CN104447681B

CN104447681B - Compound for treating cancer and its preparation method and application

Info

Publication number: CN104447681B
Application number: CN201310425570.0A
Authority: CN
Inventors: 裴剑锋; 来鲁华; 陈芳进
Original assignee: Peking University
Current assignee: Peking University
Priority date: 2013-09-17
Filing date: 2013-09-17
Publication date: 2018-05-01
Anticipated expiration: 2033-09-17
Also published as: CN104447681A; WO2015039592A1

Abstract

The present invention relates to formula

Description

Compound for treating cancer and its preparation method and application

Technical field

The present invention relates to the compound of Formulas I, its various derivative, includes its composition, and they are used to prepare and control Treat or prevent to benefit from the purposes of the medicine for the disease for adjusting 14-3-3 ζ albumen, and in particular, to the compound of Formulas I is used to make The purposes of the standby medicine for treating or preventing cancer.Compound the present invention also relates to Formulas I is used for treatment-related disease such as cancer The purposes of disease.

Background technology

The background technology of the application is described in documents below, and entire content of these documents is incorporated by reference into this Shen Please, the degree not collided with the specific descriptions in the application is reached：

1.Andrews,R.K.,Du,X.,and Berndt,M.C.(2007).The14-3-3zeta-GPIb-IX-V complex as an antiplatelet target.Drug News Perspect20,285-292.

2.Bergamaschi,A.,and Katzenellenbogen,B.S.(2012).Tamoxifen downregulation of miR-451increases14-3-3zeta and promotes breast cancer cell survival and endocrine resistance.Oncogene31,39-47.

3.Bergamaschi,A.,Frasor,J.,Borgen,K.,Stanculescu,A.,Johnson,P., Rowland,K.,Wiley,E.L.,and Katzenellenbogen,B.S.(2013).14-3-3zeta as a predictor of early time to recurrence and distant metastasis in hormone receptor-positive and-negative breast cancers.Breast Cancer Res Treat137,689- 696.

4.Boon,S.S.,and Banks,L.(2013).High-risk human papillomavirus E6oncoproteins interact with14-3-3zeta in a PDZ binding motif-dependent manner.Journal of virology87,1586-1595.

5.Cheah,P.S.,Ramshaw,H.S.,Thomas,P.Q.,Toyo-Oka,K.,Xu,X.,Martin,S., Coyle,P.,Guthridge,M.A.,Stomski,F.,van den Buuse,M.,Wynshaw-Boris,A.,Lopez, A.F.,and Schwarz,Q.P.(2012).Neurodevelopmental and neuropsychiatric behaviour defects arise from14-3-3zeta deficiency.Mol Psychiatry17,451-466.

6.Choi,J.E.,Hur,W.,Jung,C.K.,Piao,L.S.,Lyoo,K.,Hong,S.W.,Kim,S.W., Yoon,H.Y.,and Yoon,S.K.(2011).Silencing of14-3-3zeta over-expression in hepatocellular carcinoma inhibits tumor growth and enhances chemosensitivity to cis-diammined dichloridoplatium.Cancer Lett303,99-107.

7.Dai,K.,Bodnar,R.,Berndt,M.C.,and Du,X.(2005).A critical role for14- 3-3zeta protein in regulating the VWF binding function of platelet glycoprotein Ib-IX and its therapeutic implications.Blood106,1975-1981.

8.Deakin,N.O.,Bass,M.D.,Warwood,S.,Schoelermann,J.,Mostafavi-Pour,Z., Knight,D.,Ballestrem,C.,and Humphries,M.J.(2009).An integrin-alpha4-14-3- 3zeta-paxillin ternary complex mediates localised Cdc42activity and accelerates cell migration.Journal of cell science122,1654-1664.

9.Du,J.,Chen,L.,Luo,X.,Shen,Y.,Dou,Z.,Shen,J.,Cheng,L.,Chen,Y.,Li,C., Wang,H.,and Yao,X.(2012).14-3-3zeta cooperates with phosphorylated Plk1and is required for correct cytokinesis.Front Biosci(Schol Ed)4,639-650.

10.Fan,T.,Li,R.,Todd,N.W.,Qiu,Q.,Fang,H.B.,Wang,H.,Shen,J.,Zhao,R.Y., Caraway,N.P.,Katz,R.L.,Stass,S.A.,and Jiang,F.(2007).Up-regulation of14-3- 3zeta in lung cancer and its implication as prognostic and therapeutic target.Cancer Res67,7901-7906.

11.Ge,F.,Li,W.L.,Bi,L.J.,Tao,S.C.,Zhang,Z.P.,and Zhang,X.E.(2010) .Identification of novel14-3-3zeta interacting proteins by quantitative immunoprecipitation combined with knockdown(QUICK).Journal of proteome research9,5848-5858.

12.Kambach,D.M.,Sodi,V.L.,Lelkes,P.I.,Azizkhan-Clifford,J.,and Reginato,M.J.(2013).ErbB2,FoxM1and14-3-3zeta prime breast cancer cells for invasion in response to ionizing radiation.Oncogene.

13.Li,Z.,Zhao,J.,Du,Y.,Park,H.R.,Sun,S.Y.,Bernal-Mizrachi,L.,Aitken, A.,Khuri,F.R.,and Fu,H.(2008).Down-regulation of14-3-3zeta suppresses anchorage-independent growth of lung cancer cells through anoikis activation.Proceedings of the National Academy of Sciences of the United States of America105,162-167.

14.Matta,A.,DeSouza,L.V.,Ralhan,R.,and Siu,K.W.(2010).Small interfering RNA targeting14-3-3zeta increases efficacy of chemotherapeutic agents in head and neck cancer cells.Mol Cancer Ther9,2676-2688.

15.Matta,A.,Bahadur,S.,Duggal,R.,Gupta,S.D.,and Ralhan,R.(2007).Over- expression of14-3-3zeta is an early event in oral cancer.BMC Cancer7,169.

16.Maxwell,S.A.,Cherry,E.M.,and Bayless,K.J.(2011).Akt,14-3-3zeta,and vimentin mediate a drug-resistant invasive phenotype in diffuse large B-cell lymphoma.Leuk Lymphoma52,849-864.

17.Maxwell,S.A.,Li,Z.,Jaye,D.,Ballard,S.,Ferrell,J.,and Fu,H.(2009) .14-3-3zeta mediates resistance of diffuse large B cell lymphoma to an anthracycline-based chemotherapeutic regimen.The Journal of biological chemistry284,22379-22389.

18.Murata,T.,Takayama,K.,Urano,T.,Fujimura,T.,Ashikari,D.,Obinata,D., Horie-Inoue,K.,Takahashi,S.,Ouchi,Y.,Homma,Y.,and Inoue,S.(2012).14-3-3zeta,a novel androgen-responsive gene,is upregulated in prostate cancer and promotes prostate cancer cell proliferation and survival.Clin Cancer Res18,5617-5627.

19.Neal,C.L.,and Yu,D.(2010).14-3-3zeta as a prognostic marker and therapeutic target for cancer.Expert Opin Ther Targets14,1343-1354.

20.Neal,C.L.,Yao,J.,Yang,W.,Zhou,X.,Nguyen,N.T.,Lu,J.,Danes,C.G.,Guo, H.,Lan,K.H.,Ensor,J.,Hittelman,W.,Hung,M.C.,and Yu,D.(2009).14-3-3zeta overexpression defines high risk for breast cancer recurrence and promotes cancer cell survival.Cancer Res69,3425-3432.

Omi,K.,Hachiya,N.S.,Tanaka,M.,Tokunaga,K.,and Kaneko,K.(2008).14-3- 3zeta is indispensable for aggregate formation of polyglutamine-expanded huntingtin protein.Neurosci Lett431,45-50.

21.Tsukamoto,Y.,Nakada,C.,Noguchi,T.,Tanigawa,M.,Nguyen,L.T.,Uchida, T.,Hijiya,N.,Matsuura,K.,Fujioka,T.,Seto,M.,and Moriyama,M.(2010).MicroRNA- 375is downregulated in gastric carcinomas and regulates cell survival by targeting PDK1and14-3-3zeta.Cancer Res70,2339-2349.

22.Wang,Y.(2010).Breast cancer metastasis driven by ErbB2and14-3- 3zeta:A division of labor.Cell Adh Migr4,7-9.

23.Yang,X.,Cao,W.,Zhang,L.,Zhang,W.,Zhang,X.,and Lin,H.(2012) .Targeting14-3-3zeta in cancer therapy.Cancer Gene Ther19,153-159.

24.Yang,X.,Cao,W.,Zhou,J.,Zhang,W.,Zhang,X.,Lin,W.,Fei,Z.,Lin,H.,and Wang,B.(2011).14-3-3zeta positive expression is associated with a poor prognosis in patients with glioblastoma.Neurosurgery68,932-938;discussion938.

25.Yu,D.,dos Santos,C.O.,Zhao,G.,Jiang,J.,Amigo,J.D.,Khandros,E., Dore,L.C.,Yao,Y.,D'Souza,J.,Zhang,Z.,Ghaffari,S.,Choi,J.,Friend,S.,Tong,W., Orange,J.S.,Paw,B.H.,and Weiss,M.J.(2010).miR-451protects against erythroid oxidant stress by repressing14-3-3zeta.Genes Dev24,1620-1633.

26.Zang,D.,Li,X.,and Zhang,L.(2010).14-3-3zeta Overexpression and abnormal beta-catenin expression are associated with poor differentiation and progression in stage I non-small cell lung cancer.Clin Exp Med10,221-228.

27.Zhang,J.,Chen,F.,Li,W.,Xiong,Q.,Yang,M.,Zheng,P.,Li,C.,Pei,J.,and Ge,F.(2012).14-3-3zeta interacts with stat3and regulates its constitutive activation in multiple myeloma cells.PLoS One7,e29554.

28.Zhang,L.,Wang,H.,Liu,D.,Liddington,R.,and Fu,H.(1997).Raf-1kinase and exoenzyme S interact with14-3-3zeta through a common site involving lysine49.The Journal of biological chemistry272,13717-13724.

In this application, it is without being bound by theory, it is believed that benefiting from the disease of adjusting 14-3-3 ζ albumen includes various cancers. And many cancer displays are tumour.Therefore, in this application, when referring to tumour, it should be appreciated that refer to show as tumor forms Cancer.

Research shows：The 14-3-3 ζ albumen of general expression participates in the adjusting of the numerous important paths of cancer, and 14-3-3 ζ are being adjusted Play a major role in a variety of paths related with cancer occurrence and development, the height of 14-3-3 ζ is found that in kinds of tumors Expression.

14-3-3 ζ promote cancer cell multiplication：Suppress or silence 14-3-3 ζ are overexpressed and can inhibit tumour growth, its mechanism can Can be realized by activating JNK and P38/MAPK.

14-3-3 ζ promote cancer cell migration：There are some researches prove 14-3-3 ζ have an impact lung cancer metastasis, the height of 14-3-3 ζ Expression is related with the peritonaeum metastasis degree of cancer, with the migrating of oophoroma, attack, implantation capability it is related.Illustrate at present thin The core mechanism of born of the same parents' migration is Rac1（The relevant C3 creotoxins substrates 1 of Ras）Adjusting actin remodeling, but it is specific when Air-conditioning section is still unknown.

14-3-3 ζ and cancer cell-apoptosis：Research finds miR-375 by lowering high tables of the 14-3-3 ζ in stomach cancer cell Reach, can obviously reduce the survival ability of cell.MiR-375 is active by the 3 '-UTR for suppressing 14-3-3 ζ transcript mRNAs, and increase is withered Die protein activation, the activation of disturbed one 4-3-3 ζ then apoptotic proteins greatly increases, and 14-3-3 ζ take part in apoptotic proteins suppression, be to wither Die albumen and suppress albumen.Exactly 14-3-3 ζ are expressed in the height of tumour cell, tumour cell is generated anoikis suppression, and And decline the apoptotic proteins such as Bad, Bax, make tumour cell from apoptosis.

