ethoxy] propoxy} acetate.
Step 3. tert-butyl 2- [3- (2-hydroxyethoxy) propoxy] acetate
To a solution of tert-butyl 2- {3- [2- (benzyloxy) ethoxy] propoxy} acetate (331.00 mg, 1.020 mmol, 1.00 equiv. ) in MeOH (5 mL, 123.495 mmol, 121.04 equiv. ) was added Pd/C (300.00 mg, 2.819 mmol, 2.76 equiv. ) . The resulting mixture was maintained under hydrogen and stirred at 40 ℃ overnight. The reaction was filtered and concentrated to afford the tert-butyl 2- [3- (2-hydroxyethoxy) propoxy] acetate.
Step 4. tert-butyl 2- [3- (2-iodoethoxy) propoxy] acetate
To a solution of tert-butyl 2- [3- (2-hydroxyethoxy) propoxy] acetate (236.00 mg, 1.007 mmol, 1.00 equiv. ) in DCM (5 mL, 78.650 mmol, 78.08 equiv. ) was added Ph3P (290.62 mg, 1.108 mmol, 1.1 equiv. ) , imidazole (82.29 mg, 1.208 mmol, 1.2 equiv. ) , I2 (281.23 mg, 1.108 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1h. The reaction was quenched with Na2S2O3 (15 mL) . The resulting mixture was extracted with DCM (3*15 mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (0-30%ethyl acetate/petroleum ether) to afford the tert-butyl 2- [3- (2-iodoethoxy) propoxy] acetate.
Step 5. tert-butyl 2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-
yl} ethoxy) propoxy] acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 157.19 equiv. ) was added tert-butyl 2- [3- (2-iodoethoxy) propoxy] acetate (155.62 mg, 0.452 mmol, 1.1 equiv. ) , K2CO3 (284.03 mg, 2.055 mmol, 5 equiv. ) . The resulting mixture was stirred at 50 ℃ for 4 h. The reaction was quenched with H2O (15 mL) . The resulting mixture was extracted with EtOAc (3 x 100 mL) . The combined organic layers were washed with brine (5 x 100 mL) then concentrated. The residue obtained was purified by TLC (MeOH: DCM=1: 8) to afford tert-butyl 2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetate.
LC/MS: 703.39 [M+H] +.
Step 6. [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetic acid)
To a solution of tert-butyl 2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetate (190 mg) in DCM (5 mL) was added TFA (1.5 mL) . The resulting mixture was stirred at 25 ℃ for 1 h. The resulting mixture was concentrated under vacuum to afford [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetic acid) .
LC/MS: 647.33 [M+H] +.
Step 7. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) 154 yridine-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 25)
To a solution of [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) 154yridine-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetic acid (160 mg, 0.247 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (125.17 mg, 1.235 mmol, 5 equiv. ) , HATU (122.29 mg, 0.321 mmol, 1.3 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-
carboxamide (106.52 mg, 0.247 mmol, 1 equiv. ) , The resulting mixture was stirred at r.t. for 2 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3 x 15 mL) . The combined organic layers were washed with brine (5 x 20 mL) then concentrated. The residue obtained was purified by TLC (DCM : MeOH /8: 1) then further purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1NH3. H2O; Phase B: CH3CN (30%CH3CN up to 52%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 52%CH3CN in 0.1min, hold 30%in 0.9min) ; Detector, UV220 &254nm to afford 84.7 mg of Compound 25 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) 155 yridine-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 0.18 (brs, 1H) , 9.00 (s, 1H) , 8.82 (s, 1H) , 8.51 -8.68 (m, 1H) , 7.92 -8.17 (m, 1H) , 7.70 -7.81 (m, 1H) , 7.53 -7.64 (m, 3H) , 7.48 -7.52 (m, 5H) , 6.77 -7.01 (m, 2H) , 5.56 -5.73 (m, 1H) , 5.35 (s, 1H) , 5.15 (s, 1H) , 4.99 (d, J = 8.0 Hz, 1H) , 4.81 (d, J = 8 Hz, 1H) , 4.60 -4.76 (m, 2H) , 4.58 (d, J = 12.0 Hz, 1H) , 4.32 -4.50 (m, 3H) , 4.18 -4.31 (m, 1H) , 3.88 -4.02 (m, 2H) , 3.46 -3.74 (m, 9H) , 3.02 -3.10 (m, 4H) , 2.53 -2.60 (m, 4H) , 2.43 (s, 3H) , 2.00 -2.17 (m, 1H) , 1.87 -1.99 (m, 1H) , 1.74 -1.84 (m, 2H) , 1.46 (s, 7H) , 0.96 (s, 9H) .
LC/MS: 1059.52 [M+H] +.
Compound 26
Step 1. tert-butyl 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol) and tert-butyl 3- [2- (2-bromoethoxy) ethoxy] propanoate (134.37 mg, 0.452 mmol) in DMF (8 mL) was added K2CO3 (170.42 mg, 1.233 mmol) at room temperature. The resulting mixture was stirred at 50 ℃ for 2 hours. The reaction was quenched with H2O (20 mL) . The resulting mixture was extracted with
EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (4 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%DCM/MeOH) to afford tert-butyl 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoate.
LC/MS: 703.40 [M+H] +.
Step 2. 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoic acid
To a tert-butyl 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoate (190 mg, 0.270 mmol) in DCM (5 mL) was added TFA (1.5 mL, 20.195 mmol) . The resulting mixture was stirred at room temperature for 1.5 h. Then the solution was dissolved in DCM (30*2 mL) and concentrated under reduced pressure to afford 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoic acid.
LC/MS: 647.30 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 26)
To a solution of 3- (2- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) ethoxy) propanoic acid (190 mg, 0.294 mmol) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (126 mg, 0.294 mmol) in DMF (5 mL) was added HATU (145 mg, 0.382 mmol) and TEA (0.5 mL) . The mixture was stirred at room temperature overnight. The reaction solution was quenched with water and extracted with ethyl acetate, the combined organic layers was washed with water and brine and dried over Na2SO4, the solid be removed, the filtrate was concentrated under reduced pressure. The residue was purified by flash chromatography to give 100 mg of product as yellow solid. The residue was purified by Prep-HPLC with the following conditions:
column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 35%B to 45%B in 10 min, 45%B; Wavelength: 254 nm. This resulted in 50.2 mg of Compound 26 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {3- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] propanamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ: 10.18 (brs, 1H) , 8.98 (s, 1H) , 8.82 (s, 1H) , 8.50 -8.70 (m, 1H) , 8.00 -8.16 (m, 1H) , 7.90 -7.99 (m, 1H) , 7.69 -7.82 (m, 1H) , 7.51 -7.68 (m, 3H) , 7.31 -7.50 (m, 4H) , 6.86 -7.02 (m, 2H) , 5.55 -5.78 (m, 1H) , 5.33 (s, 1H) , 5.13 (s, 1H) , 4.92 -5.02 (m, 1H) , 4.77 -4.91 (m, 1H) , 4.68 (s, 2H) , 4.54 -4.62 (m, 1H) , 4.40 -4.51 (m, 2H) , 4.35 (s, 1H) , 4.10 -4.32 (m, 1H) , 3.40 -3.80 (m, 11H) , 3.00 -3.19 (m, 4H) , 2.55 -2.65 (m, 5H) , 2.30 -2.46 (m, 5H) , 2.01 -2.11 (m, 1H) , 1.86 -1.98 (m, 1H) , 1.45 (s, 6H) , 0.93 (s, 9H) .
LC/MS: [M+H] +.
Compound 27
Step 1. Methyl 4- (trifluoromethanesulfonyloxy) cyclohex-3-ene-1-carboxylate
To a solution of methyl 4-oxocyclohexane-1-carboxylate (1.0 g, 6.403 mmol) and 2, 6-di-tert-butyl-4-methylpyridine (1.97 g, 9.604 mmol) in DCM (20.0 mL) was added Tf2O (1.99 g, 7.043 mmol) at 0 ℃ under N2. The resulting mixture was stirred overnight at room temperature under N2. The resulting mixture was extracted with DCM (3 x 20 mL) . The combined organic layers were washed with brine (2x10 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column
chromatography, eluted with PE/EA (3: 1) to afford methyl 4- (trifluoromethanesulfonyloxy) cyclohex-3-ene-1-carboxylate.
Step 2. Methyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} cyclohex-3-ene-1-carboxylate
To a solution of methyl 4- (trifluoromethanesulfonyloxy) cyclohex-3-ene-1-carboxylate (1.22 g, 4.233 mmol) and 2- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane (1.35 g, 5.080 mmol) in 1, 4-dioxane (25.0 mL) and H2O (5 mL) was added K3PO4 (1.80 g, 8.466 mmol) and Pd (PPh3) 4 (0.4 g, 0.339 mmol) at room temperature under N2. The resulting mixture was stirred overnight at 100 ℃ under N2. Desired product could be detected by LCMS. The resulting mixture was extracted with EA (3 x 30 mL) . The combined organic layers were washed with brine (2x10 mL) , dried over anhydrous NaSO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE/EA (3: 1) to methyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} cyclohex-3-ene-1-carboxylate.
LC/MS: 279.15 [M+H] +.
Step 3. Methyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexane-1-carboxylate
To a solution of methyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} cyclohex-3-ene-1-carboxylate (870 mg 3.126 mmol) in MeOH (15 mL) was added Pd/C (0.3 g, 10%) . The resulting mixture was stirred at room temperature for 1.5 h under H2. The resulting mixture was filtered with diatomite and washed with MeOH. The solvent was removed under vacuum to afford methyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexane-1-carboxylate.
LC/MS: 283.20 [M+H] + .
Step 4. Methyl 4'-oxo- [1, 1'-bi (cyclohexane) ] -4-carboxylate
The solution of methyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexane-1-carboxylate (830 mg, 2.939 mmol) in TFA (15 mL) was stirred overnight at 60 ℃ in air atmosphere. The reaction mixture was concentrated under reduced pressure. Then the residue was purified by normal phase chromatography (mobile phase: EA/PE, 0 ~ 30%) to afford methyl 4'-oxo- [1, 1'-bi (cyclohexane) ] -4-carboxylate.
LC/MS: 239.20 [M+H] +.
Step 5. Methyl 4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250 mg, 0.514 mmol, 1.0 equiv. ) and methyl 4'-oxo- [1, 1'-bi (cyclohexane) ] -4-carboxylate (183.68 mg, 0.771 mmol, 1.5 equiv. ) in MeOH (10 mL) was added AcOH (20.0 mg) at room temperature in air atmosphere. The resulting mixture was stirred for 1 h at 60 ℃ in air atmosphere. To the above mixture was added NaBH3CN (51.66 mg, 0.822 mmol, 1.6 equiv. ) at room temperature. The resulting mixture was stirred for additional for 5 h at room temperature. Desired product could be detected by LCMS. The reaction was quenched with sat. NaHCO3 (aq. ) at room temperature. The resulting mixture was extracted with EA (3 x 10 mL) . The combined organic layers were dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford methyl 4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylate.
LC/MS: 709.70 [M+H] +.
Step 6. 4'- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylic acid
To a solution of methyl 4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylate (240 mg, 0.339 mmol, 1.0 equiv. ) in THF (4.5 mL) and MeOH (2.5 mL) was added LiOH (2.48 mL, 3 mol/L, 7.458 mmol, 22.0 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred overnight at room temperature in air atmosphere. Desired product could be detected by LCMS. The reaction was concentrated under reduced pressure and purified by C18 column with ACN/H2O (0.5%TFA) to afford 4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylic acid.
LC/MS: 695.70 [M+H] +.
Step 7. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [ (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-
bi (cyclohexane) ] -4-yl) formamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 27)
To a solution of 4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-carboxylic acid (190 mg, 0.273 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (129.5 mg, 0.300 mmol, 1.1 equiv. ) and HATU (155.95 mg, 0.410 mmol, 1.5 equiv. ) in DMF (4 mL) was added DIEA (141.35 mg, 1.094 mmol, 4.0 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred overnight at room temperature in air atmosphere. Desired product could be detected by LCMS. the reaction mixture was further purified by Prep-HPLC (Column: Xselect CSH C18 OBD Column 30*150mm 5μm, n; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 44%B to 64%B in 9 min, 64%B; Wave Length: 254 nm; RT1 (min) : 9; Number Of Runs: 0) to afford 100.8 mg of Compound 27 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [ (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) formamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.14 (s, 1H) , 8.99 (d, J = 1.8 Hz, 1H) , 8.82 (d, J = 1.0 Hz, 1H) , 8.56 (t, J = 6.1 Hz, 1H) , 8.03 (d, J = 8.7 Hz, 1H) , 7.75 (d, J = 8.1 Hz, 2H) , 7.68 (m, 3H) , 7.35 -7.48 (m, 4H) , 6.91 (dd, J = 8.4, 3.5 Hz, 2H) , 5.66 (m, 1H) , 5.32 (s, 1H) , 5.13 (dt, J = 3.4, 1.6 Hz, 1H) , 4.99 (m, 1H) , 4.82 (m, 1H) , 4.68 (d, J = 6.1 Hz, 2H) , 4.35 -4.52 (m, 4H) , 4.22 (m, 1H) , 3.60 -3.72 (m, 2H) , 3.08 (s, 4H) , 2.57 -2.63 (m, 4H) , 2.45 (d, J = 1.5 Hz, 3H) , 2.27 (s, 1H) , 2.18 (s, 1H) , 1.67 -1.90 (m, 8H) , 1.46 (s, 9H) , 1.27 -1.41 (m, 3H) , 1.21 (m, 1H) , 1.14 (s, 2H) , 0.83 -0.99 (m, 12H) .
LC/MS: 1107.65 [M+H] +.
Compound 28
Step 1. methyl 4- (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) cyclohexane-1-carboxylate
To a solution of methyl 4-oxocyclohexane-1-carboxylate (600 mg, 3.842 mmol, 1.00 equiv. ) in DCE (10 mL, 126.315 mmol, 32.88 equiv. ) was added 1, 4-dioxa-8-azaspiro [4.5] decane (550.08 mg, 3.842 mmol, 1 equiv. ) and a drop of CH3COOH. The mixture was stirred at rt for 1 h. Then NaBH (OAc) 3 (1628.43 mg, 7.684 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3 h. After completion, the reaction was quenched with water (30 mL) . The pH value of the solution was adjusted to 8~9 with NaHCO3. The mixture was extracted with EA (3x50 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%PE: EtOAc) . The organic layers were concentrated under reduced pressure to afford methyl 4- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} cyclohexane-1-carboxylate.
LC/MS: 284 [M+H] +.
Step 2. methyl 4- (4-oxopiperidin-1-yl) cyclohexane-1-carboxylate
A mixture of methyl 4- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} cyclohexane-1-carboxylate (470 mg, 1.660 mmol, 1.00 equiv. ) in trifluoroacetaldehyde (5 mL) equiv was stirred at 60℃ for 16 h. After completion, the organic layers were concentrated. The pH value of the solution was adjusted to 8~9 with NaHCO3. The mixture was extracted with DCM/MeOH (10: 1, 3x20 mL ) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (MeOH/DCM, 0~10%) . The organic layers were concentrated under reduced pressure to afford methyl 4- (4-oxopiperidin-1-yl) cyclohexane-1-carboxylate.
LC/MS: 240 [M+H] +.
Step 3. methyl 4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) cyclohexane-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) in MeOH (5 mL, 123.495 mmol, 89.37 equiv. ) was added methyl 4- (4-oxopiperidin-1-yl) cyclohexane-1-carboxylate (98.37 mg, 0.411 mmol, 1 equiv. ) and CH3COOH (0.01 mL, 0.175 mmol, 0.13 equiv. ) . The mixture was stirred at rt for 1 h. After that, NaBH3CN (51.66 mg, 0.822 mmol, 2 equiv. ) was added. Upon completion, the reaction was quenched with water (10 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford methyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexane-1-carboxylate.
LC/MS: 710 [M+H] +.
Step 4. 4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) cyclohexane-1-carboxylic acid
To a solution of methyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexane-1-carboxylate (104 mg, 0.147 mmol, 1.00 equiv. ) in THF (2 mL, 24.686 mmol, 168.50 equiv. ) and H2O (2 mL, 111.017 mmol, 757.79 equiv. ) was added LiOH (17.54 mg, 0.735
mmol, 5 equiv. ) . The mixture was stirred at rt for 2 h. Upon completion. The pH value of the solution was adjusted to 4~5 with HCl. The crude product was purified by reverse-phase flash with the following conditions: Column, Cat No: SO230120-2, C18, 120 g, 20~45μm, Lot: BP0002P2503; mobile phase, CH3CN: H2O (0.05%TFA) = 20%increased to CH3CN: H2O (0.05%TFA) = 70%in 40 min, hold CH3CN: H2O (0.05%TFA) = 70%in 10 min, up to CH3CN: H2O (0.05%TFA) = 95%in 2 min, hold CH3CN: H2O (0.05%TFA) = 95%in 10 min; Detector, UV 220 nm &254 nm. The organic layers were concentrated under reduced pressure to afford 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexane-1-carboxylic acid.
LC/MS: 696 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) cyclohexane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 28)
To a solution of 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexane-1-carboxylic acid (77 mg, 0.111 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609 mmol, 583.89 equiv. ) was added TEA (33.59 mg, 0.333 mmol, 3 equiv. ) , HATU (54.70 mg, 0.144 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (47.64 mg, 0.111 mmol, 1 equiv. ) . The mixture was stirred at rt for 1 h. After completion, The crude product was purified by Prep-HPLC with the following conditions: Column: CHIRALCEL OJ-3, 4.6*50mm 3um; Mobile Phase B: MEOH (0.1%DEA) ; Flow rate: 2 mL/min; Gradient: isocratic 10%B; Wavelength: 220 nm Column: Xselect CSH C18 OBD. After lyophilization, Compound 28 (2S, 4R) -4-hydroxy-1- [ (2S) -2- { [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] formamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide was afforded.
1H NMR (400 MHz, DMSO-d6) δ 10.13 (s, 1H) , 8.98 (s, 1H) , 8.82 (s, 1H) , 8.56 (t, J = 6.1 Hz, 1H) , 8.05 (s, 1H) , 7.75 (d, J = 8.1 Hz, 1H) , 7.64 -7.56 (m, 3H) , 7.50 -7.35 (m, 4H) , 6.91 (d, J = 8.9 Hz, 2H) , 5.66 (ddt, J = 16.5, 10.2, 6.0 Hz, 1H) , 5.32 (s, 1H) , 5.13 (s, 1H) , 4.99 (dt, J = 10.3, 1.4 Hz, 1H) , 4.82 (dd, J = 17.2, 1.6 Hz, 1H) , 4.68 (d, J = 5.9 Hz, 2H) , 4.54 (d, J = 9.3 Hz, 1H) ,
4.48 -4.33 (m, 2H) , 4.22 (dd, J = 15.8, 5.5 Hz, 2H) , 3.72 –3.60 (m, 2H) , 3.07 (d, J = 6.1 Hz, 4H) , 2.96 (d, J = 9.8 Hz, 2H) , 2.62 (t, J = 4.8 Hz, 3H) , 2.45 (s, 5H) , 2.16 (s, 2H) , 1.78 -2.03 (m, 10H) , 1.31 -1.46 (m, 12H) , 0.94 (s, 9H) .
LC/MS: 1108.65 [M+H] +.
Compound 29
Step 1. tert-butyl 4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidine-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.822 mmol, 1 equiv. ) in DCE (10 mL) , MeOH (10 mL) was added AcOH (74.05 mg, 1.233 mmol, 1.5 equiv. ) , 2 h later, NaBH3CN (0.08 mL, 1.233 mmol, 1.5 equiv. ) was added. The resulting mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (10 mL) . The mixture was basified to pH~9 with NaHCO3 aqueous solution and extracted with EA (3 x 50 mL) , washed with brine (3 x 70 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidine-1-carboxylate.
LC/MS: 670.4 [M+H] +.
Step 2. 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (piperidin-4-yl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidine-1-carboxylate (660 mg, 0.985 mmol, 1 equiv. ) in dioxane (10 mL) was added a solution of HCl in 1, 4-dioxane (10 mL, 329.119 mmol, 334.02 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated under vacuum to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- ( {4- [4- (piperidin-4-yl) piperazin-1-yl] phenyl} amino) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 570.3 [M+H] +.
Step 3. tert-butyl 4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 4'-bipiperidine] -1'-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- ( {4- [4- (piperidin-4-yl) piperazin-1-yl] phenyl} amino) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (135 mg, 0.237 mmol, 1 equiv. ) in MeOH (2 mL) , DCE (2 mL) was added tert-butyl 4-oxopiperidine-1-carboxylate (70.82 mg, 0.355 mmol, 1.5 equiv. ) , AcOH (21.34 mg, 0.355 mmol, 1.5 equiv. ) , 2 h later, NaBH3CN (22.34 mg, 0.355 mmol, 1.5 equiv. ) was added. The resulting mixture was stirred at 25 ℃ for 3 h. The reaction was quenched with H2O (10 mL) , the mixture was basified to pH~9 with NaHCO3 aqueous solution and extracted with DCM/MeOH (10: 1, 30 mL*3) , washed with brine (3 x 50
mL) , the organic layers were combined and concentrated under vacuum to afford tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidine] -1'-carboxylate.
LC/MS: 753.4 [M+H] +.
Step 4. 6- ( (4- (4- ( [1, 4'-bipiperidin] -4-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidine] -1'-carboxylate (285 mg, 0.379 mmol, 1 equiv. ) in dioxane (5 mL) was added HCl (gas) in 1, 4-dioxane (10 mL, 329.119 mmol, 869.53 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated under vacuum to afford the crude of 6- { [4- (4- { [1, 4'-bipiperidin] -4-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 653.4 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- ( (S) -2-isocyanato-3, 3-dimethylbutanoyl) -N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide
To a solution of (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (30 mg, 0.070 mmol, 1 equiv. ) in DCM (10 mL) was added TEA (14.10 mg, 0.140 mmol, 2 equiv. ) , Triphosgene (41.35 mg, 0.140 mmol, 2 equiv. ) . The resulting mixture was stirred at 25 ℃ for 2 h. The reaction was concentrated under vacuum. The residue was added EA, filtered, and concentrated to afford (2S, 4R) -4-hydroxy-1- [ (2S) -2-isocyanato-3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
LC/MS: 457.2 [M+H] +.
Step 6. 4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) - [1, 4'-bipiperidine] -1'-carboxamide (Compound 29)
To a solution of 6- { [4- (4- { [1, 4'-bipiperidin] -4-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (220 mg, 0.337 mmol, 1 equiv. ) in DCM (5 mL, 78.650 mmol, 233.39 equiv. ) was added triethylamine (102.30 mg, 1.011 mmol, 3 equiv. ) , (2S, 4R) -4-hydroxy-1- [ (2S) -2-isocyanato-3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (200.01 mg, 0.438 mmol, 1.3 equiv. ) . The resulting mixture was stirred at 25 ℃ for 16 h. The reaction was concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford crude and the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 10%to 64%in 10 min, then 64%to 100%in 1 min, hold 100%in 1 min, down to 10%in 1 min, hold 10%in 1 min; Wave Length: 254 nm to afford Compound 29 N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidine] -1'-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ: 10.12 (brs, 1H) , 9.02 (s, 1H) , 8.82 (s, 1H) , 8.50 -8.60 (m, 1H) , 7.95 -8.12 (m, 1H) , 7.70 -7.85 (m, 1H) , 7.50 -7.68 (m, 3H) , 7.40 (s, 4H) , 6.91 (d, J = 8.7 Hz, 2H) , 5.84 (d, J = 9.0 Hz, 1H) , 5.55 -5.80 (m, 1H) , 5.30 (s, 1H) , 5.11 (d, J = 3.6 Hz, 1H) , 4.99 (d, J = 9.6 Hz, 1H) , 4.70 -4.86 (m, 1H) , 4.66 (s, 2H) , 4.15 -4.50 (m, 5H) , 3.90 -4.10 (m, 2H) , 3.55 -3.75 (m, 2H) , 3.10 (brs, 4H) , 2.78 -2.93 (m, 2H) , 2.51 -2.60 (m, 4H) , 2.45 (s, 3H) , 2.00 -2.30 (m, 5H) , 1.55 -1.95 (m, 4H) , 1.45 (s, 7H) , 1.15 -1.40 (m, 5H) , 0.91 (s, 9H) .
LC/MS: 1109.85 [M+H] +.
Compound 30
Step 1. ethyl 1- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-4-carboxylate
To a solution of ethyl piperidine-4-carboxylate (600 mg, 3.816 mmol, 1.00 equiv. ) and 1, 4-dioxaspiro [4.5] decan-8-one (596.06 mg, 3.816 mmol, 1 equiv. ) in DCE (10 mL) was added CH3COOH (11.46 mg, 0.191 mmol, 0.05 equiv. ) . After 1 h, the NaBH (OAc) 3 (1617.74 mg, 7.632 mmol, 2 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (30 mL) . The pH value of the resulting mixture was adjusted to 7 by using NaHCO3 solution. Then extracted with DCM (3*30mL) , washed with brine (4*30mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford the ethyl 1- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-4-carboxylate.
LC/MS: 298.35 [M+H] +.
Step 2. ethyl 1- (4-oxocyclohexyl) piperidine-4-carboxylate
To a solution of ethyl 1- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-4-carboxylate (580 mg, 1.950 mmol, 1.00 equiv. ) in TFA (5 mL) . The resulting mixture was stirred at 60 ℃ for 2 h. After cooling down to rt. The reaction was concentrated. The pH value of the resulting mixture was adjusted to 8 using NaHCO3 solution. Then extracted with DCM/MeOH (10: 1, 3*30mL) , washed with brine (2*50mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford the ethyl 1- (4-oxocyclohexyl) piperidine-4-carboxylate.
Step 3. ethyl 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1 equiv. ) in MeOH (10 mL) was added CH3COOH (0.25 mg, 0.004 mmol, 0.01 equiv. ) . After 1 h, the NaBH3CN (51.66 mg, 0.822 mmol, 2 equiv. ) . The resulting mixture was stirred at 60 ℃ overnight. The reaction was quenched with H2O (50 mL) , then extracted with EA (3*50mL) . The organic layers were combined and concentrated. The residue obtained was purified by TLC (DCM/MeOH = 5/1) to afford the ethyl 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylate.
LC/MS: 724.40 [M+H] +.
Step 4. 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylic acid
To a solution of ethyl 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylate (150 mg, 0.207 mmol, 1 equiv. ) in THF (5 mL) was added LiOH. H2O (86.95 mg, 2.070 mmol, 10 equiv. ) , H2O (5 mL) . The resulting mixture was stirred at 25 ℃ overnight. The reaction was concentrated then purified by reverse phase column (A: TFA/H2O, B: CH3CN) to
afford the 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylic acid.
LC/MS: 696.40 [M+H] +.
