CN101928326B

CN101928326B - Substitution five-heterocyclic alkyl aminoacyl compound and application thereof

Info

Publication number: CN101928326B
Application number: CN200910139488.5A
Authority: CN
Inventors: 聂爱华; 顾为; 谭祖磊; 李斐; 汤立合; 陶林; 苏瑞斌; 李锦�; 马小根
Original assignee: Institute of Pharmacology and Toxicology of AMMS
Current assignee: Institute of Pharmacology and Toxicology of AMMS
Priority date: 2009-06-24
Filing date: 2009-06-24
Publication date: 2015-07-08
Anticipated expiration: 2029-06-24
Also published as: CN101928326A

Abstract

The invention relates to a substitution five-heterocyclic alkyl aminoacyl compound and application thereof, in particular to a substitution five-heterocyclic alkyl aminoacyl derivative which has IAPs inhibition activity and is expressed by a general formula I and pharmaceutically-acceptable salt or hydrate thereof, wherein the definition of each substitution group in the general formula I is described in the specifications, and a preparation method of the compound expressed by the general formula I, a medicinal composition comprising the compound expressed by the general formula I or the pharmaceutically-acceptable salt or hydrate thereof, the application of the compound expressed by the general formula I or the pharmaceutically-acceptable salt or hydrate thereof to the production of medicaments and the application of the medicaments to the treatment or prevention of solid tumors and relevant rumor diseases derived from hematologic malignancies are all shown in the specifications.

Description

Substituted aminoacyl five-membered heterocyclic compounds and application thereof

Technical Field

The invention relates to substituted aminoacyl five-membered heterocyclic derivatives, a preparation method thereof, a pharmaceutical composition containing the same and application of the derivatives serving as inhibitor of apoptosis inhibiting protein IAPs (inhibitors of Apoptosis proteins) in preparing compounds with antitumor activity.

Background

Apoptosis, otherwise known as programmed cell death, typically occurs during the development and maintenance of healthy tissue in multicellular organisms. Apoptotic pathways are known to play important roles in embryonic development, viral pathogenesis, cancer, autoimmune and neurodegenerative diseases, and other events. Alterations in the apoptotic response have been found to be associated with the development of cancer, autoimmune diseases such as systemic lupus erythematosus and multiple sclerosis and with viral infections including those associated with herpes viruses, pox viruses and adenoviruses.

The Apoptosis inhibitor protein IAPs (inhibitors of Apoptosis proteins) is an endogenous inhibitor of Caspases (Caspases), which are major regulators of Apoptosis, and selectively binds to Caspase-3, Caspase-7 and/or Caspase-9, blocking the proteolytic activity of Caspases that are essential for Apoptosis, thereby inhibiting Apoptosis. To date, there are 8 members of IAPs found in humans, including NAIP, XIAP, cIAP1, cIAP2, ILP2, Bruce, Survivin, and Livin (ML-IAP). Among these, NAIP, XIAP, cIAP1 and cIAP2 contain three BIR domains (BIR1, BIR2, BIR3), while ILP2, Bruce, Survivin and Livin contain only one BIR domain. Many IAPs have been shown to be aberrantly expressed in cancer cells, Survivin is highly expressed in a variety of malignancies, but the presence of the protein is not detected in normal cells. Other members of IAPs, although present in a variety of normal tissues, are also overexpressed in a variety of malignancies like Survivin. The over-expressed IAPs induce tumor cells to generate drug resistance to chemotherapy and inhibit apoptosis caused by chemotherapy or radiotherapy. On the other hand, Smac/Diablo proteins released from mitochondria can bind to IAPs at a site where IAPs bind to Caspase-3, -7 and Caspase-9, thereby inhibiting the anti-apoptotic activity of IAPs. Based on the function of IAP family proteins in the apoptotic pathway and the mechanism by which IAPs interact with Caspases and Smac, a number of compounds have been discovered that inhibit IAP function and have anti-tumor activity.

Although a series of IAPs inhibitors have been developed, no drugs are currently available on the market, and many compounds are also associated with limitations in terms of potency, stability or toxicity. Therefore, the development of safe and effective IAPs inhibitors with different structural types has positive social significance and good market prospect.

Disclosure of Invention

The invention aims to find a novel structural selective IAPs receptor high-affinity inhibitor which is used for inhibiting the activity of IAPs in a human body and further enhancing the proteolytic activity of Caspases, thereby achieving the aim of treating cancers.

The present inventors have discovered a series of novel compounds that bind to IAPs and promote apoptosis by modulating IAPs function, and that have pharmaceutically acceptable stability and bioavailability. The compounds can block the interaction of IAPs with Caspase-3, Caspase-7 and Caspase-9. The results of the present invention indicate that such small molecules can modulate IAPs proteins prior to apoptosis, thus demonstrating that the use of such compounds can provide clinically beneficial effects when administered with other inducers of apoptosis. The present inventors have demonstrated that the compounds of the present invention bind to mammalian XIAP-BIR3, cIAP-BIR3 domains, thereby promoting apoptosis in cancer cells. The compounds described herein have pro-apoptotic activity in a variety of cancer cell lines, such as bladder, breast, pancreatic, colon, leukemia, lung, lymphoma, multiple myeloma, and ovarian cancer cell lines, and may also be useful in other cancer cell lines and diseases in which cells are resistant to apoptosis. These results indicate that the compounds of the present invention will have therapeutic activity against solid tumors and tumors originating from hematological malignancies. In addition, the compounds of the invention are useful in preventing metastasis, invasion, inflammation of cancer cells, and other diseases characterized by anti-apoptotic cells. The inventor finds that the compound can inhibit the activity of IAPs (amyloid peptide protein), further enhance the proteolytic activity of Caspases and restore the apoptosis regulation effect of the Caspases through research. Therefore, the compounds can be used for treating tumors. The present invention has been completed based on the above findings.

Summary of the invention:

in a first aspect the present invention provides a compound of formula I:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein:

x is-CH₂-or-O-or-S-;

y is-CH₂-or-CH₂CH₂-or-CO-;

R¹selected from:

1)-H，

2)-C₁-C₆alkyl, optionally substituted with one or more halogens, and

3)-C₃-C₈cycloalkyl, optionally substituted with one or more halo;

R²selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-(C₁-C₆alkyl) -R⁷And are and

5)-Ar¹，

wherein,

R⁷selected from:

1)-OR⁸，

2)-SR⁸，

3)-NR⁸R⁹，

4)-NCONR⁸R⁹，

5)-NCNNR⁸R⁹，

6)-COOR⁸，

7)-CONR⁸R⁹and are and

8)-Ar¹，

Ar¹selected from phenyl, arylheterocyclyl and heterobicyclyl, optionally substituted with one or more groups selected from:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

R⁸and R⁹Each independently selected from:

1)-H，

2)-(C₁-C₆alkylene radical)_0-3Phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen, and the like,

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

5)-C₂-C₆alkenyl, which is optionally substituted with one or more halogens, and

6)-C₂-C₆alkynyl, optionally substituted with one or more halo;

R³selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-(C₁-C₆alkyl) -R¹⁰，

5)-Ar¹，

Wherein,

R¹⁰selected from:

1)-OR⁸，

2)-SR⁸，

3)-NR⁸R⁹，

4)-NCONR⁸R⁹，

5)-NCNNR⁸R⁹，

6)-COOR⁸，

7)-CONR⁸R⁹and are and

8)-Ar¹，

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

R⁸and R⁹Each independently selected from:

1)-H，

2)-(C₁-C₆alkylene radical)_0-3-phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen, and the like,

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo;

[ preferably, R³The carbon atoms to which they are attached may be of any optical configuration]

R⁴And R⁴⁰⁰Each independently selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-C₂-C₆alkenyl, and

5)-C₂-C₆an alkynyl group;

R⁵and R⁵⁰⁰Each independently selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-C₂-C₆alkenyl, and

5)-C₂-C₆an alkynyl group;

R⁶selected from:

1)-R¹¹，

2)-OR¹¹，

3)-NR¹¹R¹²，

4)-SOR¹¹and are and

5)-SO₂R¹¹，

wherein R is¹¹And R¹²Each independently selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-C₂-C₆an alkenyl group, which is a radical of an alkenyl group,

5)-C₂-C₆an alkynyl group,

6)-COR⁸，

7)-Ar¹，

8)-(C₁-C₆alkyl) -Ar¹，

9)-CO-Ar¹And are and

10)-CO-(C₁-C₆alkyl) -Ar¹，

Wherein,

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

R⁸and R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo. A compound according to the first aspect of the invention is a compound of formula Ia:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein X, R¹、R²、R³、R⁴、R⁴⁰⁰、R⁵、R⁵⁰⁰And R⁶As defined for the compounds of formula I in the first aspect of the invention.

A compound according to the first aspect of the invention is a compound of formula Ib:

A compound according to the first aspect of the invention is a compound of formula Ic:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein R¹、R²、R³、R⁸And R⁹As defined for the compounds of formula I in the first aspect of the invention.

A compound according to the first aspect of the invention is of formula Id:

A compound according to the first aspect of the invention which is a compound of formula Ic 1:

A compound according to the first aspect of the invention which is a compound of formula Id 1:

A compound according to the first aspect of the invention is a compound of formula Ie:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein: r¹、R²、R³And Ar¹As defined for the compounds of the formula I in the first aspect of the invention, Ar²Selected from phenyl, arylheterocyclyl and heterobicyclic radicals, optionally substituted by one or more groups selected from

And (3) group substitution:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

wherein R is⁸And R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo.

A compound according to the first aspect of the invention is a compound of formula If:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein: r¹、R²、R³And Ar¹As defined for the compounds of the formula I in the first aspect of the invention, Ar²Selected from phenyl, arylheterocyclyl and heterobicyclyl, optionally substituted with one or more groups selected from:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

wherein R is⁸And R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo.

A compound according to the first aspect of the invention which is a compound of formula Ie 1:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

wherein R is⁸And R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo.

A compound according to the first aspect of the invention which is a compound of formula If 1:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, wherein: r¹、R²、R³And Ar¹As defined for the compounds of the general formula I in the first aspect of the invention,Ar²selected from phenyl, arylheterocyclyl and heterobicyclic radicals, optionally substituted by one or more groups selected from

And (3) group substitution:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃，

5)-R⁸，

6)-OR⁸，

7)-NR⁸R⁹and are and

8)-COOR⁸，

wherein R is⁸And R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo.

A compound according to the first aspect of the invention selected from:

and isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof.

In a second aspect, the present invention provides a process for the preparation of a compound according to any one of the first aspect of the invention, comprising the steps of:

a) reacting a compound of formula II

II (e.g. formula)Compounds of the formula) with a compound of the formula H-R⁶Reaction of the compound of formula (I) to give the compound of formula (II)The compound represented by formula (I), followed by removal of the group Z to give a compound of formula (II)A compound of (a);

b) make formulaA compound of the formulaReaction of the compound of formula (I) to give the compound of formula (II)The compound represented by the formula (I) is then hydrolyzed in the presence of a base to obtain the compound represented by the formula (II)To representA compound of (1);

c) in the presence of a suitable reagent (e.g., one or more of isobutyl chloroformate, triethylamine, and N-methylmorpholine)A compound of the formulaReaction of the compound of formula (I) to give the compound of formula (II)The compound represented, followed by removal of the group-Boc, to give the compound of formula I; and optionally (c) a second set of instructions,

d) forming the product of step c) into a pharmaceutically acceptable salt;

wherein, X, Y, R¹、R²、R³、R⁴、R⁴⁰⁰、R⁵、R⁵⁰⁰And R⁶Z is selected from the group consisting of-H, -Boc, and-Cbz, as defined for the compound of formula I in any of the first aspects of the invention.