14-3-3 ζ albumen participates in the adjusting of various kinds of cell signal path, influences Apoptosis and cell cycle regulation and thin Intercellular adhesion.In recent years the research to HaCaT cell lines is found, 14-3-3 ζ albumen can inhibit the activation of p38 and JNK, is blocked The transmission of antiapoptotic signals, suppresses Apoptosis caused by radioactive ray.Found in the research to breast cancer, 14-3-3 ζ eggs P53 loss of stability is set to degrade by the PI3K/Akt MDM2 phosphorylations relied in vain, the cell for suppressing p53 inductions loses matrix The apoptosis of contact, promotes the generation of tumour.14-3-3 ζ albumen may act on cell division cycle protein Cdc25, influence its induction Cell mitogen, cell cycle regulation.14-3-3 ζ can also suppress T-cadherin, E-cadherin and γ-cadherin Deng the expression of cell adhesion molecule, promote the infiltration and transfer of tumour cell.In addition, 14-3-3 ζ albumen may additionally facilitate Cdc42 and The activation of Rac, adjusts the cell signalling of integrin, promotes Cytoskeleton, is conducive to sending out and turning for tumour cell Move.Research to lung cancer finds, lung carcinoma cell can be increased to radioactive ray and chemotherapeutics by suppressing the expression of 14-3-3 ζ albumen Sensitiveness, and the overexpression of 14-3-3 ζ is to prompt one of mouth neoplasm early stage, and be possible to tumour development and developed Play an important role in journey.Therefore, can be using 14-3-3 albumen come Clinics and Practices human diseases.

Existing result of study shows, kinds cancer such as lung (lung) cancer, liver (liver) cancer, cervical carcinoma, mammary gland (breast) cancer, prostate (prostate) cancer, lymph (diffuse large B-cell lymphoma) cancer, marrow (multiple myeloma) cancer, acute leukemia (acute promyolocytic leukemia), stomach (stomach) cancer, Neuroglia (glioma) cancer, meninx cancer (meningioma), the cancer of the esophagus (esophageal), incidence dermoid cancer (head and neck squamous cell carcinoma), oral cavity (oral) cancer, pancreas (pancreatic) cancer, ovary (ovarian) cancer and skin (skin) cancer are all related to 14-3-3 ζ albumen, namely these diseases can be by adjusting 14-3-3 ζ Albumen and prevented and treated.14-3-3 ζ albumen is in the occurrence and development process of tumour and the drug resistance of tumour cell It played an important role, can be obvious by making small molecule (such as polypeptide) and 14-3-3 ζ active-site competitive bindings Suppress the growth of cancer cell, and cause its apoptosis, so as to reach the effect of prevention and treatment cancer.Also need to find at present new Compound, its be used for by adjust 14-3-3 ζ albumen so as to treat or prevent benefit from adjust 14-3-3 ζ albumen disease, Such as above listed cancer.

The content of the invention

The present inventor is tested and is found by external binding tests, cancerous cell line test and mouse species, Formulas I Compound can be combined with 14-3-3 ζ albumen, hence it is evident that suppressed the function of 14-3-3 ζ albumen, suppressed growth of cancer cells, promote cancer thin Born of the same parents' apoptosis, suppresses the growth of tumour, so that the disease for treating and preventing and benefiting from and adjusting 14-3-3 ζ albumen can be produced（Especially Various cancers）Effect.

Therefore, the present invention includes implementation below：

The compound of 1. Formulas I of embodiment,

Wherein：

X and Y is each independently selected from the C optionally with substituent_1-2Alkylidene, oxygen atom, sulphur atom, nitrogen-atoms,

Z₁、Z₂、Z₃、Z₄、Z₅And Z₆It is each independently selected from hydrogen, fluorine, chlorine, bromine, iodine, astatine ,-NO₂、-NR⁶ ₂、-OH、-SR⁷、- SO₄NR⁸ ₂、-CF₃、-CCl₃、-CBr₃,-CN, optionally have substituent C_1-6Alkyl, the C optionally with substituent_3-10Cycloalkanes Base, optionally 3~10 circle heterocycles base of non-aromatic with substituent, the optionally C with substituent_2-10Alkenyl, optionally have take The C of Dai Ji_2-10Alkynyl, C_1-6Alkoxy, C_1-6Alkylthio group, C_1-6Alkoxy carbonyl group, C_1-6Carboxylic acid group, C_1-6Carboxylic acid ester groups, C_1-7Acyl group, C_1-7Acyloxy, C_1-7Acyloxy C_1-6Alkyl, C_6-10Aryloxy group, the C optionally with substituent_3-10Cycloalkyloxy, optionally have take The C of Dai Ji_6-10Aromatic group, the 5-10 member heteroaromatic groups optionally with substituent,

The wherein described substituent optionally having is each independently selected from fluorine, chlorine, bromine, iodine, astatine ,-NO when occurring every time₂、- NR⁹ ₂、-OH、-SR¹⁰、-SO₄NR¹¹ ₂、-CF₃、-CCl₃、-CBr₃、-CN、C_1-6Alkyl, C_3-10The 3~10 of cycloalkyl, non-aromatic Circle heterocycles base, C_2-10Alkenyl, C_2-10Alkynyl, C_1-6Alkoxy, C_1-6Alkylthio group, C_1-6Alkoxy carbonyl group, C_1-6Carboxylic acid group, C_1-6Carboxylate Base, C_1-7Acyl group, C_6-10Aryloxy group, C_3-10Cycloalkyloxy, C_6-10Aryl, 5-10 unit's heteroaryls,

Wherein R⁶、R⁷、R⁸、R⁹、R¹⁰And R¹¹Hydrogen atom, C are represented independently of one another_1-6Alkyl, C_1-6Alkoxy, C_1-6Alkane sulphur Base, C_1-6Alkoxy carbonyl group, C_1-6Carboxylic acid group, C_1-6Carboxylic acid ester groups, C_1-7Acyl group, C_3-10Cycloalkyl or C_3-8Cycloalkyloxy.

The compound of 2. embodiment 1 of embodiment, it is with following formula：

Wherein, X and Y is each independently selected from O, N, optionally S, the imido grpup with substituent, optional with substituent C_1-2Alkylidene, Z₃、Z₄And Z₁Being each independently selected from optionally has the C of substituent_1-6Alkyl, the C optionally with substituent_3-10 Cycloalkyl, the C optionally with substituent_6-10Aromatic group, the C optionally with substituent_1-6Alkoxy carbonyl group, optionally have substitution The C of base_1-6Carboxylic acid group, the C optionally with substituent_1-6Carboxylic acid ester groups, the C optionally with substituent_1-7Acyl group, optionally have take 3~10 circle heterocycles base of non-aromatic of Dai Ji and the 5-10 member heteroaromatic groups optionally with substituent, wherein Z₃And Z₄Can be It is identical or different.

（To Z₁And Z₂Restriction）The compound of 3. embodiment 1 of embodiment, wherein Z₁And Z₂In it is at least one be hydrogen, And when being hydrogen for only one, another is selected from：Phenyl, carbethoxyl group, methoxycarbonyl group.

（To Z₁And Z₂Restriction）The compound of 4. embodiment 2 of embodiment, wherein Z₁It is selected from：Optionally there is substitution The phenyl of base, carbethoxyl group, methoxycarbonyl group.

（To Z₃And Z₄Restriction）The compound of 5. embodiment 1 or 2 of embodiment, wherein the Z3 and Z4 are selected from：Optionally Aromatic group with substituent, the aromatic group are selected from：Phenyl, pyridine, thienyl, furyl, naphthyl, adamantyl, And pyrrole radicals,

Wherein when substitution, the substituent of the aromatic group is selected from fluorine, chlorine, bromine, iodine ,-NO₂、-NR¹² ₂、-OH、-SR¹³、- SO₄NR¹⁴ ₂、-CF₃、-CCl₃、-CBr₃、-CN、C_1-6Alkyl, C_3-103~10 circle heterocycles bases, the C of cycloalkyl, non-aromatic_2-10Alkene Base, C_2-10Alkynyl, C_1-6Alkoxy, C_1-6Alkylthio group, C_1-7Acyl group, C_6-10Aryloxy group, C_3-10Cycloalkyloxy, C_6-10Aromatic group, 5-10 member heteroaromatic groups, wherein R¹²、R¹³And R¹⁴Hydrogen atom, C are represented independently of one another_1-6Alkyl, C_1-6Alkoxy, C_3-8Ring Alkyl or C_3-8Cycloalkyloxy.

（To Z₅And Z₆Restriction）The compound of 6. embodiment 1 of embodiment, wherein Z5 and Z6 are selected from：Hydrogen, fluorine, chlorine, Bromine, iodine ,-CF₃、-CCl₃、-CBr₃, optionally have substituent C_1-6Alkyl.

The compound of 7. embodiment 1 or 2 of embodiment, it is selected from：

（Derivative）A kind of 8. compound of embodiment, it is the three-dimensional different of the compound of any one of embodiment 1-7 Structure body, enantiomter, officinal salt, in vivo solvate, hydrolyzable precursor.

（Composition）A kind of 9. pharmaceutical composition of embodiment, it includes the compound of any one of claim 1-8.

The pharmaceutical composition of 10. embodiment 9 of embodiment, wherein the compound of any one of claim 1-8 or its Combination is unique active ingredient or main active ingredient in described pharmaceutical composition.

（First is medicinal）Embodiment 11. is used to prepare the disease for treating or preventing and benefiting from and adjusting 14-3-3 ζ albumen The compound of any one of the claim 1-8 of medicine.

（First is medicinal）Embodiment 12. is used to treat or prevent the right for benefiting from the disease for adjusting 14-3-3 ζ albumen It is required that the compound of any one of 1-8.

The compound of any one of 13. claim 1-8 of (pharmaceutical applications) embodiment or comprising at least one of foregoing The purposes of composition, it is used to prepare the medicine for treating or preventing and benefiting from the disease for adjusting 14-3-3 ζ albumen.

The purposes of 14. embodiment 13 of (pharmaceutical applications) embodiment, wherein described benefit from adjusts 14-3-3 ζ albumen Disease is cancer.

The compound of any one of 15. claim 1-8 of (therapeutical uses) embodiment or comprising at least one of foregoing The purposes of composition, it is used to treat or prevent the disease benefited from and adjust 14-3-3 ζ albumen, and the purposes includes giving patient The compound of any one of the claim 1-8 of therapeutically effective amount includes at least one of foregoing composition, it is preferable that wherein The effective amount for the treatment of is 0.01-10mg/kg, preferably 0.3-7mg/kg, more preferably 0.5-5mg/kg, more preferably 0.8-4mg/ Kg, more preferably 0.9-2mg/kg.

The compound of any one of 16. claim 1-8 of (therapeutical uses) embodiment or comprising at least one of foregoing The purposes of composition, it is used to treat or prevent cancer, and the purposes includes giving the claim 1-8 of bacterium Any one of compound or include at least one of foregoing composition, it is preferable that the wherein described effective amount for the treatment of is 0.01- 10mg/kg, preferably 0.3-7mg/kg, more preferably 0.5-5mg/kg, more preferably 0.8-4mg/kg, more preferably 0.9-2mg/kg.

(treatment method) embodiment 17. using any one of claim 1-8 compound or include foregoing at least one The composition of item is used to treat or prevent the disease benefited from and adjust 14-3-3 ζ albumen, it includes giving bacterium's The compound of any one of claim 1-8 includes at least one of foregoing composition, it is preferable that wherein described treatment is effective Measure as 0.1-10mg/kg, preferably 0.3-7mg/kg, more preferably 0.5-5mg/kg, more preferably 0.8-4mg/kg, more preferably 0.9-2mg/kg.