Step 5. N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxamide (Compound 30)
To a solution of 1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxylic acid (140 mg, 0.201 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (101.79 mg, 1.005 mmol, 5 equiv. ) , HATU (99.45 mg, 0.261 mmol, 1.3 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (86.63 mg, 0.201 mmol, 1 equiv. ) . The mixture was stirred at 25 ℃ for 1 h. The reaction was quenched with H2O (5 mL) . The resulting mixture was extracted with EtOAc (3*15 mL) . The organic layers were combined and concentrated. The residue obtained was purified by Prep-HPLC with the following conditions: Column, XSelect CSH Prep C18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water with 10 mmol/L NH4HCO3, Mobile Phase B: CH3CN (33 %to 44%in 12 min, 44 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 33 %in 0.1 min, 33 %to 33 %in 1 min; Detector, UV 254 nm to afford the Compound 30 N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxamide (cis) .
1H NMR (400 MHz, DMSO-d6) δ 8.40 -98.90 (m, 3H) , 8.02 -8.06 (m, 1H) , 7.70 -7.77 (m, 2H) , 7.54 -7.64 (m, 3H) , 7.35 -7.45 (m, 4H) , 6.90 (d, J = 8.8 Hz, 2H) , 5.59 -5.73 (m, 1H) , 5.30 -5.34 (m, 1H) , 5.11 -5.16 (m, 1H) , 4.99 (d, J = 10.4 Hz, 1H) , 4.78 -4.87 (m, 1H) , 4.66 -4.71 (m, 2H) , 4.48 -4.55 (m, 1H) , 4.30 -4.48 (m, 3H) , 4.00 -4.30 (m, 1H) , 3.58 -3.68 (m, 2H) , 3.04 -3.09 (m, 4H) , 2.79 -2.86 (m, 2H) , 2.61 -2.67 (m, 4H) , 2.46 (s, 3H) , 2.19 -2.24 (m, 2H) , 2.11 -2.17 (m, 2H) , 1.98 -2.08 (m, 1H) , 1.60 -1.98 (m, 7H) , 1.50 -1.60 (m, 3H) , 1.46 (s, 6H) , 1.19 -1.25 (m, 4H) , 0.91 (s, 9H) .
LC/MS: 1108.65 [M+H] +.
And Compound 30 N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxamide (trans) .
1H NMR (400 MHz, DMSO-d6) δ 8.65 -9.00 (m, 2H) , 8.56 (t, J = 6.2 Hz, 1H) , 7.98 -8.15 (m, 1H) , 7.65 -7.90 (m, 2H) , 7.50 -7.64 (m, 3H ) , 7.45 -7.35 (m, 4H) , 6.91 (d, J = 8.6 Hz, 2H) , 5.73 -5.59 (m, 1H) , 5.34 -5.30 (m, 1H) , 5.15 -5.11 (m, 1H) , 5.03 -4.96 (m, 1H) , 4.87 -4.78 (m, 1H) , 4.71 -4.65 (m, 2H) , 4.52 (d, J = 9.3 Hz, 1H) , 4.48 -4.38 (m, 2H) , 4.37 -4.33 (m, 1H) , 4.26 -4.17 (m, 1H) , 3.68 -3.58 (m, 2H) , 3.12 -3.07 (m, 4H) , 2.99 -2.92 (m, 2H) , 2.60 -2.56 (m, 4H) , 2.44 (s, 3H) , 2.21 -2.16 (m, 2H) , 1.65 -2.08 (m, 10H ) , 1.50 -1.60 (m, 3H) , 1.46 (s, 6H) , 1.30 -1.40 (m, 4H) , 0.93 (s, 9H) .
LC/MS: 1108.65 [M+H] +.
Compound 31
Step 1. Tert-butyl 4- (trifluoromethanesulfonyloxy) -3, 6-dihydro-2H-pyridine-1-carboxylate
Tert-butyl 4-oxopiperidine-1-carboxylate (5 g, 25.094 mmol) , a stir bar, THF (40 ml) were added to a 250 mL three-neck round-bottom flask and stirred until homogeneous, then treated with LiHMDS (30 ml) at -78℃ under a nitrogen atmosphere. The reaction mixture was stirred at room temperature for 1 h. To the above mixture was added 1, 1, 1-trifluoro-N-phenyl-N-trifluoromethanesulfonylmethanesulfonamide (9.9 g, 27.712 mmol) at 0 ℃. The resulting mixture
was stirred overnight at room temperature. The above mixture was quenched with sat. aq. NH4Cl (100 mL) and extracted with EA (100 mL x 3) . The combined extracts were dried over anhydrous sodium sulfate, filtered, and concentrated to dryness. The residue was subjected to silica gel chromatography (PE/EA) to afford tert-butyl 4- (trifluoromethanesulfonyloxy) -3, 6-dihydro-2H-pyridine-1-carboxylate.
LC/MS : 332.20 [M+H] +.
Step 2. Tert-butyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -3, 6-dihydro-2H-pyridine-1-carboxylate
Tert-butyl 4- (trifluoromethanesulfonyloxy) -3, 6-dihydro-2H-pyridine-1-carboxylate (4 g, 12.073 mmol) and 2- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane (3.37 g, 12.677 mmol, ) , a stir bar, and 1, 4-dioxane (50 mL) and H2O (10 mL) were added to a 250 ml round-bottom flask and stirred until homogeneous, and then treated with K3PO4 (5.13 g, 24.146 mmol) and tetrakis (triphenylphosphine) palladium (0) (1.40 g, 1.207 mmol) . The resulting mixture was stirred at 100 ℃ overnight under a nitrogen atmosphere, then cooled to r.t., diluted with water (100 mL) , and extracted with EA (100 mL x 3) . The combined extracts were dried over anhydrous sodium sulfate, filtered, and concentrated to dryness to give a nearly yellow viscous oil. The oil was subjected to silica gel chromatography (0-70%EtOAc/pet ether) to afford tert-butyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -3, 6-dihydro-2H-pyridine-1-carboxylate.
LC/MS : 322.20 [M+H] +.
Step 3. Tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-1-carboxylate acetate
Tert-butyl 4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -3, 6-dihydro-2H-pyridine-1-carboxylate (1.49 g, 4.636 mmol) , a stir bar, and MeOH (25 mL) were added to a 100 ml round-bottom flask and stirred until homogeneous, then treated with 10%Pd/C (4 g, 3.755 mmol) . The resulting mixture was stirred at room temperature overnight under a hydrogen atmosphere. The resulting mixture was filtered, the filter cake was washed with MeOH. The filtrate was concentrated under reduced pressure to afford tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-1-carboxylate acetate.
LC/MS : 326.25 [M+H] +.
Step 4. 4- (piperidin-4-yl) cyclohexan-1-one
Tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperidine-1-carboxylate (1.33 g, 4.087 mmol) , a stir bar, and TFA (30 mL) were added to a 100 ml round-bottom flask and stirred until homogeneous. The resulting mixture was stirred at 60℃ overnight under a nitrogen atmosphere. The reaction mixture was concentrated under reduced pressure to afford 4- (piperidin-4-yl) cyclohexan-1-one which was used for the next step directly.
LC/MS: 182.15 [M+H] +.
Step 5. Tert-butyl 4- (4-oxocyclohexyl) piperidine-1-carboxylate
4- (piperidin-4-yl) cyclohexan-1-one (3 g, 16.549 mmol) and di-tert-butyl dicarbonate (2.98 g, 13.342 mmol) , a stir bar, and THF (20 mL) were added to a 250 mL round-bottom flask and stirred until homogeneous, then treated with TEA (2.23 g, 22.038 mmol) and DMAP (0.21 g, 1.719 mmol) . The resulting mixture was stirred at room temperature for 4 h. The resulting mixture was diluted with H2O (100 mL) and extracted with EtOAc (3 x 100 mL) . The combined extracts were washed with brine (3 x 100 mL) , dried over anhydrous Na2SO4, filtered and concentrated to dryness under reduced pressure. The residue obtained was purified by silica gel chromatography (0-60%EtOAc/petroleum ether) to afford tert-butyl 4- (4-oxocyclohexyl) piperidine-1-carboxylate.
LC/MS : 282.25 [M+H] +.
Step 6. Tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate
1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (864 mg, 1.776 mmol) and tert-butyl 4- (4-oxocyclohexyl) piperidine-1-carboxylate (749 mg, 2.652 mmol) , a stir bar, and MeOH (25 mL) were added to a 100 mL round-bottom flask and stirred until homogeneous, then treated with AcOH (10 mg) . The resulting mixture was stirred for 1 h. To the above mixture was added NaBH3CN (167.93 mg, 2.672 mmol) at room temperature. The resulting mixture was stirred for 4 h, then quenched with sat. aq NaHCO3 (50 mL) and extracted with EtOAc (100 mL x 3) . The combined extracts were dried over anhydrous sodium sulfate, filtered, and concentrated to dryness. The residue was subjected to silica gel chromatography (0-15%MeOH/DCM) to afford tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-
yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate.
LC/MS : 753.90 [M+H] +.
Step 7. 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperidin-4-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-1-carboxylate (716 mg, 0.952 mmole) , a stir bar, TFA (4 mL) and DCM (20 mL) were added to a 100 mL round-bottom flask. The resulting mixture was stirred at room temperature for 2 h. The resulting mixture was concentrated under reduced pressure and purified by C18 column with CH3CN/5mM NH4HCO3 water (5%-70%) to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperidin-4-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS (: 652.65 [M+H] +.
Step 8. N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-1-carboxamide (Compound 31)
(2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (660 mg, 1.533 mmol) and Et3N (400 mg, 4.605 mmol) , a stir bar, DCM (20 mL) were added to a 250 mL round-bottom flask and stirred until homogeneous, then treated with triphosgene (273.14 mg, 0.921 mmol) at 0 ℃ under a nitrogen atmosphere. The resulting mixture was stirred at r.t. for 1 h under a nitrogen atmosphere. The reaction mixture was concentrated in vacuo and then dissolved in EA (40 mL) and filtered. The filtrate was concentrated to dryness, then dissolved in DCM (5 mL) , added to a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperidin-4-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.307 mmol) and Et3N (65.70 mg) in DCM (5 mL) . The mixture was concentrated under reduced pressure to afford the crude product which was purified by Prep-HPLC [Column: XBridge Prep OBD C18 Column, 19*250 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 40%B to 75%B in 7 min, 75%B; Wave Length: 220
nm; RT1 (min) : 5.35 6.3; Number Of Runs: 0] to afford Compound 31 N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-1-carboxamide.
1H NMR (400 MHz, DMSO) δ 10.16 (s, 1H) , 8.99 (d, J = 3.4 Hz, 1H) , 8.83 (d, J = 1.1 Hz, 1H) , 8.57 (s, 1H) , 8.05 (s, 1H) , 7.76 (d, J = 8.3 Hz, 1H) , 7.41 -7.61 (m, 3H) , 7.40 (d, J = 2.1 Hz, 5H) , 6.92 (d, J = 8.8 Hz, 2H) , 5.62 -5.77 (m, 2H) , 5.34 (s, 1H) , 5.12 (d, J = 3.6 Hz, 1H) , 5.00 (d, J = 10.0 Hz, 1H) , 4.83 (d, J = 17.3 Hz, 1H) , 4.69 (s, 2H) , 4.39 -4.43 (m, 5H) , 4.00 (s, 2H) , 3.68 (d, J = 11.6 Hz, 2H) , 3.08 (s, 4H) , 2.52 -2.60 (m, 4H) , 2.42 -2.51 (m, 3H) , 2.19 (s, 1H) , 1.97 -2.05 (m, 1H) , 1.90 (s, 2H) , 1.71 -1.82 (m, 3H) , 1.36 -1.47 (m, 8H) , 1.24 -1.36 (m, 6H) , 0.85 -0.94 (m, 12H) .
LC/MS: 1108.65 [M+H] +.
Compound 32
Step 1. Tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate
To a solution of 1, 4-dioxaspiro [4.5] decan-8-one (1 g, 6.403 mmol) in MeOH (10 mL) was added tert-butyl piperazine-1-carboxylate (1.19 g, 6.403 mmol) and acetic acid (0.02 g, 0.320 mmol) . The resulting mixture was maintained at 60 ℃ for 1 h. After cooling down to r.t., the NaBH3CN (0.80 g, 12.806 mmol) was added. The resulting mixture was maintained at r.t. for 1h. NaHCO3 solution was added, and the resulting mixture was extracted with ethyl acetate (2 x 30 mL) . The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-15%MeOH/DCM) to afford the tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate.
LC/MS: 327.40 [M+H] +.
Step 2. 4- (Piperazin-1-yl) cyclohexan-1-one
The mixture of tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate (1.5 g, 4.595 mmol) in TFA (10 mL) was stirred at 60 ℃ overnight. The solvent was removed under vacuum and the residue was purified by silica gel column with MeOH/DCM (0%-40%) to afford the 4- (piperazin-1-yl) cyclohexan-1-one (0.8 g, 95.4%) . LC/MS : 183.15 [M+H] +.
Step 3. Tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate
To a solution of 4- (piperazin-1-yl) cyclohexan-1-one (600 mg, 3.292 mmol) , TEA (999.31 mg, 9.876 mmol) in DCM (10 mL) was added (Boc) 2O (862.13 mg, 3.950 mmol) at 0℃. The mixture was stirred at r.t. overnight. The solvent was removed under vacuum and the residue was purified by silica gel column with MeOH/DCM (0%-40%) to afford the tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate.
LC/MS: 283.20 [M+H] +.
Step 4. Tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (150 mg, 0.308 mmol) in MeOH (10 mL) was added tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate (174.10 mg, 0.616 mmol) and acetic acid (0.1 mL) . The resulting mixture was maintained at 60 ℃ for 1 h. After cooling down to r.t., the NaBH3CN (23.25 mg, 0.370 mmol) was added. The resulting mixture was maintained at r.t. for 1h. NaHCO3 solution was added, and the resulting mixture was extracted with ethyl acetate (2 x 30 mL) . The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-15%MeOH/DCM) to afford the tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate.
LC/MS: 753.60 [M+H] +.
Step 5. 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperazin-1-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate (180 mg, 0.239 mmol) in DCM (3 mL) was added TFA (3 mL) . The resulting mixture was maintained at 60℃ for 1 h. The solvent was removed under vacuum and the residue was purified by C18 column with CH3CN/NH4HCO3 water (0.5%) (5%-100%) to afford the 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperazin-1-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 653.60 [M+H] +.
Step 6. 4- (4- (4- (4- ( (2-Allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) piperazine-1-carboxamide (Compound 32)
To a solution of (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (369 mg, 0.86 mmol) and triethylamine (326 mg, 3.21 mmol) in DCM (10 mL) was added triphosgene (45 mg, 0.15 mmol) at 0 ℃. The resulting mixture was maintained under nitrogen and stirred at r.t. for 1 h. The mixture was concentrated in vacuum, and the residue was dissolved in EA (10 mL) . A small amount of undissolved material was removed by decantation. The organic layer was concentrated under reduced pressure to afford crude product. Then to a solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (4- (piperazin-1-yl) cyclohexyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (140 mg, 0.21 mmol) and triethylamine (43 mg, 0.43 mmol) in DCM (10 mL) was added the crude product slowly. The mixture was stirred at r.t. for 1h. The mixture was concentrated under reduced pressure to afford the crude product which was purified by Prep-HPLC [Column: XBridge Prep OBD C18 Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 30%B to 60%B in 7 min, 60%B; Wave Length: 254 nm; RT1 (min) : 6; Number Of Runs: 0] to afford the Compound 32 4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) piperazine-1-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.99 (s, 1H) , 8.83 (s, 1H) , 8.57 (t, J = 6.1 Hz, 1H) , 8.05 (t, J = 8.1 Hz, 1H) , 7.76 (d, J = 8.1 Hz, 1H) , 7.51 -7.66 (m, 3H) , 7.30 -7.49 (m, 5H) , 6.92 (d, J = 8.7 Hz, 2H) , 5.84 (d, J = 9.0 Hz, 1H) , 5.53 -5.74 (m, 1H) , 5.34 (s, 1H) , 5.13 (d, J = 3.6 Hz, 1H) , 5.00 (dd, J = 10.3, 1.5 Hz, 1H) , 4.83 (dd, J = 17.2, 1.6 Hz, 1H) , 4.69 (d, J = 6.0 Hz, 2H) , 4.32 -4.54 (m, 4H) , 4.15 -4.22 (m, 1H) , 3.60 -3.75 (m, 2H) , 3.10 (s, 4H) , 2.58 -2.71 (m, 4H) , 2.43 -2.47 (m, 3H) , 2.34 -2.43 (m, 4H) , 2.19 (s, 2H) , 1.92 -2.08 (m, 1H) , 1.68 -1.89 (m, 4H) , 1.32 -1.58 (m, 11H) , 0.95 (s, 10H) . LC/MS: 1109.65 [M+H] +.
Compound 33
Step 1. Methyl 2- [4- (trifluoromethanesulfonyloxy) cyclohex-3-en-1-yl] acetate
Methyl 2- (4-oxocyclohexyl) acetate (2.0 g 10.856 mmol) , 2, 6-di-tert-butyl-4-methylpyridine (3.3 g 16.284 mmol) and DCM (50 mL) were added to a 100 mL round-bottom flask with a stir bar
under N2 and at 0℃. Tf2O (4.3 g 15.198 mmol) were added after the mixture cooled to 0℃ and stirred at r.t. for 18 h. DCM were added to the mixture and then washed by water and saturated sodium chloride, the organic layer was separated and dried over anhydrous sodium sulfate. The solvent was removed under vacuum and the residue was purified by silica gel column with EA/PE from 0~50%to afford methyl 2- [4- (trifluoromethanesulfonyloxy) cyclohex-3-en-1-yl] acetate.
Step 2. Ethyl 2- (4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} cyclohex-3-en-1-yl) acetate
Methyl 2- [4- (trifluoromethanesulfonyloxy) cyclohex-3-en-1-yl] acetate (1 g 3.162 mmol) , 2- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} -4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane (840 mg 3.162 mmol) , Pd (PPh3) 4 (365 mg 0.316 mmol) , K3PO4 (1 g 4.743 mmol) , dioxane (50 mL ) and H2O (10 mL) were added to a 100 mL round-bottom flask with a stir bar. The flask was evacuated and flushed three times with nitrogen and the mixture was stirred 5.0 h at 100℃. EA were added to the mixture and then washed by water, the organic layer was separated and dried over anhydrous sodium sulfate. The solvent was removed under vacuum and the residue was purified by silica gel column with EA/PE (0~50%) to afford ethyl 2- (4- {1, 4-dioxaspiro [4.5] dec-7-en-8-yl} cyclohex-3-en-1-yl) acetate.
LC/MS: 307.40 [M+H] +.
Step 3. Ethyl 2- (4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexyl) acetate
To a 100 mL round-bottom flask containing a stirring bar, ethyl 2- (4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexyl) acetate (700 mg 2.28 mmol) , MeOH (15 mL) and Pd/C (10%) were added. The flask was evacuated and flushed three times with hydrogen and the mixture was stirred 16.0 h at room temperature. And then filtered with diatomite and washed with MeOH. The solvent was removed under vacuum to afford ethyl 2- (4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexyl) acetate.
LC/MS: 311.30 [M+H] + .
Step 4. Ethyl 2- {4'-oxo- [1, 1'-bi (cyclohexane) ] -4-yl} acetate
Methyl 2- (4- {1, 4-dioxaspiro [4.5] decan-8-yl} cyclohexyl) acetate (600 mg, 1.933 mmol) and trifluoroacetaldehyde (6 mL) were added to a round-bottom flask with a stir bar and then reacted at 60℃ for 1 h. The solvent was removed under vacuum and the residue was purified by silica gel column with DCM/MeOH from 0~20%to afford ethyl 2- {4'-oxo- [1, 1'-bi (cyclohexane) ] -4-yl} acetate.
LC/MS: 267.80 [M+H] + .
Step 5. Ethyl 2- (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250 mg, 0.514 mmol, 1 equiv. ) , ethyl 2- {4'-oxo- [1, 1'-bi (cyclohexane) ] -4-yl} acetate (410.59 mg, 1.542 mmol, 3 equiv. ) , MeOH (10 mL) and AcOH (3.09 mg) were added to a 100 mL round-bottom flask with a stir bar. Then stirred at 60℃ for 1 h. Then NaBH3CN (64.58 mg, 1.028 mmol, 2 equiv. ) were added after the mixture cooled to room temperature and then stirred at room temperature for 16 h. EA (50 mL) was added after the mixture cooled to room temperature and washed by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by silica gel column with DCM/MeOH from 0~20%to afford ethyl 2- (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetate.
LC/MS: 737.25 [M+H] +.
Step 6. (4'- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetic acid
Ethyl 2- (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetate (250 mg, 0.339 mmol, 1 equiv. ) , lithiumol (1.13 mL, 3 M in H2O) , MeOH (2 mL) and tetrahydrofuran (6 mL) were added to a round-bottom flask. And then stirred at r.t. for 2 h. The mixture was purified by C18 column with H2O (0.05%TFA) /ACN from 5~100%. The solvent was removed under vacuum to afford (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetic acid (230 mg, 95.64%yield) .
LC/MS: 709.50 [M+H] +.
Step 7. (2S, 4R) -1- ( (S) -2- (2- (4'- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 1'-
bi (cyclohexan) ] -4-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 33)
(4'- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) acetic acid (135 mg, 0.190 mmol) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (82 mg, 0.190 mmol) , HATU (109 mg, 0.285 mmol) , DIEA (74 mg, 0.570 mmol) and DMF (5 mL) were added in a round-bottom flask with a stir bar. And then stirred at r.t. for 18 h. The residue obtained was purified by Prep-HPLC with the following conditions: Column, XBridge Prep OBD C18 Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN (40 %to 70%in 7 min, 70 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 40 %in 0.1 min, 40 %to 40 %in 1 min; Detector, UV 254 nm to afford Compound 33 (2S, 4R) -1- ( (S) -2- (2- (4'- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 1'-bi (cyclohexan) ] -4-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.99 (d, J = 1.5 Hz, 1H) , 8.83 (s, 1H) , 8.53 -8.62 (m, 1H) , 8.05 (t, J = 8.0 Hz, 1H) , 7.75 -7.89 (m, 2H) , 7.51 -7.66 (m, 3H) , 7.35 -7.46 (m, 4H) , 6.92 (d, J = 8.7 Hz, 2H) , 5.55 -5.81 (m, 1H) , 5.34 (s, 1H) , 5.12 (d, J = 3.4 Hz, 1H) , 5.00 (dd, J = 10.2, 1.6 Hz, 1H) , 4.75 -4.89 (m, 1H) , 4.69 (d, J = 6.0 Hz, 2H) , 4.30 -4.60 (m, 4H) , 4.13 -4.29 (m, 1H) , 3.66 (s, 2H) , 3.08 (s, 4H) , 2.53 -2.68 (m, 4H) , 2.45 (d, J = 1.5 Hz, 3H) , 2.10 -2.21 (m, 2H) , 1.94 -2.09 (m, 2H) , 1.81 -1.93 (m, 2H) , 1.62 -1.80 (s, 4H) , 1.29 -1.55 (m, 13H) , 1.24 (s, 2H) , 1.15 (s, 2H) , 0.80 -1.00 (m, 12H) .
LC/MS: 1121.65 [M+H] +.
Compound 34
Step 1. ethyl 2- (4- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} cyclohexyl) acetate
To a solution of ethyl 2- (4-oxocyclohexyl) acetate (600 mg, 3.257mmol, 1 equiv. ) in DCE (5 mL) was added 1, 4-dioxa-8-azaspiro [4.5] decane (466.32 mg, 3.257mmol, 1equiv. ) , CH3COOH (1.96 mg, 0.033mmol, 0.01equiv. ) . The resulting mixture was stirred at r.t. for 0.5 h, then added NaBH (OAc) 3 (1380.46 mg, 6.514mmol, 2equiv. ) , stirred at r.t. overnight. The reaction was quenched with H2O (mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (2 x 100mL) then concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford ethyl 2- (4- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} cyclohexyl) acetate.
LC/MS: 312.21 [M+H] +.
Step 2. ethyl 2- [4- (4-oxopiperidin-1-yl) cyclohexyl] acetate
To a solution of ethyl 2- (4- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} cyclohexyl) acetate (600 mg, 1.927mmol, 1equiv. ) in TFA (5 mL) , The resulting mixture was stirred at 60℃ for 16 h . The reaction was concentrated and basified to pH 7 with aq. NaHCO3. The resulting mixture was extracted with EtOAc (3 x 50mL) . The combined organic layers were washed with brine (2 x 100mL) then concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 10~20%) to afford ethyl 2- [4- (4-oxopiperidin-1-yl) cyclohexyl] acetate.
LC/MS: 268.19 [M+H] +.
Step 3. ethyl 2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) -3H-pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411mmol, 1.00equiv. ) in MeOH (5 mL) was added ethyl 2- [4- (4-oxopiperidin-1-yl) cyclohexyl] acetate (142.87 mg, 0.534mmol, 1.3equiv. ) , CH3COOH (0.25 mg, 0.004mmol, 0.01equiv. ) The resulting mixture was stirred at 60℃ for 1.5 h, then added NaBH3CN (38.75 mg, 0.616mmol, 1.5equiv. ) , the resulting mixture was stirred at 60℃ for 4 h, then stirred at r.t. overnight. The reaction was quenched with H2O (mL) . The resulting mixture was extracted with EtOAc (3 x 50mL) , then concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford ethyl 2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) -3H-pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetate.
LC/MS: 738.44 [M+H] +.
Step 4. [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetic acid
To a solution of ethyl 2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetate (108 mg, 0.146mmol, 1equiv. ) in THF (3 mL) was added LiOH. H2O (61.41 mg, 1.460mmol, 10equiv. ) , H2O (3 mL) . The resulting mixture was stirred at r.t. for 2 h . The mixture/residue was neutralized to pH 7 with 2N HCl solution. The resulting mixture was filtered,
the filter cake was washed with H2O (5ml) . The filter cake was concentrated under reduced pressure, to afford [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetic acid.
LC/MS: 710.41 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 34)
To a solution of [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetic acid (90 mg, 0.127mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (64.14 mg, 0.635mmol, 5 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (60.05 mg, 0.140mmol, 1.1equiv. ) , HATU (62.67 mg, 0.165mmol, 1.3equiv. ) . The resulting mixture was stirred at r.t. for 2 h. The reaction was quenched with H2O (15mL) . The resulting mixture was extracted with EtOAc (3 x 25mL) . The combined organic layers were washed with brine (5 x 50 mL) then concentrated. The residue obtained was purified by silica gel column ( [DCM: MeOH] [8: 1] ) . then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (50%CH3CN up to 70%in 7 min, up to 100%CH3CN in 1 min, hold 100%in 1 min, down to 70%CH3CN in 0.1min, hold 50%in 0.9min) ; Detector, UV220 &254 nm to afford Compound 34 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.01 (brs, 1H) , 8.99 (s, 1H) , 8.82 -8.84 (m, 1H) , 8.52 -8.61 (m, 1H) , 8.24 (s, 1H) , 8.00 -8.11 (m, 1H) , 7.90 -7.98 (m, 1H) , 7.70 -7.78 (m, 1H) , 7.50 -7.61 (m, 3H) , 7.34 -7.46 (m, 4H) , 6.90 (d, J = 8.0 Hz, 2H) , 5.60 -5.71 (m, 1H) , 5.00 (d, J = 8.0 Hz, 1H) , 4.81 (d, J = 8.0 Hz, 1H) , 4.64 -4.70 (m, 2H) , 4.50 -4.59 (m, 1H) , 4.40 -4.44 (m, 2H) , 4.34 -4.40 (m, 1H) , 4.19 -4.27 (m, 1H) , 3.68 (s, 2H) , 3.08 -3.13 (m, 4H) , 2.97 -3.02 (m, 2H) , 2.60 -
2.64 (m, 4H) , 2.36 -2.47 (m, 6H) , 2.16 -2.24 (m, 3H) , 1.98 -2.12 (m, 5H) , 1.90 -1.93 (m, 1H) , 1.72 -1.80 (m, 2H) , 1.33 -1.50 (m, 14H) , 0.97 (s, 9H) .