The method according to the second aspect of the invention, optionally having one or more of the following features:

in step a), the reaction conditions of the first step may adopt dried tetrahydrofuran, N-dimethylformamide or dichloromethane as solvent, preferably tetrahydrofuran, and may adopt a combination of DCC + HOBt or a combination of isobutyl chloroformate + tertiary amine as reaction auxiliary reagent, preferably a combination of DCC + HOBt, and the reaction temperature may be-15 ℃ to 25 ℃, preferably 0 ℃ to 10 ℃, and the reaction may be from 1 hour to 24 hours, preferably 12 hours; the deprotection in the second step can be carried out by using a dry organic solvent dissolved with 2-4N HCl, such as tetrahydrofuran, ethyl acetate, dioxane, 4N HCl/ethyl acetate, and the temperature can be-15-25 ℃, 0 ℃ and the reaction can be carried out for 1-8 hours, 2 hours;

in step b), the reaction conditions of the first step may adopt dried tetrahydrofuran, N-dimethylformamide or dichloromethane as solvent, preferably tetrahydrofuran, DCC + HOBt combination or isobutyl chloroformate + tertiary amine combination as reaction auxiliary reagent, preferably DCC + HOBt combination, the reaction temperature may be-15 ℃ to 25 ℃, preferably 0 ℃ to 10 ℃, and the reaction may be 1 hour to 24 hours, preferably 12 hours; the second step of deprotection can adopt a mixed methanol/water solution of 1-4N NaOH or LiOH, preferably a 50% methanol solution of 1N NaOH, the reaction temperature can be-15-25 ℃, preferably 0-10 ℃, and the reaction time can be 1-24 hours, preferably 3 hours; and

in step c), the reaction conditions in the first step may adopt dried tetrahydrofuran, N-dimethylformamide or dichloromethane as solvent, preferably tetrahydrofuran, preferably isobutyl chloroformate + N-methylmorpholine as reaction auxiliary reagent, the reaction temperature may be-15 ℃ to 25 ℃, preferably 0 ℃ to 10 ℃, and the reaction may be for 1 hour to 24 hours, preferably 12 hours; the deprotection in the second step can be carried out using a dry organic solvent in which 2-4N HCl is dissolved, such as tetrahydrofuran, ethyl acetate, dioxane, preferably 4N HCl/ethyl acetate, at a temperature of-15 ℃ to 25 ℃, preferably 0 ℃, for 1 hour to 8 hours, preferably 2 hours.

The method according to the second aspect of the present invention, whereinThe compound represented by the formulaThe compound represented is substituted to obtain an isomer of the compound of formula I.

In a third aspect, the present invention provides a pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of a compound according to any one of the first aspect of the present invention, and optionally a pharmaceutically acceptable carrier or excipient.

In a fourth aspect, the invention provides the use of a compound according to any one of the first aspect of the invention in the manufacture of a medicament for the treatment and/or prophylaxis of a disease associated with the overexpression of IAPs.

The use according to the fourth aspect of the invention, wherein the disease is selected from the group consisting of bladder cancer, breast cancer, pancreatic cancer, colon cancer, leukemia, lung cancer, lymphoma, multiple myeloma, ovarian cancer and cervical cancer, such as ovarian cancer and cervical cancer in particular.

In a fifth aspect, the present invention provides a method for the treatment and/or prophylaxis of a disease associated with the overexpression of IAPs in a mammal (e.g. a human) in need thereof, which method comprises administering to said mammal (e.g. a human) a therapeutically and/or prophylactically effective amount of a compound according to any one of the first aspect of the present invention.

The use according to the fifth aspect of the invention, wherein said disease is selected from the group consisting of bladder cancer, breast cancer, pancreatic cancer, colon cancer, leukemia, lung cancer, lymphoma, multiple myeloma, ovarian cancer and cervical cancer, such as ovarian cancer and cervical cancer in particular.

Detailed description of the invention:

in one embodiment of the compounds of the present invention, wherein said X is-O-.

In one embodiment of the compounds of the invention, wherein said X is-S-.

In one embodiment of the compounds of the invention, wherein Y is-CH₂-。

In one embodiment of the compounds of the invention, wherein said Y is-CO-.

In one embodiment of the compounds of the present invention, wherein R is¹Selected from: -H, -C optionally substituted by one or more halogens₁-C₆An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is¹Selected from: -H, and-C₁-C₆An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is¹Selected from: -H, and-C₁-C₄An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is¹Selected from: -H, methyl, ethyl, propyl, isopropyl.

In one embodiment of the compounds of the present invention, wherein R is¹Selected from: -H, methyl.

In one embodiment of the compounds of the present invention, wherein R is²Selected from: -H, -C₁-C₆An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is²Selected from: -H, -C₁-C₄An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is²Selected from: -H, methyl, ethyl, propyl, isopropyl.

In one embodiment of the compounds of the present invention, wherein R is²Selected from: -H, methyl.

In one embodiment of the compounds of the present invention, wherein R is³Selected from: -H, -C₁-C₆Alkyl, -C₃-C₈Cycloalkyl, - (C)₁-C₆Alkyl) -R¹⁰Wherein R is¹⁰Selected from: -OR⁸、-SR⁸、-NR⁸R⁹、-NCONR⁸R⁹、-NCNNR⁸R⁹、-COOR⁸、-CONR⁸R⁹，R⁸And R⁹Each independently selected from: -H, - (C)₁-C₆Alkylene radical)_0-3-phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen radicals, optionally substituted by one or moreHalogen substituted-C₁-C₆Alkyl, -C optionally substituted by one or more halogens₃-C₈Cycloalkyl, -C optionally substituted by one or more halogens₂-C₆Alkenyl, and-C optionally substituted with one or more halogens₂-C₆Alkynyl.

In one embodiment of the compounds of the present invention, wherein R is³Selected from: -H, -C₁-C₆Alkyl, -C₃-C₈Cycloalkyl, - (C)₁-C₆Alkyl) -OR⁸Wherein R is⁸Selected from: -H, -C₁-C₆Alkyl, -C₂-C₆Alkenyl, and-C₂-C₆Alkynyl.

In one embodiment of the compounds of the present invention, wherein R is³Selected from: -H, -C₁-C₄Alkyl, -C₄-C₆Cycloalkyl, - (C)₁-C₄Alkyl) -OR⁸Wherein R is⁸Selected from: -C₂-C₆Alkynyl.

In one embodiment of the compounds of the present invention, wherein R is³Selected from: -H, methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, cyclopentyl, cyclohexyl, 2-propynyloxy-ethyl.

In one embodiment of the compounds of the present invention, wherein R is⁴And R⁴⁰⁰Each independently selected from: -H, -C₁-C₄An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is⁴And R⁴⁰⁰Each independently selected from: -H, methyl, ethyl, propyl, isopropyl.

In one embodiment of the compounds of the present invention, wherein R is⁴And R⁴⁰⁰Each independently selected from: -H, methyl.

In one embodiment of the compounds of the present inventionWherein R is⁵And R⁵⁰⁰Each independently selected from: -H, -C₁-C₄An alkyl group.

In one embodiment of the compounds of the present invention, wherein R is⁵And R⁵⁰⁰Each independently selected from: -H, methyl, ethyl, propyl, isopropyl.

In one embodiment of the compounds of the present invention, wherein R is⁵And R⁵⁰⁰Each independently selected from: -H, methyl.

In one embodiment of the compounds of the present invention, wherein R is⁶is-NR¹¹R¹²。

In one embodiment of the compounds of the present invention, wherein R is⁶is-NR¹¹R¹²Wherein R is¹¹And R¹²Each independently selected from: -H, -C₁-C₄alkyl-C₄-C₆Cycloalkyl, -COR⁸、-Ar¹、-(C₁-C₄Alkyl) -Ar¹、-CO-Ar¹and-CO- (C)₁-C₄Alkyl) -Ar¹。

In one embodiment of the compounds of the present invention, wherein R is⁶is-NR¹¹R¹²Wherein R is¹¹And R¹²Each independently selected from: -H, -C₁-C₄alkyl-C₄-C₆Cycloalkyl, -COR⁸、-Ar¹、-(C₁-C₄Alkyl) -Ar¹、-CO-Ar¹and-CO- (C)₁-C₄Alkyl) -Ar¹Wherein Ar is¹Is phenyl optionally substituted with one or more groups selected from: halogen, nitro, cyano, -CF₃、-R⁸、-OR⁸、-NR⁸R⁹and-COOR⁸。

In one embodiment of the compounds of the present invention, wherein R is⁶is-NR¹¹R¹²Which isIn R¹¹And R¹²Each independently selected from: -H, -C₁-C₄alkyl-C₄-C₆Cycloalkyl, -COR⁸、-Ar¹、-(C₁-C₄Alkyl) -Ar¹、-CO-Ar¹and-CO- (C)₁-C₄Alkyl) -Ar¹Wherein Ar is¹Is phenyl optionally substituted with one or more groups selected from: halogen, nitro, cyano, -CF₃、-R⁸、-OR⁸、-NR⁸R⁹and-COOR⁸Wherein R is⁸And R⁹Each independently selected from: -H, - (C)₁-C₄Alkylene radical)_0-3-phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen, optionally substituted by one or more halogens, -C₁-C₄Alkyl, -C optionally substituted by one or more halogens₄-C₆Cycloalkyl, -C optionally substituted by one or more halogens₂-C₄Alkenyl, and-C optionally substituted with one or more halogens₂-C₄Alkynyl.

In one embodiment of the compounds of the invention, wherein

X is-O-or-S-;

y is-CH₂-or-CO-;

R¹selected from: -H, -C₁-C₄An alkyl group;

R²selected from: -H, -C₁-C₄An alkyl group;

R³-H、-C₁-C₆alkyl, -C₃-C₈Cycloalkyl, - (C)₁-C₆Alkyl) -OR⁸Wherein R is⁸Selected from: -H, -C₁-C₆Alkyl, -C₂-C₆Alkenyl, and-C₂-C₆An alkynyl group;

R⁴and R⁴⁰⁰Each independently selected from:-H、-C₁-C₄an alkyl group;

R⁵and R⁵⁰⁰Each independently selected from: -H, -C₁-C₄An alkyl group;

R⁶is-NR¹¹R¹²Wherein R is¹¹And R¹²Each independently selected from: -H, -C₁-C₄alkyl-C₄-C₆Cycloalkyl, -COR⁸、-Ar¹、-(C₁-C₄Alkyl) -Ar¹、-CO-Ar¹and-CO- (C)₁-C₄Alkyl) -Ar¹Wherein Ar is¹Is phenyl optionally substituted with one or more groups selected from: halogen, nitro, cyano, -CF₃、-R⁸、-OR⁸、-NR⁸R⁹and-COOR⁸Wherein R is⁸And R⁹Each independently selected from: -H, - (C)₁-C₄Alkylene radical)_0-3-phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen, optionally substituted by one or more halogens, -C₁-C₄Alkyl, -C optionally substituted by one or more halogens₄-C₆Cycloalkyl, -C optionally substituted by one or more halogens₂-C₄Alkenyl, and-C optionally substituted with one or more halogens₂-C₄Alkynyl.

All documents cited herein are incorporated by reference in their entirety and to the extent such documents do not conform to the meaning of the present invention, the present invention shall control. Further, the various terms and phrases used herein have the ordinary meaning as is known to those skilled in the art, and even though such terms and phrases are intended to be described or explained in greater detail herein, reference is made to the term and phrase as being inconsistent with the known meaning and meaning as is accorded to such meaning throughout this disclosure.

The term "halogen" or "halo" as used herein refers to fluorine, chlorine, bromine, and iodine.

The term "hydrocarbyl" as used herein includes alkyl, alkenyl and alkynyl groups. These alkyl, alkenyl and alkynyl groups may be linear or may be branched.

The term "alkyl" as used herein is meant to include both branched and straight chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms, e.g., C₁-C₆C in alkyl₁-C₆Are defined to include groups having 1, 2, 3, 4, 5, or 6 carbons arranged in a linear or branched arrangement. C as defined above₁-C₆Examples of alkyl groups include, but are not limited to: methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, pentyl and hexyl.