(treatment method) embodiment 18. using any one of claim 1-8 compound or include foregoing at least one The composition treatment of item or pre- anti-cancer, it includes the chemical combination for giving any one of the claim 1-8 of bacterium Thing includes at least one of foregoing composition, it is preferable that the wherein described effective amount for the treatment of is 0.1-10mg/kg, preferably 0.3-7mg/kg, more preferably 0.5-5mg/kg, more preferably 0.8-4mg/kg, more preferably 0.9-2mg/kg.

A kind of 19. pharmaceutical unit dosage pharmaceutical composition of (unit dose) embodiment, said composition include 3-600mg's The compound of any one of claim 1-8, or its combination, the unit dosage form is suitable for being administered orally, percutaneous dosing, It is subcutaneously injected.

The pharmaceutical composition of 20. embodiment 19 of (formulation) embodiment, it is liquid agent, tablet, capsule and gel The form of capsule.

The pharmaceutical composition of 21. embodiment 19 of (unit dose) embodiment, in its unit dose comprising 30mg, 60mg, or the compound of any one of claim 1-8 of 120mg, or its combination.

The pharmaceutical composition of any one of 22. embodiment 19 to 21 of embodiment, wherein any one of claim 1-8 Compound or its combination as active ingredient main in described pharmaceutical composition or unique active ingredient.

Any one of 23. aforementioned embodiments 14,16 and 18 of embodiment, wherein the cancer includes：Lung cancer, liver Cancer, cervical carcinoma, breast cancer, prostate cancer, lymph cancer, bone marrow cancer, acute leukemia, stomach cancer, neuroglia cancer, meninx cancer, food Pipe cancer, incidence dermoid cancer, carcinoma of mouth, cancer of pancreas, oophoroma and cutaneum carcinoma, wherein cancer listed above can To be primary and/or metastatic.

The method that embodiment 24. prepares the compound of any one of embodiment 1 to 8, it comprises the following steps：

Wherein, condition a is alkaline condition, such as potassium carbonate and the condition of tetrabutylammonium bromide；B is alkaline condition, is such as used Sodium ethoxide；Wherein Z₁To Z₆, and X and Y are as defined in Formulas I.

Brief description of the drawings

Fig. 1 show the control of the compound 10 and gossypol of the present invention in terms of antitumous effect.In Fig. 1, horizontal seat The unit for marking concentration is micromole, and ordinate is cell mortality.

Fig. 2 show the state of the cancer cell when not adding compound, and cancer cell-apoptosis is seldom.Fig. 2 is only to add DMSO, is not added with cell state during compound 10.In figure, the point of the grey to occupy the majority is the total number of cells contaminated with Hoechst, few Several white points is with the PS of annexin V dye, that is, the cell of apoptosis.

Fig. 3 show compound concentration for 25 micromoles processing 24 it is small when after cell state.In Fig. 3, compound concentration 25 Cell state after when micromole's processing 24 is small.In figure, the point of grey is the total number of cells contaminated with Hoechst, and white point is to use film Join the PS of albumen V dyes, that is, the cell of apoptosis.Most cells are dead as seen from the figure, and only small part can dye.

Fig. 4 show control group（It is left）And administration group（It is right）Tumour growth situation in mouse.Fig. 4 is control group（It is left） And administration group（It is right）The tumour growth situation of mouse.

Embodiment

Definition

In the present invention, term " 14-3-3 ζ albumen " refers to the ζ hypotypes of 14-3-3 albumen.14-3-3 albumen is a kind of acid Property soluble protein, was separated from the brain tissue of ox in 1967 by Moor and Perez, and because it is through diethylaminoethyl cellulose Component distributed quantity in chromatography and the migration position in starch-gel electrophoresis and gain the name.14-3-3 molecular weight of albumen is about 30kD, isoelectric point are 4~5.In its natural state, 14-3-3 albumen mainly exists in the form of homotype or heterodimer, is one The protein family of a very high homology by different genes coding.In mammal, its mainly have 7 hypotypes (β, γ, ε, η, ζ, σ and τ/θ).14-3-3 ζ albumen is ζ hypotypes.

" benefiting from the disease for adjusting 14-3-3 ζ albumen " refers to express relevant disease with the height of 14-3-3 ζ albumen, passes through Make drug molecule and the protein with reference to and suppress its function so that the disease is controlled." benefit from and adjust 14-3-3 ζ The disease of albumen " includes cancer.

Existing result of study shows, kinds cancer such as lung (lung) cancer, liver (liver) cancer, cervical carcinoma, mammary gland (breast) cancer, prostate (prostate) cancer, lymph (diffuse large B-cell lymphoma) cancer, marrow (multiple myeloma) cancer, acute leukemia (acute promyolocytic leukemia), stomach (stomach) cancer, Neuroglia (glioma) cancer, meninx cancer (meningioma), the cancer of the esophagus (esophageal), incidence dermoid cancer (head and neck squamous cell carcinoma), oral cavity (oral) cancer, pancreas (pancreatic) cancer, ovary (ovarian) cancer and skin (skin) cancer etc., it is all related to 14-3-3 ζ albumen, namely these diseases can be by adjusting 14- 3-3 ζ albumen and prevented and treated.14-3-3 ζ albumen is in the occurrence and development process of tumour and the drug resistance of tumour cell It played an important role in property, can be with by making small molecule (such as polypeptide) and 14-3-3 ζ active-site competitive bindings It is obvious to suppress the growth of cancer cell, and cause its apoptosis, so as to reach the effect of prevention and treatment cancer.

Compound makes to be represented by the chemical formula in the present specification, or uses standardized denomination.

In the present specification, " halogen atom " refers to fluorine atom, chlorine atom, bromine atoms, iodine atom and astatine atom.

As the preferred embodiment of " halogen atom ", fluorine atom, chlorine atom and bromine atoms can be enumerated.

In the present specification, " the C_1-6Alkyl " refers to the straight-chain or branched-chain alkyl of carbon number 1~6, as Instantiation, can enumerate C₁Alkyl, C₂Alkyl, C₃Alkyl, C₄Alkyl, C₅Alkyl, C₆Alkyl, especially methyl, ethyl, 1- propyl group (n-propyl), 2- propyl group (isopropyl), 2- methyl isophthalic acids-propyl group (isobutyl group), 2- methyl-2-propyls (tert-butyl group), 1- butyl are (just Butyl), 2- butyl (sec-butyl), 1- amyl groups, 2- amyl groups, 3- amyl groups, 2-methyl-1-butene base, 3- methyl isophthalic acids-butyl, 2- methyl- 2- butyl, 3- methyl -2- butyl, 2,2- dimethyl -1- propyl group, 1- hexyls, 2- hexyls, 3- hexyls, 2- methyl-1-pentenes base, 3- Methyl-1-pentene base, 4- methyl-1-pentenes base, 2- methyl -2- amyl groups, 3- methyl -2- amyl groups, 4- methyl -2- amyl groups, 2- methyl -3- Amyl group, 3- methyl -3- amyl groups, 2,3- dimethyl -1- butyl, 3,3- dimethyl -1- butyl, 2,2- dimethyl -1- butyl, 2- second Base -1- butyl, 3,3- dimethyl -2- butyl, 2,3- dimethyl -2- butyl etc..

As " C_1-6The preferred embodiment of alkyl ", can enumerate methyl, ethyl, 1- propyl group, 2- propyl group, 2- methyl isophthalic acids-propyl group, 2- methyl-2-propyls, 1- butyl, 2- butyl.

In the present specification, " the C_1-6Alkylidene " refers to from " C defined above_1-6Further remove in alkyl " and appoint One hydrogen atom of meaning and the divalent group that derives, as instantiation, can enumerate C₁Alkylidene, C₂Alkylidene, C₃Alkylidene, C₄Alkylidene, C₅Alkylidene, C₆Alkylidene, especially methylene, 1,2- ethylidene, 1,1- ethylidene, 1,3- propylidene, Isosorbide-5-Nitrae- Butylidene, 1,5- pentylidene, 1,6- hexylidenes etc..

In the present specification, " singly-bound " has implication as commonly understood in the art, refers to the atom of the key both sides Direct neighbor, and be only connected by a valence link.

In the present specification, " the C_2-10Alkenyl " refers to the straight chain for having a double bond and carbon number is 2~10 Shape or branched alkenyl, as instantiation, can enumerate C_2-6Alkenyl, especially C₂Alkenyl, C₃Alkenyl, C₄Alkenyl, C₅Alkenyl, C₆ Alkenyl, C₇Alkenyl, C₈Alkenyl, C₉Alkenyl, C₁₀Alkenyl.

In the present specification, " the C_2-6Alkenyl " refers to the straight-chain for having a double bond and carbon number is 2~6 Or branched alkenyl, as instantiation, C can be enumerated₂Alkenyl, C₃Alkenyl, C₄Alkenyl, C₅Alkenyl, C₆Alkenyl, especially ethene Base, 1- acrylic, 2- acrylic (pi-allyl), 1- cyclobutenyls, 2- cyclobutenyls, 3- cyclobutenyls, pentenyl, hexenyl etc..

In the present specification, " the C_2-10Alkynyl " refers to the straight chain for having three keys and carbon number is 2~10 Shape or branched alkynyl, as instantiation, can enumerate C_2-6Alkynyl, especially C₂Alkynyl, C₃Alkynyl, C₄Alkynyl, C₅Alkynyl, C₆ Alkynyl, C₇Alkynyl, C₈Alkynyl, C₉Alkynyl, C₁₀Alkynyl.

In the present specification, " the C_2-6Alkynyl " refers to the straight-chain for having three keys and carbon number is 2~6 Or branched alkynyl, as instantiation, C can be enumerated₂Alkynyl, C₃Alkynyl, C₄Alkynyl, C₅Alkynyl, C₆Alkynyl, especially acetylene Base, 1- propinyls, 2-propynyl, 1- butynyls, 2- butynyls, 3- butynyls, pentynyl, hexin base etc..

In the present specification, " the C_3-10Cycloalkyl " refers to the monocyclic or bicyclic saturated fat that carbon number is 3~10 Fat race alkyl, as instantiation, can enumerate " C_3-8Cycloalkyl ", especially C₃Cycloalkyl, C₄Cycloalkyl, C₅Cycloalkyl, C₆Ring Alkyl, C₇Cycloalkyl, C₈Cycloalkyl, C₉Cycloalkyl, C₁₀Cycloalkyl.

In the present specification, " the C_3-8Cycloalkyl " refers to the monocyclic or bicyclic saturated fat that carbon number is 3~8 Race's alkyl, as instantiation, can enumerate C₃Cycloalkyl, C₄Cycloalkyl, C₅Cycloalkyl, C₆Cycloalkyl, C₇Cycloalkyl, C₈Cycloalkanes It is base, especially cyclopropyl, cyclobutyl, cyclopenta, cyclohexyl, suberyl, cyclooctyl, bicyclic [2.1.0] amyl group, bicyclic It is [3.1.0] hexyl, bicyclic [2.1.1] hexyl, bicyclic [4.1.0] heptyl, bicyclic [2.2.1] heptyl (norborneol alkyl), bicyclic [3.3.0] octyl group, bicyclic [3.2.1] octyl group, bicyclic [2.2.2] octyl group etc..

In this application, when certain group is described as " optionally having substituent ", which can not have hydrogen to be substituted base Substitution, or substituted selected from following substituent：Fluorine, chlorine, bromine, iodine, astatine ,-NO₂、-NR¹² ₂、-OH、-SR¹³、-SO₄NR¹⁴ ₂、- CF₃、-CCl₃、-CBr₃、-CN、C_1-6Alkyl, C_3-103~10 circle heterocycles bases, the C of cycloalkyl, non-aromatic_2-10Alkenyl, C_2-10Alkynes Base, C_1-6Alkoxy, C_6-10Aryloxy group, C_3-10Cycloalkyloxy, C_6-10Aryl, 5-10 unit's heteroaryls, wherein

As " C_3-8The preferred embodiment of cycloalkyl ", can enumerate cyclopropyl, cyclobutyl, cyclopenta.