LC/MS: 1122.60 [M+H] +
.
Compound 35
Step 1. tert-butyl 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 4'-bipiperidin] -1'-yl) acetate
To a solution of 6- { [4- (4- { [1, 4'-bipiperidin] -4-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (150 mg,
0.230 mmol, 1 equiv. ) in DMF (5 mL) was added DIEA (207.87 mg, 1.610 mmol, 7 equiv. ) , tert-butyl 2-bromoacetate (89.63 mg, 0.460 mmol, 2 equiv. ) . The resulting mixture was stirred at 60 ℃ for 12 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3 x 20 mL) , washed with brine (3 x 40 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the tert-butyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetate.
LC/MS: 767.5 [M+H] +.
Step 2. 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 4'-bipiperidin] -1'-yl) acetic acid
To a solution of tert-butyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetate (90 mg, 0.117 mmol, 1 equiv. ) in dioxane (2 mL) was added a solution of HCl in 1, 4-dioxane (6 mL, 197.471 mmol, 1682.88 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated under vacuum to afford the crude of (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetic acid.
LC/MS: 711.4 [M+H] +.
Step 3. (2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 4'-bipiperidin] -1'-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 35)
To a solution of (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetic acid (80 mg, 0.113 mmol, 1 equiv. ) in DMF (3 mL) was added DIEA (43.63 mg, 0.339 mmol, 3 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (48.45 mg, 0.113 mmol, 1 equiv. ) (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (48.45 mg, 0.113 mmol, 1 equiv. ) . The reaction was stirred at rt for 1 h. The reaction was quenched with H2O (10 mL) and extracted with EA (3
x 30 mL) , washed with brine (3 x 40 mL) , the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel column chromatography, eluted with MeOH/DCM to afford the crude and the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 38%to 53%in 10 min, then 53%to 100%in 1 min, hold 100%in 1 min, down to 38%in 1 min, hold 38%in 1 min; Wave Length: 254 nm to afford Compound 35 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ: 10.12 (brs, 1H) , 9.02 (s, 1H) , 8.82 (s, 1H) , 8.60 -8.70 (m, 1H) , 7.95 -8.12 (m, 1H) , 7.70 -7.85 (m, 2H) , 7.50 -7.68 (m, 3H) , 7.35 -7.49 (m, 4H) , 6.92 (d, J = 8.7 Hz, 2H) , 5.55 -5.80 (m, 1H) , 5.30 (brs, 1H) , 5.17 (brs, 1H) , 5.00 (d, J = 10.2 Hz, 1H) , 4.67 -4.86 (m, 1H) , 4.66 (s, 2H) , 4.20-4.55 (m, 5H) , 3.50 -3.75 (m, 2H) , 3.08 -3.10 (m, 5H) , 2.78 -2.98 (m, 5H) , 2.58 (s, 4H) , 2.42 (s, 4H) , 2.00 -2.30 (m, 7H) , 1.83 -1.99 (m, 1H) , 1.68 -1.82 (m, 4H) , 1.45 (s, 8H) , 0.91 (s, 9H) .
LC/MS: 1123.55 [M+H] +.
Compound 36
Step 1. ethyl 2- (1- (1, 4-dioxaspiro [4.5] decan-8-yl) piperidin-4-yl) acetate
To a solution of ethyl 2- (piperidin-4-yl) acetate (600 mg, 3.509 mmol, 1.00 equiv. ) in DCE (10 mL) was added 1, 4-dioxaspiro [4.5] decan-8-one (547.4 mg, 3.509 mmol, 1 equiv. ) and a drop of CH3COOH. The mixture was stirred at rt for 1 h. Then NaBH (OAc) 3 (1487.7 mg, 7.017 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3 h. After completion, the reaction was quenched with water (30 mL) . The pH value of the solution was adjusted to 8~9 with NaHCO3. The mixture was extracted with EA (3x50 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel
column (0-50%PE: EtOAc) . The organic layers were concentrated under reduced pressure to afford ethyl 2- (1- (1, 4-dioxaspiro [4.5] decan-8-yl) piperidin-4-yl) acetate.
LC/MS: 312.2 [M+H] +.
Step 2. ethyl 2- (1- (4-oxocyclohexyl) piperidin-4-yl) acetate
A mixture of ethyl 2- (1- (1, 4-dioxaspiro [4.5] decan-8-yl) piperidin-4-yl) acetate (700 mg, 2.250 mmol, 1.00 equiv. ) in trifluoroacetaldehyde (5 mL) was stirred at 60℃ for 16 h. After completion, the organic layers were concentrated. The pH value of the solution was adjusted to 8~9 with sat. aq. NaHCO3. The mixture was extracted with DCM: MeOH=10: 1 (3x20 mL ) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-10%DCM: MeOH) . The organic layers were concentrated under reduced pressure to afford ethyl 2- (1- (4-oxocyclohexyl) piperidin-4-yl) acetate.
LC/MS: 268.2 [M+H] +.
Step 3. ethyl 2- (1- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-4-yl) acetate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.823 mmol, 1.00 equiv. ) in MeOH (8 mL) was added ethyl 2- (1- (4-oxocyclohexyl) piperidin-4-yl) acetate (220 mg, 0.823 mmol, 1 equiv. ) and CH3COOH (0.03 mL) . The mixture was stirred at rt for 1 h. After that, NaBH3CN (104 mg, 1.645 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3 h. After completion, the reaction was quenched with water (10 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford ethyl 2- (1- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-4-yl) acetate.
LC/MS: 738.4 [M+H] +.
Step 4. 2- (1- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-4-yl) acetic acid
To a solution of ethyl 2- (1- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-4-yl) acetate (300 mg, 0.407 mmol, 1.00 equiv. ) in THF (5 mL) and H2O (5 mL) was added LiOH (84 mg, 2.035 mmol, 5 equiv. ) . The mixture was stirred at rt for 2 h. After completion. The pH value of the solution was adjusted to 4~5 with HCl. The crude product was purified by reverse-phase flash with the following conditions: Column, Cat No: SO230120-2, C18, 120 g, 20~45μm, Lot: BP0002P2503; mobile phase, CH3CN: H2O (0.05%TFA) = 20%increased to CH3CN: H2O (0.05%TFA) = 70%in 40 min, hold CH3CN: H2O (0.05%TFA) = 70%in 10 min, up to CH3CN: H2O (0.05%TFA) = 95%in 2 min, hold CH3CN: H2O (0.05%TFA) = 95%in 10 min; Detector, UV 220 nm &254 nm. The organic layers were concentrated under reduced pressure to afford 2- (1- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-4-yl) acetic acid.
LC/MS: 710.4 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-4-yl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 36)
To a solution of [1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-4-yl] acetic acid (250 mg, 0.352 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (178.18 mg, 1.760 mmol, 5 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (151.63 mg, 0.352 mmol, 1 equiv. ) , HATU (174.07 mg, 0.458 mmol, 1.3 equiv. ) . The resulting mixture was stirred at r.t. for 2 h. The reaction was quenched with H2O (15mL) . The resulting mixture was extracted with EtOAc (3 x 25mL) . The combined organic layers were washed with brine (5 x 50 mL) then concentrated. The residue obtained was purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water with 10mmol/L NH4HCO3+0.1%NH3H2O; Phase B: CH3CN (34%CH3CN up to 44%in 9 min, up to
100%CH3CN in 1 min, hold 100%in 1 min, down to 44%CH3CN in 0.1min, hold 34%in 0.9min) ; Detector, UV220 &254nm to afford Compound 36 (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-4-yl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.01 (brs, 1H) , 8.99 (s, 1H) , 8.80 (s, 1H) , 8.40 –8.69 (m, 1H) , 7.90 –8.19 (m, 1H) , 7.71 –7.90 (m, 2H) , 7.47 –7.68 (m, 3H) , 7.19 –7.34 (m, 4H) , 6.77 -7.08 (m, 2H) , 5.52 –5.81 (m, 1H) , 5.32 (s, 1H) , 4.90 –5.19 (m, 2H) , 4.83 (d, J = 12.0 Hz, 1H) , 4.61 –4.71 (m, 2H) , 4.55 (d, J = 8.0 Hz, 1H) , 4.30 –4.50 (m, 3H) , 4.03 –4.25 (m, 1H) , 3.59 –3.92 (m, 2H) , 2.99 –3.17 (m, 4H) , 2.71 –2.97 (m, 2H) , 2.54 –2.70 (m, 4H) , 2.43 (s, 3H) , 2.12 –2.28 (m, 3H) , 1.99 –2.11 (m, 3H) , 1.69 –1.93 (m, 6H) , 1.52 –1.67 (m, 3H) , 1.30 –1.51 (m, 8H) , 1.20 –1.30 (m, 2H) , 1.07 –1.17 (m, 2H) , 0.82 –1.00 (m, 9H) .
LC/MS: 1122.60 [M+H] +
Compound 37
Step 1. 2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-1-yl) acetic acid
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperidin-4-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (140 mg, 0.215 mmol, 1.0 equiv. ) in MeOH (8 mL) was added glyoxalate (15.90 mg, 0.215 mmol, 1 equiv. ) . The resulting mixture was stirred for 1 h at room temperature. To the above mixture was added NaBH3CN (13.50 mg, 0.215 mmol, 1 equiv. ) . The resulting mixture was stirred for 5 minutes at room temperature. The reaction mixture was purified by C18 column with ACN/H2O (0.05%TFA) to afford [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-1-yl] acetic acid.
LC/MS: 710.70 [M+H] +.
Step 2. (2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperidin-1-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 37)
To a solution of [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-1-yl] acetic acid (150 mg, 0.211 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (100.08 mg, 0.232 mmol, 1.1 equiv. ) , and HATU (120.51 mg, 0.317 mmol, 1.5 equiv. ) in DMF (1 mL) was added DIEA (109.24 mg, 0.844 mmol, 4.0 equiv. ) . The resulting mixture was stirred for 2 h at room temperature. The reaction was purified by C18 column with ACN/H2O (0.05%TFA) . The reaction mixture was further purified by Prep-HPLC (Column: XBridge Prep OBD C18 Column, 19*250 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 40%B to 75%B in 7.5 min, 75%B; Wavelength: 220 nm; RT1 (min) : 5.87.; Number Of Runs: 0) to afford Compound 37. (yield of (2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-1-yl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.16 (s, 1H) , 8.90 -9.10 (m, 1H) , 8.83 (s, 1H) , 8.50 -8.64 (m, 1H) , 8.05 (s, 1H) , 7.75 -7.83 (m, 2H) , 7.61 -7.70 (m, 3H) , 7.36 -7.48 (m, 4H) , 6.91 -7.00 (m, 2H) , 5.67 -5.75 (m, 1H) , 5.34 (s, 1H) , 5.12 -5.20 (m, 1H) , 4.95 -5.10 (m, 1H) , 4.83 (d, J = 17.3 Hz, 1H) , 4.68 (s, 2H) , 4.53 -4.34 (m, 3H) , 4.15 -4.26 (m, 1H) , 3.50 -3.80 (m, 2H) , 3.07 (s, 4H) , 2.85 (s, 3H) , 2.55 -2.60 (m, 4H) , 2.50 -2.43 (m, 3H) , 1.70 -2.17 (m, 11H) , 1.47 -1.60 (m, 8H) , 1.15 -1.30 (m, 7H) , 0.94 -1.05 (m, 10H) .
LC/MS: 1122.50 [M+H] +.
Compound 38
Step 1. Tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate
To a solution of 1, 4-dioxaspiro [4.5] decan-8-one (1 g, 6.403 mmol) in MeOH (10 mL) was added tert-butyl piperazine-1-carboxylate (1.19 g, 6.403 mmol) and acetic acid (0.02 g, 0.320 mmol) . The resulting mixture was maintained at 60 ℃ for 1 h. After cooling down to r.t., the NaBH3CN (0.80 g, 12.806 mmol) was added. The resulting mixture was maintained at r.t. for 1h. The resulting mixture was extracted with ethyl acetate (2 x 30 mL) . The organic layers
were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-15%MeOH/DCM) to afford the tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate.
LC/MS: 327.40 [M+H] +.
Step 2. 4- (piperazin-1-yl) cyclohexan-1-one
The mixture of tert-butyl 4- {1, 4-dioxaspiro [4.5] decan-8-yl} piperazine-1-carboxylate (1.5 g, 4.595 mmol) in trifluoroacetaldehyde (10 mL) was stirred at 60 ℃ overnight. The solvent was removed under vacuum and the residue was purified by silica gel column with MeOH/DCM (0%~40%) to afford the 4- (piperazin-1-yl) cyclohexan-1-one.
LC/MS: 183.15 [M+H] +.
Step 3. Tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate
To a solution of 4- (piperazin-1-yl) cyclohexan-1-one (600 mg, 3.292 mmol) , 4- (piperazin-1-yl) cyclohexan-1-one (600 mg, 3.292 mmol) , TEA (999.31 mg, 9.876 mmol) in DCM (10 mL) was added (Boc) 2O (862.13 mg, 3.950 mmol) at 0 ℃. The mixture was stirred at r.t. overnight. The solvent was removed under vacuum and the residue was purified by silica gel column with MeOH/DCM (0%-40%) to afford the tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate.
LC/MS: 283.20 [M+H] +.
Step 4. Tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (150 mg, 0.308 mmol) in MeOH (10 mL) was added tert-butyl 4- (4-oxocyclohexyl) piperazine-1-carboxylate (174.10 mg, 0.616 mmol) and acetic acid (0.1 mL) . The resulting mixture was maintained at 60 ℃ for 1 h. After cooling down to r.t., the NaBH3CN (23.25 mg, 0.370 mmol) was added. The resulting mixture was maintained at r.t. for 1 h. The resulting mixture was extracted with ethyl acetate (2 x 30 mL) . The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-40%MeOH/DCM) to afford the tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-
oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate.
LC/MS: 753.60 [M+H] +.
Step 5. 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperazin-1-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of tert-butyl 4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazine-1-carboxylate (180 mg, 0.239 mmol) in DCM (3 mL) was added trifluoroacetaldehyde (3 mL) . The resulting mixture was maintained at 60℃ for 1h. The solvent was removed under vacuum and the residue was purified by C18 column with CH3CN/NH4HCO3 water (5%-100%) to afford the 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperazin-1-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 653.60 [M+H] +.
Step 6. [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazin-1-yl] acetic acid
1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- [ (4- {4- [4- (piperazin-1-yl) cyclohexyl] piperazin-1-yl} phenyl) amino] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (280 mg, 0.429 mmol, 1.0 equiv. ) , glyoxylate (32 mg, 0.348 mmol, 0.81 equiv. ) , MeOH (10 mL) were added to a 10 mL schlenk tube with a stir bar and stirred at room temperature for 1 h. The mixture was cooled by an ice-water bath. NaBH3CN (27 mg, 0.0430 mmol, 1.00 equiv. ) was added, and stirred at room temperature for 2 h. Dilute directly with methanol and filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with Acetonitrile/Acid water to afford the product which was used for the next step directly.
LC/MS: 711.65 [M+H] +.
Step 7. (2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperazin-1-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 38)
[4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperazin-1-yl] acetic acid (280 mg, 0.394 mmol, 1 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (169.59 mg, 0.014 mmol, 1 equiv. ) and dimethylformamide (3 ml) were dissolved in 100 mL round-bottom flask then HATU (224.65 mg, 0.591 mmol) and DIEA (168 mg, 1.300 mmol) added to the reaction solution. The reaction mixture was stirred at room temperature for 2 h. LCMS showed the reaction was completed and the reaction mixture was further purified by Prep-HPLC (Column: XBridge Shield RP18 OBD Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 46%B to 66%B in 7 min, 66%B; Wave Length: 254 nm; RT1 (min) : 4.78; Number Of Runs: 0) afford Compound 38 (2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperazin-1-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
LC/MS: 1123.65 [M+H] +.
Compound 39
Step 1. methyl 4- ( (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) methyl) cyclohexane-1-carboxylate
To a solution of methyl 4-formylcyclohexane-1-carboxylate (950 mg, 5.588 mmol, 1.00 equiv. ) in DCE (10 mL) was added 1, 4-dioxa-8-azaspiro [4.5] decane (799 mg, 5.588 mmol, 1 equiv. ) and a drop of CH3COOH. The mixture was stirred at rt for 1 h. Then NaBH (OAc) 3 (2.37 g, 11.176 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3 h. After completion, the reaction was quenched with water (30 mL) . The pH value of the solution was adjusted to 8~9 with sat. aq. NaHCO3. The mixture was extracted with EA (3 x 50 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%PE: EtOAc) . The organic layers were concentrated under reduced
pressure to afford methyl 4- ( (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) methyl) cyclohexane-1-carboxylate.
LC/MS: 298.2 [M+H] +.
Step 2. methyl 4- ( (4-oxopiperidin-1-yl) methyl) cyclohexane-1-carboxylate
A mixture of methyl 4- ( (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) methyl) cyclohexane-1-carboxylate (1.1 g, 3.699 mmol, 1 equiv. ) in trifluoroacetaldehyde (20 mL, 204.032 mmol, 55.16 equiv. ) was stirred at 60 ℃ for 12 h. The mixture was concentrated under reduced pressure to removed TFA. The pH value of the solution was adjusted to 8~9 with NaHCO3. The mixture was extracted with EA (300 mL x 2) , dried over Na2SO4, and filtered. The solvent was removed under vacuum to give the methyl 4- [ (4-oxopiperidin-1-yl) methyl] cyclohexane-1-carboxylate.
LC/MS: 254 [M+H] +.
Step 3. methyl 4- ( (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) methyl) cyclohexane-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (350 mg, 0.719 mmol, 1 equiv. ) in DCE (10 mL, 126.315 mmol, 175.61 equiv. ) was added 4- [ (4-oxopiperidin-1-yl) methyl] cyclohexane-1-carboxylic acid (258.21 mg, 1.079 mmol, 1.5 equiv. ) and AcOH (0.01 mL, 0.175 mmol, 0.24 equiv. ) . The mixture was stirred at rt for 1 h. After that, NaBH3CN (90.41 mg, 1.438 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3 h. After completion, the reaction was quenched with water (20 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford methyl 4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) methyl] cyclohexane-1-carboxylate.
LC/MS: 724 [M+H] +.
Step 4. 4- ( (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) methyl) cyclohexane-1-carboxylic acid
To a solution of methyl 4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) methyl] cyclohexane-1-carboxylate (270 mg, 0.373 mmol, 1 equiv. ) in THF (2 mL) and H2O (2 mL) was added LiOH (89.32 mg, 3.730 mmol, 10 equiv. ) . The mixture was stirred at r.t. for 3 h. After completion, the crude product was purified by reverse-phase flash with the following conditions: Column, Cat No: SO230120-2, C18, 120 g, 20~45μm, Lot: BP0002P2503; mobile phase, CH3CN: H2O (0.05%TFA) = 20%increased to CH3CN: H2O (0.05%TFA) = 70%in 40 min, hold CH3CN: H2O (0.05%TFA) = 70%in 10 min, up to CH3CN: H2O (0.05%TFA) = 95%in 2 min, hold CH3CN: H2O (0.05%TFA) = 95%in 10 min; Detector, UV 220 nm &254 nm. The organic layers were concentrated under reduced pressure to afford 4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) methyl] cyclohexane-1-carboxylic acid.
LCMS: 710 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (4- ( (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) methyl) cyclohexane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 39)
To a solution of 4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) methyl] cyclohexane-1-carboxylic acid (250 mg, 0.352 mmol, 1 equiv. ) in DMF (5 mL, 64.609 mmol, 183.46 equiv. ) was added TEA (106.91 mg, 1.056 mmol, 3 equiv. ) , HATU (174.07 mg, 0.458 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (151.63 mg, 0.352 mmol, 1 equiv. ) . The mixture was stirred at rt for 1 h. After completion. The crude product was purified by Prep-HPLC with the following conditions: Column: CHIRALPAK AS-3, 3.0*50 mm, 3μm; Mobile Phase B: MEOH (0.1%DEA) ; Flow rate: 2 mL/min; Gradient: isocratic 10%B; Wavelength: 220 nm. After lyophilization, it was afforded Compound 39 (2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-
yl) methyl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.57 (t, J = 6.0 Hz, 1H) , 8.05 (s, 1H) , 7.75 (d, J = 8.0 Hz, 1H) , 7.50 -7.73 (m, 3H) , 7.46 -7.35 (m, 4H) , 6.91 (d, J = 8.7 Hz, 2H) , 5.66 (ddt, J = 16.4, 10.2, 6.0 Hz, 1H) , 5.33 (s, 1H) , 5.14 (dd, J = 3.5, 1.5 Hz, 1H) , 4.99 (dd, J = 10.3, 1.4 Hz, 1H) , 4.87 -4.78 (m, 1H) , 4.68 (d, J = 5.9 Hz, 2H) , 4.52 (dd, J = 9.3, 4.2 Hz, 1H) , 4.48 -4.38 (m, 2H) , 4.35 (s, 1H) , 4.22 (dd, J = 15.9, 5.4 Hz, 1H) , 3.65 (q, J = 11.1, 10.0 Hz, 2H) , 3.08 (s, 4H) , 2.85 (s, 2H) , 2.61 (d, J = 5.1 Hz, 4H) , 2.45 (s, 3H) , 2.26 -2.41 (m, 1H) , 2.20 -2.11 (m, 2H) , 2.04 (d, J = 6.3 Hz, 2H) , 1.53 -1.97 (m, 9H) , 1.24 -1.52 (m, 12H) , 0.73 -1.00 (m, 9H) .
LC/MS: 1122.55 [M+H] +.
Compound 40
Step 1. Methyl 1- {1, 4-dioxaspiro [4.5] decan-8-ylmethyl} piperidine-4-carboxylate
1, 4-Dioxaspiro [4.5] decane-8-carbaldehyde (500 mg, 2.938 mmol, 1.0 equiv. ) , methyl piperidine-4-carboxylate (420.62 mg, 2.938 mmol, 1.0 equiv. ) , AcOH (60.21 mg) and MeOH (10 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 1 hour. The mixture was cooled by ice water, NaBH3CN (369.21 mg, 5.8763 mmol, 2.0 equiv. ) was added, and stirred at room temperature for overnight. EA was added after the mixture cooled to room temperature and washed by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and
concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS: 298.20 [M+H] +
Step 2. Methyl 1- [ (4-oxocyclohexyl) methyl] piperidine-4-carboxylate
Methyl 1- {1, 4-dioxaspiro [4.5] decan-8-ylmethyl} piperidine-4-carboxylate (442 mg) and TFA (6 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature overnight. The reaction mixture was concentrated under reduced pressure to afford the product which was used for next step directly.
LC/MS: 254.30 [M+H] +.
Step 3. Methyl 1- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.0 equiv. ) , methyl 1- [ (4-oxocyclohexyl) methyl] piperidine-4-carboxylate (624.79 mg, 2.466 mmol, 6.0 equiv. ) , MeOH and K2CO3 were added to a 10 mL schlenk tube with a stir bar and stirred at room temperature for 2 min, then the mixture was filtered. The filtrate, AcOH (20.7 mg) and MeOH (15 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 1.5 hours. The mixture was cooled by an ice-water bath, NaBH3CN (2.58 mg, 0.042 mmol, 2.0 equiv. ) was added, and stirred at 60 ℃ for 1 hour. EA was added after the mixture was cooled to room temperature and washed by saturated aq. NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS: 724.65 [M+H] +.
Step 4. 1- [ (4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-carboxylic acid
Methyl 1- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-
carboxylate (130 mg, 0.180 mmol, 1.0 equiv. ) , THF (6 mL) , MeOH (2 mL) and LiOH (0.72 mL, 3 mol/L, 2.160 mmol, 12.0 equiv. ) were added to a 10 mL schlenk tube with a stir bar and stirred at room temperature for overnight. The mixture was concentrated under reduced pressure and purified by C18 column with CH3CN/H2O (0.05%TFA) to afford the product.
LC/MS: 710.70 [M+H] +.
Step 5. N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-carboxamide (Compound 40)
1- [(4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-carboxylic acid (42 mg, 0.059 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (25.47 mg, 0.059 mmol, 1.0 equiv. ) , HATU (33.74 mg, 0.088 mmol, 1.5 equiv. ) , DIEA (30.59 mg, 0.236 mmol, 4.0 equiv. ) and DMF (3 mL) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. LCMS showed the reaction was completed and the reaction mixture was further purified by Prep-HPLC (Column: XBridge Prep OBD C18 Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 30%B to 50%B in 11 min, 50%B; Wavelength: 254 nm; RT1 (min) : 10.6; Number of Runs: 0) to afford Compound 40.
1H NMR (300 MHz, DMSO-d6) δ 10.16 (s, 1H) , 9.00 (s, 1H) , 8.83 (s, 1H) , 8.59 (t, J = 6.1 Hz, 1H) , 8.05 (t, J = 7.9 Hz, 1H) , 7.77 (t, J = 9.6 Hz, 2H) , 7.57 -7.61 (m, 3H) , 7.34 -7.47 (m, 4H) , 6.92 (d, J = 8.8 Hz, 2H) , 5.55 -5.81 (m, 1H) , 5.34 (s, 1H) , 5.15 (d, J = 3.4 Hz, 1H) , 4.95 -5.05 (m, 1H) , 4.76 -4.88 (m, 1H) , 4.69 (d, J = 6.0 Hz, 2H) , 4.53 (d, J = 9.3 Hz, 1H) , 4.44 (q, J = 7.2 Hz, 2H) , 4.35 (s, 1H) , 4.10 -4.28 (m, 1H) , 3.65 (s, 2H) , 3.09 (s, 4H) , 2.80 -2.99 (m, 2H) , 2.59 (s, 3H) , 2.45 (s, 3H) , 2.28 -2.41 (m, 1H) , 2.10 -2.23 (m, 3H) , 2.04 (t, J = 9.5 Hz, 1H) , 1.75 -1.96 (m, 3H) , 1.52 -1.72 (m, 6H) , 1.53 (s, 11H) , 1.24 (s, 3H) , 0.94 (s, 9H) .
LC/MS: 1122.55 [M+H] +.