The term "alkenyl" as used herein refers to an unsaturated, straight or branched chain hydrocarbon radical containing the specified number of carbon atoms, wherein at least two carbon atoms are connected to each other by a double bond, and having either the E or Z configuration, and combinations thereof. E.g. C₂-C₆C in alkenyl₂-C₆Are defined to include groups having 2, 3, 4, 5 or 6 carbons arranged in a linear or branched arrangement with at least two carbon atoms connected by a double bond. C₂-C₆Examples of alkenyl groups include ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, and the like.

The term "alkynyl" as used herein refers to an unsaturated, straight or branched chain hydrocarbon radical containing the specified number of carbon atoms, and wherein at least two carbon atoms are joined together by a triple bond. E.g. C₂-C₆C in alkynyl₂-C₆Are defined to include groups having 2, 3, 4, 5, or 6 carbon atoms in the chain, at least two of which are linked together by a triple bond. Examples of such alkynyl groups include, but are not limited to: ethynyl, 1-propynyl, 2-propynyl and the like.

The term "cycloalkyl" as used herein refers to a monocyclic saturated aliphatic hydrocarbon group containing the specified number of carbon atoms therein, e.g. C₃-C₈C in cycloalkyl₃-C₈Are defined to include groups having 3, 4, 5, 6, 7, or 8 carbons in a single ring arrangement. C as defined above₃-C₈Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl.

The term "cycloalkenyl" as used herein refers to a monocyclic unsaturated aliphatic hydrocarbon group having the specified number of carbon atoms therein, e.g., C₃-C₈C in cycloalkenyl₃-C₈Are defined to include groups having 3, 4, 5, 6, 7, or 8 carbons in a single ring arrangement. C as defined above₃-C₈Examples of cycloalkenyl groups include, but are not limited to, cyclopentenyl and cyclohexenyl.

The term "halo" or "halogen" as used herein refers to fluoro, chloro, bromo and iodo.

The term "haloalkyl" as used herein means an alkyl group as defined above wherein each hydrogen atom may be replaced successively by a halogen atom. Examples of haloalkyl groups include, but are not limited to: -CH₂F、-CHF₂and-CF₃。

The term "aryl", as used herein, whether alone or in combination with another group, refers to a carbocyclic aromatic monocyclic group containing 6 carbon atoms that may be further fused to a second 5-or 6-membered carbocyclic group that may be aromatic, saturated or unsaturated. Aryl groups include, but are not limited to, phenyl, 2, 3-indanyl, 1-naphthyl, 2-naphthyl, and tetrahydronaphthyl. The fused aryl group may be attached to a cycloalkyl ring or another group at an appropriate position on the aromatic ring.

The term "biphenyl" as used herein refers to two phenyl groups bonded together through any one of the available sites on the phenyl ring. The biphenyl group can be covalently linked to other groups from any available position on the phenyl ring.

The term "heteroaryl" as used herein refers to a monocyclic or bicyclic ring system having up to ten atoms, wherein at least one ring is aromatic and contains 1 to 4 heteroatoms selected from O, N and S. Heteroaryl substituents may be attached through one of the ring carbon atoms or through one of the heteroatoms. Examples of heteroaryl groups include, but are not limited to, thienyl, benzimidazolyl, benzo [ b ] thienyl, furyl, benzofuryl, pyranyl, isobenzofuryl, benzopyranyl, 2H-pyrrolyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, indolizinyl, isoindolyl, 3H-indolyl, indazolyl, purinyl, 4H-quinolizinyl, isoquinolyl, quinolyl, 2, 3-naphthyridinyl, 1, 5-naphthyridinyl, quinoxalyl, quinazolinyl, cinnolinyl, pteridinyl, isothiazolyl, isobenzodihydropyranyl, chromanyl, isoxazolyl, furoryl, indolinyl, isoindolinyl, thiazolo [4, 5-b ] pyridine, and fluorescent derivatives.

The term "heterocycle", "heterocyclic" or "heterocyclyl" as used herein means a 5, 6 or 7 membered non-aromatic ring system containing 1 to 4 heteroatoms selected from O, N and S. Examples of heterocycles include, but are not limited to, pyrrolidinyl, tetrahydrofuranyl, piperidinyl, pyrrolinyl, piperazinyl, imidazolidinyl, morpholinyl, imidazolinyl, pyrazolidinone, pyrazolinyl.

The term "heterobicyclic group" as used herein, refers to a heterocyclic ring as defined above fused to another ring, which may be a heterocyclic ring, an aryl group or any other ring defined herein. Examples of such heterobicyclics include, but are not limited to, coumarin, benzo [ d ] [1, 3] dioxetane, 2, 3-dihydrobenzo [ b ] [1, 4] dioxole, and 3, 4-dihydro-2H-benzo [ b ] [1, 4] dioxole (dioepine).

The present invention encompasses the compounds of the general formula I of the first aspect as well as the compounds of its various preferred or specific embodiments, and also encompasses the various isomers of these compounds. Some of the compounds of the invention may exist in the form of optical isomers or tautomers and the invention includes all the forms in which they exist, in particular the pure isomers. The different isomeric forms may be separated or resolved from the other isomeric forms by conventional means, or an isomer may be obtained by conventional synthetic methods or stereospecific or asymmetric syntheses. Since the compounds of formula I of the present invention are intended for pharmaceutical use, it will be appreciated that they are preferably provided in pure form, for example, at least 60% pure, more suitably at least 75% pure, even more suitably at least 85% pure, and most preferably at least 98% pure (% means weight percent).

The invention also relates to suitable pharmaceutically acceptable salts, solvates or hydrates of the compounds of the invention.

The compounds of the present invention may be used in the form of pharmaceutically acceptable salts derived from inorganic or organic acids. The phrase "pharmaceutically acceptable salt" refers to salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without excessive toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, s.m.berge, et al.j.pharmaceutical Sciences, 1977, 66: pharmaceutically acceptable salts are described in detail in 1. The salts may be prepared in situ during the final isolation and purification of the compounds of the invention or separately by reacting the free base functionality of the compounds of the invention with a suitable organic acid. Pharmaceutically acceptable salts of the compounds of the present invention include, but are not limited to, salts of the compounds of formula I with inorganic acids such as hydrochloric acid, sulfuric acid, phosphoric acid, phosphorous acid, hydrobromic acid, and nitric acid; and salts of the compounds of formula I with various organic acids such as maleic acid, fumaric acid, malic acid, fumaric acid, succinic acid, tartaric acid, citric acid, acetic acid, trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, lactic acid, methanesulfonic acid, p-toluenesulfonic acid, benzoic acid, cinnamic acid, mandelic acid, palmitic acid, salicylic acid, and the like. In addition, pharmaceutically acceptable salts of the compounds of the present invention include, but are not limited to, salts prepared from the compounds of formula I and inorganic bases, such as sodium salts, potassium salts, lithium salts, ammonium salts, calcium salts, magnesium salts, iron salts, zinc salts, copper salts, manganese salts, aluminum salts, and the like; salts derived from organic bases include, but are not limited to, salts of primary, secondary, and tertiary amines, where substituted amines include naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins, and the like. The free base forms of the compounds of the invention differ slightly from their respective salt forms in certain physical properties (such as solubility in polar solvents), but for the purposes of the present invention, each acid salt is equivalent to their respective free base form (see, e.g., s.m. berge, et al., "Pharmaceutical Salts," j.pharm.sci., 66: 1-19(1977), which is incorporated herein by reference).

Some of the compounds of the present invention may be crystallized or recrystallized using water or various organic solvents, in which case various solvates may be formed. The present invention includes those solvates, including hydrates, in stoichiometric amounts, as well as compounds containing variable amounts of water formed when prepared by the low pressure sublimation drying method. In general, for the purposes of the present invention, the solvate forms with pharmaceutically acceptable solvents such as water, ethanol, etc. are equivalent to the non-solvate forms.

The compounds of the invention and their pharmaceutically acceptable salts may also be prodrugs or forms which release the active ingredient after metabolic changes in the body. The selection and preparation of suitable prodrug derivatives is well known to those skilled in the art.

The term "pharmaceutically acceptable carrier" or "excipient" as used herein refers to any adjuvant, carrier, excipient, glidant, sweetener, diluent, preservative, dye/colorant, flavor enhancer, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, solvent, emulsifier or encapsulating agent, such as a liposome, cyclodextrin, encapsulated polymeric delivery system, or polyethylene glycol matrix, which is acceptable for use in a subject, preferably a human.

The present invention relates to the use of compounds of general formula I, all possible isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof for the manufacture of a medicament which can be used for the treatment and/or prevention of associated diseases caused by the overexpression of IAPs. Such diseases include, but are not limited to, the following: bladder cancer, breast cancer, pancreatic cancer, colon cancer, leukemia, lung cancer, lymphoma, multiple myeloma, ovarian cancer and cervical cancer, such as ovarian cancer and cervical cancer, among others.

In another aspect, the compound of formula I of the present invention or a pharmaceutically acceptable salt thereof can be used alone or in combination with a pharmaceutically acceptable carrier or excipient in the form of a pharmaceutical composition, and when used in the form of a pharmaceutical composition, an effective dose of the compound of formula I of the present invention or a pharmaceutically acceptable salt or hydrate thereof and one or more pharmaceutically acceptable carriers or diluents are usually combined to prepare a suitable administration form or dosage form, and such procedure includes mixing, granulating, compressing or dissolving the components by a suitable manner. Accordingly, the present invention provides a pharmaceutical composition comprising a compound of formula I, all possible isomers, prodrugs, pharmaceutically acceptable salts, solvates or hydrates thereof, and at least one pharmaceutically acceptable carrier.

Pharmaceutical compositions of the compounds of the invention may be administered in any of the following ways: oral, aerosol inhalation, rectal, nasal, vaginal, topical, parenteral such as subcutaneous, intravenous, intramuscular, intraperitoneal, intrathecal, intraventricular, intrasternal or intracranial injection or infusion, or by means of an explanted reservoir, with oral, intramuscular, intraperitoneal or intravenous administration being preferred.

The compounds of the present invention or pharmaceutical compositions containing them may be administered in unit dosage form. The administration dosage form can be liquid dosage form or solid dosage form. The liquid dosage forms can be true solutions, colloids, microparticles, emulsions, and suspensions. Other dosage forms such as tablet, capsule, dripping pill, aerosol, pill, powder, solution, suspension, emulsion, granule, suppository, lyophilized powder for injection, clathrate, implant, patch, liniment, etc.

The pharmaceutical composition of the present invention may further comprise conventional carriers, wherein the pharmaceutically acceptable carriers include, but are not limited to: ion exchangers, aluminum oxide, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycerol, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulosic substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, beeswax, lanolin and the like. The carrier may be present in the pharmaceutical composition in an amount of 1% to 98% by weight, typically about 80% by weight. For convenience, the local anesthetic, preservative, buffer, etc. may be dissolved directly in the vehicle.

Oral tablets and capsules may contain excipients such as binding agents, for example syrup, acacia, sorbitol, tragacanth, or polyvinylpyrrolidone, fillers such as lactose, sucrose, corn starch, calcium phosphate, sorbitol, glycine, lubricants such as magnesium stearate, talc, polyethylene glycol, silica, disintegrants such as potato starch, or acceptable wetting agents such as sodium lauryl sulfate. The tablets may be coated by methods known in the art of pharmacy.

Oral liquids may be prepared as suspensions, solutions, emulsions, syrups or elixirs in water and oil, or as dry products, supplemented with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents, sorbitol, cellulose methyl ether, glucose syrup, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, aluminum stearate gelatin, hydrogenated edible fats and oils, emulsifying agents such as lecithin, sorbitan monooleate, gum arabic; or a non-aqueous carrier (which may comprise an edible oil), such as almond oil, an oil such as glycerol, ethylene glycol, or ethanol; preservatives, e.g. methyl or propyl p-hydroxybenzoates, sorbic acid. Flavoring or coloring agents may be added if desired.