In the present specification, " the C_6-10Aryl " refers to the aromatic hydrocarbon cyclic group that carbon number is 6~10, As instantiation, C can be enumerated₆Aryl, C₇Aryl, C₈Aryl, C₉Aryl, C₁₀Aryl, especially phenyl, 1- naphthyls, 2- naphthalenes Base, indenyl, azulenyl etc..

As " C_6-10The preferred embodiment of aryl ", can enumerate phenyl and naphthyl.

In the present specification, " hetero atom " refers to nitrogen-atoms, oxygen atom or sulphur atom.

In the present specification, the atomicity that " 5~10 unit's heteroaryl " refers to form ring is 5~10, its cyclization is former Contain 1~5 heteroatomic aromatic cyclic groups in son, as instantiation, can enumerate furyl, thienyl, pyrrole radicals, Imidazole radicals, triazolyl, tetrazole radical, thiazolyl, pyrrole radicals, oxazolyl, isoxazolyls, isothiazolyl, furazanyl, thiadiazolyl group, Oxadiazolyl, pyridine radicals, pyrazinyl, pyridazinyl, pyrimidine radicals, triazine radical, purine radicals, pteridyl, quinolyl, isoquinolyl, naphthalene Piperidinyl, quinoxalinyl, cinnoline base, quinazolyl, phthalazinyl, imidazopyridyl, Imidazothiazole base, Mi Zuo Bing oxazolyls, Benzothiazolyl, benzoxazolyl, benzimidazolyl, indyl, isoindolyl, indazolyl, pyrrolopyridinyl, thieno pyrrole Piperidinyl, furopyridyl, diazosulfide base, Ben Bing oxadiazolyl, Pyridopyrimidine base, benzofuranyl, benzothiophene Base, thienofuran base etc..

As the preferred embodiment of " 5~10 unit's heteroaryl ", furyl, thienyl, pyrrole radicals, imidazole radicals, thiazole can be enumerated Base, pyrazolyl, oxazolyl, isoxazolyls, isothiazolyl, pyridine radicals, pyrimidine radicals.

In the present specification, " 3~10 circle heterocycles base of non-aromatic " refers to：

(a) ring member nitrogen atoms number for 3~10,

(b) ring member nitrogen atoms include 1~2 hetero atom,

(c) in ring optionally containing 1~2 double bond,

(d) in ring optionally containing 1~3 carbonyl, sulfinyl or sulfonyl,

(e) monocyclic or dicyclic non-aromatic cyclic group,

When containing nitrogen-atoms in ring member nitrogen atoms, can also have a substituent or hydrogen atom on nitrogen-atoms.

As the instantiation of " 3~10 circle heterocycles base of non-aromatic ", aziridinyl, azetidinyl, pyrrole can be enumerated Cough up alkyl, piperidyl, nitrogen heterocyclic heptyl, Azacyclooctane base, piperazinyl, Diazesuberane base, diazocine alkyl, Diazabicyclo [2.2.1] heptyl, morpholinyl, thio-morpholinyl, 1,1- dioxidothiomorpholinyls, Oxyranyle, oxa- ring Butane group, tetrahydrofuran base, dioxolane base, THP trtrahydropyranyl, dioxane base, tetrahydro-thienyl, tetrahydrochysene thiophene Mutter base, oxazole alkyl, thiazolidinyl etc..

As the preferred embodiment of " 3~10 yuan of non aromatic heterocyclyls ", aziridinyl, azetidinyl, pyrrole can be enumerated Cough up alkyl, piperidyl, nitrogen heterocyclic heptyl, piperazinyl, dioxy nitrogen heterocyclic heptyl, morpholinyl, thio-morpholinyl, 1,1- dioxies Thio-morpholinyl, tetrahydrofuran base, THP trtrahydropyranyl.

In the present specification, " the C_1-6Alkoxy " refers in " C defined above_1-6Alkyl " end is bonded aerobic original The group of son, as instantiation, can enumerate methoxyl group, ethyoxyl, 1- propoxyl group (positive propoxy), 2- propoxyl group (isopropyl oxygen Base), 2- methyl isophthalic acids-propoxyl group (isobutoxy), 2- methyl -2- propoxyl group (tert-butoxy), 1- butoxy (n-butoxy), 2- Butoxy (sec-butoxy), 1- amoxys, 2- amoxys, 3- amoxys, 2-methyl-1-butene epoxide, 3- methyl isophthalic acids-butoxy, 2- methyl -2- butoxy, 3- methyl -2- butoxy, 2,2- dimethyl -1- propoxyl group, 1- hexyloxies, 2- hexyloxies, the own oxygen of 3- Base, 2- methyl-1-pentenes epoxide, 3- methyl-1-pentenes epoxide, 4- methyl-1-pentenes epoxide, 2- methyl -2- amoxys, 3- methyl -2- Amoxy, 4- methyl -2- amoxys, 2- methyl -3- amoxys, 3- methyl -3- amoxys, 2,3- dimethyl -1- butoxy, 3, 3- dimethyl -1- butoxy, 2,2- dimethyl -1- butoxy, 2- ethyl -1- butoxy, 3,3- dimethyl -2- butoxy, 2, 3- dimethyl -2- butoxy etc..

As " C_1-6The preferred embodiment of alkoxy ", can enumerate methoxyl group, ethyoxyl, 1- propoxyl group, 2- propoxyl group, 2- first Base -1- propoxyl group, 2- methyl -2- propoxyl group, 1- butoxy, 2- butoxy.

In the present specification, " the C_1-6Alkylthio group " refers in " C defined above_1-6Alkyl " end is bonded with sulphur original The group of son, as instantiation, can enumerate methyl mercapto, ethylmercapto group, 1- rosickyite base (positive rosickyite base), 2- rosickyite base (isopropyl sulphur Base), 2- methyl isophthalic acids-rosickyite base (isobutylthio), 2- methyl -2- rosickyite bases (tertiary butylthio), 1- butylthios (positive butylthio), 2- fourths Sulfenyl (secondary butylthio), 1- penta sulfenyls, 2- penta sulfenyls, 3- penta sulfenyls, 2-methyl-1-butene sulfenyl, 3- methyl isophthalic acids-butylthio, 2- first Base -2- butylthios, 3- methyl -2- butylthios, 2,2- dimethyl -1- rosickyite base, the own sulfenyls of 1-, the own sulfenyls of 2-, the own sulfenyls of 3-, 2- first Base -1- penta sulfenyls, 3- methyl-1-pentenes sulfenyl, 4- methyl-1-pentenes sulfenyl, 2- methyl -2- penta sulfenyls, 3- methyl -2- penta sulfenyls, 4- first Base -2- penta sulfenyls, 2- methyl -3- penta sulfenyls, 3- methyl -3- penta sulfenyls, 2,3- dimethyl -1- butylthios, 3,3- dimethyl -1- fourth sulphur Base, 2,2- dimethyl -1- butylthios, 2- ethyl -1- butylthios, 3,3- dimethyl -2- butylthios, 2,3- dimethyl -2- butylthios etc..

As " C_1-6The preferred embodiment of alkylthio group ", can enumerate methyl mercapto, ethylmercapto group, 1- rosickyite base (positive rosickyite base), 2- third Sulfenyl (isopropyisulfanyl), 2- methyl isophthalic acids-rosickyite base (isobutylthio), 2- methyl -2- rosickyite bases (tertiary butylthio), 1- butylthios (positive butylthio), 2- butylthios (secondary butylthio).

In the present specification, " the C_3-10Cycloalkyloxy " refers in " C defined above_3-10Cycloalkyl " end is bonded There is the group of oxygen atom, as instantiation, " C can be enumerated_3-8Cycloalkyloxy ", C₃Cycloalkyloxy, C₄Cycloalkyloxy, C₅Cycloalkanes oxygen Base, C₆Cycloalkyloxy, C₇Cycloalkyloxy, C₈Cycloalkyloxy, C₉Cycloalkyloxy, C₁₀Cycloalkyloxy.

In the present specification, " the C_3-8Cycloalkyloxy " refers in " C defined above_3-8Cycloalkyl " end is bonded aerobic original The group of son, as instantiation, can enumerate C₃Cycloalkyloxy, C₄Cycloalkyloxy, C₅Cycloalkyloxy, C₆Cycloalkyloxy, C₇Cycloalkanes oxygen Base, C₈It is cycloalkyloxy, especially ring propoxyl group, cyclobutoxy group, cyclopentyloxy, cyclohexyloxy, cycloheptyl epoxide, ring octyloxy, bicyclic [2.1.0] amoxy, bicyclic [3.1.0] hexyloxy, bicyclic [2.1.1] hexyloxy, bicyclic [4.1.0] epoxide in heptan, bicyclic [2.2.1] Epoxide in heptan (norbornane epoxide), bicyclic [3.3.0] octyloxy, bicyclic [3.2.1] octyloxy, bicyclic [2.2.2] octyloxy etc..

As " C_3-8The preferred embodiment of cycloalkyloxy ", can enumerate ring propoxyl group, cyclobutoxy group, cyclopentyloxy.

In the present specification, " single C_1-6Alkyl amino " refers to 1 hydrogen atom in amino with defined above “C_1-6Group obtained by alkyl " substitution, as instantiation, can enumerate methylamino, ethylamino, 1- propylcarbamics (positive third Base amino), 2- propylcarbamics (isopropylamino), 2- methyl isophthalic acids-propylcarbamic (isobutylamino), 2- methyl-2-propyl ammonia Base (tert-butylamino), 1- butylaminos (n-butylamino), 2- butylaminos (s-butylamino), 1- pentyl aminos, 2- penta Base amino, 3- pentyl aminos, 2-methyl-1-butene base amino, 3- methyl isophthalic acids-butylamino, 2- methyl -2- butylaminos, 3- first Base -2- butylaminos, 2,2- dimethyl -1- propylcarbamics, 1- hexylaminos, 2- hexylaminos, 3- hexylaminos, 2- methyl - 1- pentyl aminos, 3- methyl-1-pentene bases amino, 4- methyl-1-pentene bases amino, 2- methyl -2- pentyl aminos, 3- methyl -2- penta Base amino, 4- methyl -2- pentyl aminos, 2- methyl -3- pentyl aminos, 3- methyl -3- pentyl aminos, 2,3- dimethyl -1- fourths Base amino, 3,3- dimethyl -1- butylaminos, 2,2- dimethyl -1- butylaminos, 2- ethyl -1- butylaminos, 3,3- diformazans Base -2- butylaminos, 2,3- dimethyl -2- butylaminos etc..

In the present specification, " two C_1-6Alkyl amino " refer to 2 hydrogen atoms in amino respectively with identical or Different " C defined above_1-6Group obtained by alkyl " substitution, as instantiation, can enumerate N, N- dimethylaminos, N, N- diethylaminos, N, bis--n-propyl aminos of N-, N, N- diisopropylaminoethyls, N, N- di-n-butyls amino, N, N- diisobutyls Amino, N, N- di-sec-butyls amino, N, N- di-t-butyls amino, N- ethyl-N-methylaminos, N- n-propyl-N- methylaminos, N- isopropyl-N- methylaminos, N- normal-butyl-N- methylaminos, N- isobutyl-N-methylaminos, N- sec-butyl-N- methyl ammonia Base, N- t-butyl-N-methylaminos etc..

In the present specification, " the C_1-7Acyl group " includes " C_2-7Acyl group " and " C₁Acyl group ", wherein " C₁Acyl group " refers to aldehyde Base, " C_2-7Acyl group ".