Compound 41
Step 1. 4- ( ( (tert-butyldimethylsilyl) oxy) methyl) piperidine
To a solution of piperidin-4-ylmethanol (1 g, 8.682 mmol, 1.00 equiv. ) in DCM (10 mL) was added imidazole (0.71 g, 10.418 mmol, 1.2 equiv. ) and TBSCl (1.44 g, 9.550 mmol, 1.1 equiv. ) . The resulting mixture was stirred at 25 ℃ for 1h. The reaction was concentrated. The residue obtained was purified by silica gel chromatography (0-20%MeOH/DCM) to afford the 4- { [ (tert-butyldimethylsilyl) oxy] methyl} piperidine.
LC/MS: 230.15 [M+H] +.
Step 2. methyl 4- (4- ( ( (tert-butyldimethylsilyl) oxy) methyl) piperidin-1-yl) cyclohexane-1-carboxylate
To a solution of 4- { [ (tert-butyldimethylsilyl) oxy] methyl} piperidine (2.0 g, 8.717 mmol, 1.00 equiv. ) and methyl 4-oxocyclohexane-1-carboxylate (1.36 g, 8.717 mmol, 1 equiv. ) in DCE (20
mL) was added CH3COOH (0.03 g, 0.500 mmol, 0.06 equiv. ) . After 1 h, NaBH (OAc) 3 (3.69 g, 17.434 mmol, 2 equiv. ) . The resulting mixture was stirred at 25 ℃ overnight. The reaction was quenched with H2O (30 mL) . The pH value of the resulting mixture was adjusted to 7 used NaHCO3 solution. Then extracted with DCM (3*30mL) . The organic layers were combined and concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford the methyl 4- (4- { [ (tert-butyldimethylsilyl) oxy] methyl} piperidin-1-yl) cyclohexane-1-carboxylate.
Step 3. methyl 4- (4- (hydroxymethyl) piperidin-1-yl) cyclohexane-1-carboxylate
To a solution of methyl 4- (4- { [ (tert-butyldimethylsilyl) oxy] methyl} piperidin-1-yl) cyclohexane-1-carboxylate (2.7 g, 7.305 mmol, 1.00 equiv. ) in 1, 4-dioxane (15 mL) was added a solution of HCl in 1, 4-dioxane (20 mL, 658.238 mmol, 90.11 equiv. ) . The resulting mixture was stirred at 25 ℃ for 3 h. The reaction was concentrated. The residue obtained was purified by silica gel chromatography (MeOH/DCM, 0~10%) to afford the methyl 4- [4- (hydroxymethyl) piperidin-1-yl] cyclohexane-1-carboxylate.
Step 4. methyl 4- (4-formylpiperidin-1-yl) cyclohexane-1-carboxylate
To a cosolvent of DMSO (305.97 mg, 3.916 mmol, 2 equiv. ) and DCM (10 mL) was added (COCl) 2 (372.79 mg, 2.937 mmol, 1.5 equiv. ) . The mixture was stirred at -78 ℃ under nitrogen atmosphere. After 30 min, the solution of methyl 4- [4- (hydroxymethyl) piperidin-1-yl] cyclohexane-1-carboxylate (500 mg, 1.958 mmol, 1.00 equiv. ) in DCM was added. After 1h, TEA (990.67 mg, 9.790 mmol, 5 equiv. ) was added. The resulting mixture was stirred at 25 ℃ for 1 h. The reaction was concentrated. The residue obtained was purified by silica gel chromatography (0-50%ethyl acetate/petroleum ether) to afford the methyl 4- (4-formylpiperidin-1-yl) cyclohexane-1-carboxylate.
Step 5. methyl 4- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) piperidin-1-yl) cyclohexane-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.823 mmol, 1.00 equiv. ) in MeOH (10 mL) was added methyl 4- (4-formylpiperidin-1-yl) cyclohexane-1-carboxylate (208 mg, 0.823 mmol, 1 equiv. ) and CH3COOH (0.02 mL) . The mixture was stirred at rt for 1 h. After that, NaBH3CN (104 mg, 1.644 mmol, 2 equiv. ) was added. The mixture was stirred at rt for 3
h. After completion, the reaction was quenched with water (10 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford methyl 4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexane-1-carboxylate.
LC/MS: 724.4 [M+H] +
Step 6. 4- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) piperidin-1-yl) cyclohexane-1-carboxylic acid
To a solution of methyl 4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexane-1-carboxylate (300 mg, 0.414 mmol, 1 equiv. ) in THF (2 mL) and H2O (2 mL) was added LiOH (99.24 mg, 4.140 mmol, 10 equiv. ) . The mixture was stirred at rt for 3 h. After completion, the crude product was purified by reverse-phase flash with the following conditions: Column, Cat No: SO230120-2, C18, 120 g, 20~45μm, Lot: BP0002P2503; mobile phase, CH3CN: H2O (0.05%TFA) = 20%increased to CH3CN: H2O (0.05%TFA) = 70%in 40 min, hold CH3CN: H2O (0.05%TFA) = 70%in 10 min, up to CH3CN: H2O (0.05%TFA) = 95%in 2 min, hold CH3CN: H2O (0.05%TFA) = 95%in 10 min; Detector, UV 220 nm &254 nm. The organic layers were concentrated under reduced pressure to afford 4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexane-1-carboxylic acid.
LC/MS: 710 [M+H] +.
Step 7. (2S, 4R) -1- ( (S) -2- (4- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) piperidin-1-yl) cyclohexane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 41)
To a solution of 4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexane-1-carboxylic acid (180 mg, 0.254 mmol, 1.00 equiv. ) in DMF (5 mL, 64.609
mmol, 254.81 equiv. ) was added TEA (76.97 mg, 0.762 mmol, 3 equiv. ) , HATU (125.33 mg, 0.330 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (109.17 mg, 0.254 mmol, 1 equiv. ) . The mixture was stirred at r.t. for 1 h. After completion, The crude product was purified by Prep-HPLC with the following conditions: Column: Kinetex EVO C18, 21.2*250mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 40%B to 60%B in 7 min, 60%B; Wavelength: 254 nm; RT1 (min) : 5.6. After lyophilization, it was afforded Compound 41 (2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.13 (s, 1H) , 8.98 (s, 1H) , 8.82 (s, 1H) , 8.55 (t, J = 6.1 Hz, 1H) , 8.04 (t, J = 7.9 Hz, 1H) , 7.72 (dd, J = 19.4, 8.7 Hz, 1H) , 7.48 -7.66 (m, 4H) , 7.48 -7.35 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.66 (ddt, J = 16.4, 10.2, 6.0 Hz, 1H) , 5.31 (s, 1H) , 5.12 (t, J = 3.2 Hz, 1H) , 4.99 (dd, J = 10.2, 1.5 Hz, 1H) , 4.88 -4.78 (m, 1H) , 4.68 (d, J = 6.0 Hz, 2H) , 4.52 (t, J = 10.3 Hz, 1H) , 4.43 (dq, J = 10.1, 6.0, 4.9 Hz, 2H) , 4.35 (s, 1H) , 4.22 (dd, J = 16.0, 5.4 Hz, 1H) , 3.66 (q, J = 10.9 Hz, 2H) , 3.09 (s, 4H) , 2.75 -3.00 (m, 2H) , 2.40 -2.46 (m, 3H) , 2.10 -2.26 (m, 3H) , 1.95 -2.10 (m, 3H) , 1.76 -1.95 (m, 5H) , 1.62 -1.76 (m, 3H) , 1.27 -1.57 (m, 13H) , 1.01 -1.18 (m, 3H) , 0.86 -0.98 (m, 9H) . LC/MS: 1122 [M+H] +.
Compound 42
Step 1. 4- ( ( (tert-butyldimethylsilyl) oxy) methyl) cyclohexan-1-one
To a solution of 4- (hydroxymethyl) cyclohexan-1-one (500 mg, 3.901 mmol, 1.00 equiv. ) in DCM (5 mL) was added TBSCl (705.57 mg, 4.681 mmol, 1.2 equiv. ) and imidazole (371.81 mg, 5.461 mmol, 1.4 equiv. ) . The mixture was stirred at r.t. for 4 h. After completion, the reaction was quenched with water (10 mL) . The mixture was extracted with EA (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%PE: EtOAc) . The organic layers were concentrated under reduced pressure to afford 4- { [ (tert-butyldimethylsilyl) oxy] methyl} cyclohexan-1-one.
LC/MS: 243 [M+H] +.
Step 2. ethyl 1- (4- ( ( (tert-butyldimethylsilyl) oxy) methyl) cyclohexyl) piperidine-4-carboxylate
To a solution of 4- { [ (tert-butyldimethylsilyl) oxy] methyl} cyclohexan-1-one (716 mg, 2.953 mmol, 1.00 equiv. ) in DCE (10 mL) and CH3COOH (0.01 mL, 0.175 mmol, 0.06 equiv. ) . The mixture was stirred at r.t. for 1 h. After that, NaBH3CN (371.19 mg, 5.906 mmol, 2 equiv. ) was added and stirred at r.t. for 3 h. After completion, the reaction was quenched with water (10 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford ethyl 1- (4- { [ (tert-butyldimethylsilyl) oxy] methyl} cyclohexyl) piperidine-4-carboxylate.
LC/MS: 384 [M+H] +.
Step 3. ethyl 1- (4- (hydroxymethyl) cyclohexyl) piperidine-4-carboxylate
To a solution of ethyl 1- (4- { [ (tert-butyldimethylsilyl) oxy] methyl} cyclohexyl) piperidine-4-carboxylate (662 mg, 1.726 mmol, 1.00 equiv. ) ethyl 1- (4- { [ (tert-butyldimethylsilyl) oxy] methyl} cyclohexyl) piperidine-4-carboxylate (662 mg, 1.726 mmol, 1.00 equiv. ) was added a solution of HCl in dioxane (4 M) . The mixture was stirred at r.t. for 2 h. The mixture was concentrated under reduced pressure to removed dioxane. The solvent was removed under vacuum to give the ethyl 1- [4- (hydroxymethyl) cyclohexyl] piperidine-4-carboxylate (220 mg, 47.33%) as white oil. LC/MS: 270 [M+H] +.
Step 4. ethyl 1- (4-formylcyclohexyl) piperidine-4-carboxylate
To a solution of DMSO (116.02 mg, 1.484 mmol, 2 equiv. ) in DCM (10 mL, 157.300 mmol, 211.87 equiv. ) was added (COCl) 2 (141.35 mg, 1.113 mmol, 1.5 equiv. ) N2 atmosphere. The mixture was stirred at -78 ℃ for 30 min. ethyl 1- [4- (hydroxymethyl) cyclohexyl] piperidine-4-carboxylate (200 mg, 0.742 mmol, 1.00 equiv. ) was added to the mixture. The mixture was stirred at -78 ℃ for 1 h. TEA (375.63 mg, 3.710 mmol, 5 equiv. ) was added to the mixture, the mixture was stirred at r.t. for 1 h. After completion, the reaction was quenched with water (50 mL) . The mixture was extracted with EA (3x50 m) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%PE: EtOAc) . The combined organic layers were concentrated under reduced pressure to afford ethyl 1- (4-formylcyclohexyl) piperidine-4-carboxylate.
LC/MS: 268 [M+H] +.
Step 5. ethyl 1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) piperidine-4-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (223 mg, 0.458 mmol, 1 equiv. ) in MeOH (10 mL) was added ethyl 1- (4-formylcyclohexyl) piperidine-4-carboxylate (122.54 mg, 0.458 mmol, 1 equiv. ) and AcOH (0.01 mL, 0.175 mmol, 0.38 equiv. ) . The mixture was stirred at r.t. for 1 h. After that, NaBH3CN (57.60 mg, 0.916 mmol, 2 equiv. ) was added and stirred at r.t. for 3 h. After completion, the reaction was quenched with water (10 mL) . The mixture was extracted with DCM (3 x 20 mL) and the organic layer was dried over Na2SO4 and filtered. The organic layers were concentrated. The crude product was purified with silica gel column (0-50%DCM: EtOH) . The organic layers were concentrated under reduced pressure to afford ethyl 1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxylate.
LC/MS: 738 [M+H] +.
Step 6. 1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) piperidine-4-carboxylic acid
To a solution of ethyl 1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxylate (228 mg, 0.309 mmol, 1 equiv. ) in THF (3 mL, 37.029 mmol, 119.85 equiv. ) and H2O (3 mL, 166.525 mmol, 538.98 equiv. ) was added LiOH (73.99 mg, 3.090 mmol, 10 equiv. ) . The mixture was stirred at r.t. for 3 h. After completion, the pH value of the solution was adjusted to 4~5 with concentrated HCl. The crude product was purified by reverse-phase flash with the following conditions: Column, Cat No: SO230120-2, C18, 120 g, 20~45μm, Lot: BP0002P2503; mobile phase, CH3CN: H2O (0.05%TFA) = 20%increased to CH3CN: H2O (0.05%TFA) = 70%in 40 min, hold CH3CN: H2O (0.05%TFA) = 70%in 10 min, up to CH3CN: H2O (0.05%TFA) = 95%in 2 min, hold CH3CN: H2O (0.05%TFA) = 95%in 10 min; Detector, UV 220 nm &254 nm. The organic layers were concentrated under reduced pressure to afford 1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxylic acid.
LC/MS: 710 [M+H] +.
Step 7. N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxamide (Compound 42)
To a solution of 1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxylic acid (150 mg, 0.211 mmol, 1.00 equiv. ) in DMF (5 mL) was added TEA (64.14 mg, 0.633 mmol, 3 equiv. ) , HATU (104.44 mg, 0.274 mmol, 1.3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (90.98 mg, 0.211 mmol, 1 equiv. ) . The mixture was stirred at r.t. for 1 h. After completion, the crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 45%B to 55%B in 10 min, 55%B; Wave Length: 254 nm; RT1 (min) : 9. After lyophilization, it was afforded Compound 42 N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.14 (s, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.56 (t, J = 6.0 Hz, 1H) , 8.04 (s, 1H) , 7.78 -7.68 (m, 2H) , 7.50 -7.64 (m, 3H) , 7.45 -7.35 (m, 4H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.66 (ddt, J = 16.5, 11.1, 6.1 Hz, 1H) , 5.32 (s, 1H) , 5.13 (s, 1H) , 4.99 (d, J = 10.8 Hz, 1H) , 4.82 (d, J = 17.2 Hz, 1H) , 4.68 (d, J = 5.9 Hz, 2H) , 4.51 (d, J = 9.2 Hz, 1H) , 4.48 -4.38 (m, 2H) , 4.35 (s, 1H) , 4.22 (dd, J = 16.0, 5.5 Hz, 1H) , 3.57 -3.71 (m, 2H) , 3.08 (s, 4H) , 2.81 (d, J = 10.3 Hz, 2H) , 2.41 -2.47 (m, 6H) , 2.06 -2.37 (m, 5H) , 1.61 -2.06 (m, 6H) , 1.34 -1.60 (m, 13H) , 1.11 -1.29 (m, 2H) , 0.76 -1.04 (m, 9H) .
LC/MS: 1122.6 [M+H] +.
Compound 43
Step 1. methyl 2- (4-oxopiperidin-1-yl) acetate
To a stirred mixture of 4-piperidinone (600 mg, 6.052 mmol, 1.00 equiv. ) and K2CO3 (2.51 g, 18.156 mmol, 3 equiv. ) in ACN (20 mL) were added methyl 2-bromoacetate (1.39 g, 9.078 mmol, 1.5 equiv. ) dropwise at room temperature. The resulting mixture was stirred for 16 h at 80 ℃. The resulting mixture was filtered, the filter cake was washed with acetonitrile (4x10 mL) . The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford methyl 2- (4-oxopiperidin-1-yl) acetate.
LC/MS: 172.15 [M+H] +.
Step 2. methyl 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) acetate
A mixture of methyl 2- (4-oxopiperidin-1-yl) acetate (140.73 mg, 0.822 mmol, 2 equiv. ) and 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.411 mmol, 1.00 equiv. ) and added CH3COOH (0.1 equiv. ) in methanol (5 mL) was stirred overnight at 60 ℃. The mixture was allowed to cool down to room temperature. To the above mixture was added NaBH3CN (51.66 mg, 0.822 mmol, 2 equiv. ) in portions over 5 min at room temperature. The resulting mixture was stirred for additional 2 h at 60 ℃. The resulting mixture was extracted with DCM (3 x 50 mL) . The combined organic layers were washed with brine (1x50 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford methyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetate.
LC/MS: 642.55 [M+H] +.
Step 3. 2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) acetic acid
To a stirred mixture of methyl 2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetate (206 mg, 0.321 mmol, 1.00 equiv. ) in THF (3 mL) was added LiOH (3 mL, 4M) dropwise at room temperature. The resulting mixture was stirred for 3 h at room temperature. The mixture was acidified to pH 5 with HCl (2 M) . The precipitated solids were collected by filtration and washed with water (1x3 mL) . The filter cake was concentrated under reduced pressure to afford (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetic acid.
LC/MS: 628.40 [M+H] +.
Step 4. (2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 43)
To a stirred mixture of (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetic acid (133 mg, 0.212 mmol, 1.00 equiv. ) and HATU (104.73 mg, 0.276 mmol, 1.3 equiv. ) in DMF (2 mL) were added TEA (64.32 mg, 0.636 mmol, 3 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (91.22 mg, 0.212 mmol, 1.0 equiv. ) in portions at room temperature. The resulting mixture was stirred for 1 h at room temperature. The resulting mixture was extracted with EA (3 x 50 mL) . The combined organic layers were washed with water (1x50 mL) , dried over anhydrous Na2SO4. The residue was purified by reverse flash chromatography with the following conditions: Column: Kinetex EVO C18 Column, 30*150, 5um; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 30%B to 50%B in 9 min, 50%B; Wavelength: 254 nm. The product was concentrated under vacuum to afford Compound 43 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.94 (s, 1H) , 8.83 (s, 1H) , 8.62 (t, J = 6.1 Hz, 1H) , 8.04 (d, J = 8.1 Hz, 1H) , 7.72 -7.86 (m, 2H) , 7.51 -7.65 (m, 3H) , 7.37 -7.49 (m, 4H) , 6.89 (d, J = 8.8 Hz, 2H) , 5.60 -5.74 (m, 1H) , 5.33 (s, 1H) , 5.15 (d, J = 3.4 Hz, 1H) , 4.96 -5.04 (m, 1H) , 4.78 -4.88 (m, 1H) , 4.69 (d, J = 5.8 Hz, 2H) , 4.35 -4.54 (m, 4H) , 4.23 -4.33 (m, 1H) , 3.57 -3.73 (m, 2H) , 2.98 -3.16 (m, 5H) , 2.84 -2.96 (m, 2H) , 2.57 -2.71 (m, 4H) , 2.45 (s, 3H) , 2.01 -2.29 (m, 4H) , 1.75 -1.98 (m, 3H) , 1.39 -1.54 (m, 9H) , 0.96 (s, 9H) .
LC/MS: 1040.55 [M+H] +.
Compound 44
Step 1. Methyl 3- (1, 4-dioxa-8-azaspiro [4.5] decan-8-yl) propanoate
1, 4-Dioxa-8-azaspiro [4.5] decane (1 g, 6.984 mmol) , K2CO3 (1.46 g, 10.488 mmol) and methyl 3-bromopropanoate (1.52 g, 9.102 mmol) were dissolved in 20 mL acetone in a 50 mL flask and stirred at 70℃ for 1 h. Desired product was found by LCMS. The solvent was removed under vacuum and the residue was purified by silica gel column with MeOH/DCM (0~15%) to afford the product.
Step 2. Methyl 3- (4-oxopiperidin-1-yl) propanoate
The solution of methyl 3- {1, 4-dioxa-8-azaspiro [4.5] decan-8-yl} propanoate (1.6 g, 6.978 mmol, 1 equiv. ) in trifluoroacetaldehyde (10 mL) was heated at 60℃ overnight. The mixture was allowed to cool down to r.t. . The resulting mixture was concentrated under reduced pressure to afford to afford crude methyl 3- (4-oxopiperidin-1-yl) propanoate which was used for next step without purification.
Step 3. Methyl 3- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) piperidin-1-
yl) propanoate
Methyl 3- (4-oxopiperidin-1-yl) propanoate (456.80 mg, 2.468 mmol, 4 equiv. ) , 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (300 mg, 0.617 mmol, 1.00 equiv. ) and acetic acid (3.70 mg, 0.062 mmol, 0.1 equiv. ) were dissolved in 6 mL MeOH in a 40 mL vial and stirred at 70℃ for 1 h. Following this, the mixture was cooled to r.t. . NaBH3CN (77.49 mg, 1.234 mmol, 2 equiv. ) was added and stirred at r.t. for 1h. LCMS showed the reaction succeed and the mixture diluted with 100 mL EA and washed with sat. aq. NaHCO3. The organic layer was combined and dried by Na2SO4, then concentrated. The residue was purified by silica gel column with MeOH/DCM from 0~15%to afford methyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanoate.
Step 4. 3- (4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanoic acid
Methyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanoate (370 mg, 0.564 mmol, 1.0 equiv. ) , THF (12 mL) , MeOH (4 mL) and LiOH (2.26 mL, 3 mol/L, 6.768 mmol, 12.0 equiv. ) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature overnight. The mixture was concentrated under reduced pressure and purified by C18 column with MeOH/H2O (0.05%TFA) to afford the product.
LC/MS: 642.60 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 44)
3- (4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanoic acid (120 mg, 0.167 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (80.51 mg, 0.167 mmol, 1.0 equiv. ) , HATU (106.64 mg, 0.280 mmol, 1.5 equiv. ) , DIEA (96.66 mg, 0.748 mmol, 4.0 equiv. ) and DMF (4 mL) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature
for 1 hour. LCMS showed the reaction was completed and the reaction mixture was purified by Prep-HPLC (Column: CHIRALPAK IB N-3, 4.6*100mm, 3um; Mobile Phase B: MeOH (0.1%DEA) ; Flow rate: 2 mL/min; Gradient: isocratic 50%B; Wave Length: 220 nm) afford Compound 44 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.16 (s, 1H) , 8.93 (s, 1H) , 8.83 (s, 2H) , 8.64 (t, J = 6.1 Hz, 1H) , 8.04 (t, J = 7.9 Hz, 1H) , 7.75 (d, J = 8.1 Hz, 1H) , 7.52 -7.68 (m, 3H) , 7.25 -7.49 (m, 4H) , 6.84 (d, J = 8.7 Hz, 2H) , 5.57 -5.76 (m, 1H) , 5.34 (s, 1H) , 5.16 (d, J = 3.4 Hz, 1H) , 4.95 -5.08 (m, 1H) , 4.78 -4.95 (m, 1H) , 4.69 (d, J = 6.0 Hz, 2H) , 4.52 -4.62 (m, 1H) , 4.40 -4.52 (m, 2H) , 4.37 (s, 1H) , 4.16 -4.28 (m, 1H) , 3.58 -3.74 (m, 2H) , 2.97 (s, 6H) , 2.60 (s, 4H) , 2.42 (s, 5H) , 2.12 -2.28 (m, 3H) , 1.86 -1.99 (m, 6H) , 1.52 -1.74 (m, 2H) , 1.46 (s, 6H) , 0.97 (s, 9H) .
LC/MS: 1054.50 [M+H] +.
Compound 45
Step 1. tert-butyl 9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecane-3-carboxylate
Tert-butyl 9-oxo-3-azaspiro [5.5] undecane-3-carboxylate (4.4 g, 16.442 mol) and 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (4 g, 8.221 mmol) , a stir bar, and DCE (80 mL) were added to a 250 mL round-bottom flask and stirred until homogeneous, then treated with AcOH (60 mg,
0.999 mmol) . The resulting mixture was stirred for 1 h at room temperature. To the above mixture was added NaBH3CN (5.23 g, 24.663 mmol) . The resulting mixture was stirred overnight at 60℃ under a nitrogen atmosphere. The resulting mixture was cooled to r.t. and quenched with sat. aq NaHCO3 (40 mL) , then extracted with CH2Cl2 (3 x 20 mL) . The combined extracts were washed with brine (2 x 20 mL) , dried over anhydrous Na2SO4. The residue obtained was purified by silica gel column chromatography (0-15%MeOH/DCM) to afford tert-butyl 9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecane-3-carboxylate.
LC/MS: 738.75 [M+H] +.
Step 2. 6- { [4- (4- {3-azaspiro [5.5] undecan-9-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecane-3-carboxylate (5.8 g, 7.860 mmol) , a stir bar, DCM (50 mL) were added to a 250 ml round-bottom flask and stirred until homogeneous, and then treated with HCl in 1, 4-dioxane (70 ml) at room temperature. The resulting mixture was stirred at r.t. for 2 h and then concentrated under reduced pressure, the residue was diluted with DCM/MeOH (10/1) (1 L) . The pH value of the solution was adjusted to 9 with NaHCO3 saturated solution. Then the resulting mixture was extracted with DCM/MeOH (10/1, 1 L x 3) . The combined extracts were washed with H2O (500 ml) , dried over anhydrous Na2SO4, filtered and concentrated under vacuum to afford 6- { [4- (4- {3-azaspiro [5.5] undecan-9-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one. The crude product was used in the next step directly without further purification.
LC/MS: 638.65 [M+H] +.
Step 3. (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid
6- { [4- (4- {3-azaspiro [5.5] undecan-9-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (1.3 g, 2.038 mmol) , glyoxylic acid monohydrate (0.21 g, 2.242 mmol) , a stir bar, and MeOH (13 mL) were added to a 100 ml round-bottom flask. The resulting mixture was stirred at room temperature for 1 h. To the
above mixture was added NaBH3CN (0.13 g, 2.038 mmol) at room temperature. The resulting mixture was stirred for 5 minutes. The reaction was purified by C18 column with CH3CN/5mM NH4HCO3 Water (5%-60%) to afford (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid.
LC/MS: 696.30 [M+H] +
Step 4. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 45)
(9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid (1.6 g, 2.299 mmol) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (0.99 g, 2.299 mmol) , a stir bar, DMF (28 mL) were added to a 250 ml round-bottom flask and stirred until homogeneous, and then treated with DIEA (0.89 g, 6.897 mmol) and HATU (0.96 g, 2.529 mmol) . The resulting mixture was stirred at room temperature for 2 h. The reaction was purified by C18 column with ACN/H2O (6.5 mM NH4HCO3+Ammonia Hydroxide (pH=10) ) to afford Compound 45 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO) δ 10.11 (s, 1H) , 8.98 (d, J = 2.4 Hz, 1H) , 8.82 (d, J = 2.4 Hz, 1H) , 8.60 (s, 1H) , 8.03 (d, J = 8.4 Hz, 1H) , 7.78 -7.84 (m, 2H) , 7.56 -7.75 (m, 3H) , 7.40 -7.42 (m, 4H) , 6.90 (d, J = 7.5 Hz, 2H) , 5.56 -5.82 (m, 1H) , 5.33 (s, 1H) , 5.16 -5.15 (m, 1H) , 5.00 (d, J = 10.2 Hz, 1H) , 4.84 (d, J = 16.8 Hz, 1H) , 4.69 (d, J = 5.7 Hz, 2H) , 4.40 -4.50 (m, 4H) , 4.26 -4.35 (m, 1H) , 3.42 -3.60 (m, 2H) , 2.87 -3.10 (m, 6H) , 2.58 -2.71 (m, 4H) , 2.45 -2.50 (m, 6H) , 2.10 -2.18 (m, 1H) , 2.05 (d, J = 7.5 Hz, 1H) , 1.91 (s, 1H) , 1.46 -1.72 (m, 13H) , 1.35 -1.33 (m, 5H) , 1.34 -1.47 (m, 3H) , 0.94 -1.06 (m, 7H) .