Suppositories may contain conventional suppository bases which are solid at room temperature and melt at body temperature to release the drug, such as cocoa butter, other glycerides or beeswax.

For parenteral administration, the liquid dosage forms are generally prepared from the compound and a sterile vehicle. The carrier is preferably water. The compound can be dissolved in the carrier or made into suspension solution according to the concentration of the carrier and the drug, and the compound is firstly dissolved in water when made into the solution for injection, filtered and sterilized and then filled into a sealed bottle or ampoule.

When applied topically to the skin, the compounds of the present invention may be formulated in the form of a suitable ointment, lotion, or cream in which the active ingredient is suspended or dissolved in one or more carriers. Among the vehicles that may be used in the ointment formulation include, but are not limited to: mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyethylene oxide, polypropylene oxide, emulsifying wax and water; lotions and creams may employ carriers including, but not limited to: mineral oil, sorbitan monostearate, tween 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

The term "composition" as used herein is intended to encompass a product comprising the specified ingredients in the specified amounts, as well as any product which results, directly or indirectly, from combination of the specified ingredients in the specified amounts.

The term "therapeutically effective amount" as used herein refers to a dose which, upon administration to the subject, e.g., a mammal, e.g., a human, produces a desired physiological response, particularly a physiological response associated with the diseases described herein. The term "therapeutically effective amount" is also used in a similar sense.

Depending on the mode of administration, the composition may contain 0.1% by weight, or more suitably 10-60% by weight of the active ingredient. However, where the components comprise unit doses, each unit preferably contains from 1 to 500 mg of active ingredient.

It is further noted that the specific dosage and method of administration of the compounds of the present invention for each individual patient will depend upon a variety of factors including the age, body weight, sex, physical condition, nutritional status, the activity level of the compounds, the time of administration, the metabolic rate, the severity of the condition, and the judgment of the attending physician. The dosage is preferably between 0.01 and 100mg/Kg body weight/day.

It will be appreciated that the optimum dosage and interval for administration of a compound of formula I will be determined by the nature of the compound and the external conditions, such as form, route and site of administration, and that such optimum dosage may be determined by conventional techniques. It will also be appreciated that the optimal course of treatment, i.e. the daily dosage of a compound of formula I over a nominal period of time, may be determined by methods well known in the art.

The present invention relates to a process for the preparation of a compound of formula I or a pharmaceutically acceptable salt, solvate or hydrate thereof. In the following general description of the process, the specific definitions of the variables or substituents are as defined for the compounds of the first aspect of the invention.

The compounds of the invention may be formed by coupling a compound represented by formula II:

wherein Z is selected from the group including, but not limited to: -H, -Boc, -Cbz, preferably Z is-Boc.

Preferred but not limiting structures for the compounds of formula II are as follows:

the IIa and IIb structures can be synthesized by directly using commercially available L-proline, 4-S-hydroxy-L-proline and Boc anhydride. The synthesis of IIc, IId, IIe, IIf is via the following scheme 1:

scheme 1

The general procedure for scheme 1 above is: selecting unsubstituted native cysteine, serine, or by R₅、R₅₀₀Substituted native cysteine, serine, by analogous routes as described above, with formaldehyde, acetaldehyde, acetone or with R₄、R₄₀₀And reacting the substituted aldehyde and ketone compounds to obtain the substituted/unsubstituted five-membered heterocyclic alkane with S, O atoms.

Using the Ie compound as an example, scheme 2 below illustrates how the invention proceeds from compounds of formula II to compounds of formula I.

Scheme 2

The general procedure for scheme 2 above is: after the amine fragment 2ii and the acid fragment 2iv are synthesized respectively, the condensation is carried out with IIc in sequence to obtain 2vii, and then the deprotection is carried out to obtain the final product Ie.

Scheme 3 below illustrates how the invention proceeds from compounds of formula II to compounds of formula I, using If compounds as an example.

Scheme 3

The general procedure of scheme 3 above differs from scheme 2 only in that after intermediate 2v is obtained, intermediate 3i is obtained by a suitable reduction method, followed by sequential condensation to give 3iii and deprotection to give If.

The compounds of formula I can be synthesized individually by conventional methods, or in pools (at least two, or 5-1000, preferably 10-100 compounds per pool) by combinatorial chemistry, mixed-split methods or parallel synthesis, either in liquid phase or solid phase. The various starting materials for the reaction are either prepared by the skilled worker on the basis of their knowledge, or can be prepared by methods known from the literature, or are commercially available. The intermediates, starting materials, reagents, reaction conditions, etc. used in the above reaction schemes may be appropriately modified according to the knowledge of those skilled in the art. For more detailed information on the preparation of the compounds of the formula I, reference is made to the detailed description below.

The specific implementation mode is as follows:

the invention is further illustrated by the following specific intermediates and examples, but it should be understood that these intermediates and examples are for illustrative purposes only and are not to be construed as limiting the invention in any way.

The present invention has been described generally and/or specifically with respect to materials used in testing and testing methods. Although it is made for the purpose of achieving the inventionMany materials and methods of operation are known in the art, but the invention is described in as much detail as possible. It will be apparent to those skilled in the art that the materials and methods of operation used in the present invention are well known in the art, unless otherwise specified. The melting point of the compound was determined by a RY-1 melting point apparatus, with no correction by thermometer. Mass spectra were determined on a MicromassZabSpec high resolution mass spectrometer (resolution 1000).¹H-NMR was measured by JNM-ECA-400 superconducting NMR instrument, operating frequency¹H-NMR 400MHz。

In the context of the present invention, the following abbreviations are used:

boc: a tert-butoxycarbonyl group;

cbz: a benzyloxycarbonyl group;

DCM: dichloromethane;

DIPEA: diisopropylethylamine;

DMF: n, N-dimethylformamide;

DCC: cyclohexyl carbodiimide

HOBt: 1-hydroxybenzotriazole;

TEA: triethylamine;

NMF: n-methylmorpholine;

THF: tetrahydrofuran;

ClCOOiBu: isobutyl chloroformate;

LiAlH₄: lithium aluminum hydride;

NaBH₄: sodium borohydride;

TLC: thin layer chromatography.

Intermediate preparation example 1: synthesis of Compounds of formula IIc

The reaction process comprises the following steps:

step 1)

Dissolving 12.1g (0.1mol) of L-cysteine in 60mL of hot water, slowly pouring 10mL of 36% formaldehyde aqueous solution, shaking up, standing overnight, filtering the precipitated crystals the next day, and recrystallizing with ethanol and water to obtain 12.8g of needle-like white crystals, wherein the yield is 96.2%, and the m.p: 196 ℃ and 197 ℃.

Step 2)

In an ice bath, 0.10mol of L-thiazolealkanoic acid was dissolved in 50mL of a 2N aqueous solution (0.10mol) of sodium hydroxide, and 24.4g (0.11mol) of (Boc) was added under stirring₂And slowly dripping the mixed solution of O and 50mL of acetone, and continuously stirring for 2h after the addition is finished. The reaction mixture was diluted with 200mL of water, extracted 3 times with 80mL of 3X ethyl acetate, and the organic phase was discarded. Adjusting pH value of the water phase to 2 with 1mol/L hydrochloric acid under ice bath, extracting with 80mL of multiplied by 3 ethyl acetate for 3 times, combining organic phases, drying with anhydrous sodium sulfate, removing solvent under reduced pressure, and recrystallizing with petroleum ether and ethyl acetate to obtain white crystals.

Intermediate preparation example 2: synthesis of Compounds of formula IId

The reaction process comprises the following steps:

step 1)

L-cysteine (12.2g, 100mmol) was dissolved in 100ml of anhydrous methanol, HCl was introduced until the starting material was completely dissolved, aeration was continued for 2 hours or more, and stirring was continued overnight after the aeration. The next day the solvent was evaporated down under reduced pressure, the residue was taken up in methanol and evaporated down again, repeating 2 times to carry away the HCl. The solvent was evaporated to dryness to give a crude product, which was recrystallized from methanol-ether. The yield thereof was found to be 91%. m.p: 143 ℃ and 145 ℃.

Step 2)

3.5g (20mmol) of the intermediate 1ii and 40ml of acetone are mixed and refluxed for 10 minutes, 10ml of methanol is added to completely dissolve the solid in the system, the mixture is refluxed for 0.5 hour, cooled and crystallized, and the mother liquor is concentrated and crystallized after the crystals are filtered out. Colorless crystals were obtained in 94% yield. m.p: 165 ℃ and 168 ℃.

Step 3)

4.03g (19mmol) of intermediate 1iii mixed with 1.92g TEA were dissolved in 20ml DCM and 4.15g (19mmol) of Boc was added dropwise with stirring₂O in 20ml DCM, reaction for 3h at room temperature, 20ml 10% citric acid, 20ml water washing, spin drying directly for the next step.

Step 4)

Dissolving intermediate 1iv in 25ml methanol, adding 25ml 2N NaOH aqueous solution, reacting at room temperature for 4h, detecting by TLC after the raw material disappears, removing methanol under reduced pressure, diluting with water to 50ml, washing off excess Boc with 20ml × 3 diethyl ether₂And O, adjusting the pH value to 2 by using 1mol/L hydrochloric acid under phase ice bath, precipitating a large amount of white solid, filtering and washing to obtain an intermediate IId.

Intermediate preparation example 3: synthesis of Compounds of formula IIe

The reaction process comprises the following steps:

step 1)

10.5g (100mmol) of L-serine and 10ml (120mmol) of 37% aqueous formaldehyde solution were dissolved in 50ml of an aqueous solution of 2N NaOH, and the mixture was stirred overnight in an ice bath. 24.0g (Boc) in the next day ice bath₂And dripping a solution of O (110mmol) dissolved in 40ml of acetone into the reaction solution, continuously stirring for 1h, diluting the reaction solution with 350ml of water, extracting with 120ml of ethyl acetate for 3 times, discarding an organic phase, adjusting the pH value of an aqueous phase to 2 with HCl, extracting with 120ml of diethyl ether for three times, collecting the organic phase, evaporating to dryness under reduced pressure to obtain IIe, standing overnight to obtain a solid, and recrystallizing with ethyl acetate-petroleum ether to obtain a prismatic crystal. The yield thereof was found to be 94%.

Intermediate preparation example 4: synthesis of Compounds of formula IIf

The reaction process comprises the following steps:

the synthesis of the compounds of the formula IIf can be referred to the synthesis of the compounds of the formula IIe, except that the starting material is exchanged for L-threonine.

Intermediate preparation example 5: synthesis of intermediate 2ii useful for preparing the Compound of example 1

The reaction process comprises the following steps:

step 1)

12.1g (100mmol) of phenethylamine and 11.1g (110mmol) of triethylamine are dissolved in 200mL of dry dichloromethane, a solution of 14.5g (100mmol) of benzoyl chloride in 100mL of dichloromethane is carefully added dropwise in ice bath, the temperature is slowly raised to room temperature after the dropwise addition, and the reaction is carried out for 2 hours. The reaction solution is extracted by ethyl acetate-water partition, and the organic phase is collected and anhydrous Na₂SO₄Drying, decompressing and spin-drying to obtain a white solid, and directly using the white solid for the next reaction after drying without separation.

Step 2)

Carefully add 5.0g (approximately 130mmol) LiAlH in portions to 400mL of dry THF under ice-bath₄After the addition is finished, after the temperature is stabilized to 0 ℃, dropwise adding a solution of the product obtained in the previous step dissolved in 200mL of dry THF into the system, and slowly dropwise adding to ensure that the system releases heat and deflates stably. After dropping, the system was refluxed overnight. The reaction was cooled in an ice bath and saturated ammonium chloride solution was carefully added dropwise to destroy the remaining LiAlH₄. Filtration and the filter cake washed with ethyl acetate until product free. The organic phases are combined, the solvent is removed under reduced pressure, and then the N-phenethylbenzylamine is obtained by column chromatography (the mobile phase is ethyl acetate-petroleum ether with corresponding proportion).