In the present specification, " the C_2-7Acyl group " refers to be bonded with " C defined above_1-6The carbonyl of alkyl ", as Instantiation, can enumerate such as acetyl group, propiono, iso-propionyl, bytyry, isobutyryl, valeryl, isovaleryl, three Methyl acetyl etc..

In the present specification, " the C_1-6Alkoxy carbonyl " refers to be bonded with " C defined above_1-6The carbonyl of alkoxy " Base, as instantiation, can enumerate such as methoxycarbonyl, ethoxy carbonyl, 1- propoxycarbonyls, 2- propoxycarbonyls, 2- Methyl -2- propoxycarbonyls etc..

In the present specification, " optionally the having substituent " refers to：" optionally the position that can substitute with appoint Meaning combination has one or more substituents ", the instantiation of substituent, can enumerate example as the substituent as defined above Such as halogen atom, hydroxyl, thio hydroxyl, nitro, cyano group, formoxyl, carboxyl, amino, silicyl, mesyl, C_1-6Alkyl, C_2-6Alkenyl, C_2-6Alkynyl, C_3-8Cycloalkyl, C_6-10Aryl, 5~10 unit's heteroaryls, 3~10 yuan of non aromatic heterocyclyls, C_1-6Alkane Epoxide, C_1-6Alkylthio group, C_3-8Cycloalkyloxy, list C_1-6Alkyl amino, two C_1-6Alkyl amino, C_2-7Acyl group or C_2-7Alkoxy carbonyl Deng.Wherein, C_1-6Alkyl, C_2-6Alkenyl, C_2-6Alkynyl, C_3-8Cycloalkyl, C_6-10Aryl, 5~10 unit's heteroaryls, 3~10 yuan it is non-aromatic Fragrant race's heterocyclic radical, C_1-6Alkoxy, C_1-6Alkylthio group, C_3-8Cycloalkyloxy, list C_1-6Alkyl amino, two C_1-6Alkyl amino, C_2-7Acyl Base and C_2-7Alkoxy carbonyl can also have 1~3 group being selected from the group in substituent independently of one another.

In the present specification, " the C_1-6Carboxylic acid group " refers to there is formula "-RCOOH " or the group of "-COOH ", wherein R Can be C_1-6Alkyl.

In the present specification, " the C_1-6Carboxylic acid ester groups " refers to formula "-R^aCOOR^b" or "-COOR^b" group, Wherein R^aCan be C_1-6Alkyl, R^bCan be C_1-6Alkyl.

In the present specification, " the C_1-7Acyloxy C_1-6Alkyl " refers to formula " R^aCOOR^b- " or " HCOOR^b" Group, wherein R^aCan be C_1-6Alkyl, R^bCan be C_1-6Alkyl.

Alloisomerism, which is herein defined as its sensu lato implication, has identical structural formula, but its various group The isomery of the compound spatially arranged in a different manner, it is all in particular such as in mono-substituted hexamethylene, wherein substituting Base can be in the various possible rotational conformations of axial direction or equatorial position, and ethane derivative.However, since fixed takes Different spatial arrangements of the Dai Ji in double bond or ring and there are another type of alloisomerism, phase to typically refer to geometrical isomerism Or cis-trans isomerization.Term " stereoisomer " is used and therefore represented according to its broadest implication in this application All compounds indicated above.

The application refers to " enantiomter ", " officinal salt ", " solvate " and " internal hydrolyzable precursor " With the normally understood implication of those skilled in the art.

Present invention relates in general to the compound of Formulas I, its various derivative, includes its composition, and they are used for Prepare the purposes for treating or preventing the medicine for benefiting from the disease for adjusting 14-3-3 ζ albumen, and in particular, to the compound of Formulas I It is used to prepare the purposes for the medicine for treating or preventing cancer.Compound the present invention also relates to Formulas I is used for treatment-related disease Such as the purposes of cancer.

The compound of Formulas I is as follows,

Wherein：

X and Y is each independently selected from C1-2 alkylidenes, oxygen atom, sulphur atom, the nitrogen-atoms optionally with substituent,

The compound of the present invention can be prepared by the methodology of organic synthesis disclosed herein below, can also pass through it It is prepared by its method.

Compound of formula I can be prepared into acid-addition salts or base addition salts.

The acid-addition salts for the compound of formula I that can be mentioned that include the salt formed with inorganic acid, such as hydrochloride and hydrobromate； And the salt formed with organic acid, such as formates, acetate, maleate, benzoate, tartrate and fumarate.

The acid-addition salts of compound of formula I can be by making free alkali or its salt, enantiomter or shielded derivative Thing is reacted and formed with the appropriate acid of monovalent or more equivalents.Reaction can in salt is insoluble in solvent therein or medium or Salt, which is dissolved in solvent therein, to carry out, such as water, twoAlkane, ethanol, tetrahydrofuran, the mixture of ether or solvent, these are molten Agent can be removed in vacuum or be removed by lyophilized.Reaction can be metathesis process, can also be carried out on ion exchange resin.

Some compound of formula I have chirality.

Some compound of formula I can exist in the form of tautomerism or enantiomerism.All these forms are included in In the scope of the present invention.The racemic of compound can be mixed by using routine techniques such as fractional crystallization or chirality HPLC Thing is separated, so as to separate various optical isomers.Selectively, can be by the way that the reaction bar of racemization will not be being caused Make that there is the raw material of suitable optical activity to react under part, simple enantiomter is made.

The compound of the present invention

In a particular embodiment, compound of the invention can be selected from following compound 1-20：

The preparation method of compound

The compound of the present invention can be prepared by following steps：

Wherein, condition a is alkaline condition, such as potassium carbonate and the condition of tetrabutylammonium bromide；B is alkaline condition, is such as used Sodium ethoxide, wherein Z₁To Z₆, and X and Y are as defined in Formulas I.

Specifically, the step of prepare compound can be as follows：

Condition and reagent：A) alkaline condition, K₂CO₃, TBAB；B) alkaline condition, EtONa

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS2（1.0eq.）With And glycol dibromide（1.0eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add ethyl acetate extraction, Organic layer is separated, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

By intermediate 4（1eq.）It is dissolved in suitable ethanol, sequentially adds（3eq.）Sodium ethoxide and aromatic aldehyde （2eq.）, be heated to reflux 4 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound.

, can be with prepare compound 1 to compound 14 using this specific method.

Specifically, the step of prepare compound also can be as follows：

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS2（1.0eq.）With And raw material 3（1.0eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add ethyl acetate extraction, separated Machine layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

, can be with prepare compound 15 to compound 16 using this specific method.

Specifically, the step of prepare compound also can be as follows：

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS2（1.0eq.）With And iodomethane（3.0eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add ethyl acetate extraction, separated Machine layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

By intermediate 4（1eq.）It is dissolved in suitable tetrahydrofuran, adds 2- aminoethanols（2eq.）Triethylamine （2eq.）Solution, be heated to reflux 3 it is small when, separate out solid after cooling.Crude product ethyl alcohol recrystallization, obtains mesh intermediate 5.

By intermediate 5（1eq.）It is dissolved in suitable ethanol, sequentially adds（3eq.）Sodium ethoxide and aromatic aldehyde （2eq.）, be heated to reflux 4 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound 17（Chemical combination The synthesis of thing 20 is identical with this law）.

, can be with prepare compound 17 and compound 20 using this specific method.

Specifically, the step of prepare compound also can be as follows：

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, and raw material 2 （1.5eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 24 it is small when, add ethyl acetate extraction, separate organic layer, do It is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 3.

By intermediate 3（1eq.）It is dissolved in suitable ethanol, sequentially adds sodium ethoxide（3eq.）And aromatic aldehyde（2eq.）, Be heated to reflux 6 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound 19.

, can be with prepare compound 18 and compound 19 using this specific method.

The therapeutical uses of compound

The compound of the present invention can be used for treating the disease benefited from and adjust 14-3-3 ζ albumen, can be used for preparation and control Treat or prevent to benefit from the medicine for the disease for adjusting 14-3-3 ζ albumen.

It is without being bound by theory, it is believed that compound of the invention is when for treating disease, with internal 14-3-3 ζ albumen knots Close, cause a large amount of apoptosis of cancer cell dead, so as to reach the effect for suppressing growth of cancer cells and killing cancer cell.

When using the compounds for treating disease of the present invention, compound, its solid of the Formulas I of bacterium are given Isomers, enantiomter, officinal salt, solvate, internal hydrolyzable precursor include at least one of foregoing combination Thing, wherein the effective amount of the treatment is preferably 0.01-10mg/kg, more preferably preferably 0.3-7mg/kg, 0.5-5mg/kg, more It is preferred that 0.8-4mg/kg, more preferably 0.9-2mg/kg.

Discovery based on more than, for people of the weight at 30 kilograms to double centner, can will use the compound of the present invention Unit dose drug composition is made.The pharmaceutical composition of unit dose includes the active constituents of medicine of 0.3mg to 1g.It is it is preferred that single The pharmaceutical composition of position dosage includes 3mg to 1000mg, especially, the active constituents of medicine of 3mg to 600mg.More preferably unit The pharmaceutical composition of dosage includes the active constituents of medicine of 10mg to 500mg.The pharmaceutical composition of more preferably unit dose includes The active constituents of medicine of 50mg to 300mg.The medicine that more preferably pharmaceutical composition of unit dose includes 100mg to 200mg is lived Property component.

Above-described dosage is the daily dosage of patient.However, depending on used medical compounds, treated Individual and the illness that goes wrong, which is variable.

Therefore the theme of the present invention is the compound of formula (I) as defined above, its stereoisomer, enantiomerism Body, officinal salt, solvate, internal hydrolyzable precursor are used to benefit from adjusting comprising at least one of foregoing composition The chemotherapy (medication alone or in combination) of the disease of 14-3-3 ζ albumen, especially cancer.

Such medicine for cancer chemotherapeutic can be to be used alone or in combination.

The present invention product can particularly be administered alone or with chemotherapy or radiotherapy or a combination thereof combine to Medicine, for example, with other medicine administering drug combinations.

Such other medicines can be usually used antitumor drug.

Pharmaceutical composition

Compound, its enantiomter or its officinal salt of the present invention can be used alone, can also be with suitable medicine The form of thing preparation uses, enteral or parenteral administration.Another aspect of the present invention provides pharmaceutical composition, it includes preferably few In 80%, the compound of the invention of 50% weight is more preferably less than, and be mixed with inertia pharmaceutically acceptable diluent, lubricant or load Body.

The example of diluent, lubricant and carrier has：

- for tablet and lozenge：Lactose, starch, talcum, stearic acid；

- for capsule：Tartaric acid or lactose；

- for parenteral solution（Liquid agent）：Water, alcohol, glycerine, vegetable oil；

- for suppository：Natural or fixed oil or wax.

The present invention also provides the method for preparing this pharmaceutical composition, and this method includes：Each component is mixed or is compounded in Together, and each component of mixing is made to form tablet or suppository, Jiang Gecheng is distributed into capsule, or component is dissolved to form injection Liquid.

For the purposes mentioned by the application, method, medicine and composition, the dosage of compound and the formulation of administration are certain Can with changed using compound, mode of administration and expected therapeutic purposes.However, when the compound of the present invention is with about During the daily dosage administration of 0.01-10mg/kg the weight of animals, satisfied result is generally yielded.These dosage can be by portioning agent Amount is given with daily 1 to 4 time, can also be given in the form of sustained release.For the mankind, per total daily dose can scope for 3mg extremely 600mg, more preferably 10mg to 100mg, the unit dosage forms suitable for oral administration include 2mg to 600mg compounds and mix There are solid-state or liquid pharmaceutical acceptable carrier, lubricant or diluent.

Some compounds of the invention can be deposited in the form of tautomerism, enantiomerism, alloisomerism or geometrical isomerism All these forms are included within the scope of the invention.Can be by using routine techniques such as fractional crystallization or chirality HPLC separates the racemic mixture of compound, so that separating optical isomers.Selectively, can be by will not The starting material for making to have suitable optical activity under the reaction condition of racemization is caused to react, the optics for being made simple is different Structure body.