LC/MS: 1108.65 [M+H] +.
Compound 46
Step 1. tert-butyl 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonane-2-carboxylate
To a stirred solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (500 mg, 1.028 mmol, 1.00 equiv. ) and tert-butyl 7-oxo-2-azaspiro [3.5] nonane-2-carboxylate (491.83 mg, 2.056 mmol, 2 equiv. ) in DCE was added AcOH (0.62 mg, 0.010 mmol, 0.01 equiv. ) at room temperature under an air atmosphere. The resulting mixture was stirred for 1 h at room temperature under an air atmosphere. To the above mixture was added NaBH (OAc) 3 (653.36 mg, 3.084 mmol, 3 equiv. ) at room temperature. The resulting mixture was stirred for additional 12 h at 60 ℃. The reaction was quenched with sat. NaHCO3 (aq. ) at room temperature. The resulting mixture was extracted with DCM (3 x 10 mL) . The combined organic layers were washed with brine (2x10 mL) , dried over anhydrous MgSO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with DCM /MeOH (10: 1) to afford tert-butyl 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonane-2-carboxylate.
LC/MS: 710 [M+H] +.
Step 2. 6- { [4- (4- {2-azaspiro [3.5] nonan-7-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a stirred solution of tert-butyl 7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonane-2-carboxylate (690 mg, 0.972 mmol, 1.00 equiv. ) in DCM was added a solution of HCl in 1, 4-dioxane (5 mL, 164.559 mmol, 169.30 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 2 h at room temperature in air atmosphere. The resulting mixture was concentrated under vacuum to afford 6- { [4- (4- {2-azaspiro [3.5] nonan-7-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one as a crude product, which was used in the next step directly without further purification.
LC/MS: 610 [M+H] +.
Step 3. tert-butyl 2- (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetate
To a stirred solution of 6- { [4- (4- {3-azaspiro [5.5] undecan-9-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (580 mg, 0.909 mmol, 1.00 equiv. ) and tert-butyl 2-bromoacetate (371.06 mg, 1.902 mmol, 2 equiv. ) in DMF was added DIEA (368.79 mg, 2.853 mmol, 3 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for overnight at room temperature in air atmosphere. The reaction was quenched with Water at room temperature. The resulting mixture was extracted with EtOAc (3 x 10 mL) . The combined organic layers were washed with brine (2x10 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford tert-butyl 2- (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetate.
LC/MS: 724 [M+H] +.
Step 4. (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonan-2-yl) acetic acid
To a stirred solution of tert-butyl 2- (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonan-2-yl) acetate (178 mg, 0.246 mmol, 1.00 equiv. ) in DCM was added HCl (gas) in 1, 4-dioxane (2 mL, 65.824 mmol, 267.70 equiv. ) in portions at room temperature in air atmosphere. The resulting mixture was stirred for 4 h at room temperature in air atmosphere. The resulting mixture was concentrated under vacuum to afford (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonan-2-yl) acetic acid, which was used in the next step directly without further purification. LC/MS: 668 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-
azaspiro [3.5] nona-2-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 46)
To a stirred solution of (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nonan-2-yl) acetic acid (160 mg, 0.240 mmol, 1.00 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (113.48 mg, 0.264 mmol, 1.1 equiv. ) in DMF was added HATU (182.20 mg, 0.480 mmol, 2 equiv. ) and DIEA (139.34 mg, 1.080 mmol, 4.5 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 2 h at room temperature in air atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 10 mL) . The combined organic layers were washed with brine (2x 8 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford the crude product. The crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Prep OBD C18 Column, 30*150 mm, 5 μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 27%B to 37%B in 15 min, 37%B; Wave Length: 254 nm; RT1 (min) : 14.350 to afford title Compound 46: (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nona-2-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.11 (s, 1H) , 8.97 (s, 1H) , 8.82 (s, 1H) , 8.59 (t, J = 6.0 Hz, 1H) , 8.04 (t, J = 7.8 Hz, 1H) , 7.75 (d, J = 8.1 Hz, 1H) , 7.55 -7.67 (m, 4H) , 7.38 -7.45 (m, 4H) , 6.89 (d, J = 9 Hz, 2H) , 5.65 (ddd, J = 16.4, 10.5, 5.7 Hz, 1H) , 5.31 (s, 1H) , 5.14 (s, 1H) , 4.98 (dd, J = 10.2, 1.5 Hz, 1H) , 4.86 (dd, J = 17.1, 1.8 Hz, 1H) , 4.68 (d, J = 4.8 Hz, 2H) , 4.47 (d, J = 10.2 Hz, 1H) , 4.38 -4.44 (m, 1H) , 4.34 -4.37 (m, 1H) , 4.27 -4.29 (m, 1H) , 3.61 (q, J = 10.5, 9.6 Hz, 2H) , 3.00 -3.18 (m, 9H) , 2.60 (s, 4H) , 2.45 (s, 3H) , 2.00 -2.28 (m, 3H) , 1.81 -1.95 (m, 3H) , 1.62 -1.75 (m, 2H) , 1.46 (s, 6H) , 1.37 (d, J = 12.9 Hz, 2H) , 1.24 (d, J = 12.3 Hz, 3H) , 0.94 (s, 9H) .
LC/MS: 1080.60 [M+H] +.
Compound 47
Step 1. tert-butyl 2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonane-7-carboxylate
To a stirred solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (550 mg, 1.130 mmol, 1.00 equiv. ) and tert-butyl 2-oxo-7-azaspiro [3.5] nonane-7-carboxylate (541.01 mg, 2.260 mmol, 2 equiv. ) in DCE (10 mL, 126.315 mmol, 111.75 equiv. ) added AcOH (0.68 mg, 0.011 mmol, 0.01 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 1 h at room temperature in air atmosphere. To the above mixture was added NaBH (OAc) 3 (479.13 mg, 2.260 mmol, 2 equiv. ) at room temperature. The resulting mixture was stirred for additional 2 h at room temperature. The reaction was quenched with sat. NaHCO3 (aq. ) at room temperature. The resulting mixture was extracted with CH2Cl2 (3 x 20 mL) . The combined organic layers were washed with brine (2 x 10 mL) , dried over anhydrous MgSO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford tert-butyl 2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonane-7-carboxylate.
LC/MS: 710 [M+H] +.
Step 2. 6- { [4- (4- {7-azaspiro [3.5] nonan-2-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a stirred solution of tert-butyl 2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonane-7-carboxylate (709 mg, 0.999 mmol, 1.00 equiv. ) in DCM was added TFA (10 mL, 134.630 mmol, 134.80 equiv. ) in portions at room temperature. The resulting mixture was stirred for 2 h at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford 6- { [4- (4- {7-azaspiro [3.5] nonan-2-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 610 [M+H] +.
Step 3. tert-butyl 2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate
To a stirred solution of 6- { [4- (4- {7-azaspiro [3.5] nonan-2-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400
mg, 0.656 mmol, 1.00 equiv. ) and tert-butyl 2-bromoacetate (255.90 mg, 1.312 mmol, 2 equiv. ) in DMF was added DIEA (211.95 mg, 1.640 mmol, 2.5 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 2 h at room temperature in air atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 8 mL) . The combined organic layers were washed with brine (2x8 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford tert-butyl 2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate.
LC/MS: 724 [M+H] +.
Step 4. (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid
To a stirred solution of tert-butyl 2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate (120 mg, 0.166 mmol, 1.00 equiv. ) in DCM was added TFA (2 mL, 26.926 mmol, 162.44 equiv. ) in portions at room temperature in air atmosphere. The resulting mixture was stirred overnight at room temperature in air atmosphere. Desired product could be detected by LCMS. The resulting mixture was concentrated under vacuum to afford (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid. The crude product was used in the next step directly without further purification.
LC/MS: 668 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 47)
To a stirred solution of (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid (110 mg, 0.165 mmol, 1.00 equiv. ) and (2S, 4R) -1- [ (2S) -2-amino-3, 3-
dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (78.01 mg, 0.182 mmol, 1.1 equiv. ) in DMF was added HATU (81.42 mg, 0.215 mmol, 1.3 equiv. ) and DIEA (85.15 mg, 0.660 mmol, 4 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 2 h at room temperature in air atmosphere. Desired product could be detected by LCMS. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 6 mL) . The combined organic layers were washed with brine (2x8 mL) , dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford the crude product. The crude product was purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 19*250 mm, 10 μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 45%B to 75%B in 7 min, 75%B; Wave Length: 254 nm; RT1 (min) : 6.5 to afford title Compound 47: (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.12 (s, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.59 (t, J = 6.0 Hz, 1H) , 8.04 (t, J = 9.0 Hz, 1H) , 7.74 -7.84 (m, 2H) , 7.55 -7.65 (m, 3 H) , 7.38 -7.52 (m, 4 H) , 6.90 (d, J = 9.3 Hz, 2H) , 5.54 -5.82 (m, 1H) , 5.08 -5.44 (m, 2H) , 4.99 (dd, J = 10.2, 1.5 Hz, 1H) , 4.77 -4.89 (m, 1H) , 4.68 (d, J = 4.8 Hz, 2H) , 4.32 -4.56 (m, 4H) , 4.14 -4.31 (m, 1H) , 3.42 -3.78 (m, 3H) , 3.09 (s, 4H) , 2.79 -3.04 (m, 2H) , 2.61 -2.77 (m, 1H) , 2.45 (s, 4H) , 2.37 (s, 6H) , 2.00 -2.15 (m, 1H) , 1.83 -1.95 (m, 3H) , 1.52 -1.68 (m, 6H) , 1.46 (s, 6 H) , 0.94 (s, 9 H) .
LC/MS: 1081.05 [M+H] +.
Compound 48
Step 1. Tert-butyl 3- (4- (isopropoxycarbonyl) cyclohex-1-en-1-yl) azetidine-1-carboxylate
To a stirred solution of tert-butyl 3-iodoazetidine-1-carboxylate (3.03 g, 10.703 mmol, 1.00 equiv. ) , and ethyl 4- (4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) cyclohex-3-ene-1-carboxylate (3.30 g, 11.773 mmol, 1.1 equiv. ) in isopropyl alcohol (20 mL) were added sequentially nickel (II) iodide (334.46 mg, 1.070 mmol, 0.1 equiv. ) and (1R, 2R) trans-2-amino cyclohexanol hydrochloride (0.16 g, 1.070 mmol, 0.1 equiv. ) at 26 ℃. The reaction mixture was degassed with nitrogen for 15 min and stirred for 10 min at 26 ℃, followed by the addition of NaHMDS (21.41 mL, 21.406 mmol, 2 equiv. ) at 26 ℃. The reaction mixture was heated to 80 ℃ and stirred for 16 h at the same temperature. The reaction mixture was quenched with saturated ammonium chloride
solution (100 mL) and extracted with EtOAc (3 x 100 mL) . The organic layer was dried over (anh) Na2SO4, filtered and concentrated under reduced pressure. The crude was purified by silica chromatography column using a linear gradient of petroleum ether/EtOAc to yield tert-butyl 3- (4- (isopropoxycarbonyl) cyclohex-1-en-1-yl) azetidine-1-carboxylate.
Step 2. Tert-butyl 3- [4- (isopropoxycarbonyl) cyclohexyl] azetidine-1-carboxylate
To a solution of tert-butyl 3- [4- (isopropoxycarbonyl) cyclohex-1-en-1-yl] azetidine-1-carboxylate (1.65 g 5.102 mmol) in MeOH (15 mL) was added Pd/C (0.53 g, 10%) . The resulting mixture was stirred at room temperature for 1.5 h under H2. The resulting mixture was filtered with diatomite and washed with MeOH. The solvent was removed under vacuum to afford tert-butyl 3- [4- (isopropoxycarbonyl) cyclohexyl] azetidine-1-carboxylate.
LC/MS: 326.25 [M+H] +
Step 3. Tert-butyl 3- [4- (hydroxymethyl) cyclohexyl] azetidine-1-carboxylate
To a solution of tert-butyl 3- [4- (isopropoxycarbonyl) cyclohexyl] azetidine-1-carboxylate (1.44 g, 4.425 mmol, 1.0 equiv. ) in THF (15 mL) was added LiBH4 (6.64 mL, 13.275 mmol, 3.0 equiv., 2 mol/L) dropwise at 0 ℃ under N2. The resulting mixture was stirred at 50 ℃ for 3.5 h. The reaction was quenched with sat. NH4Cl (aq. ) at room temperature. The resulting mixture was extracted with EA (3 x 20 mL) . The combined organic layers were washed with brine (2x10 mL) , dried over anhydrous NaSO4. After filtration, the filtrate was concentrated under reduced pressure to afford tert-butyl 3- [4- (hydroxymethyl) cyclohexyl] azetidine-1-carboxylate.
LC/MS: 270.15 [M+H] +.
Step 4. Tert-butyl 3- (4-formylcyclohexyl) azetidine-1-carboxylate
To a solution of bis (carbon monoxide) (2.36 g, 18.56 mmol, 5.0 equiv. ) was added DCM (40 mL) . The resulting mixture was stirred at -78 ℃ for 20 minutes. To the above mixture was added DMSO (2.03 g, 25.98 mmol, 7 equiv. ) at -78 ℃. The resulting mixture was stirred for additional for 20 minutes at -78 ℃. To the above mixture was added tert-butyl 3- [4- (hydroxymethyl) cyclohexyl] azetidine-1-carboxylate (1.0 g, 3.71 mmol, 1.0 equiv. ) and DCM (10 mL) at -78 ℃. The resulting mixture was stirred for additional for 30 minutes at -78 ℃. To the above mixture was added TEA (3.76 g, 37.12 mmol, 10.0 equiv. ) at -78 ℃. The resulting mixture was stirred for additional for 30 minutes at room temperature. The reaction was quenched with
H2O at room temperature. The resulting mixture was extracted with EA (3 x 60 mL) . The combined organic layers were washed with brine (2x20 mL) and HCl (0.5 mol/L, 2x20 mL) , dried over anhydrous NaSO4. After filtration, the filtrate was concentrated under reduced pressure to afford tert-butyl 3- (4-formylcyclohexyl) azetidine-1-carboxylate.
LC/MS: 268.15 [M+H] +.
Step 5. Tert-butyl 3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidine-1-carboxylate
To a solution of 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.822 mmol, 1.0 equiv. ) and tert-butyl 3- (4-formylcyclohexyl) azetidine-1-carboxylate (439.59 mg, 1.644 mmol, 2 equiv. ) in MeOH (15 mL) was added AcOH (20.0 mg) at room temperature in air atmosphere. The resulting mixture was stirred for 1 h at 60 ℃ in air atmosphere. To the above mixture was added NaBH3CN (82.66 mg, 1.315 mmol, 1.6 equiv. ) at room temperature. The resulting mixture was stirred for additional for 4 h at room temperature. Desired product could be detected by LCMS. The reaction was quenched with sat. NaHCO3 (aq. ) at room temperature. The resulting mixture was extracted with EA (3 x 20 mL) . The combined organic layers were dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2Cl2 /MeOH (10: 1) to afford tert-butyl 3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidine-1-carboxylate.
LC/MS: 738.40 [M+H] +.
Step 6. 6- { [4- (4- { [4- (Azetidin-3-yl) cyclohexyl] methyl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
To a solution of tert-butyl 3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidine-1-carboxylate (390 mg, 0.528 mmol, 1.0 equiv. ) in DCM (20 mL) was added TFA (4 mL) . The resulting mixture was stirred at room temperature for 1 h in air atmosphere. The reaction was concentrated under reduced pressure. The residue was purified by C18 column with ACN/H2O (0.05%NH4HCO3) to afford 6- { [4- (4- { [4- (azetidin-3-
yl) cyclohexyl] methyl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 638.65 [M+H] +.
Step 7. {3- [4- ( {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetic acid
To a solution of 6- { [4- (4- { [4- (azetidin-3-yl) cyclohexyl] methyl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (220 mg, 0.345 mmol, 1.0 equiv. ) in MeOH (8 mL) was added glyoxalate (25.54 mg, 0.345 mmol, 1 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 1 h at room temperature in air atmosphere. To the above mixture was added NaBH3CN (21.68 mg, 0.345 mmol, 1 equiv. ) at room temperature. The resulting mixture was stirred for additional for 5 minutes at room temperature. The reaction was purified by C18 column with ACN/H2O (0.5%TFA) to afford {3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetic acid.
LC/MS: 696.70 [M+H] +.
Step 8. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 48)
To a solution of {3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetic acid (150 mg, 0.216 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (102.09 mg, 0.238 mmol, 1.1 equiv. ) and HATU (122.94 mg, 0.324 mmol, 1.5 equiv. ) in DMF (4 mL) was added DIEA (111.44 mg, 0.864 mmol, 4.0 equiv. ) at room temperature in air atmosphere. The resulting mixture was stirred for 2 h at room temperature in air atmosphere. Desired product could be detected by LCMS. the reaction mixture was further purified by Prep-HPLC (Column: XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile
Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 42%B to 62%B in 11 min, 62%B; Wavelength: 254 nm; RT1 (min) : 10.67; Number Of Runs: 0) to afford Compound 48. (yield of (2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (s, 1H) , 9.00 (s, 1H) , 8.83 (s, 1H) , 8.62 (t, J = 6.0 Hz, 1H) , 8.05 (s, 1H) , 7.76 (d, J = 8.0 Hz, 1H) , 7.58 –7.61 (m, 4H) , 7.42 (s, 4H) , 6.92 (d, J = 8.7 Hz, 2H) , 5.74 -5.60 (m, 1H) , 5.33 (s, 1H) , 5.16 (d, J = 3.4 Hz, 1H) , 5.00 (m, 1H) , 4.83 (d, J = 17.2 Hz, 1H) , 4.51 -4.69 (m, 2H) , 4.26 -4.41 (m, 6H) , 3.50 -3.66 (m, 3H) , 2.92 -3.08 (m, 8H) , 2.46 (s, 7H) , 2.08 (m, 3H) , 1.64 -1.90 (m, 4H) , 1.47 (s, 8H) , 1.35 (s, 1H) , 0.94 (s, 10H) , 0.82 (s, 4H) .
LC/MS: 1108.60 [M+H] +.
Compound 49
Step 1. Methyl 2- (1- (4- (hydroxymethyl) cyclohexyl) azetidin-3-yl) acetate
4- (Hydroxymethyl) cyclohexan-1-one (992.3 mg, 7.742 mmol) , methyl 2- (azetidin-3-yl) acetate (1 g, 7.742 mmol) and AcOH (46 mg, 0.774 mmol) were dissolved in 15 mL MeOH in a 40 mL vial and stirred at 70℃ for 1 h. Then the mixture was cooled to r.t. NaBH3CN (973 mg, 15.485 mmol) was added and stirred at r.t. for 1h. LCMS showed the reaction successful, and the mixture was diluted with 100 mL EA and washed with NaHCO3 (aq. ) , the organic layer was combined and dried by Na2SO4, concentrated. The residue was purified by silica gel column with MeOH/DCM (0~15%) to afford the product.
Step 2. Methyl 2- (1- (4-formylcyclohexyl) azetidin-3-yl) acetate
Methyl 2- [1- (4-formylcyclohexyl) azetidin-3-yl] acetate (600 mg, 93.09%) To a stirred solution/mixture of oxalyl chloride (854.61 mg, 6.732 mmol, 2.5 equiv. ) in DCM (10 mL) was added DMSO (1052.22 mg, 13.465 mmol, 5 equiv. ) dropwise/in portions at -78 ℃ under N2 atmosphere. The resulting mixture stirred at -78℃ for 20 mins. Following this, methyl 2- {1- [4- (hydroxymethyl) cyclohexyl] azetidin-3-yl} acetate (650 mg, 2.693 mmol, 1.00 equiv. ) in 2 mL DCM was added at -78 ℃ and stirred for 30 mins. The reaction was quenched with TEA (1907.81 mg, 18.851 mmol, 7 equiv. ) at -78℃ and stirred at r.t. for 30 mins. The resulting mixture was extracted with DCM (3 x 30 mL) and washed with water, dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to afford methyl 2- [1- (4-formylcyclohexyl) azetidin-3-yl] acetate which was used to the next step without purification.
Step 3. Methyl 2- (1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) azetidin-3-yl) acetate
Methyl 2- [1- (4-formylcyclohexyl) azetidin-3-yl] acetate (860.70 mg, 3.598 mmol, 7 equiv. ) , 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250 mg, 0.514 mmol, 1 equiv. ) and acetic acid (3.70 mg, 0.062 mmol, 0.1 equiv. ) were dissolved in methanol (10 mL) in a 40 mL vial and stirred at 70℃ for 1 h. Then the mixture was cooled to r.t. . NaBH3CN (64.58 mg, 1.028 mmol, 2 equiv. ) was added and stirred at r.t. for 1h. LCMS showed the reaction succeed and the mixture diluted with 100 mL EA and washed with NaHCO3 (aq. ) , the organic layer was combined and dried by Na2SO4 and concentrated. The residue was purified by silica gel column with MeOH/DCM (0~15%) to afford methyl 2- {1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-3-yl} acetate.
LC/MS: 710.15 [M+H] +.
Step 4. {1- [4- ( {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-3-yl} acetic acid
Methyl 2- {1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-3-yl} acetate (170 mg, 0.239 mmol, 1.0 equiv. ) , THF (6 mL) , MeOH (2 mL) and LiOH (0.96 mL, 3
mol/L, 2.868 mmol, 12.0 equiv. ) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature for overnight. The mixture was concentrated under reduced pressure and purified by C18 column with MeOH/H2O (0.05%TFA) to afford the product.
LC/MS: 696.70 [M+H] +.
Step 5. (2S, 4R) -1- ( (S) -2- (2- (1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) azetidin-3-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 49)
{1- [4- ( {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-3-yl} acetic acid (100 mg, 0.144 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (61.88 mg, 0.144 mmol, 1.0 equiv. ) , HATU (81.96 mg, 0.216 mmol, 1.5 equiv. ) , DIEA (74.29 mg, 0.576 mmol, 4.0 equiv. ) and DMF (4 mL) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature for 2 hours. LCMS showed the reaction was completed and the reaction mixture was purified by Prep-HPLC (Column: XBridge Prep Phenyl OBD Column, 19*250 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3) , Mobile Phase B: ACN; Flow rate: 25 mL/min; Gradient: 65%B to 85%B in 7 min, 85%B; Wave Length: 254 nm; RT1 (min) : 3.9; to afford Compound 49 (2S, 4R) -1- ( (S) -2- (2- (1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) azetidin-3-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.02 (s, 1H) , 9.02 (S, 1H) , 8.87 (s, 1H) , 8.70 (s, 1H) , 8.45 (t, J = 6.0 Hz, 1H) , 7.92 (t, J = 7.7 Hz, 1H) , 7.76 -7.8 (m, 1H) , 7.41 -7.63 (m, 3H) , 7.22 -7.35 (m, 4H) , 6.79 (d, J = 8.8 Hz, 2H) , 5.58 -5.82 (m, 1H) , 5.45 -5.64 (m, 1H) , 5.20 (s, 1H) , 5.01 (d, J = 3.5 Hz, 1H) , 4.78 -4.95 (m, 1H) , 4.62 -4.76 (m, 2H) , 4.56 (d, J = 5.9 Hz, 1H) , 4.20 -4.44 (m, 3H) , 4.15 -4.22 (m, 1H) , 3.56 -3.72 (m, 2H) , 3.15 -3.28 (m, 2H) , 3.13 (s, 4H) , 2.54 (s, 2H) , 2.33 (d, J = 4.0 Hz, 6H) , 2.00 (d, J = 8.1 Hz, 4H) , 1.85 -1.96 (m, 2H) , 1.53 -1.82 (m, 3H) , 1.42 -1.50 (m, 6H) , 1.20 -1.38 (m, 5H) , 0.81 (s, 9H) , 0.71 (s, 4H) .
LC/MS: 1108.65 [M+H] +.
Compound 51
Step 1. Ethyl 4- [3- (hydroxymethyl) azetidin-1-yl] cyclohexane-1-carboxylate
To a solution of azetidin-3-ylmethanol (800 mg, 9.183 mmol) in MeOH (10 mL) was added ethyl 4-oxocyclohexane-1-carboxylate (2.3 g, 13.774 mmol) and acetic acid (0.01 mL) . The resulting mixture was maintained at 60 ℃ for 1h. After cooling down to r.t., the NaBH3CN (692.4 mg, 11.020mmol) was added. The resulting mixture was maintained at r.t. for 1h. The resulting mixture was extracted with ethyl acetate (2 x 30 mL) and washed with NaHCO3 (aq. ) . The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-40%MeOH/DCM) to afford the ethyl 4- [3- (hydroxymethyl) azetidin-1-yl] cyclohexane-1-carboxylate.
LC/MS: 228.10 [M+H] +.
Step 2. Ethyl 4- {3- [ (methanesulfonyloxy) methyl] azetidin-1-yl} cyclohexane-1-carboxylate
To a solution of the ethyl 4- [3- (hydroxymethyl) azetidin-1-yl] cyclohexane-1-carboxylate (400 mg, 1.657 mmol) and TEA (335.4 mg, 3.314 mmol) in DCM (10 mL) was added methanesulfonyl chloride (208.8 mg, 1.823 mmol) at 0 ℃. The mixture was stirred at room temperature for 1h. The resulting mixture was diluted with water and extracted with DCM (2 x 20 mL) . The organic layers were combined, dried over anhydrous sodium sulfate, filtered and concentrated. The residue obtained was purified by silica gel chromatography (0-20%MeOH/DCM) to afford the ethyl 4- {3- [ (methanesulfonyloxy) methyl] azetidin-1-yl} cyclohexane-1-carboxylate.
LC/MS: 306.30 [M+H] +.
Step 3. Methyl 4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylate
To a solution of the 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (300 mg, 0.617 mmol) was added methyl 4- {3- [ (methanesulfonyloxy) methyl] azetidin-1-yl} cyclohexane-1-carboxylate (188.29 mg, 0.617 mmol) and Cs2CO3 (302.26 mg, 0.925 mmol) . The mixture was stirred at 70 ℃ overnight. The residue was purified by C18 column with CH3CN/NH4HCO3 water (5%-100%) to afford the methyl 4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylate.
LC/MS: 696.65 [M+H] +.
Step 4. 4- [3- ( {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylic acid
To a solution of methyl 4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylate (300 mg, 0.431 mmol) in THF (10 mL) and MeOH (2.5 mL) was added lithium hydroxide (2.16 mL, 6.465 mmol, 3M in H2O) . The mixture was stirred at r.t.
overnight. The residue was purified by C18 column with CH3CN/NH4HCO3 water (5%-100%) to afford the 4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylic acid.