Intermediate preparation example 6: synthesis of intermediate 2iv useful for preparing the Compound of example 1

The reaction process comprises the following steps:

Boc-L-Ala-OH was prepared as Boc-L-thiazoloic acid.

The preparation of 2S-cyclohexyl-L-glycine methyl ester hydrochloride is the same as the preparation of L-cysteine methyl ester hydrochloride.

Step 1)

Controlling the temperature in an ice bath to be below 0 ℃, dissolving 1.89g (10mmol) of Boc-alanine in dichloromethane, carefully adding DCC2.27g, HOBt1.49g, 2.07g (10mmol) of 2S-cyclohexyl-L-glycine methyl ester hydrochloride and triethylamine 1.01g in sequence, reacting overnight at room temperature, filtering to remove white precipitate DCU, washing an organic phase with 10% citric acid, saturated saline water, 4% sodium bicarbonate water solution and saturated saline water in sequence, and washing with anhydrous Na₂SO₄Drying, spin-drying dichloromethane solvent under reduced pressure to obtain light yellow oily substance, adding ethyl acetate solvent, standing at low temperature to precipitate white precipitate DCU, filtering to remove DCU, spin-drying ethyl acetate under reduced pressure, adding ethyl acetate, standing at low temperature for several times to remove most DCU, and performing silica gel column chromatography to obtain white solid 2.73 with yield of 80%.

Step 2)

Dissolving about 2.70g of the intermediate 2iii in 15mL of methanol, adding 10mL of 2N NaOH, reacting at room temperature for 2 hours, detecting the reaction process by TLC, adjusting the pH to be nearly neutral by 1N HCl, removing part of methanol by decompression, adding water for dilution, filtering insoluble impurities, acidifying the filtrate by 1N HCl in an ice bath, immediately precipitating white solid, filtering and collecting, washing by water until the washing solution is nearly neutral, drying, and recrystallizing by methanol-water to obtain 2.05g of colorless needle-shaped crystals with the yield of 79%.

Intermediate preparation example 6: synthesis of intermediate 3ii useful for preparing the Compound of example 1

The reaction process comprises the following steps:

step 1)

Controlling the temperature of an ice salt bath to be below 0 ℃, carefully adding 10.6g (50mmol) of N-phenethylbenzylamine and 7.4g (55mmol) of HOBt into dichloromethane in which 11.5g (50mmol) of Boc-thiazoloic acid is dissolved, carefully dropwise adding 11.3g (55mmol) of dichloromethane solution of DCC, reacting at low temperature for half an hour after dropwise adding, stirring at room temperature for overnight, filtering to remove white precipitate DCU, washing an organic phase with 10% of citric acid, saturated saline water, 4% of sodium bicarbonate water solution and saturated saline water in sequence, and washing with anhydrous Na₂SO₄Drying, spin-drying dichloromethane solvent under reduced pressure to obtain light yellow oily substance, adding ethyl acetate solvent, standing at low temperature to precipitate white precipitate DCU, filtering to remove DCU, spin-drying ethyl acetate under reduced pressure, adding ethyl acetate, standing at low temperature for several times to remove most DCU, and performing silica gel column chromatography to obtain white solid 17.5 with yield of 83%.

Step 2)

N₂Under the protection condition, 12.7g (30mmol) of the raw material in the previous step is dissolved in anhydrous THF, the temperature of an ice salt bath is controlled below 0 ℃, NaBH is carefully added₄2.4g (30mmol), 3.81g I was carefully added dropwise after half an hour₂After the addition, the reaction was continued for 2 hours, and the mixture was refluxed for 48 hours, and the progress of the reaction was checked by TLC.

After the reaction was complete, saturated NH was carefully added₄Discharging no bubble at 50 deg.C in Cl aqueous solution, dissolving insoluble precipitate with 2N NaOH, extracting water layer with methyl tert-butyl ether, mixing organic phases, and anhydrous Na₂SO₄Drying, silica gel column chromatography to obtain white powdery intermediate 3i about 9.2g, yield 75%.

Step 3)

Controlling the temperature of the ice salt bath below 0 ℃, and dissolving 9.18g of the white solid obtained in the previous step in a small amount of CH₂Cl₂50mL of ice-cold 4N HCl/EtOAc solution are carefully added and the reaction is carried out at 0-4 ℃ for 2 hoursA large amount of white precipitate was precipitated, and the intermediate 3ii was filtered off to obtain about 7.6g, i.e., a yield of 97%.

Example 1

Reaction scheme

Step 1)

Intermediate 2iv 328mg was dissolved in 5mL of anhydrous THF with temperature controlled below-15 deg.C, and 110. mu. L N-methylmorpholine was added carefully followed by 140. mu.L of isobutyl chloroformate. Half an hour later, intermediate 3ii 348mg and 110. mu. L N-methylmorpholine in 3mL DMF were added and stirring continued at-15 ℃ for half an hour, reaction at room temperature for 2h, and solvent was removed by concentration under reduced pressure. The residue was dissolved in 25mL ethyl acetate and washed with 5% NaHCO₃Water, 1NHCl and water 25mL each, followed by anhydrous Na₂SO₄Drying and silica gel column chromatography gave 3iii321mg as a white solid in 48% yield.

Step 2)

Dissolving 440mg of the intermediate 3iii in dichloromethane, controlling the temperature below 0 ℃ in an ice bath, adding 10mL of ice-cold 2N HCl/EtOAc solution, reacting at low temperature for half an hour, continuously stirring at room temperature for about 2 hours, precipitating a white solid, detecting the reaction completion by TLC, filtering and collecting 215mg of the white solid, wherein the yield is 98%.

The total yield of the two steps is 47 percent. All the following examples describe yields which refer to the total yield of the two steps of condensation and deprotection.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.13-1.17(8H，m，(CH₂-CH₂)×2)，1.28-1.30(2H，m，CH₂)，1.72(3H，d，J＝6.8Hz，CH₃)，2.0(1H，m，CH)，2.3-.25(2H，d，J＝4.6Hz，CH₂)，2.6(2H，t，J＝2Hz，CH₂)，2.6-2.9(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝7.2Hz，CH₂)，3.6(2H，s，CH₂)，3.59(2H，d，J＝7.6Hz，CH₂)，3.89-3.92(1H，m，CH×2)，4.10(1H，d，J＝4.4Hz，CH)，4.86(1H，s，NH)，7.2-7.5(10H，m，ArH)，8.06(2H，t，J＝7.6Hz，NH₂)，8.16(2H，s，HCl)，MS m/e：523.5([M+1]⁺).

Example 2

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 67%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.18(3H，d，J＝6.8Hz，CH3)，2.6-2.9(2H，d，J＝4.8Hz，CH2)，2.8(2H，t，J＝2Hz，CH2)，3.5(2H，t，J＝7.6Hz，CH2)，3.6-3.7(2H，d，J＝7.6Hz，CH2)，3.76(2H，d，J＝7.6Hz，CH2)，3.8(1H，m，CH)，3.99(1H，m，CH)，4.5(2H，s，CH2)，6.96(1H，s，NH)，7.2-7.5(10H，m，ArH)8.06(2H，t，J＝7.6Hz，NH2)，8.6(1H，s，HCl)，MSm/e：455.2([M+1]⁺).

Example 3

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (R) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and using 326mg of (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide instead of 3ii, yield 35%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.85(3H，d，J＝6.8Hz，CH₃)，1.20-1.38(8H，m，(CH₂-CH₂)×2)，1.58-1.62(2H，m，CH₂)，2.79(2H，d，J＝4.8Hz，CH₂)，2.90-2.92(1H，m，CH)，3.43(2H，t，J＝2Hz，CH₂)，3.5(2H，t，J＝7.6Hz，CH₂)，3.63(2H，d，J＝7.6Hz，CH₂)，3.9(1H，s，CH)，4.11-4.12(1H，m，CH)，4.36(2H，s，CH₂)，4.75(1H，d，J＝4.4Hz，CH)，5.08(1H，s，NH)，7.28-7.32(10H，m，ArH)，8.25(2H，t，J＝7.6Hz，NH₂)，8.78(1H，s，HCl)，MSm/e：537.3([M+1]⁺).

Example 4

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and (R) -N-phenethylthiazolidine-4-carboxamide 236mg was used instead of 3ii, yield 28%.

¹H-NMR(400 MHz，DMSO-d₆)，(ppm)：0.86(6H，d，J＝6.4Hz，CH₃×2)，1.18(3H，d，J＝6.8Hz，CH₃)，1.96-1.99(1H，m，CH)，2.69(2H，d，J＝4.8Hz，CH₂)，3.67(2H，d，J＝7.6Hz，CH₂)，3.88-3.92(1H，m，CH)，4.12(1H，d，J＝4.4Hz，CH)，4.2(2H，d，J＝5.4Hz，CH₂)，4.96(1H，s，NH)，7.25-7.28(5H，m，ArH)，7.5(1H，t，J＝6.5Hz，NH)，8.86(2H，t，J＝7.6Hz，NH₂)，9.4(1H，s，HCl)，MS m/e：407.5([M+1]⁺).

Example 5

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and (R) -N-phenethyl-N- (thiazolidin-4-yl-methyl) benzamide 326mg was used instead of 3ii, yield 36%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.86(6H，d，J＝6.4Hz，CH₃×2)，1.18(3H，d，J＝6.8Hz，CH₃)，1.96-1.99(1H，m，CH₂)，2.72(2H，s，CH₂)，3.12-3.15(2H，m，CH₂)，4.55(2H，d，J＝5.4Hz，CH₂)，3.89-3.91(2H，m，CH×2)，4.21(1H，d，J＝4.4Hz，CH)，4.96(1H，s，NH)，7.46-7.49(10H，m，ArH)，8.06(2H，t，J＝7.6Hz，NH₂)，9.40(1H，s，HCl)，MS m/e：483.2([M+1]⁺).

Example 6

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and using 326mg of (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide instead of 3ii, yield 49%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.83-0.85(2H，m，CH₂)，1.28-1.30(8H，m，(CH₂-CH₂)×2)，1.91(3H，d，J＝6.8Hz，CH₃)，2.6(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝2Hz，CH₂)，2.94-2.96(1H，m，CH)，3.2(2H，t，J＝7.6Hz，CH₂)，3.49-3.51(2H，d，J＝7.6Hz，CH₂)，4.36-4.39(2H，m，CH×3)，4.5(2H，s，CH₂)，6.96(1H，s，NH)，7.2-7.5(10H，m，ArH)，8.22(2H，t，J＝7.6Hz，NH₂)，8.72(1H，s，HCl)，MS m/e：537.3([M+1]⁺).

Example 7

The compound of this example was prepared by reference to the synthesis of example 1 except that 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and 222mg of (R) -N-benzylthiazolidine-4-carboxamide was used instead of 3ii, yield 52%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.91(6H，d，J＝6.4Hz，CH₃×2)，1.19(3H，d，J＝6.8Hz，CH₃)，1.38(3H，s，CH₃)，1.98-2.10(1H，m，CH)，2.5(2H，d，J＝4.8Hz，CH₂)，3.16(2H，d，J＝7.6Hz，CH₂)，3.35-3.36(2H，m，CH×2)，3.9(1H，d，J＝4.4Hz，CH)，4.2(2H，d，J＝5.4Hz，CH₂)，4.5(1H，s，NH)，7.5(1H，t，J＝16Hz，NH)，7.28-7.32(5H，m，ArH)，8.06(1H，t，J＝7.6Hz，NH)，9.36(1H，s，HCl)，MS m/e：407.3([M+1]⁺).

Example 8

The compound of this example was prepared by reference to the synthesis of example 1 except using 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-tert-butyl-acetic acid instead of 2iv and using 326mg of (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide instead of 3ii, yield 49%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.05(9H，d，J＝6.4Hz，CH₃×3)，1.18(3H，d，J＝6.8Hz，CH₃)，1.33-1.34(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝2Hz，CH₂)，2.89-2.91(2H，m，CH×2)，3.5(2H，t，J＝10.4Hz，CH₂)，3.79(2H，d，J＝7.6Hz，CH₂)，4.1(1H，d，J＝4.4Hz，CH)，4.36(2H，s，CH₂)，5.13(1H，s，NH)，7.26-7.35(10H，m，ArH)，8.06(2H，t，J＝7.6Hz，NH₂)，8.62(1H，s，HCl)，MS m/e：511.4([M+1]⁺).