By method similar as above, example compound of the invention can be obtained.Those skilled in the art can To be readily appreciated that, a variety of reagents known in the art can be used for the compound to form Formulas I.

These pharmaceutical compositions can by oral administration, intestines and stomach or by being locally applied to skin or mucous membrane local administration, or Through intravenous or intramuscular administration.

These compositions can be solid or liquid, and can be all medicine shapes usually used in physianthropy Formula, for example, simple or sweet tablet tablet, alkyl, lozenge, gel capsule, drops, granule, injectable formulation, ointment, Creme or gelling agent；They are prepared according to conventional methods.In this application, active ingredient can be mixed to usual and used In the auxiliary material of these pharmaceutical compositions, such as talcum, Arabic gum, lactose, starch, magnesium stearate, cocoa butter, it is water-based or Non-aqueous carrier, animal or plant source property lipid, paraffin derivative, glycol, various wetting agents, dispersant or emulsifying agent or anti-corrosion Agent.

Embodiment

In order to be expressly understood, by explanation and way of example, this hair is described with exemplary Compounds and methods for It is bright.However, to those skilled in the art, when the teaching of close examination compound of the present invention, technique and method, Without departing from the content purport or scope in the case of, the modifications and changes that can be done to it are obvious.

Embodiment 1（Synthetic example）

Following prepare compound 1：

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS2（1.0eq.）With And glycol dibromide（1.0eq.）With TBAB (tetrabutylammonium bromide)（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add Enter ethyl acetate extraction, separate organic layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

By intermediate 4（1eq.）It is dissolved in suitable ethanol, sequentially adds（3eq.）Sodium ethoxide and benzaldehyde（2eq.）, Be heated to reflux 4 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound, is yellow solid, production Rate 87.2%.Compound¹HNMR data are as shown in table 1.

Chemical reaction process is as follows：

Condition and reagent：A) alkaline condition, K2CO3, TBAB;B) alkaline condition, EtONa；The Ar in the preparation of compound 1 For phenyl.

Embodiment 2-14（Synthetic example）

It is similar with synthetic example 1, synthesis compound 2-14.Wherein the characterize data of compound 1-14 and yield etc. are as follows Shown in table 1.

Table 1:The composite result data of compound 1-14

Embodiment 15-16（Synthetic example）

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS2（1.0eq.）With And raw material 3 as shown below（1.0eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add ethyl acetate extraction Take, separate organic layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

By intermediate 4（1eq.）It is dissolved in suitable ethanol, sequentially adds（3eq.）Sodium ethoxide and aromatic aldehyde（2eq.）, Be heated to reflux 4 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound 15.

, can be with prepare compound 15 to compound 16 using this specific method.

Reaction equation is as follows：

Embodiment 17 and 20（Synthetic example）

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, CS₂（1.0eq.）With And iodomethane（3.0eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 10 it is small when, add ethyl acetate extraction, separated Machine layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 4.

By intermediate 4（1eq.）It is dissolved in suitable tetrahydrofuran, adds 2- ethylaminoethanols（2eq.）Triethylamine （2eq.）Solution, be heated to reflux 3 it is small when, separate out solid after cooling.Crude product ethyl alcohol recrystallization, obtains intermediate 5.

By intermediate 5（1eq.）It is dissolved in suitable ethanol, sequentially adds（3eq.）Sodium ethoxide and aromatic aldehyde（2eq.）, Be heated to reflux 4 it is small when, separate out solid after cooling.Crude product is recrystallized with alcohol-water, obtains target compound 17（Compound 20 Synthesis is identical with this law）.

, can be with prepare compound 17 and compound 20 using this specific method.

Reaction equation is as follows：

Embodiment 18-19（Synthetic example）

By 2,4- pentanediones（1eq.）It is added in suitable quantity of water, sequentially adds potassium carbonate（2.5eq.）, and it is as shown below Raw material 2（1.5eq.）And TBAB（0.1eq.）.Be stirred at room temperature reaction 24 it is small when, add ethyl acetate extraction, separated Machine layer, it is dry.After rotary evaporation removes organic layer, ethyl alcohol recrystallization, obtains intermediate 3.

, can be with prepare compound 18 and compound 19 using this specific method.

Chemical equation is as follows：

Table 2:The generated data of compound 15-20

In order to prove that the compounds for treating of Formulas I benefits from purposes of disease of adjusting 14-3-3 ζ albumen etc., carry out following Experiment.

Embodiment 21（In vitro test）

SPR test principles describe, SPR and BIACORE T100 brief introductions（In vitro test）

Technical principle:

SPR refers to surface plasma body resonant vibration (Surface Plasmon Resonance).Surface plasma body resonant vibration is to use In the optics professional technique that characterization surface refraction coefficient changes, surface mentioned here is usually the interface between solid phase and liquid phase. Surface plasma resonance technology had significant progress at past 10 years, and application field includes film, self assembled monolayer Formation and property, the reciprocation between protein, nucleosides, pharmaceuticals, surfactant equimolecular.SPR can be with real-time monitored To superficial phenomena such as molecule combination, film formation, and high sensitivity, high selectivity non-specific binding minimum at the same time can be provided Signal.SPR can be interacted with the label-free biomolecule that obtains in real time, between different pharmaceutical or drug modification structure and biomolecule Interaction, molecule interact and separated speed, molecule interact when reach balance, the size of co-acting force Deng important information.

Application field:Life science, food security, environment measuring are biomedical；Toxin and antibiotic quickly detect；Egg White matter group；Drug screening and related drugs dynamics detect in real time；The inspection of the special peptide fragment of biomolecule and related coupled molecule Survey；Viral and pathogenic molecule protein and acceptor research；Molecular recognition, immunological regulation, immunoassays etc..

Biacore T100 are the systems of new generation of newest release in 2005, increase thermokinetics research application newly, are the whole world Currently the only one is taken the instrument that surface plasma resonance technology (SPR) carries out real-time thermokinetics research, with the instrument The affinity and dynamics data of interaction of molecules and reaction at different temperatures can easily be obtained.Surface etc. from Sub-resonance can be between real-time tracking biomolecule interaction, and do not have to any label.First by a kind of biology point during experiment Son is fixed on sensor chip surface, and the molecule interacted therewith is dissolved in solution flows through chip surface.Detector can track Molecule in detection solution is combined, is dissociated the change of whole process with the molecule of chip surface.3 cores of Biacore systems Part is sensor chip, SPR Systems for optical inspection, microjet chuck.

The technical parameter of setting：

1. sample type：Micromolecular compound, polypeptide, protein, oligonucleotides, oligosaccharide, lipoid, bacteriophage, virus And cell

2. sample capacity：+ 8 Reagent Tube of 2x96 orifice plates

3. sampling volume：5-750ul

4. index of refraction scope：1.33-1.40

5. sample recycles：Automatically

6. sample introduction controls：Automatically

7. instrument temperature control：Automatically

8. instrument deaerates：Automatically

9. analysis time：1-5 minutes/sample

10. circulate tankage：~0.02ul

11. flow cell number：4

12. remove ground control online：Automatically

13. detectable limit molecular weight：100Da

14.GxP certifications：It is

The source of reagent

SPR T100 binding tests steps, as a result, and discuss

1st, pre- binding tests

Since chip surface carries negative electricity, the molecule that be coupled on chip must become positively charged, and just be conducive to point Son is adsorbed onto chip surface, in favor of the completion of covalent coupling reaction.Pre- binding tests are exactly to dissolve a sample in different PH Buffer solution in, it is taken different amounts of electric charge.Then, sample is flowed through into chip surface, the combination for observing it and chip is bent Line, so that it may judge suitable condition.Adsorption test under two kinds of different conditions, first peak show under this condition, to adsorb Saturation is quickly reached, the amount so adsorbed is limited, and second in rising trend always, is conducive to be coupled more egg on chip In vain.In real work, also a variety of adsorption curves can be encountered, it is necessary to judge optimum condition according to actual conditions.By egg The white buffer solution with different PH dilutes 10-40 times, flows through chip surface with the flow velocity of 5 μ l/min, observation albumen and chip Absorption result.

2nd, it is coupled

A) 100 μ l NHS are taken（N- hydroxysuccinimides, N-hydroxysuccinimide, NHS, from Sigma- Aldrich）With 100 μ l EDC（Ethyl-dimethylamino-propyl-carbodiimide, 1-ethyl-3- (3- Dimethylaminopropyl) carbodimide hydrochloride, EDC, from Sigma-Aldrich）Mixing, 12000rpm is centrifuged 5 minutes；

B) coupling protein of certain volume is taken, by the dilution proportion combined in advance, 150 μ l are needed after dilution, takes 100 μ l hydrochloric acid Monoethanolamine (ethanolamine hydrochloridem, from Sigma-Aldrich）.Above reagent equilibrates to room temperature 25 and takes the photograph Family name's degree or so, high speed centrifugation five minutes；

C) above-mentioned three kinds of reagents are placed on specimen holder, set sample-adding flow velocity as 10 μ l/min, inject mode sample introductions, NHS/EDC, sample, diethanolamine hydrochloride difference loading 100,100, and 60 μ l.

D) terminate to can obtain trial curve.

3rd, combine

Micromolecular compound is diluted to debita spissitudo with suitable buffer solution, loading after centrifugation, observes compound and egg White cohesive process.If necessary to measure the bonding state under different condition, chip can be carried out after single injected sampling terminates Regeneration.Then next sample is gone up again.

4th, chip regenerates

The regeneration condition that antigen-antibody combines is fairly simple, is simply rinsed with the 10mM glycine (glycine) of PH2.5 Can.If coupling is albumen, situation is more complicated.First, regeneration condition will can remove conjugate from chip, Secondly the albuminous degeneration being incorporated on chip cannot be allowed to inactivate.After chip regeneration, not requiring nothing more than baseline be able to will be returned to initially Value, also requires again upper identical sample, binding ability is not in be decreased obviously.

5th, obtain combining the kinetic constant with dissociation if desired, at least need to carry out the combination examination of five groups of various concentrations Test.

6th, result of the test is analyzed, prints test report.

7th, instrument maintenance, experiment finish, and running buffer changes filtering and the ultra-pure water or HES-EP buffer solutions that deaerate into （Acetate buffer, from Reagecon companies）.Chip, which is changed to, safeguards chip, after carrying out at least twice of prime, operation standby。

It is as follows using each compound and the dissociation constant of 14-3-3 ζ albumen of SPR measurements：

Compound	K_D±SD(μM)
		1	1.17±0.5
2	100.0

3	3.65±1.7
		4	42.81±8.3
5	0.12±0.1
		6	36.75±1.3
7	1.45±1.2
		8	100.0
9	0.76±0.8
		10	0.42±0.4
11	9.10±2.2
		12	14.22±5.8
15	0.33±0.2
		16	1.96±0.9
17	1.55±0.7
		18	1.10±0.8
19	2.57±1.4
		20	1.32±1.0

Tested by SPR, it is found that the compound 1-20 of the present invention can be with 14-3-3 ζ albumen directly in conjunction with wherein having three A compound（5,10,15）And the combination solution of 14-3-3 ζ albumen leaves constant and is less than 1 micromole, belongs to stronger combination.Say There occurs stronger interaction with 14-3-3 ζ really for bright compound of the invention.