LC/MS: 682.40 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 51)
To a solution of 4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexane-1-carboxylic acid (140 mg, 0.205 mmol) in DMF (5 mL) was added (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (88.41 mg, 0.205 mmol) , TEA (62.33 mg, 0.615 mmol) and BOP (136.22 mg, 0.307 mmol) . The resulting mixture was maintained at r.t. overnight. The mixture was purified by Prep-HPLC [Column: XBridge Prep OBD C18 Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 18%B to 46%B in 7 min, 46%B; Wave Length: 254 nm; RT1 (min) : 6.1; Number Of Runs: 0] to afford the Compound 51 (2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [3- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) azetidin-1-yl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 8.94 -9.05 (m, 1H) , 8.58 (s, 1H) , 7.33 -7.46 (m, 4H) , 7.07 -7.29 (m, 5H) , 6.83 -7.05 (m, 2H) , 5.50 (s, 1H) , 5.04 -5.30 (m, 3H) , 4.69 -4.92 (m, 2H) , 4.10 -4.63 (m, 7H) , 2.93 -3.30 (m, 12H) , 2.58 -2.92 (m, 5H) , 2.20 -2.41 (m, 4H) , 1.55 -2.10 (m, 8H) , 1.17 -1.49 (m, 9H) , 0.72 -1.09 (m, 11H) .
LC/MS: 1094.55 [M+H] +.
Compound 52
Step 1. Methyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) cyclobut-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutene-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) 246yclobut-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (250 mg, 0.514 mmol, 1 equiv. ) , methyl 3- (4-oxopiperidin-1-yl) 246yclobutene-1-carboxylate (759.81 mg, 3.598 mmol, 7 equiv. ) , acetic acid (3.09 mg, 0.051 mmol, 0.1 equiv. ) and methanol (10 mL) were added to a 100 mL round-bottom flask with a stir bar and stirred at 60℃ for 1 h. Then NaBH3CN (64.58 mg, 1.028 mmol, 2 equiv. ) were added after the mixture being cooled to room temperature. And the reaction was stirred at room temperature for 6 h. EA were added to the mixture and washed by saturated sodium bicarbonate and saturated sodium chloride, the organic layer was separated and then dried over anhydrous sodium sulfate. The solvent was removed under vacuum and the residue was purified
by silica gel column with DCM/MeOH (0~20%) to afford methyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) cyclobut-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutene-1-carboxylate.
LC/MS: 682.60 [M+H] +.
Step 2. 3- (4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutane-1-carboxylic acid
Methyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutane-1-carboxylate (170 mg, 0.249 mmol, 1 equiv. ) , tetrahydrofuran (6.00 mL, 83.101 mmol, 333.74 equiv. ) , methanol (2.00 mL) , and LiOH (0.83 mL, 3 M in H2O) were added to a 100 mL round-bottom flask. And the reaction was stirred at r.t. for 2 h. The residue was purified by C18 column with H2O (0.5%TFA) /ACN from 5~100%. The solvent was removed under vacuum to afford 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutane-1-carboxylic acid.
LC/MS: 668.60 [M+H] +.
Step 3. (2S, 4R) -4-hydroxy-1- [ (2S) -2- { [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutyl] formamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 52)
3- (4- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutane-1-carboxylic acid (110 mg, 0.165 mmol, 1 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (70.9 mg, 0.165 mmol, 1 equiv. ) , HATU (59.5 mg, 0.247 mmol, 1.5 equiv. ) , dimethyl formamide (4 mL) and DIEA (63.8 mg, 0.495 mmol, 3 equiv. ) were added in a round-bottom flask with a stir bar. And then stirred at r.t. for 18 h. The residue obtained was purified by Prep-HPLC with the following conditions: Column, XBridge Shield RP18 OBD Column, 19*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN (30 %to 55%in 11 min, 55 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 30 %in 0.1 min, 30 %to 30 %in 1 min; Detector, UV 254 nm to afford Compound 52 (2S, 4R) -4-hydroxy-1- [ (2S) -2- { [3-
(4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutyl] formamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (300 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.99 (s, 1H) , 8.83 (s, 1H) , 8.58 (t, J = 6.0 Hz, 1H) , 8.05 (t, J = 7.9 Hz, 1H) , 7.70 -7.87 (m, 2H) , 7.53 -7.65 (m, 3H) , 7.34 -7.50 (m, 4H) , 6.92 (d, J = 8.8 Hz, 2H) , 5.57 -5.76 (m, 1H) , 5.34 (s, 1H) , 5.15 (d, J = 3.5 Hz, 1H) , 4.95 -5.05 (m, 1H) , 4.78 -4.87 (m, 1H) , 4.69 (d, J = 6.1 Hz, 2H) , 4.55 (d, J = 9.3 Hz, 1H) , 4.51 -4.33 (m, 3H) , 4.17 -4.28 (m, 1H) , 3.67 (s, 2H) , 3.08 (s, 4H) , 2.83 (s, 3H) , 2.63 (s, 4H) , 2.45 (s, 3H) , 2.16 (m, 2H) , 1.98 -2.09 (m, 2H) , 1.83 -1.98 (m, 3H) , 1.61 -1.80 (m, 4H) , 1.47 (s, 8H) , 1.24 (s, 1H) , 0.94 (d, J = 2.9 Hz, 9H) .
LC/MS: 1080.55 [M+H] +.
Compound 53
Step 1. Tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidine-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (500 mg, 1.028 mmol) , tert-butyl 4-oxopiperidine-1-carboxylate (614 mg, 3.084 mmol) , AcOH (6.17 mg, 0.103 mmol) and MeOH (25 mL) were added to a 100 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 1 h. Then NaBH3CN (130 mg 2.056 mmol) were added after the mixture was cooled to room temperature and then stirred at room temperature for 3 h. EA were added to the mixture and then washed by saturated sodium bicarbonate and saturated sodium chloride, the organic layer was separated and then dried over
anhydrous sodium sulfate. The solvent was removed under vacuum and the residue was purified by silica gel column with DCM/MeOH from 0~20%to afford tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidine-1-carboxylate.
LC/MS: 670.55 [M+H] +.
Step 2. 1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- ( {4- [4- (piperidin-4-yl) piperazin-1-yl] phenyl} amino) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidine-1-carboxylate (500 mg, 0.746 mmol, 1 equiv. ) , TFA (1 mL) and DCM (10 mL) were added to a round-bottom flask. Then the reaction was stirred at r.t. for 2 h. The residue was purified by C18 column with H2O (0.05%NH4HCO3) /ACN from 5~100%to afford 1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -6- ( {4- [4- (piperidin-4-yl) piperazin-1-yl] phenyl} amino) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 570.55 [M+H] +.
Step 3. Tert-butyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) azetidine-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- ( {4- [4- (piperidin-4-yl) piperazin-1-yl] phenyl} amino) -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (350 mg, 0.614 mmol, 1 equiv. ) , tert-butyl 3-oxoazetidine-1-carboxylate (631.04 mg, 3.684 mmol, 6 equiv. ) , acetic acid (3.69 mg, 0.061 mmol, 0.1 equiv. ) and methanol (10 mL) were added to a 100 mL round-bottom flask with a stir bar. Then the reaction was stirred at 60℃ for 1 h. Then NaBH3CN (77.21 mg, 1.228 mmol, 2 equiv. ) were added after the mixture being cooled to room temperature and then the reaction was stirred at room temperature for 16 h. EA were added to the mixture and then washed by saturated sodium bicarbonate and saturated sodium chloride, the organic layer was separated and then dried over anhydrous sodium sulfate. The solvent was removed under vacuum and the residue was purified by silica gel column with DCM/MeOH (0~20%) to afford tert-butyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) azetidine-1-carboxylate.
LC/MS: 725.45 [M+H] +.
Step 4. 6- [ (4- {4- [1- (Azetidin-3-yl) piperidin-4-yl] piperazin-1-yl} phenyl) amino] -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) azetidine-1-carboxylate (268 mg, 0.370 mmol, 1 equiv. ) , TFA (1 mL) and DCM (10 mL) were added to a round-bottom flask with a stirring bar. And then stirred at r.t. for 2 h. The residue was purified by C18 column with H2O (0.05%NH4HCO3) /ACN from 5~100%and then 100%ACN and then 100%MeOH to afford 6- [ (4- {4- [1- (azetidin-3-yl) piperidin-4-yl] piperazin-1-yl} phenyl) amino] -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one.
LC/MS: 625.40 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) azetidine-1-carbonylamino] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 53)
(2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (379.03 mg, 0.880 mmol, 5 equiv. ) , TEA (302.86 mg, 2.992 mmol, 17 equiv. ) and DCM (10 mL) were added to a 50 mL round-bottom flask with a stir bar under N2 and at 0℃. Triphosgene (182.86 mg, 0.616 mmol, 3.5 equiv. ) were added after the mixture were cooled to the 0℃ and then stirred at r.t. for 1.5 h. The solvent was removed at vacuum. EA was added and the solid was filtered. The solid obtained was added to the solution of 6- [ (4- {4- [1- (azetidin-3-yl) piperidin-4-yl] piperazin-1-yl} phenyl) amino] -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (110 mg, 0.176 mmol, 1 equiv. ) and TEA (302.86 mg, 2.992 mmol, 17 equiv. ) in DCM (1.5 mL) and the reaction was stirred at r.t. for 20 h. The residue obtained was purified by Prep-HPLC (XBridge Prep Phenyl OBD Column, 19*250 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3) , Mobile Phase B: ACN (38%to 58%in 7 min, 58 %to 100 %in 0.1 min, 100 %to 100 %in 2 min, 100%to 38 %in 0.1 min, 38%to 38%in 1 min; Detector, UV 254 nm) to afford Compound 53 (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) azetidine-1-
carbonylamino] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (s, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.49 -8.65 (m, 1H) , 8.05 (s, 1H) , 7.75 (d, J = 8.2 Hz, 1H) , 7.55 -7.65 (m, 3H) , 7.57 (s, 4H) , 6.85 -6.99 (m, 2H) , 5.77 -5.82 (m, 1H) , 5.58 -5.71 (m, 1H) , 5.32 (s, 1H) , 5.13 (d, J = 3.6 Hz, 1H) , 4.99 (m, 1H) , 4.82 (m, 1H) , 4.68 (d, J = 6.0 Hz, 2H) , 4.31 -4.47 (m, 2H) , 4.23 (m, 2H) , 3.90 (t, J = 7.6 Hz, 1H) , 3.75 -3.85 (m, 2H) , 3.66 (s, 4H) , 3.30 (s, 1H) , 3.08 (s, 4H) , 2.99 (s, 1H) , 2.72 -2.84 (m, 2H) , 2.56 -2.69 (m, 3H) , 2.42 -2.58 (m, 3H) , 2.17 -2.29 (m, 1H) , 2.04 (t, J = 10.4 Hz, 1H) , 1.96 -1.86 (m, 1H) , 1.68 -1.72 (m, 4H) , 1.46 (s, 8H) , 0.93 (s, 9H) .
LC/MS: 1081.60 [M+H] +.
Compound 54
Step 1. Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (3.0 g, 6.165 mmol, 1.0 equiv. ) , tert-butyl 3-oxoazetidine-1-carboxylate (3.17 g, 18.495 mmol, 3.0 equiv. ) , AcOH (20.7 mg) and MeOH (40 mL) were added to a 100 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 2 hours. The mixture was cooled by ice water, NaBH3CN (0.77 g, 12.330 mmol, 2.0 equiv. ) was added, and stirred at room temperature for 2 hours. EA was added after the mixture cooled to room temperature and washed
by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS : 642.15 [M+H] +.
Step 2. 6- ( {4- [4- (Azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate (2.0 g) , TFA (4 mL) and DCM (20 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 2 hours. The reaction mixture was concentrated under reduced pressure and purified by C18 column with MeCN/H2O (0.05%NH4HCO3) to afford the product.
LC/MS: 542.30 [M+H] +.
Step 3. Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidine] -1'-carboxylate
6- ( {4- [4- (azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (1.0 g, 1.846 mmol, 1.0 eq) , tert-butyl 3-oxoazetidine-1-carboxylate (1.9 g, 11.076 mmol, 6.0 eq) , AcOH (20.7 mg) and MeOH (10 mL) were added to a 50 mL round-bottom flask with a stir bar with a stir bar and stirred at 60 ℃ for 2 hours. The mixture was cooled by ice water, NaBH3CN (0.23 g, 3.692 mmol, 2.0 eq) was added, and stirred at room temperature for 1 hour. EA was added after the mixture cooled to room temperature and washed by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS : 697.40 [M+H] +.
Step 4. 6- { [4- (4- { [1, 3'-Biazetidin] -3-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidine] -1'-
carboxylate (700 mg) . TFA (2.0 mL) and DCM (10.0 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. The mixture was concentrated under reduced pressure and was purified by C18 column with MeCN/H2O (0.05%NH4HCO3) to afford the product.
LC/MS: 597.55 [M+H] +.
Step 5. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide
6- { [4- (4- { [1, 3'-biazetidin] -3-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (150 mg, 0.251 mmol, 1.0 equiv. ) , glyoxalate (18.61 mg, 0.251 mmol, 1.0 equiv. ) and MeOH (10 mL) were added to 50 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. The mixture was cooled by ice water, NaBH3CN (15.80 mg, 0.251 mmol, 1.0 equiv. ) was added, and stirred at room temperature for 2 min. The reaction mixture was concentrated under reduced pressure and purified by C18 column with MeOH/H2O (0.5%TFA) to afford the product.
LC/MS : 655.60 [M+H] +.
Step 6. (2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 54)
(3- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) acetic acid (180 mg, 0.275 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (59.18 mg, 0.138 mmol, 0.5 equiv. ) , BOP (182.38 mg, 0.413 mmol, 1.5 equiv. ) , TEA (111.27 mg, 1.100 mmol, 4.0 equiv. ) and DMF (3 mL) were added to a 10 mL schlenk tube with a stir bar and stirred at room temperature for 1 hour. The mixture was further purified by Prep-HPLC (Column: Xselect CSH C18 OBD Column 30*150mm 5μm, n; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 25%B to 40%B in 11 min, 40%B; Wave Length: 254 nm; RT1 (min) : 10.72; Number Of Runs: 0) to afford Compound 54 (2S, 4R) -4-
hydroxy-1- [ (2S) -2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.02 (s, 1H) , 8.95 -9.08 (m, 1H) , 8.86 (s, 1H) , 8.70 (s, 1H) , 8.18 -8.52 (m, 1H) , 7.96 -8.13 (m, 1H) , 7.91 (d, J = 8.1 Hz, 1H) , 7.40 -7.62 (m, 4H) , 7.28 (s, 4H) , 6.82 -6.95 (m, 2H) , 5.55 -5.77 (m, 1H) , 5.21 (s, 1H) , 5.04 (d, J = 3.4 Hz, 1H) , 4.82 -4.92 (m, 1H) , 4.56 (d, J = 6.1 Hz, 2H) , 4.13 -4.60 (m, 5H) , 3.50 -3.78 (m, 3H) , 3.02 -3.15 (m, 8H) , 2.97 (d, J = 4.4 Hz, 4H) , 2.42 -2.45 (m, 3H) , 2.25 (s, 4H) , 1.34 (s, 8H) , 1.10 -1.13 (m, 3H) , 0.82 (s, 9H) .
LC/MS : 1067.60 [M+H] +.
Compound 55
Step 1. Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (3.0 g, 6.165 mmol, 1.0 equiv. ) , tert-butyl 3-oxoazetidine-1-carboxylate (3.17 g, 18.495 mmol, 3.0 equiv. ) , AcOH (20.7 mg) and MeOH (40 mL) were added to a 100 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 2 hours. The mixture was cooled in an ice-water bath. Then NaBH3CN (0.77 g, 12.330 mmol, 2.0 equiv. ) was added, and stirred at room temperature for 2 hours. EA was added after the mixture being cooled to room temperature and washed by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate,
filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS: 642.15 [M+H] +.
Step 2. 6- ( {4- [4- (Azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate (2.0 g) , TFA (4 mL) and DCM (20 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 2 hours. The reaction mixture was concentrated under reduced pressure and purified by C18 column with MeCN/H2O (0.05%NH4HCO3) to afford the product.
LC/MS : 542.30 [M+H] +.
Step 3. Tert-butyl 3- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidine-1-carboxylate
6- ( {4- [4- (Azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.738 mmol, 1.0 equiv. ) , tert-butyl 3-formylazetidine-1-carboxylate (273.56 mg, 1.476 mmol, 2.0 equiv. ) , AcOH (20.7 mg) and MeOH (8 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 2 hours. The mixture was cooled in an ice-water bath. Then NaBH3CN (92.81 mg, 1.476 mmol, 2.0 equiv. ) was added, and the reaction was stirred at room temperature for 1.5 hours. EA was added after the mixture being cooled to room temperature and washed by saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS : 711.65 [M+H] +.
Step 4. 6- [ (4- {4- [1- (azetidin-3-ylmethyl) azetidin-3-yl] piperazin-1-yl} phenyl) amino] -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidine-1-carboxylate (360 mg) and HCl-1, 4-dioxane (20 mL) were added to a 100 mL round-bottom flask
with a stir bar and stirred at room temperature for 1 hour. The reaction mixture was purified by C18 gel column with H2O (NH4HCO3, 0.05%) /THF/MeOH to afford the product.
LC/MS: 611.55 [M+H] +.
Step 5. {3- [ (3- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidin-1-yl} acetic acid
6- [ (4- {4- [1- (Azetidin-3-ylmethyl) azetidin-3-yl] piperazin-1-yl} phenyl) amino] -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.327 mmol, 1.0 equiv. ) , glyoxalate (24.24 mg, 0.327 mmol, 1.0 equiv. ) and MeOH (10 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 2 hours. NaBH3CN (20.58 mg, 0.327 mmol, 1.0 equiv. ) was added, and stirred at room temperature for 2 min. The reaction mixture was purified by C18 gel column with MeCN/H2O (0.05%TFA) to the product.
LC/MS : 669.60 [M+H] +.
Step 6. (2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 55)
{3- [ (3- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidin-1-yl} acetic acid (70 mg, 0.105 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (36.05 mg, 0.105 mmol, 1.0 equiv. ) , HATU (54.11 mg, 0.420 mmol, 1.5 equiv. ) , DIEA (3.86 mg, 0.028 mmol, 4.0 equiv. ) and DMF (3 mL) were added to a 10 mL schlenk tube with a stir bar and stirred at room temperature for 1 hour. The mixture was further purified by Prep-HPLC (Column: Xselect CSH C18 OBD Column 30*150mm 5μm, n; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 36%B to 56%B in 9 min, 56%B; Wave Length: 254 nm; RT1 (min) : 8.35; Number Of Runs: 0) to afford Compound 55 (2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-
yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.14 (s, 1H) , 8.97 (s, 1H) , 8.81 (s, 1H) , 8.58 -8.70 (m, 1H) , 7.96 -8.11 (m, 1H) , 7.73 (d, J = 8.1 Hz, 1H) , 7.55 (s, 4H) , 7.39 (s, 4H) , 6.86 -6.96 (m, 2H) , 5.55 -5.74 (m, 1H) , 5.32 (s, 1H) , 5.14 (d, J = 3.4 Hz, 1H) , 4.88 -5.15 (m, 1H) , 4.74 -4.86 (m, 1H) , 4.66 (d, J = 6.0 Hz, 2H) , 4.31 -4.53 (m, 4H) , 4.20 -4.28 (m, 1H) , 3.69 -3.60 (m, 1H) , 3.56 -3.75 (m, 2H) , 3.10 -3.52 (m, 2H) , 3.05 (d, J = 8.9 Hz, 5H) , 2.76 -2.99 (m, 5H) , 2.76 (t, J = 6.4 Hz, 2H) , 2.43 (s, 4H) , 2.33 (s, 5H) , 1.87 -2.10 (m, 2H) , 1.44 (s, 6H) , 1.22 (s, 1H) , 0.91 (s, 9H) .
LC/MS : 1081.50 [M+H] +.
Compound 56
Step 1. Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate
1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (3.0 g, 6.165 mmol, 1.0 equiv. ) , tert-butyl 3-oxoazetidine-1-carboxylate (3.17 g, 18.495 mmol, 3.0 equiv. ) , AcOH (20.7 mg) and MeOH (40 mL) were added to a 100 mL round-bottom flask with a stir bar and stirred at 60 ℃ for 2 hours. The mixture was cooled in an ice-water bath. Then NaBH3CN (0.77 g, 12.330 mmol, 2.0 equiv. ) was added, and
the reaction was stirred at room temperature for 2 hours. EA was added after the mixture being cooled to room temperature. The organic layer was washed with saturated NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS : 642.15 [M+H] +.
Step 2. 6- ( {4- [4- (Azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidine-1-carboxylate (2.0 g) , TFA (4 mL) and DCM (20 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 2 hours. The reaction mixture was concentrated under reduced pressure and purified by C18 column with MeCN/H2O (0.05%NH4HCO3) to afford the product.
LC/MS: 542.30 [M+H] +.
Step 3. Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidine] -1'-carboxylate
6- ( {4- [4- (azetidin-3-yl) piperazin-1-yl] phenyl} amino) -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (1.0 g, 1.846 mmol, 1.0 equiv. ) , tert-butyl 3-oxoazetidine-1-carboxylate (1.9 mg, 11.076 mmol, 6.0 equiv. ) , AcOH (20.7 mg) and MeOH (10 mL) were added to a 50 mL round-bottom flask with a stir bar. The reaction was stirred at 60 ℃ for 2 hours. The mixture was cooled in an ice-water batch. NaBH3CN (0.23 g, 3.692 mmol, 2.0 equiv. ) was added, and stirred at room temperature for 1 hour. EA was added after the mixture being cooled to room temperature and washed with saturated aq. NaHCO3 (10 mL) , then dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS: 697.40 [M+H] +.
Step 4. 6- { [4- (4- { [1, 3'-Biazetidin] -3-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one
Tert-butyl 3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidine] -1'-carboxylate (700 mg) , TFA (2.0 mL) and DCM (10.0 mL) were added to a 50 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. The mixture was concentrated under reduced pressure and the residue was purified by C18 column with MeCN/H2O (0.05%NH4HCO3) to afford the product.
LC/MS: 597.55 [M+H] +.
Step 5. Methyl 3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) propanoate
6- { [4- (4- { [1, 3'-Biazetidin] -3-yl} piperazin-1-yl) phenyl] amino} -1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (280 mg, 0.469 mmol, 1.0 equiv. ) , methyl acrylate (40.39 mg, 0.469 mmol, 1.0 equiv. ) and MeOH (10 mL) were added to a round-bottom flask with a stir bar. The reaction was stirred at 60 ℃ for 1 hour. The reaction mixture was purified by silica gel column with DCM/MeOH to afford the product.
LC/MS: 683.65 [M+H] +.
Step 6. 3- (3- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) propanoic acid
Methyl 2- {1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-3-yl} acetate (107 mg, 0.157 mmol, 1.0 equiv. ) , THF (6 mL) , MeOH (2 mL) and LiOH (0.63 mL, 3 mol/L, 1.884 mmol, 12.0 equiv. ) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure. The residue was purified by C18 column with MeOH/H2O (0.05%TFA) to afford the product.
LC/MS: 669.65 [M+H] +.
Step 7. (2S, 4R) -1- ( (S) -2- (3- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 3'-
biazetidin] -1'-yl) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide (Compound 56)
3- (3- {4- [4- ( {1- [6- (2-Hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) propanoic acid (80 mg, 0.120 mmol, 1.0 equiv. ) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (41.20 mg, 0.096 mmol, 0.8 equiv. ) , BOP (79.36 mg, 0.180 mmol, 1.5 equiv. ) , TEA (48.42 mg, 0.480 mmol, 4.0 equiv. ) and DMF (3 mL) were added to a 25 mL round-bottom flask with a stir bar and stirred at room temperature for 1 hour. The mixture was further purified by Prep-HPLC (Column: XBridge Shield RP18 OBD Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3+0.1%NH3. H2O) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 40%B to 52%B in 10 min, 52%B; Wave Length: 254 nm; RT1 (min) : 6.38; Number Of Runs: 0) to afford Compound 56 (2S, 4R) -1- ( (S) -2- (3- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 3'-biazetidin] -1'-yl) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide.
1H NMR (300 MHz, DMSO-d6) δ 10.00 (s, 1H) , 9.01 (s, 1H) , 8.86 (s, 1H) , 8.70 (s, 1H) , 8.44 (t, J = 5.9 Hz, 1H) , 8.21 (d, J = 9.3 Hz, 1H) , 7.92 (t, J = 7.8 Hz, 1H) , 7.63 (d, J = 8.1 Hz, 1H) , 7.44 (s, 3H) , 7.22 -7.35 (m, 4H) , 6.85 -3.96 (m, 2H) , 5.54 -5.75 (m, 1H) , 5.20 -5.42 (m, 1H) , 5.19 (s, 1H) , 5.01 (s, 1H) , 4.72 -4.87 (m, 1H) , 4.58 -4.70 (m, 2H) , 4.56 (d, J = 6.1 Hz, 4H) , 4.24 (s, 1H) , 3.63 (s, 2H) , 3.41 (s, 2H) , 3.11 (s, 3H) , 2.97 (s, 5H) , 2.78 (d, J = 7.8 Hz, 3H) , 2.32 (s, 3H) , 2.26 (s, 1H) , 2.07 -2.25 (m, 2H) , 1.88 (s, 3H) , 1.34 (s, 6H) , 1.12 (s, 5H) , 0.83 (s, 8H) .
LC/MS: 1081.60 [M+H] +.
Compound 57
Step 1. 3-azaspiro [5.5] undecan-9-one
To a solution of tert-butyl 9-oxo-3-azaspiro [5.5] undecane-3-carboxylate (3 g, 11.220 mmol) in DCM (5 mL) was added TFA (4 mL) , The resulting mixture was stirred at r.t for 1 h . The reaction was monitored by LCMS. The reaction was concentrated to afford 3-azaspiro [5.5] undecan-9-one.
LC/MS: 168.13 [M+H] +.
Step 2. ethyl 2- {9-oxo-3-azaspiro [5.5] undecan-3-yl} acetate
To a solution of 3-azaspiro [5.5] undecan-9-one (1.5 g, 8.969 mmol, 1 equiv) in DMF (5 mL) was added K2CO3 (6.20 g, 44.845 mmol, 5 equiv) , ethyl bromoacetate (1.20 g, 7.175 mmol, 0.8 equiv) . The resulting mixture was stirred at r.t for 16 h. The reaction was monitored by LCMS. The reaction was quenched with H2O (25mL) . The resulting mixture was extracted with EtOAc (3 x
50 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) to afford ethyl 2- {9-oxo-3-azaspiro [5.5] undecan-3-yl} acetate.
LC/MS: 254.17 [M+H] +.