Example 9

The compound of this example was prepared by reference to the synthesis of example 1 except that 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2 iv. yield 31%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.91(6H，d，J＝6.4Hz，CH₃×2)，1.36(3H，d，J＝6.8Hz，CH₃)，1.9(1H，m，CH)，1.99(2H，d，J＝1.6Hz，CH₂)，2.51(2H，t，J＝2Hz，CH₂)，3.09(2H，d，J＝4.8Hz，CH₂)，3.12(2H，t，J＝7.2Hz，CH₂)，3.19(3H，s，CH₃)，3.21(1H，s，NH)，3.6(2H，s，CH₂)，3.39-3.41(1H，m，CH×2)，4.90(1H，d，J＝4.4Hz，CH)，4.39(2H，d，J＝7.6Hz，CH₂)，7.2-7.5(10H，m，ArH)，7.78(1H，t，J＝7.6Hz，NH)，8.92(2H，s，HCl×2)，MS m/e：497.4([M+1]⁺).

Example 10

The compound of this example was prepared by reference to the synthesis of example 1 except that 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-tert-butyl-acetic acid was used instead of 2iv and (R) -N-phenethylthiazolidine-4-carboxamide 236mg was used instead of 3ii, yield 50%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.01(9H，d，J＝6.8Hz，CH₃×3)，1.18(3H，d，J＝6.8Hz，CH₃)，1.28(2H，d，J＝7.6Hz，CH₂)，2.69(2H，t，J＝2Hz，CH₂)，2.73-2.75(2H，d，J＝4.8Hz，CH₂)，3.23(2H，d，J＝4.4Hz，CH₂)，4.11-4.12(2H，m，CH×2)，4.24(1H，t，J＝16Hz，NH)，4.26(1H，d，J＝4.4Hz，CH)，4.53(1H，s，NH)，7.21-7.25(5H，m，ArH)，8.24(2H，t，J＝7.6Hz，NH₂)，8.46(1H，s，HCl)，MS m/e：421.2([M+1]⁺).

Example 11

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and using 236mg of (R) -N-phenethylthiazolidine-4-carboxamide instead of 3ii, yield 49%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.16-1.18(8H，m，(CH₂-CH₂)×2)，1.30(3H，d，J＝6.8Hz，CH₃)，1.68-1.71(2H，m，CH₂)，1.68-1.71(1H，m，CH)，2.68(2H，t，J＝2Hz，CH₂)，2.86(2H，d，J＝4.8Hz，CH₂)，3.20-3.22(2H，m，CH₂)，3.57(2H，d，J＝7.6Hz，CH₂)，3.76-3.89(2H，m，CH×2)，4.45(1H，d，J＝4.4Hz，CH)，4.56(1H，s，NH)，7.22-7.29(5H，m，ArH)，8.18(1H，t，J＝16Hz，NH)，8.27(2H，t，J＝7.6Hz，NH₂)，8.62(1H，s，HCl)，MS m/e：447.3([M+1]⁺).

Example 12

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2 iv. yield 45%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.91(6H，d，J＝6.4Hz，CH₃×2)，1.32(3H，d，J＝6.8Hz，CH₃)，1.91(2H，d，J＝1.6Hz，CH₂)，2.01-2.03(1H，m，CH)，3.09(2H，t，J＝2Hz，CH₂)，3.20(2H，t，J＝7.2Hz，CH₂)，3.42(2H，d，J＝4.8Hz，CH₂)，3.93(1H，d，J＝4.4Hz，CH)，4.33(2H，s，CH₂)，4.41(2H，d，J＝7.6Hz，CH₂)，4.87-4.89(2H，m，CH×2)，7.78(1H，s，NH)，7.29-7.46(10H，m，ArH)，8.29(2H，s，2HCl)，8.69(2H，t，J＝7.6Hz，NH₂)，MS m/e：483.3([M+1]⁺).

Example 13

The compound of this example was prepared by reference to the synthesis of example 1 except that 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and (R) -N-phenethylthiazolidine-4-carboxamide 236mg was used instead of 3ii, yield 52%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.91(6H，d，J＝7.2Hz，CH₃×2)，1.19(3H，d，J＝6.8Hz，CH₃)，2.10-2.11(1H，m，CH)，2.75(2H，t，J＝2Hz，CH₂)，2.89-2.91(2H，d，J＝4.8Hz，CH₂)，3.34-3.35(2H，m，CH₂)，3.9(3H，s，CH₃)，4.15(2H，d，J＝7.6Hz，CH₂)，4.25-4.27(2H，m，CH×2)，4.56(1H，d，J＝4.4Hz，CH)，5.03(1H，s，NH)，7.23-7.28(5H，m，ArH)，8.82(1H，t，J＝7.6Hz，NH)，9.44(1H，t，NH)，MS m/e：421.5([M+1]⁺).

Example 14

The compound of this example was prepared by reference to the synthesis of example 1 except that 342mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-cyclohexyl-acetic acid was used instead of 2iv and (R) -N-phenethylthiazolidine-4-carboxamide 236mg was used instead of 3 ii.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.85(3H，d，J＝6.8Hz，CH₃)，1.13-1.24(8H，m，(CH₂-CH₂)×2)，1.32-1.34(2H，m，CH₂)，1.68(3H，s，CH₃)，2.0(1H，m，CH)，2.6(2H，t，J＝2Hz，CH₂)，2.83(2H，d，J＝4.8Hz，CH₂)，3.2(2H，m，CH₂)，3.34(2H，d，J＝7.6Hz，CH₂)，3.83(1H，d，J＝4.4Hz，CH)，4.54-4.63(2H，m，CH×2)，5.07(1H，s，NH)，7.22-7.29(5H，m，ArH)，8.13(1H，t，J＝16Hz，NH)，8.81(1H，t，J＝7.6Hz，NH)，9.25(1H，s，HCl)，MSm/e：461.3([M+1]⁺).

Example 15

The compound of this example was prepared by reference to the synthesis of example 1 except that 342mg of (R) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-cyclohexyl-acetic acid was used instead of 2iv and (R) -N-phenethylthiazolidine-4-carboxamide 236mg was used instead of 3ii, yield 53%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.83(3H，d，J＝6.8Hz，CH₃)，1.07-1.12(8H，m，(CH₂-CH₂)×2)，1.38-1.40(2H，m，CH₂)，2.50(2H，t，J＝2Hz，CH₂)，2.69-2.71(1H，m，CH)，2.9(2H，d，J＝4.8Hz，CH₂)，3.24-3.29(2H，m，CH₂)，3.29(3H，s，CH₃)，3.6-3.7(2H，d，J＝7.6Hz，CH₂)，3.85-3.87(2H，m，CH×2)，3.91(1H，d，J＝4.4Hz，CH)，4.61(1H，s，NH)，7.20-7.29(5H，m，ArH)，7.9(1H，t，J＝16Hz，NH)，8.97(1H，t，J＝7.6Hz，NH)，9.26(1H，s，HCl)，MS m/e：461.3([M+1]⁺).

Example 15

The compound of this example was prepared by reference to the synthesis of example 1 except using 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-isopropyl-acetic acid instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide 326mg instead of 3ii, yield 43%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.91(6H，d，J＝7.2Hz，CH₃×2)，1.35(3H，d，J＝6.8Hz，CH₃)，2.51(3H，s，CH₃)，2.06-2.08(1H，m，CH)，2.83(2H，d，J＝4.8Hz，CH₂)，2.9(2H，d，J＝7.6Hz，CH₂)，3.54(2H，t，J＝2.1Hz，CH₂)，3.89-3.92(1H，m，CH×2)，4.1(1H，d，J＝4.4Hz，CH)，4.56(2H，t，J＝7.6Hz，CH₂)，4.89(2H，s，CH₂)，5.18(1H，s，NH)，7.2-7.5(10H，m，ArH)，8.06(1H，t，J＝7.6Hz，NH)，MS m/e：511.3([M+1]⁺).

Example 17

The compound of this example was prepared by reference to the synthesis of example 1 except using 316mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-tert-butyl-acetic acid instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide 326mg instead of 3ii, yield 36%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.85(2H，d，J＝4.8Hz，CH₂)，1.19(9H，d，J＝6.8Hz，CH₃×3)，1.38(3H，d，J＝6.8Hz，CH₃)，1.59(3H，s，CH₃)，2.50(2H，t，J＝2.0Hz，CH₂)，2.73(2H，t，J＝7.6Hz，CH₂)，2.89(2H，d，J＝7.6Hz，CH₂)，3.53-3.57(1H，m，CH×2)，4.13(1H，d，J＝4.4Hz，CH)，4.36(2H，s，CH₂)，5.12(1H，s，NH)，7.29-7.32(10H，m，ArH)，8.93(1H，t，J＝7.6Hz，NH)，9.40(1H，s，HCl)，MS m/e：525.5([M+1]⁺).

Example 18

The compound of this example was prepared by reference to the synthesis of example 1 except that 342mg of (R) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-cyclohexyl-acetic acid was used instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide was used instead of 3ii, yield 46%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.93(3H，d，J＝6.8Hz，CH₃)，1.25-1.29(8H，m，(CH₂-CH₂)×2)，1.56-1.59(2H，m，CH₂)，1.89(1H，s，CH)，2.6(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝2.0Hz，CH₂)，3.4(3H，s，CH₃)，3.5(2H，t，J＝7.6Hz，CH₂)，3.76(2H，d，J＝7.6Hz，CH₂)，3.89-3.92(1H，m，CH×2)，4.36(1H，d，J＝4.4Hz，CH)，4.57(2H，s，CH₂)，4.96(1H，s，NH)，7.28-7.35(10H，m，ArH)，8.26(1H，t，J＝7.6Hz，NH)，9.40(1H，s，HCl)，MSm/e：551.7([M+1]⁺).

Example 19

The compound of this example was prepared by reference to the synthesis of example 1 except using 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-tert-butyl-acetic acid instead of 2iv and 222mg of (R) -N-benzylthiazolidine-4-carboxamide instead of 3ii, yield 56%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.01(9H，d，J＝6.8Hz，CH₃×3)，1.18(3H，d，J＝6.8Hz，CH₃)，1.29(2H，d，J＝4.8Hz，CH₂)，3.02(2H，d，J＝7.6Hz，CH₂)，3.35-3.46(1H，m，CH×2)，3.02(1H，d，J＝4.4Hz，CH)，4.16-4.19(2H，m，CH₂)，4.36(1H，s，NH)，7.5-7.8(5H，m，ArH)，8.2(1H，t，J＝16Hz，NH)，8.49(2H，t，J＝7.6Hz，NH₂)，9.21(1H，s，HCl)，MSm/e：407.2([M+1]⁺).

Example 20

The compound of this example was prepared by referring to the synthesis method of example 1, except that 274mg of (S) -2- (2- (tert-butoxycarbonylamino) acetylamino) -3-methylbutyric acid was used instead of 2 iv. The yield thereof was found to be 41%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.86(6H，d，J＝6.4Hz，CH₃×2)，1.96-1.98(1H，m，CH)，2.35(2H，d，J＝1.6Hz，CH₂)，2.6(2H，t，J＝2Hz，CH₂)，2.67(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝7.2Hz，CH₂)，3.6(2H，s，CH₂)，3.67(2H，d，J＝6.4Hz，CH₂)，3.86-3.90(1H，m，CH)，4.1(1H，d，J＝4.4Hz，CH)，4.96(1H，s，NH)，7.2-7.5(10H，m，ArH)，8.7(2H，t，J＝7.6Hz，NH₂)，9.40(1H，s，HCl)，MS m/e：469.3([M+1]⁺).