Embodiment 22（In vivo studies a, Apoptosis assay）

Apoptosis assay is carried out with compound 10, using gossypol as control, is the description of the experiment below, ties Fruit, and discuss（In vivo studies a）

Test principle：

Apoptosis is one of essential characteristic of cell, it is in the embryonic development of body, tissue repair, the stabilization of interior environment Etc. play a very important role.In normal cell, phosphatidylserine（PS）Only it is distributed in cell membrane lipid bilayer Inner side, and in Apoptosis early stage, the phosphatidylserine in cell membrane（PS）By turning on one's side laterally in adipose membrane.Annexin V is the Ca dependence cardiolipin binding proteins that a kind of molecular weight is 35-36kD, can be with being turned in apoptosis process outside film PS high-affinities are specifically bound.By the use of the AnnexinⅤ that marked FITC as fluorescence probe, flow cytometer or glimmering is utilized Light microscope can detect the generation of Apoptosis., can be left in 350nm and Hoechst is a kind of DNA dyestuffs that can pass through cell membrane Right ultraviolet excitation, sends indigo color fluorescence at 461nm.Therefore the cell with annexin V-FITC is to be withered The cell died, and the cell for catching Hoechst is total cell concentration, by counting different cell numbers, so as to count Calculate the cell proportion that apoptosis occurs.

Test reagent：

DMEM culture mediums（Gibco by life technologies,REF:11995-065）, trypsase, Hank ' s Buffer solution（By standard configuration）, apoptosis detection kit（apoptosis detection kit）（Mainly include annexin V- FITC（BD Pharmingen,Cat:556547）, Hoechst33342）Deng.

Test material：

Hela cells (human cervical carcinoma cell system).

Test apparatus and apparatus：

High intension microscope, incubator, water-bath, 24 orifice plates etc..

Test procedure：

1. cultivate Hela cells；

2. after cell is all adherent, the rush apoptosis compound for adding various concentrations is handled；

After when 3. processing 24 is small, old culture medium is discarded, is washed twice with Hank ' s buffer solutions；

4. adding 10uM Hoechst dyestuffs, 30min in 37 DEG C of incubators is placed on；

5.Hank ' s buffer solutions are washed twice；

6. combination buffer is washed once；

7. annexin V-FITC and combination buffer are carried out 1:Culture dish is added after 10 mixing；

8. it is placed on 20min in 37 DEG C of incubators；

9. washing dyestuff off with Hank ' s, the DEME culture mediums of respective amount are added；

10. high intension microscope photographing；

11. count and analyze data.

Wherein Hoechst dyestuffs and combination buffer are included in apoptosis detection kit.

The result of experiment is as shown in Fig. 1,2,3.

From figure 1 it appears that it is substantially better than gossypol using the cancer cell death rate of compound 10, namely its is antitumor Positive effect is more excellent.（Antitumoral compounds in Fig. 1 are exactly compound 10.）

Fig. 2, which is shown, is not added with compound 10, cancer cell states during using only DMSO.The grey to occupy the majority in fig. 2 Point is the total number of cells contaminated with Hoechst, and a small number of white points is with the PS of annexin V dye, that is, the cell of apoptosis. From figure 2 it can be seen that survivaling cell when being not added with compound is very more, cancer cell-apoptosis obtains considerably less.

Fig. 3 be show 10 concentration of compound for 25 micromoles processing 24 when small after cell state, be illustrated that in Fig. 3 The parallel test of Fig. 2 experiments, except that with the addition of compound 10 in DMSO.The point of grey is contaminated with Hoechst in figure Total number of cells（Reflect the number of survivaling cell）, white point is with the PS of annexin V dye, that is, the cell of apoptosis. Most cells are dead as can be seen from Fig. 3, and only small part can dye.

The result shows that：Compound 10 has the function that significantly to promote cancer cell-apoptosis (using gossypol as with reference to chemical combination Thing), compound 10 kills cancer cell by way of promoting cancer cell-apoptosis.

More than experiment prove that compound 10 has the function that strong carcinogenic cells apoptosis, this 14-3-3 with document report ζ albumen be apoptosis inhibitory protein be consistent, of the invention compound by suppressing 14-3-3 ζ albumen, relieve the albumen pair The suppression of Apoptosis, so as to promote a large amount of apoptosis of cancer cell dead, has reached suppression cell growth and has killed cancer cell Effect.

Embodiment 23（In vivo studies b, Carbazole alkaloid experiment）

MTT cell tests principles, method and result（In vivo studies）

The principle of cell tests

This project use mtt assay screening compounds activity, MTT full name for 3- (4,5)-dimethylthiahiazo (- 2-y1) -3,5-di-phenytetrazoliumromide, entitled 3- (4,5- dimethylthiazole -2) -2, the 5- hexichol of Chinese chemistry Base tetrazole bromide, trade name：Tetrazolium bromide.It is a kind of dyestuff of yellow color.Mtt assay is also known as MTT colorimetric methods, is that a kind of detection is thin Born of the same parents are survived and the method for growth.Its testing principle is that the succinate dehydrogenase in living cells mitochondria can reduce exogenous MTT First a ceremonial jade-ladle, used in libation is crystallized for the bluish violet of water-insoluble（Formazan）And be deposited in cell, and dead cell is without this function.Dimethyl is sub- Sulfone（DMSO）The first a ceremonial jade-ladle, used in libation in cell can be dissolved, its absorbance value is measured at 490nm wavelength with enzyme-linked immunosorbent assay instrument, can be indirect Reflect number of viable cells.In the range of certain cell number, it is directly proportional to cell number that MTT crystallizes the amount to be formed.This method is extensive Activity determination, large-scale screening anti-tumor medicine, cell toxicity test and tumour for some bioactie agents are put Penetrate sensitivity testing etc..Its feature is high sensitivity, economy.MTT has carcinogenicity, with when it is careful, it is saturating preferably to wear that Bright book film gloves.The MTT needs being made into are sterile, and MTT is very sensitive to bacterium；Toward 96 orifice plate added-time as far as possible the illumination on operation console Lamp is turned off.PBS is used when preparing MTT（pH=7.4）Dissolving.PBS is a kind of buffer solution.PBS is formulated：NaCl8g+KCl0.2g+ Na₂HPO₄1.44g+KH₂PO₄0.24g tune pH7.4, constant volume 1L.In general, the MTT concentration in this method is 5mg/ml.Therefore, can be with MTT0.5 grams is weighed, is dissolved in the phosphate buffer of 100ml（PBS）Or without in phenol red culture medium, with 0.22 μm of membrane filtration with The bacterium in solution is removed, 4 DEG C is put and is kept in dark place.During preparation and preservation, the most handy aluminium-foil paper of container encases.

The method of cell tests

Test procedure is as follows：

Attached cell

1st, logarithmic phase cell is collected, adjusts concentration of cell suspension, 100ul is added per hole, bed board makes cell tune density to be measured To 1000-10000 cells/wells,（Edge hole is filled with sterile PBS）.

2nd, 5%CO2,37 DEG C of incubations, bottom hole is paved with to cell monolayer（96 hole flat undersides）, the medicine of concentration gradient is added, it is former On then, after cell attachment can dosing, or two hours, or time half a day, but we are often in noon before that day bed board, morning next day Dosing.General 5-7 gradient, per hole 100ul, if 3-5 multiple holes.It is recommended that setting 5, otherwise it is difficult to react truth.

3rd, 5%CO2, when 37 DEG C of incubation 16-48 are small, is observed under inverted microscope.

4th, 20ulMTT solution is added per hole（5mg/ml, i.e. 0.5%MTT）, continue to cultivate 4h.If medicine can be anti-with MTT Should, it can first centrifuge and discard nutrient solution afterwards, carefully rush 2-3 after with PBS, add the nutrient solution containing MTT.

5th, culture is terminated, carefully sucks nutrient solution in hole.

6th, 150ul dimethyl sulfoxide (DMSO)s are added per hole, low-speed oscillation 10min on shaking table is put, crystal is fully dissolved.In enzyme Join the light absorption value in each hole of measurement at immune detector OD490nm.

7 while zeroing hole is set（Culture medium, MTT, dimethyl sulfoxide (DMSO)）, control wells（Cell, the medicine of same concentrations are molten Solve medium, nutrient solution, MTT, dimethyl sulfoxide (DMSO)）.

Suspension cell

1st, logarithmic phase cell is collected, adjusts 1 × 106/ml of concentration of cell suspension, 1640 will 1. supplied in order（Without blood Clearly）Culture medium 40ul；2. plus Actinomycin D（Actinomycin D, Sigma-Aldrich, Inc. are toxic）10ul is trained Nutrient solution dilute lg/ml, need prerun find optimum dilution degree, 1:10-1:20)；3. need detectable substance 10ul；4. cell suspension 50ul （I.e. 5 × 10⁴A cells/well）, common 100ul is added to 96 orifice plates（Edge hole is filled with sterile water）.Control is set per plate（Add 100ul (storing liquid 100ul1640）.

2nd, 37 DEG C, when 5%CO2 incubations 16-48 is small are put, is observed under inverted microscope.

3rd, 10ul MTT solution is added per hole（5mg/ml, i.e. 0.5%MTT）, continue to cultivate 4h.（Suspension cell recommends WST-1, step 4 is can skip after cultivating 4h）, direct enzyme-linked immunosorbent assay instrument OD570nm（630nm is calibrated）Measure the extinction in each hole Value.

4th, centrifuge（1000 turns of 10min）, supernatant is carefully sopped up, 100ul dimethyl sulfoxide (DMSO)s are added per hole, put low speed on shaking table 10min is vibrated, crystal is fully dissolved.In enzyme-linked immunosorbent assay instrument OD570nm（630nm is calibrated）Measure the extinction in each hole Value.

5 while zeroing hole is set（Culture medium, MTT, dimethyl sulfoxide (DMSO)）, control wells（Cell, the medicine of same concentrations are molten Solve medium, nutrient solution, MTT, dimethyl sulfoxide (DMSO)）, every group of 3 multiple holes of setting.

Points for attention：

(1) appropriate cell implantation concentrations are selected.

(2) serum interference is avoided：The nutrient solution of general hyclone of the choosing less than 10% is tested.After colour generation as far as possible Exhaust residual culture in hole.

(3) blank control is set：The blank control for being not added with cell and only adding nutrient solution parallel with experiment.Other test procedures are kept Unanimously, last colorimetric is returned to zero with blank.

(4) MTT experiments absorbance finally will be between 0-0.7, and super go beyond the scope is not just linear relation.

(5) it is MTT with 96 orifice plate culture cells and surveys cell viability, adds 200ul1640,20ul MTT according to what is said on book, 150ul DMSO.Nutrient solution will be removed as far as possible before DMSO by adding, and colorimetric estimation is carried out easy to DMSO dissolving first Zan particles.

(6) general to be advisable per 4000, hole cell, i.e. for cell concentration in 20000/ml, MTT adds 20ul, acts on four hours After wash supernatant off, be careful not to wash off first Zan, then add 150ul DMSO per hole, vibrated 10 minutes on decolorization swinging table, Then light absorption value is surveyed.

The result of cell tests（Compound 1 to 10）

Tested more than, from test result as it can be seen that the compound 1-10 of the present invention has suppression for various cancer cells Effect.And it was found that compound 10 is best to the inhibition of cancer cell, to Hela cells（Human cervical carcinoma cell system）IC₅₀ It is 3.2 micromoles, to SGC cells（Human gastric cancer cell line）IC₅₀It is 17.90 micromoles, there is good inhibiting effect, it is right hep G2（Bel7402）IC₅₀It is 29.54 micromoles, and for the cell U937 close to normal somatic cell（People's macrophage Cell line）IC₅₀It is 899 micromoles, illustrates that suppression and killing of the compound 10 to cancer cell have preferable selectivity and peace Quan Xing.

Embodiment 24（Animal experiment）

The description of mouse test, step, as a result, and discuss（Using compound 10 as representing, the chemical combination of the present invention is tested Control of the thing to rat liver cancer）

Mouse Acute Toxicity trial test

Date：On June 22nd, 2013

9:00 pair of 6 mouse is divided into 3 groups, A groups（10mg/kg）, B groups（20mg/kg）With C groups（Blank control）, claim respectively Weight is taken, tries to achieve average value, configures medicine, each group mouse injection medicine is reached required concentration.