Step 3. ethyl 2- (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetate
To a solution of 1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (200 mg, 0.390 mmol, 1 equiv) in MeOH (10 mL) was added ethyl 2- {9-oxo-3-azaspiro [5.5] undecan-3-yl} acetate (197.68 mg, 0.780 mmol, 2 equiv) , AcOH (40 mg) . The resulting mixture was stirred at 60 ℃ for 1.5 h, then added NaBH3CN (36.78 mg, 0.585 mmol, 1.5 equiv) . The resulting mixture was stirred at 60 ℃ for 4 h. The reaction was monitored by LCMS. On completion. The reaction was quenched with H2O (5mL) . The resulting mixture was extracted with EtOAc (3 x 25 mL) . The combined organic layers were washed with brine (2 x 30 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) to afford ethyl 2- (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetate.
LC/MS: 750.44 [M+H] +.
Step 4. (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid
To a solution of ethyl 2- (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetate (500 mg, 0.667 mmol, 1 equiv) in H2O (5 mL) and THF (5 mL) was added LiOH. H2O (139.87 mg, 3.335 mmol, 5 equiv) . The resulting mixture was stirred at r.t for 16 h. The reaction was monitored by LCMS. On completion, the residue was purified by reverse flash chromatography with the following conditions: column. C18 silica gel; mobile phase, MeCN in water, 0%to 50%gradient in 30min detector, UV254, 220 nm. This resulted in (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-
yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid.
LC/MS: 722.41 [M+H] +.
Step 5. (2S, 4R) -1- [ (2S) -2- [2- (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetamido] -3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 57)
To a solution of (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetic acid (160 mg, 0.222 mmol, 1 equiv) in DMF (5 mL, 64.608 mmol, 291.50 equiv) was added NMI (109.19 mg, 1.332 mmol, 6 equiv) , TCFH (186.56 mg, 0.666 mmol, 3 equiv) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (95.43 mg, 0.222 mmol, 1 equiv) , the resulting mixture was stirred at r.t for 1 h, The reaction was monitored by LCMS. On completion. the reaction was quenched with H2O (25mL) . The resulting mixture was extracted with EtOAc (3 x 50 mL) . The combined organic layers were washed with brine (3 x 100 mL) then concentrated, the residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) , then purified by Prep-HPLC with the following conditions: Column, Sun Fire Prep C18, 19*150mm 5umH PrepC-001 (T) ; mobile phase, Phase A: water (10 mmol/L NH4HCO3+0.1%NH3. H2O) ; Phase B: CH3CN (40%CH3CN up to 70%in 7 min, up to 70%CH3CN in 1 min, hold 100%in 1 min, down to 40%CH3CN in 0.1min, hold 40%in 0.9min) ; Detector, UV220 &254nm to afford (2S, 4R) -1- [ (2S) -2- [2- (9- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetamido] -3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.11 (brs, 1H) , 9.00 (s, 1H) , 8.82 (s, 1H) , 8.63 (t, J = 61 Hz, 1H) , 7.93 (brs, 1H) , 7.82 (d, J = 10 Hz, 1H) , 7.69 (d, J = 8.2 Hz, 1H) , 7.52 -7.61 (m, 2H) , 7.43 (q, J = 8.3 Hz, 4H) , 6.90 (d, J = 8.4 Hz, 2H) , 5.58 -5.78 (m, 1H) , 5.16 (d, J = 3.2 Hz, 1H) , 5.06 (s, 1H) , 5.01 (d, J = 10 Hz, 1H) , 4.85 (d, J = 17.1 Hz, 1H) , 4.70 -4.82 (m, 1H) , 4.50 -4.65 (m, 2H) , 4.32 -4.47 (m, 3H) , 4.10 -4.30 (m, 1H) , 3.52 -3.70 (m, 2H) , 3.06 (s, 4H) , 2.90 -3.03 (m, 3H) , 2.73 -2.85 (m, 1H) , 2.62 (s, 4H) , 2.40 -2.44 (m, 7H) , 2.13 -2.22 (m, 2H) , 1.99 -2.10 (m,
2H) , 1.86 -1.98 (m, 2H) , 1.61 -1.75 (m, 3H) , 1.42 -1.60 (m, 4H) , 1.28 -1.40 (m, 4H) , 1.01 -1.12 (m, 2H) , 0.94 (s, 9H) , 0.87 (t, J = 7.2 Hz, 3H) .
LC/MS: 1134.55 [M+H] +.
Compound 58
Step 1. 7-azaspiro [3.5] nonan-2-one
To a solution of tert-butyl 2-oxo-7-azaspiro [3.5] nonane-7-carboxylate (3 g, 12.536 mmol, 1 equiv) in DCM (5 mL) was added TFA (4 mL) , The resulting mixture was stirred at r.t for 1 h . The reaction was monitored by LCMS. The reaction was concentrated to afford 7-azaspiro [3.5] nonan-2-one.
LC/MS: 140.10 [M+H] +.
Step 2. ethyl 2- {2-oxo-7-azaspiro [3.5] nonan-7-yl} acetate
To a solution of 7-azaspiro [3.5] nonan-2-one (1.6 g, 11.494 mmol, 1 equiv) in DMF (5 mL) was added K2CO3 (7.94 g, 57.470 mmol, 5 equiv) , ethyl bromoacetate (1.54 g, 9.195 mmol, 0.8 equiv) . The resulting mixture was stirred at r.t for 1 h. The reaction was quenched with H2O (15mL) . The resulting mixture was extracted with EtOAc (3 x 40 mL) . The combined organic layers were washed with brine (2 x 50 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) to afford ethyl 2- {2-oxo-7-azaspiro [3.5] nonan-7-yl} acetate.
LC/MS: 226.14 [M+H] +.
Step 3. ethyl 2- (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate
To a solution of 1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -6- { [4- (piperazin-1-yl) phenyl] amino} -2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-3-one (400 mg, 0.780 mmol, 1 equiv) in MeOH (10 mL) was added ethyl 2- {2-oxo-7-azaspiro [3.5] nonan-7-yl} acetate (404.33 mg, 1.794 mmol, 2.3 equiv) , AcOH (40 mg, 0.666 mmol, 0.85 equiv) . The resulting mixture was stirred at 60 ℃ for 1.5 h, then added NaBH3CN (73.55 mg, 1.170 mmol, 1.5 equiv) . The resulting mixture was stirred at 60 ℃ for 4 h. The reaction was monitored by LCMS. The reaction was quenched with H2O (5mL) . The resulting mixture was extracted with EtOAc (3 x 25 mL) . The combined organic layers were washed with brine (2 x 30 mL) then concentrated. The residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) to afford ethyl 2- (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate.
LC/MS: 722.41 [M+H] +.
Step 4. (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid
To a solution of ethyl 2- (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetate (500 mg, 0.693 mmol, 1 equiv) in H2O (3 mL) and THF (3 mL)
was added LiOH. H2O (145.31 mg, 3.465 mmol, 5 equiv) . The resulting mixture was stirred at r.t for 16 h. The reaction was monitored by LCMS. On completion, the residue was purified by reverse flash chromatography with the following conditions: column. C18 silica gel; mobile phase, MeCN in water, 0%to 50%gradient in 30min detector, UV254, 220 nm. This resulted in (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid.
LC/MS: 694.38 [M+H] +.
Step 5. (2S, 4R) -1- [ (2S) -2- [2- (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetamido] -3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (Compound 58)
To a solution of (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetic acid (200 mg, 0.288 mmol, 1 equiv) in DMF (5 mL) was added NMI (142.00 mg, 1.728 mmol, 6 equiv) , TCFH (242.63 mg, 0.864 mmol, 3 equiv) , (2S, 4R) -1- [ (2S) -2-amino-3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide (124.11 mg, 0.288 mmol, 1 equiv) , the resulting mixture was stirred at r.t for 1 h, The reaction was monitored by LCMS. On completion. the reaction was quenched with H2O (10mL) . The resulting mixture was extracted with EtOAc (3 x 30 mL) . The combined organic layers were washed with brine (3 x 50 mL) then concentrated, the residue obtained was purified by silica gel chromatography (0-10%MeOH/DCM) , then purified by Prep-HPLC with the following conditions: Column: XBridge Shield RP18 OBD Column, 30*150 mm, 5μm; Mobile Phase A: Water (10 mmol/L NH4HCO3) , Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 42%B to 62%B in 7 min, 62%B; Wave Length: 254 nm; RT1 (min) : 5.18; Number Of Runs: 0 to afford Compound 58 (2S, 4R) -1- [ (2S) -2- [2- (2- {4- [4- ( {1- [ (7R) -7-ethyl-7-hydroxy-5H, 6H-cyclopenta [b] pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetamido] -3, 3-dimethylbutanoyl] -4-hydroxy-N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide.
1HNMR (400 MHz, DMSO-d6) δ 10.12 (brs, 1H) , 9.00 (s, 1H) , 8.82 (s, 1H) , 8.63 (t, J = 6.1 Hz, 1H) , 7.93 (brs, 1H) , 7.84 (d, J = 9.6 Hz, 1H) , 7.69 (d, J = 8.4 Hz, 1H) , 7.59 (brs, 2H) , 7.42 (q, J = 8.4 Hz, 4H) , 6.92 (d, J = 9.6 Hz, 2H) , 5.65 -5.75 (m, 1H) , 5.16 (d, J = 3.2 Hz, 1H) , 5.07 (s, 1H) ,
4.99 (d, J = 10.8 Hz, 1H) , 4.85 (d, J = 17.2 Hz, 1H) , 4.70 -4.76 (m, 1H) , 4.56 -4.63 (m, 1H) , 4.38 -4.52 (m, 4H) , 4.27 -4.36 (m, 1H) , 3.61 -3.65 (m, 2H) , 3.08 (m, 4H) , 2.92 -3.01 (m, 2H) , 2.72 -2.90 (m, 2H) , 2.62 -2.70 (m, 1H) , 2.46 (s, 3 H) , 2.41 -2.44 (m, 2H) , 2.29 -2.38 (m, 6H) , 2.14 -2.22 (m, 1H) , 2.00 -2.10 (m, 2H) , 1.82 -1.96 (m, 4H) , 1.59 -1.76 (m, 1H) , 1.58 -1.62 (m, 2H) , 1.48 -1.56 (m, 4H) , 0.94 (s, 9H) , 0.87 (t, J = 7.2 Hz, 3H) .
LC/MS: 1106.50 [M+H] +.
Compound 59
Step 1. tert-butyl 8- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -2-azaspiro [4.5] decane-2-carboxylate
To the solution of tert-butyl 8-oxo-2-azaspiro [4.5] decane-2-carboxylate (78.10 mg, 308.28 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4-
(piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (71.51 μL, 513.80 μmol) in order. The mixture was stirred at 25 ℃ for 2 h. Then NaBH (OAc) 3 (108.90 mg, 513.80 μmol) was added and the resulting mixture was stirred for additional 14 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 724.5 [M+H] +.
Step 2. 6- ( (4- (4- (2-azaspiro [4.5] decan-8-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl 8- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -2-azaspiro [4.5] decane-2-carboxylate (29.2 mg, 40.34 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 624.3 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- (8- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -2-azaspiro [4.5] decan-2-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 59)
To the solution of 6- ( (4- (4- (2-azaspiro [4.5] decan-8-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (25 mg, 37.92 μmol) in DMAC (2 mL) was added ( (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (27.64 mg, 48.09 μmol) , DIPEA (20.94 uL, 120.23 μmol) , HOAt (6.55 mg, 48.09 μmol) , HATU (18.29 mg, 48.09 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Boston Green ODS 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 22 %-52 %, 12 min) to afford Compound 59.
LCMS: MS (ESI) m/z 590.8 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.27 (br, 1H) , 9.01 (s, 1H) , 8.85 (s, 1H) , 8.60 -8.54 (m, 1H) , 8.08 -7.97 (m, 1H) , 7.76 (d, J = 8.0 Hz, 1H) , 7.63 -7.61 (m, 3H) , 7.50 -7.35 (m, 4H) , 7.26 -7.24 (m, 1H) , 7.02 (d, J = 8.8 Hz, 2H) , 5.74 -5.60 (m, 1H) , 5.30-5.21 (m, 1H) , 5.01 (d, J = 9.2 Hz 1H) , 4.83 (dd, J = 1.2, 17.2 Hz 1H) , 4.69 (d, J = 4.4 Hz, 2H) , 4.58 (d, J = 9.2 Hz, 1H) , 4.51 -4.42 (m, 1H) , 4.29 (br, 1H) , 3.82 (d, J = 12.0 Hz, 3H) , 3.43 -3.10 (m, 10H) , 3.08 -2.87 (m, 3H) , 2.86 -2.67 (m, 2H) , 2.47 (m, 3H) , 2.12 -1.91 (m, 3H) , 1.83 -1.53 (m, 5H) , 1.52-1.47 (m, 8H) , 1.42 -1.29 (m, 4H) , 1.28 -1.18 (m, 3H) , 0.98 (s, 9H) .
Compound 60
Step 1. tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -8-azaspiro [4.5] decane-8-carboxylate
To the solution of tert-butyl 2-oxo-8-azaspiro [4.5] decane-8-carboxylate (31.24 mg, 123.31 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-
yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (43.56 mg, 205.52 μmol) was added and the resulting mixture was stirred for additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 724.9 [M+H] +.
Step 2. 6- ( (4- (4- (8-azaspiro [4.5] decan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -8-azaspiro [4.5] decane-8-carboxylate (67.1 mg, 92.69 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 624.5 [M+H] +.
Step 3. (2S, 4R) -N- ( (1S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -8-azaspiro [4.5] decan-8-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 60)
To the solution of 6- ( (4- (4- (8-azaspiro [4.5] decan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (57.82 mg, 92.69 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (63.92 mg, 111.23 μmol) , DIPEA (48.44 uL, 278.07 μmol) , HOAt (15.14 mg, 111.23 μmol) , HATU (42.29 mg, 111.23 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Boston Green ODS 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 22%-42 %, 10 min) to afford Compound 60.
LCMS: MS (ESI) m/z 591.1 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.20 (br s, 1H) , 9.56 (br s, 1H) , 9.06 -8.96 (m, 1H) , 8.85 (s, 1H) , 8.52 (d, J = 5.2 Hz, 1H) , 8.02 (t, J = 7.6 Hz, 1H) , 7.75 (m, 1H) , 7.68 -7.60 (m, 3H) , 7.47 -7.38 (m, 4H) , 7.25 (d, J = 8.8 Hz, 1H) , 7.01 (m, 2H) , 5.74 -5.59 (m, 1H) , 5.23-5.17 (m, 1H) , 4.99 (d, J = 10.8 Hz, 1H) , 4.82 (d, J = 18.0 Hz, 1H) , 4.68 (m, 2H) , 4.58 (m, 1H) , 4.48 -4.43 (m, 1H) , 4.28 (m, 1H) , 3.63 -3.23 (m, 13H) , 3.13 (m, 2H) , 2.99 -2.74 (m, 4H) , 2.47 (m, 3H) , 2.20 -1.71 (m, 5H) , 1.53 (m, 2H) , 1.46 (s, 6H) , 1.43 -1.26 (m, 5H) , 1.26 -1.17 (m, 3H) , 1.16 -1.04 (m, 1H) , 1.01 -0.93 (m, 9H) .
Compound 61
Step 1. tert-butyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -3-azaspiro [5.5] undecane-3-carboxylate
To the solution of tert-butyl 9-oxo-3-azaspiro [5.5] undecane-3-carboxylate (82.97 mg, 310.33 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (31.19 mg, 308.28μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (103.56 mg, 488.63 μmol) was added and the resulting mixture was stirred for additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 738.9 [M+H] +.
Step 2. 6- ( (4- (4- (3-azaspiro [5.5] undecan-9-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl 9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -3-azaspiro [5.5] undecane-3-carboxylate (32.4 mg, 43.91 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 638.4 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- (9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -3-azaspiro [5.5] undecan-3-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 61)
To the solution of 6- ( (4- (4- (3-azaspiro [5.5] undecan-9-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (28 mg, 43.90 μmol) in DMAC (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (25.23 mg, 43.90 μmol) , DIPEA (22.94 uL, 131.70 μmol) , HOAt (7.17 mg, 52.68 μmol) , HATU (20.03 mg, 52.68 μmol)
in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Boston Green ODS 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 17 %-47 %, 12 min) to afford Compound 61.
LCMS: MS (ESI) m/z 598.1 [M/2 + H] +.
1H NMR (400MHz, DMSO-d6) δ 10.14 (br s, 1H) , 8.99 (s, 1H) , 8.82 (s, 1H) , 8.53 (t, J = 8.8 Hz, 1H) , 8.11 -8.00 (m, 1H) , 7.74 (d, J = 7.6 Hz, 1H) , 7.64 -7.53 (m, 3H) , 7.47 -7.37 (m, 4H) , 7.26 (d, J = 7.2 Hz, 1H) , 6.90 (d, J = 8.8 Hz, 2H) , 5.73 -5.59 (m, 1H) , 5.33 (s, 1H) , 5.25 -5.12 (m, 2H) , 4.99 (d, J = 10.6 Hz, 1H) , 4.82 (d, J = 18.4 Hz, 1H) , 4.68 (d, J = 4.4 Hz, 2H) , 4.58 (d, J = 9.2 Hz, 1H) , 4.48 -4.38 (m, 1H) , 4.32 -4.24 (m, 1H) , 3.62 -3.52 (m, 2H) , 3.49 -3.44 (m, 2H) , 3.07 (m, 4H) , 2.88 -2.77 (m, 2H) , 2.61 (m, 4H) , 2.48 –2.43 (m, 3H) , 2.25 -2.14 (m, 1H) , 2.09 -1.98 (m, 1H) , 1.81 -1.67 (m, 2H) , 1.66 -1.49 (m, 3H) , 1.46 (s, 6H) , 1.42 -1.26 (m, 5H) , 1.27 -1.09 (m, 5H) , 1.09 -0.98 (m, 4H) , 0.95 (s, 9H) .
Compound 62
Step 1. tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -6-azaspiro [3.4] octane-6-carboxylate
To the solution of tert-butyl 2-oxo-6-azaspiro [3.4] octane-6-carboxylate (77.78 mg, 345.25 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (93.56 mg, 441.44 μmol) was added and the resulting mixture was stirred for additional 1 h. LCMS ( (ES23824-13-P1c) showed 67.0 %of desired product was detected. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with
DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 696.4 [M+H] +.
Step 2. 6- ( (4- (4- (6-azaspiro [3.4] octan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -6-azaspiro [3.4] octane-6-carboxylate 62.5 mg, 89.82 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 596.4 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -6-azaspiro [3.4] octan-6-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 62)
To the solution of 6- ( (4- (4- (6-azaspiro [3.4] octan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (53.51 mg, 89.82 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (61.94 mg, 107.79 μmol) , DIPEA (46.94 uL, 269.46 μmol) , HOAt (14.67 mg, 107.79 μmol) , HATU (40.98 mg, 107.79 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Boston Green ODS 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 22 %-42 %, 10 min) to afford Compound 62.
LCMS: MS (ESI) m/z 577.0 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.21 (br s, 1H) , 9.99 -9.69 (m, 1H) , 9.01 (s, 1H) , 8.85 (s, 1H) , 8.61 –8.51 (m, 1H) , 8.02 (t, J = 8.0 Hz, 1H) , 7.75 (m, 1H) , 7.67 –7.59 (m, 3H) , 7.47 -7.35 (m, 4H) , 7.24 (d, J = 9.2 Hz, 1H) , 7.01 (d, J = 8.4 Hz, 2H) , 5.75 –5.60 (m, 1H) , 5.27 –5.14 (m, 1H) , 4.99 (d, J = 10.0 Hz, 1H) , 4.82 (m, 1H) , 4.68 (m, 2H) , 4.58 (m, 1H) , 4.52 –4.41 (m, 1H) , 4.28 (m, 1H) , 3.92 -3.73 (m, 2H) , 3.50 -3.32 (m, 8H) , 3.30 -3.15 (m, 3H) , 3.06 -2.84 (m, 4H) , 2.83 -2.65
(m, 2H) , 2.47 (s, 3H) , 2.29 -2.00 (m, 5H) , 1.98 -1.69 (m, 3H) , 1.46 (s, 6H) , 1.43 –1.28 (m, 2H) , 1.27 –1.19 (m, 2H) , 1.00 -0.94 (s, 9H) .
Compound 63
Step 1. tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -7-azaspiro [3.5] nonane-7-carboxylate
To the solution of tert-butyl 2-oxo-7-azaspiro [3.5] nonane-7-carboxylate (79.51 mg, 332.25 μmol) in DCM (2 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (103.56 mg, 488.63 μmol) was added and the resulting mixture was stirred for
additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 710.5 [M+H] +.
Step 2. 6- ( (4- (4- (7-azaspiro [3.5] nonan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl 2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -7-azaspiro [3.5] nonane-7-carboxylate (52 mg, 72.67 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 610.5 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -7-azaspiro [3.5] nonan-7-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 63)
To the solution of 6- ( (4- (4- (7-azaspiro [3.5] nonan-2-yl) piperazin-1-yl) phenyl) amino) -2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (47 mg, 72.73 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (50.15 mg, 87.28 μmol) , DIPEA (38.00 uL, 218.19 μmol) , HATU (33.19 mg, 87.28 μmol) , HOAt (11.88 mg, 87.28 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Boston Green ODS 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 20 %-40 %, 12 min) to afford Compound 63.
LCMS: MS (ESI) m/z 584.1 [M/2 + H] +.
1H NMR (400MHz, MeOD) δ 8.89 (s, 1H) , 8.80 (s, 1H) , 8.42 (s, 1H) , 7.96 (t, J = 8.0 Hz, 1H) , 7.76 (d, J = 8.4 Hz, 1H) , 7.64 (d, J = 7.6 Hz, 1H) , 7.57 (d, J = 8.8 Hz, 2H) , 7.53 -7.48 (m, 1H) , 7.47 (d, J = 1.6 Hz, 3H) , 7.46 -7.42 (m, 1H) , 6.98 (d, J = 9.2 Hz, 2H) , 5.77 -5.65 (m, 1H) , 5.40 -5.33 (m, 1H) , 5.03 (d, J = 10.4 Hz, 1H) , 4.81 (d, J = 6.8 Hz, 3H) , 4.75 (d, J = 6.8 Hz, 2H) , 4.59 -
4.55 (m, 2H) , 4.46 (s, 1H) , 3.87 -3.81 (m, 1H) , 3.80 -3.74 (m, 1H) , 3.51 -3.38 (m, 3H) , 3.26 (m, 4H) , 3.17 -2.93 (m, 3H) , 2.79 (m, 4H) , 2.49 (s, 3H) , 2.26 -2.05 (m, 3H) , 2.03 -1.93 (m, 1H) , 1.85 -1.71 (m, 2H) , 1.57 (s, 6H) , 1.56 -1.41 (m, 3H) , 1.41 -1.26 (m, 4H) , 1.11 (s, 9H) .
Compound 64
Step 1. tert-butyl methyl (6-oxohexyl) carbamate
To the solution of anhydrous DCM (8 mL) and DMSO (405.31 μL, 5.19 mmol) was added (COCl) 2 (189.20 μL, 2.16 mmol) dropwise at -78 ℃. The mixture was stirred at -78 ℃ for 30 min followed by the solution of tert-butyl (6-hydroxyhexyl) (methyl) carbamate (100 mg, 432.28 μmol) in DCM (2 mL) dropwise. The solution was stirred for another 30 min and then added Et3N (601.68 μL, 4.32 mmol) . The suspension was warmed until 25 ℃ and stirred for 2 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was washed
with aqueous HCl (5 %, 10 mL) , aqueous saturated NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo to afford the title compound.
1H NMR (400 MHz, DMSO-d6) δ = 9.66 (s, 1H) , 3.32 (s, 3H) , 2.74 (br, 4H) , 1.38 (br, 15H) .
Step 2. tert-butyl (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) (methyl) carbamate
To the solution of tert-butyl methyl (6-oxohexyl) carbamate (70.69 mg, 308.28 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (43.56 mg, 205.52 μmol) was added and the resulting mixture was stirred for additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 700.5 [M+H] +.
Step 3. 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (6- (methylamino) hexyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) (methyl) carbamate (71 mg, 101.45 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound. LCMS: MS (ESI) m/z 600.4 [M+H] +.
Step 4. (2S, 4R) -N- ( (S) -3- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 64)
To the solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (6- (methylamino) hexyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (60 mg, 94.30 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-
carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (68.99 mg, 120.05 μmol) , DIPEA (52.27 uL, 300.12 μmol) , HOBt (16.22 mg, 120.05 μmol) , EDCI (23.01 mg, 120.05 μmol) in order. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Welch Xtimate C18 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 19 %-49 %, 7min) to afford Compound 64.
LCMS: MS (ESI) m/z 578.5 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.16 (br, 1 H) , 8.98 (s, 1H) , 8.82 (s, 1H) , 8.51 (t, J = 8.4 Hz, 1H) , 8.04 (br, 1H) , 7.75 (d, J = 8.0 Hz, 1H) , 7.64 -7.50 (m, 3H) , 7.46 -7.35 (m, 4H) , 7.30 -7.23 (m, 1H) , 6.91 (d, J = 8.8 Hz, 2H) , 5.71 -5.61 (m, 1H) , 5.34 (br, 1H) , 5.25 -5.16 (m, 1H) , 4.99 (dd, J = 1.2, 10.4 Hz, 1H) , 4.84 -4.80 (m, 1H) , 4.69 -4.67 (m, 2H) , 4.57 (d, J = 9.2 Hz, 1H) , 4.44 (t, J = 8.0 Hz, 1H) , 4.28 (br, 1H) , 3.66 -3.49 (m, 3H) , 3.29 -3.22 (m, 1H) , 3.21 -3.12 (m, 1H) , 3.06 (m, 4H) , 2.97 -2.78 (m, 4H) , 2.75 (s, 1H) , 2.45 (m, 7H) , 2.30 -2.22 (m, 2H) , 2.09 -2.00 (m, 1H) , 1.80 -1.74 (m, 1H) , 1.46 (m, 6H) , 1.43 -1.31 (m, 6H) , 1.28 -1.08 (m, 6H) , 0.97 (s, 9H) .
Compound 65
Step 1. tert-butyl (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) (methyl) carbamate
To the solution of tert-butyl methyl (5-oxopentyl) carbamate (26.55 mg, 123.31 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (43.56 mg, 205.52 μmol) was added and the resulting mixture was stirred for additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 686.4 [M+H] +.
Step 2. 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (5- (methylamino) pentyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) (methyl) carbamate (50.4 mg, 73.48 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 586.4 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 65)
To the solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (5- (methylamino) pentyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (45 mg, 73.48 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (49.87 mg, 86.79 μmol) , DIPEA (37.79 uL, 216.97 μmol) , HOBt (11.73 mg, 86.79 μmol) , EDCI (16.64 mg, 86.79 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was
directly purified by reversed-phase HPLC (column: Welch Xtimate C18 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 19 %-49 %, 7min) to afford Compound 65.