Example 21

The compound of this example was prepared by reference to the synthesis of example 1 except that 302mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-tert-butyl-acetic acid was used instead of 2 iv.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.94(9H，d，J＝6.4Hz，CH₃×3)，1.3(3H，d，J＝6.8Hz，CH₃)，1.9(2H，d，J＝1.6Hz，CH₂)，2.87(2H，t，J＝2Hz，CH₂)，3.11(2H，d，J＝4.8Hz，CH₂)，3.25(2H，t，J＝7.2Hz，CH₂)，3.36(2H，d，J＝7.6Hz，CH₂)，3.48(2H，s，CH₂)，4.02-4.03(1H，m，CH×2)，4.42(1H，d，J＝4.4Hz，CH)，4.68(1H，s，NH)，7.27-7.31(10H，m，ArH)，7.75(2H，t，J＝7.6Hz，NH₂)，8.25(2H，s，2HCl)，MS m/e：497.4([M+1]⁺).

Example 22

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and using 236mg of (R) -N-phenethylthiazolidine-4-carboxamide instead of 3ii, yield 47%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.14-1.16(8H，m，(CH₂-CH₂)×2)，1.36(3H，d，J＝6.8Hz，CH₃)，1.65-1.68(4H，m，CH₂×2)，2.0(1H，m，CH)，2.6(2H，t，J＝2Hz，CH₂)，2.75(2H，d，J＝7.6Hz，CH₂)，3.25-3.26(2H，m，CH₂)，3.91-3.93(2H，m，CH×2)，4.1(1H，d，J＝4.4Hz，CH)，4.3(1H，t，J＝7.2Hz，NH)，4.71(1H，s，NH)，7.22-7.29(5H，m，ArH)，8.06(2H，t，J＝7.6Hz，NH₂)，8.76(1H，s，HCl)，MS m/e：447.2([M+1]⁺).

Example 23

The compound of this example was prepared by referring to the synthesis method of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2 iv. The yield thereof was found to be 46%.

1H-NMR(400MHz，DMSO-d6)，(ppm)：1.39(3H，d，J＝6.8Hz，CH₃)，1.91(2H，d，J＝1.6Hz，CH₂)，3.10-3.39(6H，m，CH₂×3)，，3.60(2H，s，CH₂)，3.94(2H，t，J＝6.3Hz，CH₂)，4.11(2H，s，CH₂)，4.50-4.60(3H，m，CH，CH₂)，4.99(1H，s，CH)，7.26-7.71(10H，m，ArH)，8.25(2H，t，J＝7.6Hz，HCl.NH₂)，8.75(1H，s，CONH)，MS m/e：441.5([M+1]⁺).

Example 24

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and 236mg of (R) -N-phenethylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 40%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.18(3H，d，J＝6.8Hz，CH₃)，2.6(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝2.0Hz，CH₂)，3.4(3H，s，CH₃)，3.5(2H，t，J＝7.2Hz，CH₂)，3.65(2H，d，J＝7.6Hz，CH₂)，3.76-3.79(2H，m，CH₂)，3.89-3.92(1H，m，CH×2)，4.5(2H，s，CH₂)，6.96(1H，s，NH)，7.25-7.29(10H，m，ArH)，8.76(1H，t，J＝7.6Hz，NH)，9.40(1H，s，HCl)，MSm/e：365.3([M+1]⁺).

Example 25

The compound of this example was prepared by reference to the synthesis of example 1 except that 342mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylmethylamino) propionylamino) -2-cyclohexyl-acetic acid was used instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide was used instead of 3ii, yield 44%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.18-1.19(8H，m，(CH₂-CH₂)×2)，1.33(3H，d，J＝6.8Hz，CH₃)，1.39-1.41(2H，m，CH₂)，2.0(1H，m，CH)，2.51(2H，d，J＝4.8Hz，CH₂)，2.8(2H，t，J＝2.0Hz，CH₂)，2.95(2H，d，J＝7.6Hz，CH₂)，3.4(3H，s，CH₃)，3.5(2H，t，J＝7.2Hz，CH₂)，3.51-3.53(1H，m，CH)，4.1(1H，d，J＝4.4Hz，CH)，4.38(2H，s，CH₂)，4.68(1H，s，NH)，7.28-7.30(10H，m，ArH)，8.89(1H，t，J＝7.6Hz，NH)，9.31(1H，br，HCl)，MS m/e：551.5([M+1]⁺).

Example 26

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and 264mg of (R) -N-benzyl-N-propylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 35%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.84-0.86(3H，t，J＝7.2Hz，CH₃)，1.37-1.41(5H，m，CH₂，CH₃)，2.91-2.93(2H，m，CH₂)，3.36-3.37(3H，m，CH₂，CH)，4.30-4.34(2H，m，CH₂)，4.55-4.66(2H，m，CH₂)，4.66-4.93(1H，m，CH)，5.17-5.18(2H，s，CH₂)，7.21-7.40(5H，m，C₆H₅)，8.12-8.16(2H，br，J＝15.6Hz，NH₂)，8.642-8.671(1H，m，CONH)，MS m/e：393.1[M+H]⁺。

Example 27

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and 264mg of (R) -N-benzyl-N-propylthiazolidine-4-carboxamide was used instead of 3ii, 32% yield.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.78-1.42(14H，m，CH₂，CH₃×4)，2.07(1H，m，CH)，2.95-3.66(3H，t，CH₂，CH)，3.19-3.32(2H，m，CH₂)，4.23-4.26(1H，m，CH)，4.38-4.51(3H，m，CH₂，CH)，4.81-4.82(2H，s，CH₂)，7.26-7.34(5H，m，C₆H₅)，8.30(2H，br，NH₂)，8.58(1H，s，CONH)，MS m/e：435.0[M+H]⁺。

Example 28

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and (R) -N-benzyl-N-butylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 48%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.81-0.84(3H，t，J＝5.6Hz，CH₃)，1.09-1.63(7H，m，CH₂×2，CH₃)，2.89-2.91(1H，-H)，3.22-3.39(5H，m，CH₂×2，CH)，3.92-4.03(2H，m，CH₂)，4.302-4.341(2H，m，CH₂)，4.68-5.21(2H，m，CH₂)，7.20-7.40(5H，m，C₆H₅)，8.252(2H，br，NH₂)，8.69(1H，s，CONH)，MS m/e：407.0[M+H]⁺。

Example 29

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and (R) -N-benzyl-N-butylthiazolidine-4-carboxamide 276mg was used instead of 3ii, yield 52%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.84-0.91(9H，m，CH₃×3)，1.17-1.30(7H，m，CH₂×2，CH₃)，1.99-2.06(1H，m，CH)，2.954-2.983(1H，m，CH)，3.32-3.35(4H，m，CH₂×2)，3.59-3.60(1H，m，CH)，4.50-4.54(3H，m，CH₂，CH)，5.13-5.18(2H，m，CH₂)，7.21-7.40(5H，m，C₆H₅)，8.22(2H，br，NH₂)，8.68-8.70(1H，m，CONH)，MS m/e：449.1[M+H]⁺。

Example 30

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and 288mg of (R) -N-benzyl-N-pentylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 39%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.81-0.84(3H，t，J＝6.8Hz，CH₃)，1.09-1.66(9H，m，CH₂×3，CH₃)，2.91-2.94(1H，m，CH)，3.37-3.39(4H，m，CH₂×2)，3.93-4.03(2H，m，CH₂)，4.55-4.57(2H，m，CH₂)，4.93-4.95(1H，m，CH)，5.12-5.22(2H，m，CH₂)，7.21-7.40(5H，m，C₆H₅)，8.10-8.26(2H，br，NH₂)，8.69(1H，s，CONH)，MSm/e：421.1[M+H]⁺。

Example 31

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and 288mg of (R) -N-benzyl-N-pentylthiazolidine-4-carboxamide was used instead of 3ii, the yield was 21%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.81-0.93(9H，m，CH₃×3)，1.18-1.32(9H，m，CH₂×4，-CH₃)，2.06-2.08(1H，m，CH)，2.951-2.980(1H，m，CH₃)，3.32-3.39(4H，m，CH₂×2)，4.49-4.55(3H，m，CH，CH₂)，4.81-4.85(1H，m，CH)，5.12-5.18(2H，m，CH₂)，7.21-7.40(5H，m，C₆H₅)，8.23(2H，br，NH₂)，8.67-8.68(1H，s，CONH)，MS m/e：463.1[M+H]⁺。

Example 32

The compound of this example was prepared by referring to the synthesis procedure of example 1, except that 246mg of (S) -2- (2- (tert-butoxycarbonylamino) propionylamino) acetic acid was used instead of 2iv and 304mg of (R) -N-benzyl-N-cyclohexylthiazolidine-4-carboxamide was used instead of 3 ii. The yield thereof was found to be 36%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：1.15-1.66(13H，m，CH₃，CH₂×5)，3.01-3.34(1H，m，CH)，3.63(1H，m，CH)，4.01-4.03(4H，m，CH₂×2)，4.40-4.56(3H，m，CH₂，CH)，4.93-4.95(2H，m，CH₂)，7.17-7.25(5H，m，C₆H₅)，8.23(2H，br，NH₂)，8.67(1H，s，CONH)，MS m/e：433.1[M+H]⁺。

Example 33

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and 304mg of (R) -N-benzyl-N-cyclohexylthiazolidine-4-carboxamide was used instead of 3ii, the yield was 27%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.89-2.07(19H，m，CH₂×5，CH₃×3)，2.05-2.07(1H，m，CH)，2.98-3.01(1H，m，CH)，3.39-3.40(3H，m，CH₂，CH)，3.62-3.63(1H，m，CH)，4.48-4.60(3H，m，CH₂，CH)，5.17-5.19(2H，m，CH₂)，7.18-7.46(5H，m，C₆H₅)，8.22(2H，br，NH₂)，8.61-8.69(1H，s，CONH)，MS m/e：475.3[M+H]⁺。

Example 34

The compound of this example was prepared by reference to the synthesis of example 1 except that 288mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-isopropyl-acetic acid was used instead of 2iv and (R) -N-benzyl-N-phenethylthiazolidine-4-carboxamide was used instead of 3ii, the yield was 31.5%.

¹H-NMR(400MHz，DMSO-d₆)，ppm：0.91-0.93(6H，m，CH₃×2)，1.30-1.32(3H，m，CH₃)，2.05-2.06(1H，m，CH)，2.82-2.96(4H，m，CH₂×2)，3.31-3.36(1H，m，CH)，3.36(2H，m，CH₂)，4.01-4.03(1H，m，CH)，4.57-4.59(3H，m，CH₂，CH)，5.10-5.18(2H，m，CH₂)，7.26-7.31(10H，m，C₆H₅×2)，8.10(2H，br，NH₂)，8.66-8.68(1H，d，J＝8.4Hz，CONH)，MS m/e：497.1[M+H]⁺。

Example 35

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (S) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and 310mg of (R) -N-benzyl-N-phenethyloxazolidine-4-carboxamide instead of 3ii, yield 42%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.85-1.15(5H，m，CH₂×2，CH)，1.15(3H，d，J＝6.8Hz，CH₃)，1.55-1.66(6H，m，CH₂×3)，2.65-2.75(1H，m，CH)，2.80-2.99(1H，m，CH)，3.40-3.51(2H，m，CH₂)，3.60-3.71(2H，m，CH₂)，3.97(1H，t，J＝7.2Hz，CH)，4.27-4.33(2H，m，CH₂)，4.58-4.68(1H，m，CH)，4.89-4.98(1H，m，CH)，5.08(1H，t，J＝5.6Hz，CH)，7.09-7.35(10H，m，ArH)，7.52(1H，d，J＝9.2Hz，CONH)，8.41(3H，br，HCl.NH₂)，MS m/e：521.6([M+1]⁺).