9:30 injections finish

10:After 00 administration, there is face portion in two groups of mouse, tail blacks, expiratory dyspnea（Hurriedly）, to environmental stimuli without Reaction.B groups are even more serious compared with A groups.

14:First dead mouse of 00A groups, the symptom of second mouse have slight relief.First mouse symptom of B groups has Slight relief, second mouse are more serious.

20:The not dead mouse symptom of 30A groups continues to alleviate, and sensitiveer to environmental stimuli, tail black turns light；B groups One mouse symptom continues to alleviate, sensitiveer to environmental stimuli, and tail black turns light；Second mouse does not improve, tail Black, be short of breath, it is slow in reacting.

Date：On June 23rd, 2013

18:The not dead mouse of 30A groups is in good condition.One mouse of B groups is in good condition；Another dead mouse.

Date：On June 24th, 2013

9:00 two groups of not dead mouse are in good condition.

18:00 two groups of not dead mouse are in good condition.

Date：On June 25th, 2013

9:00 two groups of not dead mouse are in good condition.

Date：On June 26th, 2013

9:00 two groups of not dead mouse are in good condition.

Trial test conclusion：By experimental observation understand compound to the acute fatal concentration of mouse should 5mg/kg to Between 10mg/kg, so it is experiment to take 5mg/kg, tri- groups of 7mg/kg, 10mg/kg respectively in formal toxicological test drug concentration Group.

Mouse Acute Toxicity formal test

Date：On June 27th, 2013

9：00

12 mouse are divided into four groups, every group 3, carry out weighing body weight.Blank group 3, is injected intraperitoneally 200 μ l DMSO；Change Three groups of compound dosing group, is injected intraperitoneally compound DMSO solution, final concentration of 5mg/kg, 7mg/kg, 10mg/kg by every group 3. After injection, blank group, 5mg/kg groups, 7mg/kg group mouse, animation is good, and hair is neat, eyes glow, without not Good reaction；10mg/kg group mouse, appearance activity is abnormal, few dynamic slow-witted sleeping, insensitive, the obvious intoxicating phenomenon such as hyporeactive.

Date：On June 27th, 2013

21:00

Blank group, 5mg/kg groups, 7mg/kg group mouse, animation is good, and fur is neat, eyes glow, without bad anti- Should, no death；10mg/kg group mouse, poisoning symptom are eased, no death.

On June 28 9:00

Blank group, 5mg/kg groups, 7mg/kg group mouse, animation is good, and fur is neat, eyes glow, without bad anti- Should, no death；10mg/kg group mouse, dead 1, remaining 2 animations are good.

On June 28 21:00

Blank group, 5mg/kg groups, 7mg/kg group mouse, animation is good, and fur is neat, eyes glow, without bad anti- Should, no death；10mg/kg group mouse, animation is good, and fur is neat, and eyes glow, has no adverse reaction.

On June 29 9:00, mouse performance such as 6.28.

On June 29 21:00, mouse performance such as 6.28.

On June 30 9:00, mouse performance such as 6.28.

On July 59：00 observation

Blank group, 5mg/kg group mouse animations are normal；7mg/kg group mouse, dead one, remaining 2 animations Well；10mg/kg group mouse, animation are good.

Anatomic observation：

Remaining 10 mouse are put to death, are dissected, every group takes two dissections, observes the character of internal organ.Blank group and 5mg/kg group mouse, internal organ character are normal；Kidney, the color of heart is normal, scarlet, no toxicity performance；Liver is normal color, Shape is normal, undistorted, does not have toxicity symptom.7mg/kg and 10mg/kg group mouse, liver surface have stain, are a bit darkish in color.

Formal test conclusion：

The dosage of the 5mg/kg of compound 10 does not have mouse toxic side effect, 7mg/kg and 10mg/kg to the poisonous pair of mouse Effect；Dissection test proves that the dosage of 7mg/kg and 10mg/kg have toxicological effect to its internal organ；Weight detection is carried out, from body Re-detection finds out that compound 10 does not influence the weight of mouse significantly.

According to acute toxic test as a result, choosing 2 groups of transplanting liver tumour mouse, every group 6, one group is used as control, another Group injects 1mg/kg compounds 10 daily.

Fig. 4 is control group（It is left）And administration group（It is right）Tumour growth situation in mouse.

Mouse test conclusion：The compound 10 of the present invention is can be seen that 5mg/kg's from the data and Fig. 4 of this experiment There is no toxic side effect, 7mg/kg and 10mg/kg below dosage to mouse to mouse toxic side effect；Dissection test proves, 7mg/ The dosage of kg and 10mg/kg has toxicological effect to its internal organ；Weight detects, it can be seen that compound is not bright to the weight of mouse Aobvious influence.The mouse medication with liver cancer is given using the dosage successive administration of 1mg/kg within 15 days, finds the mouse of medication group Tumour block size is significantly less than control group, and tumor control rate reaches about 65% (being calculated by knurl weight), and for control group Extend the service life of mouse.

Claims

1. the compound of Formulas I,

Wherein：

X and Y is each independently selected from the C optionally with substituent_1-2Alkylidene, oxygen atom, sulphur atom and nitrogen-atoms；

Z₁And Z₂In it is at least one be hydrogen, and when being hydrogen for only one, another is selected from phenyl, carbethoxyl group and methoxycarbonyl group；

Z₃And Z₄Selected from the following group optionally with substituent：Phenyl, pyridine, thienyl, furyl, naphthyl, adamantyl and Pyrrole radicals, and when substitution, the substituent is selected from fluorine, chlorine, bromine, iodine ,-NO₂、-NR¹² ₂、-OH、-SR¹³、-SO₄NR¹⁴ ₂、- CF₃、-CCl₃、-CBr₃、-CN、C_1-6Alkyl, C_3-10Cycloalkyl, C_2-10Alkenyl, C_2-10Alkynyl, C_1-6Alkoxy, C_1-6Alkylthio group, C_1-7Acyl group, C_6-10Aryloxy group, C_3-10Cycloalkyloxy and C_6-10Aromatic group, wherein R¹²、R¹³And R¹⁴Represent that hydrogen is former independently of one another Son, C_1-6Alkyl, C_1-6Alkoxy, C_3-8Cycloalkyl or C_3-8Cycloalkyloxy；

Z₅And Z₆It is selected from：Hydrogen, fluorine, chlorine, bromine, iodine ,-CF₃、-CCl₃、-CBr₃With the C optionally with substituent_1-6Alkyl；

The wherein described substituent optionally having is each independently selected from fluorine, chlorine, bromine, iodine ,-NO when occurring every time₂、-NR⁹ ₂、- OH、-SR¹⁰、-SO₄NR¹¹ ₂、-CF₃、-CCl₃、-CBr₃、-CN、C_1-6Alkyl, C_3-10Cycloalkyl, C_2-10Alkenyl, C_2-10Alkynyl, C_1-6 Alkoxy, C_1-6Alkylthio group, C_1-6Alkoxy carbonyl group, C_1-6Carboxylic acid group, C_1-6Carboxylic acid ester groups, C_1-7Acyl group, C_6-10Aryloxy group, C_3-10Cycloalkanes Epoxide, C_6-10Aryl,

Wherein R⁶、R⁷、R⁸、R⁹、R¹⁰And R¹¹Hydrogen atom, C are represented independently of one another_1-6Alkyl, C_1-6Alkoxy, C_1-6Alkylthio group, C_1-6Alkoxy carbonyl group, C_1-6Carboxylic acid group, C_1-6Carboxylic acid ester groups, C_1-7Acyl group, C_3-10Cycloalkyl or C_3-8Cycloalkyloxy；

The compound of wherein described Formulas I excludes：

CompoundWherein Ar for furyl, thienyl, p-methoxyphenyl, to Dimethylaminobenzene Base, p-methylphenyl, phenyl, rubigan, o-methoxyphenyl, m-methoxyphenyl, 3,4- bis- (methoxyl group) phenyl,Between chlorphenyl, Chloro-O-Phenyl, p-bromophenyl, p-fluorophenyl, 3,4-OCH₂OC₆H₃-, 3- pyridine radicals；

CompoundWherein R¹For rubigan, R²For phenyl or p-methylphenyl or p-methoxyphenyl；Or Person R¹For p-methylphenyl, R²For phenyl；

CompoundWherein Ar is C₆H₅、p-CH₃OC₆H₄、p-CH₃C₆H₄Or

CompoundWherein n=1, R or R¹/R²For 4-tBuC₆H₄、3-CH₃C₆H₄Or 4-ClC₆H₄/2- ClC₆H₄；

CompoundWherein Ar is 4-N (CH₃)₂C₆H₄、C₆H₅、4-CH₃OC₆H₄、3-CH₃OC₆H₄、4- CH₃C₆H₄、4-FC₆H₄、3-NO₂C₆H₄Or furyl；

2. the compound of claim 1, it is selected from：

3. a kind of compound, it is the stereoisomer or officinal salt of the compound of any one of claim 1-2.

4. the compound of claim 3, the stereoisomer is enantiomter.

5. a kind of pharmaceutical composition, it includes the compound of any one of claim 1-4 or its combination.

6. a kind of pharmaceutical composition, it includes selected from following any one compound or its combination：

7. the pharmaceutical composition of claim 5 or 6, wherein the compound or its combination are unique in described pharmaceutical composition Active ingredient or main active ingredient.

8. a kind of unit dose drug composition, said composition includes the compound of any one of the claim 1-4 of 3-600mg Or its combination.

9. a kind of unit dose drug composition, said composition include 3-600mg selected from following any one compound or its Combination

10. the unit dose drug composition of claim 8, it includes 30mg, 60mg, the claim of 120mg, or 300mg The compound of any one of 1-4 or its combination.

11. the unit dose drug composition of claim 9, it includes 30mg, 60mg, the chemical combination of 120mg, or 300mg Thing or its combination.

12. the unit dose drug composition of any one of claim 8-11, wherein the compound or its combination are used as institute State active ingredient main in pharmaceutical composition or unique active ingredient.

13. the unit dose drug composition of any one of claim 8-11, it is suitable for being administered orally, percutaneous dosing, skin Lower injection.

14. the unit dose drug composition of any one of claim 8-11, it is liquid agent, tablet, suppository or capsule The form of agent.

15. the unit dose drug composition of claim 14, the capsule are gel capsules.

16. the compound of any one of claim 1-4 is used to treat or prevent to benefit from preparation adjusts 14-3-3 ζ albumen Purposes in the medicine of disease.

17. the purposes of claim 16, wherein the disease for benefiting from adjusting 14-3-3 ζ albumen is cancer.

18. the purposes of claim 17, wherein the cancer includes：Lung cancer, liver cancer, cervical carcinoma, breast cancer, prostate cancer, leaching Bar cancer, bone marrow cancer, acute leukemia, stomach cancer, neuroglia cancer, meninx cancer, the cancer of the esophagus, incidence dermoid cancer, mouth Chamber cancer, cancer of pancreas, oophoroma and cutaneum carcinoma, wherein cancer listed above can be primary and/or metastatic.

19. it is used to treat or prevent in preparation selected from following any compound and benefits from the disease for adjusting 14-3-3 ζ albumen Purposes in medicine

20. the purposes of claim 19, wherein the disease for benefiting from adjusting 14-3-3 ζ albumen is cancer.

21. the purposes of claim 20, wherein the cancer includes：Lung cancer, liver cancer, cervical carcinoma, breast cancer, prostate cancer, leaching Bar cancer, bone marrow cancer, acute leukemia, stomach cancer, neuroglia cancer, meninx cancer, the cancer of the esophagus, incidence dermoid cancer, mouth Chamber cancer, cancer of pancreas, oophoroma and cutaneum carcinoma, wherein cancer listed above can be primary and/or metastatic.