LCMS: MS (ESI) m/z 572.1 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (br, 1H) , 8.97 (br, 1H) , 8.82 (s, 1H) , 8.51 (dd, J = 8.4, 11.2 Hz, 1H) , 8.03 (t, J = 6.4 Hz, 1H) , 7.76 -7.74 (m, 1H) , 7.64 -7.52 (m, 3H) , 7.46 -7.34 (m, 4H) , 7.29 -7.22 (m, 1H) , 6.90 (d, J = 8.0 Hz, 2H) , 5.71 -5.61 (m, 1H) , 5.40 (br, 1H) , 5.27 -5.15 (m, 1H) , 4.99 (d, J = 10.4 Hz, 1H) , 4.87 -4.78 (m, 1H) , 4.69 -4.67 (m, 2H) , 4.58 (d, J = 9.2 Hz, 1H) , 4.45 (t, J = 7.2 Hz, 1H) , 4.28 (br, 1H) , 3.61 -3.53 (m, 3H) , 3.29 -3.24 (m, 1H) , 3.19 -3.14 (m, 1H) , 3.06 (m, 4H) , 2.93 -2.80 (m, 4H) , 2.75 (s, 1H) , 2.45 (m, 7H) , 2.30 -2.20 (m, 2H) , 2.09 -1.99 (m, 1H) , 1.80 -1.74 (m, 1H) , 1.46 (s, 6H) , 1.43 -1.30 (m, 6H) , 1.25 -1.08 (m, 4H) , 0.97 (s, 9H) .
Compound 66
Step 1. tert-butyl (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butyl) (methyl) carbamate
To the solution of tert-butyl methyl (4-oxobutyl) carbamate (24.82 mg, 123.31 μmol) in DCM (5 mL) was added 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one (50 mg, 102.76 μmol) and Et3N (42.91 μL, 308.28 μmol) in order. The mixture was stirred at 25 ℃ for 1 h. Then NaBH (OAc) 3 (43.56 mg, 205.52 μmol) was added and the resulting mixture was stirred for additional 1 h. Upon completion, the reaction was quenched with H2O (10 mL) . The organic layer was diluted with DCM (10 mL) , washed with saturated aqueous NaHCO3 (3 × 10 mL) , brine (10 mL) , dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column, eluting with DCM/MeOH (9/1) to afford the title compound.
LCMS: MS (ESI) m/z 672.4 [M+H] +.
Step 2. 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (4- (methylamino) butyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride
To the solution of tert-butyl (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butyl) (methyl) carbamate (52.7 mg, 78.44 μmol) in 1, 4-dioxane (5 mL) was added HCl/1, 4-dioxane (4 M, 15 mL) . The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the mixture was concentrated in vacuo and the residue was dried under vacuum to afford the title compound.
LCMS: MS (ESI) m/z 572.2 [M+H] +.
Step 3. (2S, 4R) -N- ( (S) -3- ( (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide (Compound 66)
To the solution of 2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -6- ( (4- (4- (4- (methylamino) butyl) piperazin-1-yl) phenyl) amino) -1, 2-dihydro-3H-pyrazolo [3, 4-d] pyrimidin-3-one hydrochloride (47 mg, 78.44 μmol) in DMF (2 mL) was added (S) -3- ( (2S, 4R) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamido) -3- (4- (4-methylthiazol-5-yl) phenyl) propanoic acid (53.07 mg, 92.35 μmol) , DIPEA (40.22 uL, 230.88 μmol) , HOBt (12.48 mg, 92.35 μmol) , EDCI (17.70 mg, 92.35 μmol) in order. The mixture was stirred at 25 ℃ for 2 hr. Upon completion, the reaction mixture was directly purified by reversed-phase HPLC (column: Welch Xtimate C18 150*30mm*5um; mobile phase: [water (FA) -ACN] ; B %: 19 %-49 %, 7min) to afford Compound 66.
LCMS: MS (ESI) m/z 565.2 [M/2 + H] +.
1H NMR (400 MHz, DMSO-d6) δ 10.14 (br, 1H ) , 8.96 (br, 1H) , 8.82 (s, 1H) , 8.50 (dd, J = 4.0, 8.0 Hz, 1H) , 8.05 -8.03 (m, 1H) , 7.75 (d, J = 8.0 Hz, 1H) , 7.64 -7.50 (m, 3H) , 7.41 -7.40 (m, 4H) , 7.31 -7.21 (m, 1H) , 6.89 (d, J = 8.4 Hz, 2H) , 5.71 -5.61 (m, 1H) , 5.33 (br, 1H) , 5.26 -5.16 (m, 1H) , 4.99 (d, J = 10.0 Hz, 1H) , 4.87 -4.78 (m, 1H) , 4.69 -4.68 (m, 2H) , 4.58 (d, J = 9.2 Hz, 1H) , 4.49 -4.39 (m, 1H) , 4.28 (br, 1H) , 3.64 -3.53 (m, 3H) , 3.29 -3.27 (m, 1H) , 3.22 -3.16 (m, 1H) , 3.05 (m, 4H) , 2.97 -2.80 (m, 4H) , 2.76 (s, 1H) , 2.45 -2.44 (m, 7H) , 2.30 -2.22 (m, 2H) , 2.09 -1.99 (m, 1H) , 1.81 -1.74 (m, 1H) , 1.46 (s, 6H) , 1.42 -1.27 (m, 6H) , 1.25 -1.17 (m, 2H) , 0.97 (s, 9H) .
II. Biological Evaluation
ZNL-02-096 (structure below) as disclosed in Li et al., “Development and Characterization of a Wee1 Kinase Degrader” Cell Chemical Biology, 2020, 27 (1) 57-65 was chosen as reference compound.
Example B1: WEE1 Degradation Assay
Cell Culture
Human T lymphoblast MOLT-4 (ATCC, CRL1582) were cultured in RPMI 1640 (Gibco, 11875119) media, supplemented with 10%FBS (Gibco, 10099-141) . All cell lines were maintained at 37 ℃ in an atmosphere of 5%CO2.
Protein Extraction and Western Blotting and DC50 determination
Whole cell lysates were prepared using cell Lysis Buffer (CST, 9803) supplemented with Protease inhibitor, phosphatase inhibitor and PMSF solution. The concentration of protein was quantified using a Pierce Bicinchoninic Acid (BCA) Protein Concentration Kit (Solarbio, PC0020) . Equal amounts of protein were loaded for PAGE and transfer samples onto a PVDF membrane using Bio-Rad's Trans blot. The blots were incubated with a primary antibody (anti-WEE1, and anti-α-Tubulin) , followed by HRP conjugated to an appropriate secondary antibody. Compounds 29, 31, 32 40, 42, 49, 51, and 56 had a DC50 of > 1.0 μM to ≤ 10 μM and a Dmax %of < 50%.
Cell Viability Assay and Growth Inhibition Determination
Cell viability was measured using a CellTiter-Glo Assay kit (Promega, G7573) according to the manufacturer’s protocol. MOLT-4 were seeded into 384-well plates and treated with various increasing doses of compound for 72 hours. Take cells plate from incubator and equilibrate the plate and Read on Envision 2104 Multi-plate reader using luminescence detector.
Representative degradation activity and cell viability from exemplary compounds is presented in Table 4.
Note: DC50 data are designated within the following ranges:
A: ≤ 0.010 μM C: > 0.10 μM to ≤ 1.0 μM
B: > 0.010 μM to ≤ 0.10 μM D: > 1.0 μM to ≤ 10 μM
Note: Dmax data are designated within the following ranges:
A: ≥ 80% B: ≥ 50%; < 80% C: < 50%
Note: GI50 data are designated within the following ranges:
A: ≤ 0.10 μM B: > 0.10 μM to ≤ 1.0 μM C: > 1.0 μM to ≤ 10 μM
Note: The “-” indicates not determined.
Table 4
Example B2: Compound Solubility
Solubility Determination
30 μL of stock solution (10 mM) of each sample were placed in order into their proper 96-well rack. 970 μL of FaSSGF, FaSSIF or FeSSIF were added into each vial of the cap-less Solubility Sample plate. The assay was performed in duplicate. One magnetic stirring rod was added to each vial and sealed using a molded PTFE/Silicone plug. The Solubility Sample plate was then transferred to the Eppendorf Thermomixer Comfort plate shaker and shaken at 37℃ at 1100 RPM for 2 hours. After completion of the 2 hours, plugs were removed and the magnetic stirring rods were removed using a magnet, the samples from the Solubility Sample plate were transferred into the filter plate. Using a Vacuum Manifold, all the samples were filtered. Aliquot of 5 μL were taken from the filtrate followed by addition of 5 μL DMSO and 490 μL of acetonitrile containing internal standards (IS, 100 nM Alprazolam, 500 nM Labetalol and 2 μM Ketoprofen) with 0.1%formic acid. The dilution factor was changed according to the solubility values and the LC-MS signal response. The plate was placed into the well plate autosampler. The samples were evaluated by LC-MS/MS analysis. LC-MS/MS data was analyzed with SCIEX Analyst software. All calculations were carried out using Microsoft Excel.
Compounds of Formula (I) as disclosed herein show significantly improved FaSSGF, FassiF, FessiF solubility when compared with reference ZNL-02-096.
Representative solubility of exemplary compounds is presented in Table 5.
Table 5
Example B3: Metabolic Stability Determination
Liver Microsomes Stability
Two separate experiments were performed as follows: a) with Cofactors (NADPH and UDPGA) and b) without Cofactors (NADPH and UDPGA) . The final concentration of human and mouse microsomes was 0.5 mg/mL. The reaction was started with the addition of 4 μL of 100 μM test compound solution or control compound solution at the final concentration of 1 μM and carried out at 37 ℃. Aliquots of 50 μL were taken from the reaction solution at 0, 15, 30, 45 and 60 min. The reaction was stopped by the addition of 4 volumes of cold acetonitrile with IS (3%formic acid, 100 nM alprazolam, 200 nM labetalol, 200 nM caffeine and 2 μM ketoprofen) . Samples were centrifuged at 3, 220 g for 40 minutes. Aliquot of 100 μL of the supernatant was mixed with 100 μL of ultra-pure H2O and then used for LC-MS/MS analysis. LC-MS/MS data was analyzed with SCIEX Analyst software. Peak areas were determined from extracted ion chromatograms. All calculations were carried out using Microsoft Excel. The slope value, k, was determined by linear regression of the natural logarithm of the remaining percentage of the parent drug vs. incubation time curve.
Representative liver microsome stability for exemplary compounds is presented in Table 6.
Table 6
Hepatocyte Metabolic Stability
10 mM stock solutions of test compound (s) and positive control were prepared in DMSO. Incubation medium and hepatocyte thawing medium were placed in a 37℃ water bath and allowed to warm for at least 15 minutes prior to use. A vial of cryopreserved hepatocytes was removed from storage. After thawing, the vial was sprayed with 70%ethanol and transferred to a biosafety cabinet. Hepatocytes were transferred into a 50 mL conical tube containing thawing medium and placed into a centrifuge and spun at 100 g for 10 minutes. Upon completion of spin, the thawing medium was aspirated, and the hepatocytes were resuspended. Cells were counted and viable cell density was determined using AOPI staining solution. LC-MS/MS data was analyzed with SCIEX Analyst software. Peak areas were determined from extracted ion chromatograms. All calculations were carried out using Microsoft Excel. The in vitro half-life (t1/2) of parent compound was determined by regression analysis of the percent parent disappearance vs. time curve.
Representative hepatocyte stability for exemplary compounds is presented in Table 7.
Table 7. Compound stability with human and mouse hepatocytes
Example B4: In Vivo Pharmacokinetic Determinations
Compound Pharmacokinetics in Mice
The study was conducted in male CD-1 mice with an age of 6-8 weeks. Animals received a single intravenous injection at the dose of 2.0 mg/kg. A dose of 2.0 mg/kg was prepared as 0.4 mg/ml solutions in 5%DMSO/45%PEG400/50%Water tested in mice.
Blood samples were taken from three different mice at each of the following time points: 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, 24 h. Blood samples (30 μL) were taken from mice and transferred to EDTA tubes. Blood was centrifuged at 4000 g for 5 minutes in a 4℃ centrifuge to obtain plasma. All plasma samples were stored in a freezer at -75±15℃ prior to analysis.
The desired serial concentrations of working solutions were achieved by diluting stock solution of analyte with 50%acetonitrile in water solution. Four quality control samples at 1 ng/mL, 2 ng/mL, 50 ng/mL and 800 ng/mL for plasma were prepared independently of those used for the calibration curves. These QC samples were prepared on the day of analysis in the same way as calibration standards. 33 μL standards, 33 μL QC samples and 33 μL unknown samples (30 μL plasma with 3 μL blank solution) were added to 200 μL of acetonitrile containing IS mixture for precipitating protein respectively. Then the samples were vortexed for 30 s. The supernatant was diluted with water at a rate of 1: 2, 10 μL of the diluted supernatant was injected into the LC/MS/MS system for quantitative analysis.
WinNonlin (PhoenixTM, version 8.3) software was used for pharmacokinetic calculations and the following pharmacokinetic parameters were calculated from the plasma concentration versus time data: T1/2, AUC∞, Cl, and VDss.
Compounds of Formula (I) as disclosed herein show significantly improved half-life and AUC when compared with reference ZNL-02-096.
Representative pharmacokinetic parameters for exemplary compounds are presented in Table 8.
Table 8. Compound pharmacokinetic properties in vivo
Example B5: Evaluation of Anti-proliferation Effects of Compounds in TOV21G Cell Line by Cell Viability Assay
Cell Culture
TOV-21G cell line was maintained in culture conditions at 37℃ in an atmosphere with 5%CO2
in air. The tumor cells were routinely subcultured. The cells growing in an exponential growth phase were harvested and counted for plating.
Cell Plating
(1) . Count cells by haemocytometer with Trypan blue staining.
(2) . Adjust the cell concentration to proper cell density (4000/per 96-well) .
(3) . Plate 90 μL of cell suspension into the assay plates according to the plate map. Add 90 μL of assay medium into the Blank wells.
(4) . Incubate the plates at 37℃, 5%CO2, 95%air and 100%relative humidity overnight.
Compound Stock Plate Preparation
Preparation of compound stock plate (1000X stock plates) : Serially dilute the stock solution from highest concentration (10 μM) down to lowest (1.52 μM) in DMSO.
Compound Plate (10X) Preparation and Compound Treatment
(1) . 10X concentrate compound plate preparation: Add 198 μL of assay medium into each well of the V-bottom plate; then transfer 2 μL of the stock compound solution of each concentration from the stock plate. Add 2 μL of DMSO into the Blank and Control wells. Pipette up and down to mix well. This V-plate is designated as the 10X concentrate compound plate.
(2) . For day 0 plate, add 10 μL of the DMSO-medium into the Control wells. The final DMSO concentration was 0.1%. Then directly proceed to cell viability assay.
(3) . Compound Treatment: add 10 μL compound-medium of each well from the 10X concentrate compound plate into the cells in 96-well assay plate according to the plate map. For Vehicle and Blank group, Add 10 μL of the DMSO-medium into the wells. The final DMSO concentration was 0.1%.
(4) . Return the assay plate into incubator and incubate for 3 days.
CellTiter-Glo Luminescent Cell Viability Assay
The procedures were performed according to the Promega CellTiter-Glo Luminescent Cell Viability Assay Kit manual (Promega-G7573) .
(1) . Thaw the CellTiter-Glo buffer and equilibrate to room temperature prior to use.
(2) . Equilibrate the lyophilized CellTiter-Glo Substrate to room temperature prior to use.
(3) . Transfer the entire liquid volume of CellTiter-Glo Buffer into the amber bottle containing CellTiter-Glo Substrate to reconstitute the lyophilized enzyme/substrate mixture. This forms the CellTiter-Glo Reagent.
(4) . Mix by gently vortexing to obtain a homogeneous solution.
(5) . Equilibrate the plate and its contents to room temperature for approximately 30 minutes.
(6) . Add 50 μL (equal to the half volume of culture medium present in each well) CellTiter-
Glo Reagent in each well. Cover plates with aluminum foil to protect from light.
(7) . Mix contents for 2 minutes on an orbital shaker to induce cell lysis.
(8) . Allow the plate to incubate at room temperature for 10 minutes to stabilize luminescent signal.
(9) . Record luminescence on the 2104 EnVision plate reader.
Data Analysis
Inhibition rate (IR) of the tested compounds was determined by the following formula: IR (%) = (1– (RLU compound –RLU blank) / (RLU control –RLU blank) ) *100%. The inhibitions of different dose of compound were calculated in Excel file, and then were used to plot inhibition curves and evaluate related parameters, such as Bottom (%) , Top (%) and Relative IC50. The data were interpreted by GraphPad Prism.
Representative cell viability from exemplary compounds is presented in Table 9:
Table 9: Anti-proliferation Effects of Compounds in TOV21G Cell Line
III. Preparation of Pharmaceutical Dosage Forms
Example C1: Oral capsule
The active ingredient is a compound of Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt or solvate thereof. A capsule for oral administration is prepared by mixing 1-1000 mg of active ingredient with starch or other suitable powder blend. The mixture is incorporated into an oral dosage unit such as a hard gelatin capsule, which is suitable for oral administration.
Example C2: Solution for injection
The active ingredient is a compound of Table (I) Table (2) , Table (3) , Formula (I) , Formula (I-a) , or Formula (I-b) , or a pharmaceutically acceptable salt thereof, and is formulated as a solution in sesame oil at a concentration of 50 mg-eq/mL.
The examples and embodiments described herein are for illustrative purposes only and various modifications or changes suggested to persons skilled in the art are to be included within the spirit and purview of this application and scope of the appended claims.
Claims (48)
- A compound, or a pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, having the structure of Formula (I) :
wherein,R1, R2, R3, and R4 are each independently selected from hydrogen, optionally substituted straight or branched C1-6 alkyl, optionally substituted C1-3 alkoxy, halogen, amino, hydroxyl, and cyano; orR1 and R2, R2 and R3, or R3 and R4 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted heterocycloalkyl ring;L is -Ak1-Z1-Ak2-Z2-Ak3-*;Ak1 is selected from - (CR5R6) k-;Ak2 is selected from - (CR5R6) m-;Ak3 is selected from - (CR5R6) n-;each of k, m, and n is selected from 0 to 10R5 and R6 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C1-3 alkyl, and C3 cycloalkyl;Z1 and Z2 are each independently selected from bond, -O-, -NR7-, -S-, heterocyclene, and cycloalkylene;R7 is hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced;wherein at least one of Z1 and Z2 is not bond; orL is a polyethylene glycol containing moiety; andwherein the point of bonding to the VHL ligand is indicated by the asterisk; andU is a VHL ligand. - The compound of claim 1, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein R1 is optionally substituted branched C1-6 alkyl, wherein the alkyl is optionally substituted with hydroxy; andR2, R3, and R4 are hydrogen.
- The compound of claim 1 or claim 2, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein R1 is
- The compound of claim 1, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein R1 and R2 together with the intervening atoms to which they are attached form an optionally substituted 5-6 membered cycloalkyl or optionally substituted 5-6 membered heterocyclyl ring, wherein the cycloalkyl and heterocyclyl are optionally substituted with one or more substituents selected from C1-6 alkyl and hydroxyl.
- The compound of claim 1 or claim 4, wherein R1 and R2 together with the intervening atoms to which they are attached form
- The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) n-O– (CR5R6) m-*, wherein n is selected from 1 to 10, and m is selected from 1 to 10;and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-6, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is – (CH2) n-O– (CH2) m-*, wherein n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is - (CR5R6) n-NR7– (CR5R6) m-*, wherein n is selected from 1 to 10, m is selected from 0 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-5, or 8, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is – (CH2) n-NR7– (CH2) m-*, wherein n is selected from 1 to 10, m is selected from 0 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is – (CR5R6) k-O–– (CR5R6) n-O– (CR5R6) m-*, wherein k is selected from 1 to 10, n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-5, or 10, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein the -Ak1-Z1-Ak2-Z2-Ak3-*is – (CH2) k-O–– (CH2) n-O– (CH2) m-*, wherein k is selected from 1 to 10, n is selected from 1 to 10, and m is selected from 1 to 10; and the point of bonding to the VHL ligand indicated by the asterisk.
- The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein X1 and X2 at each instance are independently CH or N; andwherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 12, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 12, or 13, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 12, or 14, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 12-14, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 12, or 16, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 18, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 19, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 20, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 18-20, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18-20, or 22 or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 18-23, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 24, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18-20, 22, or 24, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 26, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18-22, 24, 26, or 27, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 28, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18-20, 22-24, or 26-28, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 18, or 30, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or 12-31, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
- The compound of any one of claims 1-5, 12-14, 16, 17, or 32, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
- The compound of any one of claims 1-5, or 12, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, 12, or 34, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein the point of bonding to the VHL ligand is indicated by the asterisk. - The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein -Ak1-Z1-Ak2-Z2-Ak3-*is selected from a molecular fragment illustrated below:
wherein each R7 is independently hydrogen, optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced, with the point of bonding to the VHL ligand indicated by the asterisk. - The compound of claim 36, whereinis
- The compound of claim 36 or 37, wherein R7 is hydrogen or methyl.
- The compound of any one of claims 1-5, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein L is a polyethylene glycol containing moiety selected from:– (O-CH2-CH2) n-O-and – (O-CH2-CH2) n-N (R23) -*, wherein n is selected from 1 to 20, and R23 is optionally substituted alkyl, or optionally substituted alkyl wherein 1 or more -CH2-units are replaced with O provided that two adjacent -CH2-units are not both replaced ; and the point of bonding to the recruitment motif indicated by the asterisk.
- The compound of any one of claims 1-39 or pharmaceutically salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein U is selected from:
- The compound of any one of claims 1-40 or pharmaceutically salt, solvate, prodrug, stereoisomer, or tautomer thereof, wherein U is selected from
- The compound of claim 1, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, selected from:(2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (2- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] ethoxy) butanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1-yl] butoxy) propanamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (4- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) propoxy) butanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- [ (6- [4- [4- ( [1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl] amino) phenyl] piperazin-1- yl] hexyl) oxy] acetamido] -3, 3-dimethylbutanoyl] -N- [ [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl] pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (2- ( (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) ethoxy) methoxy) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (5- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} pentyl) oxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) methoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (3- (5- (4- (4- (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-ylamino) phenyl) piperazin-1-yl) pentyloxy) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) butanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} butoxy) methoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [5- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) pentanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} propoxy) ethoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [6- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) hexanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [3- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) propoxy] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {3- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} ethoxy) ethoxy] propanamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [ (4'- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 1'-bi (cyclohexane) ] -4-yl) formamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- { [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin- 1-yl) cyclohexyl] formamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidine] -1'-carboxamide;N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-4-carboxamide;N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidine-1-carboxamide;4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) piperazine-1-carboxamide;(2S, 4R) -1- ( (S) -2- (2- (4'- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 1'-bi (cyclohexan) ] -4-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclohexyl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 4'-bipiperidin] -1'-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [1- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-4-yl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- {2- [4- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) piperidin-1-yl] acetamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (2- (4- (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclohexyl) piperazin-1-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) methyl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- [ (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} cyclohexyl) methyl] piperidine-4-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- ( {4- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) piperidin-1-yl] cyclohexyl} formamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;N- [ (2S) -1- [ (2S, 4R) -4-hydroxy-2- ( { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} carbamoyl) pyrrolidin-1-yl] -3, 3-dimethyl-1-oxobutan-2-yl] -1- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] piperidine-4-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) propanamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (9- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -3-azaspiro [5.5] undecan-3-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (7- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -2-azaspiro [3.5] nona-2-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (2- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} -7-azaspiro [3.5] nonan-7-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [4- ( {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} methyl) cyclohexyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (2- (1- (4- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) cyclohexyl) azetidin-3-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;1- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) cyclobutyl) -N- ( (S) -1- ( (2S, 4R) -4-hydroxy-2- ( (4- (4-methylthiazol-5-yl) benzyl) carbamoyl) pyrrolidin-1-yl) -3, 3-dimethyl-1-oxobutan-2-yl) piperidine-4-carboxamide;(2S, 4R) -1- ( (S) -2- (4- (3- ( (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) methyl) azetidin-1-yl) cyclohexane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- { [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin-1-yl) cyclobutyl] formamido} -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [3- (4- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} piperidin- 1-yl) azetidine-1-carbonylamino] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- [2- (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} - [1, 3'-biazetidin] -1'-yl) acetamido] -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -4-hydroxy-1- [ (2S) -2- (2- {3- [ (3- {4- [4- ( {1- [6- (2-hydroxypropan-2-yl) pyridin-2-yl] -3-oxo-2- (prop-2-en-1-yl) pyrazolo [3, 4-d] pyrimidin-6-yl} amino) phenyl] piperazin-1-yl} azetidin-1-yl) methyl] azetidin-1-yl} acetamido) -3, 3-dimethylbutanoyl] -N- { [4- (4-methyl-1, 3-thiazol-5-yl) phenyl] methyl} pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (3- (3- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) - [1, 3'-biazetidin] -1'-yl) propanamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (2- (9- (4- (4- ( (2-allyl-1- ( (R) -7-ethyl-7-hydroxy-6, 7-dihydro-5H-cyclopenta [b] pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -3-azaspiro [5.5] undecan-3-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -1- ( (S) -2- (2- (2- (4- (4- ( (2-allyl-1- ( (R) -7-ethyl-7-hydroxy-6, 7-dihydro-5H-cyclopenta [b] pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -7-azaspiro [3.5] nonan-7-yl) acetamido) -3, 3-dimethylbutanoyl) -4-hydroxy-N- (4- (4-methylthiazol-5-yl) benzyl) pyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- (8- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -2-azaspiro [4.5] decan-2-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (1S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -8-azaspiro [4.5] decan-8-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- (9- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -3-azaspiro [5.5] undecan-3-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -6-azaspiro [3.4] octan-6-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- (2- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) -7-azaspiro [3.5] nonan-7-yl) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- ( (6- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) hexyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide;(2S, 4R) -N- ( (S) -3- ( (5- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) pentyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide; and(2S, 4R) -N- ( (S) -3- ( (4- (4- (4- ( (2-allyl-1- (6- (2-hydroxypropan-2-yl) pyridin-2-yl) -3-oxo-2, 3-dihydro-1H-pyrazolo [3, 4-d] pyrimidin-6-yl) amino) phenyl) piperazin-1-yl) butyl) (methyl) amino) -1- (4- (4-methylthiazol-5-yl) phenyl) -3-oxopropyl) -1- ( (S) -2- (1-fluorocyclopropane-1-carboxamido) -3, 3-dimethylbutanoyl) -4-hydroxypyrrolidine-2-carboxamide.
- A pharmaceutical composition comprising a compound of Formula (I) , or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, as described in any one of claims 1-42.
- A compound of any one of claims 1-42, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, for use in a method of treatment of a human or animal body.
- A compound of any one of claims 1-42, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, for use in a method of treatment of cancer or neoplastic disease.
- Use of a compound of any one of claims 1-42, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, in the manufacture of a medicament for the treatment of cancer or neoplastic disease.
- A method of treating cancer in a patient in need thereof comprising administering to the patient a compound of Formula (I) as described in any one of claims 1-42, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof.
- A method of treating cancer in a patient in need thereof comprising administering to the patient a pharmaceutical composition comprising a compound of Formula (I) as described in any one of claims 1-42, or pharmaceutically acceptable salt, solvate, prodrug, stereoisomer, or tautomer thereof, and a pharmaceutically acceptable excipient.
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