Example 36

The compound of this example was prepared by reference to the synthesis of example 1 except using 328mg of (R) -2- ((S) -2- (2- (tert-butoxycarbonylamino) propionylamino) -2-cyclohexyl-acetic acid instead of 2iv and 310mg of (R) -N-benzyl-N-phenethyloxazolidine-4-carboxamide instead of 3ii, yield 47%.

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：0.90-1.18(5H，m，CH₂×2，CH)，1.37(3H，d，J＝6.8Hz，CH₃)，1.55-1.68(6H，m，CH₂×3)，2.66-2.75(1H，m，CH)，2.88-3.00(1H，m，CH)，3.40-3.50(2H，m，CH₂)，3.60-3.70(2H，m，CH₂)，3.93(1H，t，J＝7.2Hz，CH)，4.32-4.49(2H，m，CH₂)，4.67-4.78(1H，m，CH)，4.98-5.05(1H，m，CH)，5.17(1H，t，J＝5.6Hz，CH)，7.10-7.40(10H，m，ArH)，8.18(3H，br，HCl.NH₂)，8.59(1H，d，J＝9.2Hz，CONH)，MS m/e：521.6([M+1]⁺).

Example 37

The compound of this example was prepared by reference to the synthesis of example 1, except that 328mg of (2S, 3S) -2- ((S) -2- (tert-butoxycarbonylamino) propionylamino) -3- (propyl-2-ynyloxy) butanoic acid was used instead of 2iv and 236mg of (R) -N-p-methylbenzylthiazolidine-4-carboxamide was used instead of 3 ii. The yield is 35%

¹H-NMR(400MHz，DMSO-d₆)，(ppm)：1.16(3H，d，J＝5.6Hz，CH₃)，1.31(3H，d，J＝6.8Hz，CH₃)，2.27(3H，s，CH₃)，3.01-3.05(1H，m，CH)，3.31-3.44(3H，m，CH₂，CH)，3.88-3.91(1H，m，CH)，3.97(1H，s，CH)，4.18-4.25(4H，m，CH₂×2)，4.64(1H，d，J＝9.2Hz，CH)，4.73(2H，t，J＝6.8Hz，CH₂)，5.08(1H，d，J＝9.2Hz，CH)，7.11-7.15(4H，m，Ar-H)，8.18(3H，br，HCl.NH2)，8.35(1H，s，CONH)，8.71(1H，d，J＝5.6Hz，CONH)MS m/e：447.1([M+1]⁺).

Pharmacological test example 1: XIAP-BIR3 and cIAP1-BIR3 inhibitory Activity assay

A fixed concentration of fluorescent tracer (SM-F2, IAPs inhibitor) was mixed with the target protein, with increasing protein concentration until saturation. The equilibrium dissociation constants (K) for SM-F2 binding to XIAP-BIR3 (residues 241-356) and cIAP1-BIR3 (residues 253-363), respectively, were determined by monitoring the total fluorescence polarization intensity of the mixture_d). Fluorescence polarization values were determined in 96-well fluorescence detection microplates (Mierofluor 2, black, very low background, round bottom) (Thermo Scientific) using a microplate reader (tecanu.s., Research Triangle Park, NC). To each well of assay buffer (containing 100mM dipotassium phosphate, pH 7.5, 100. mu.g/ml bovine gamma-globulin, 0.02% sodium azide, 4% DMSO, Invitrogen) was added SM-F2 and the corresponding target protein at increasing concentrations, 2nM (for binding to XIAP-BIR 3) and 1nM (for binding to cIAP1-BIR 3), respectively, in a final volume of 125. mu.L. The 96-well plate was incubated at room temperature for 3 hours with gentle shaking to ensure equilibrium was reached. The polarization was measured at an excitation wavelength of 485nm and an emission wavelength of 530nm, and the measured values were expressed in millipolarization units (mP). Equilibrium dissociation constant (K) was calculated by fitting a type S (sigmoidal) dose-dependent FP increase as a function of protein concentration using Graphpad Prism 5.0 Software (Graphpad Software, San Diego, Calif.)_d)。

Determination of K of inhibitors by dose-dependent competitive binding assays of inhibitors_iThe value is obtained. In this experiment, the concentration of inhibitor was gradually diluted, with different concentrationsDegree of inhibitor was separately combined with a fixed concentration of fluorescent tracer (SM-F2) at a fixed concentration (for the above-described determination of K)_d2-3 times the value) of the target protein. mu.L of DMSO solutions of the test compounds at various concentrations and 120. mu.L of buffer solution containing the preincubated target protein/SM-F2 complex (100mM dipotassium phosphate, pH 7.5, 100. mu.g/ml bovine gamma-globulin, 0.02% sodium azide, 4% DMSO, Invitrogen) were added to 96-well plates, incubated at room temperature for 3 hours and gently shaken. For the determination of competitive binding of test compounds to SM-F2 and XIAP-BIR3, the final concentrations of XIAP-BIR3 and SM-F2 in the system were 10nM and 2nM, respectively; whereas for binding to cIAP1-BIR3, the final concentrations of cIAP1-BIR3 and SM-F2 in the system were 3nM and 1nM, respectively. Each 96-well plate contains a negative control group containing only the target protein/SM-F2 complex (inhibition equivalent to 0%) and a positive control group containing only free SM-F2 (inhibition equivalent to 100%). The FP value was determined in the same manner as described above. IC of test Compound₅₀Determined by nonlinear regression fitting of the competition curve. K of competitive inhibitors_iCalculated by the following equation:

K_i＝[I]₅₀/([L]₅₀/K_d+[P]₀/K_d+1)

in the formula, [ I ]]₅₀Indicates the concentration of inhibitor at 50% inhibition, [ L ]]₅₀Is the labeled ligand (SM-F2) concentration at 50% inhibition, [ P ]]₀Is the protein concentration at 0% inhibition, K_dIs the equilibrium dissociation constant.

The results of the XIAP-BIR3 and cIAP1-BIR3 inhibitory activity assays are shown in Table 1.

Table 1: XIAP-BIR3 and cIAP1-BIR3 inhibitory Activity assay results

Pharmacological test example 2: determination by the thiazole blue (MTT) methodVarious synthetic target compounds in vitro Survival inhibiting effect on ovarian cancer and cervical cancer cells

Survival inhibitory effect on ovarian cancer:collecting ovarian cancer SKOV3 cells at logarithmic growth phase at concentration of 1 × 10⁵Adding into 96-well plate, adding 200 μ l cell-containing culture medium into each well, standing at 37 deg.C and 5% CO₂After incubation in an incubator for 16h, 100. mu.M of each synthesized target compound was added sequentially, while DMEM medium was used as a blank control, and MTT (5mg/ml) was added at 20. mu.l/well, 37 ℃, 5% CO, 48h after dosing₂After 4h incubation in the incubator, the supernatant was discarded and 150. mu.l of DMSO was added to each well to determine the OD absorbance at 490nm, and 4 wells were assayed in parallel for each concentration.

(ii) an inhibitory effect on the survival of cervical cancer cells:log phase cells were collected, cell suspension concentration was adjusted, 200ul was added to each well, and test cells were plated to 10000/well (marginal wells filled with sterile PBS). Blank control, positive control and negative control were set. 5% CO₂Incubate at 37 ℃ until the cell monolayer is well-bottom (96-well flat bottom plate) and add the drug at a certain concentration. 5% CO₂Incubated at 37 ℃ for 48 hours and observed under an inverted microscope. 20ul of MTT solution (5mg/ml, i.e.0.5% MTT) was added to each well and incubation was continued for 4 h. The culture was terminated and the culture medium in the wells was carefully aspirated. Add 150. mu.l dimethyl sulfoxide into each well, and shake for 10min at low speed on a shaking bed to dissolve the crystals sufficiently. The absorbance of each well was measured at OD 490nm in an ELISA detector. And setting a zero setting hole (culture medium, MTT and dimethyl sulfoxide) and a control hole (cells, a drug dissolving medium with the same concentration, a culture solution, MTT and dimethyl sulfoxide).

From the above raw data, the growth Inhibition Ratio (IR) was calculated by the following formula.

The results of the survival inhibitory effect on ovarian cancer showed that all compounds had between 10% and 70% inhibition (100 μm) of ovarian cancer cells. The results of the survival inhibitory effect on cervical cancer cells are shown in Table 2, wherein the inhibitory rates (100 μm) of other compounds not listed in Table 2 on cervical cancer cells were all between 10% and 70%.

Table 2: EXAMPLE inhibition ratio (100 μm) of cervical cancer cells

Claims

1. A compound of the general formula I:

and isomers, pharmaceutically acceptable salts thereof, wherein:

x is-S-;

y is-CH₂-；

R¹Selected from:

1)-H，

2)-C₁-C₆alkyl, optionally substituted with one or more halogens;

R²selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

R³selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

R⁴and R⁴⁰⁰Each independently selected from:

1)-H，

R⁵and R⁵⁰⁰Each independently selected from:

1)-H，

R⁶selected from:

1)-NR¹¹R¹²，

wherein R is¹¹And R¹²Each independently selected from:

1)-H，

2)-C₁-C₆an alkyl group, a carboxyl group,

3)-C₃-C₈a cycloalkyl group,

4)-(C₁-C₆alkyl) -Ar¹，

Wherein,

Ar¹is phenyl, optionally substituted by one or more groups selected from

Generation:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃。

2. the compound of claim 1, wherein R³The attached carbon atoms may be in any optical configuration.

3. A compound of formula Id:

and isomers, pharmaceutically acceptable salts thereof, wherein R¹、R²、R³The definition as claimed in claim 1 for the compounds of the formula I, R⁸And R⁹Each independently selected from:

1)-H，

3)-C₁-C₆alkyl, optionally substituted with one or more halogens,

4)-C₃-C₈cycloalkyl, optionally substituted with one or more halogens,

6)-C₂-C₆alkynyl, optionally substituted with one or more halo.

4. A compound of formula If:

and isomers, pharmaceutically acceptable salts thereof, wherein:

R¹、R²、R³and Ar¹The definition of the compounds of the formula I as claimed in claim 1,

Ar²is phenyl, optionally substituted with one or more groups selected from:

1) the halogen(s) are selected from the group consisting of,

2) the nitro group(s),

3) the cyano group(s),

4)-CF₃。

5. a compound of the general formula I:

and isomers, pharmaceutically acceptable salts thereof, wherein:

x is-S-;

y is-CH₂-；

R¹Selected from: -H, -C₁-C₄An alkyl group;

R²selected from: -H, -C₁-C₄An alkyl group;

R³selected from: -H, -C₁-C₆Alkyl, -C₃-C₈Cycloalkyl, - (C)₁-C₆Alkyl) -OR⁸Wherein R is⁸Selected from: -H, -C₁-C₆Alkyl, -C₂-C₆Alkenyl, and-C₂-C₆An alkynyl group;

R⁴and R⁴⁰⁰Each independently selected from: -H, -C₁-C₄An alkyl group;

R⁶is-NR¹¹R¹²Wherein R is¹¹And R¹²Each independently selected from: -H, -C₁-C₄Alkyl-, C₄-C₆Cycloalkyl, -COR⁸、-Ar¹、-(C₁-C₄Alkyl) -Ar¹、-CO-Ar¹and-CO- (C)₁-C₄Alkyl) -Ar¹Wherein Ar is¹Is phenyl optionally substituted with one or more groups selected from: halogen, nitro, cyano, -CF₃Wherein R is⁸Selected from: -H, - (C)₁-C₄Alkylene radical)_0-3-phenyl, wherein the phenyl is optionally substituted by one or more groups selected from-C₁-C₄Alkyl and halogen, optionally substituted by one or more halogens, -C₁-C₄An alkyl group.

6. A compound selected from:

and isomers and pharmaceutically acceptable salts thereof.

7. A pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of a compound according to any one of claims 1-6 and optionally a pharmaceutically acceptable carrier or excipient.

8. Use of a compound according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment and/or prophylaxis of a disease associated with overexpression of XIAP-BIR3 or cIAP1-BIR3, wherein the disease is selected from ovarian cancer and cervical cancer.