Compound as WRN inhibitor and application thereof

Abstract

The invention belongs to the field of medical chemistry, relates to a compound serving as a WRN inhibitor and application of the compound, and particularly provides a compound shown in a formula (I) or pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug of the compound, processes for their preparation, pharmaceutical compositions containing these compounds and the use of these compounds or compositions for the treatment of diseases associated with WRN. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the field of medicinal chemistry, and particularly relates to compounds as WRN inhibitors or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, methods for preparing them, pharmaceutical compositions containing these compounds, and the use of these compounds or compositions for treating diseases related to WRN. Background Art

[0002] Microsatellites are a class of short tandem repeat DNA sequences in the genome, generally composed of 1-6 nucleotides. Due to the difference in the number of repeats of the core repeat unit, microsatellites usually have population polymorphisms. Compared with normal tissues, the phenomenon that a new microsatellite allele appears at a certain microsatellite locus in tumors due to the insertion or deletion of repeat units is called microsatellite instability (MSI). The occurrence of MSI is due to the functional defect of DNA mismatch repair (dMMR) in tumor tissues. With the continuous deepening of preclinical and clinical studies, MSI accompanied by DNA mismatch repair defects is an important tumor marker in clinical practice. Clinically, dMMR and MSI-H usually have the same clinical significance. Cancers with MSI include: colorectal cancer, gastric cancer, endometrial cancer, ovarian cancer, cholangiocarcinoma, urethral cancer, brain cancer and skin cancer. Among them, colorectal cancer is the most common (MSI-H≈15%). In a number of clinical trials, PD-1 monoclonal antibodies represented by pembrolizumab have been found to have good therapeutic effects on MSI-H tumors. However, there are still a significant proportion of MSI patients who do not respond to immunotherapy in the research, and some MSI-H tumor patients develop primary resistance to immunotherapy. The clinical needs of this part of patients have not been met at present.

[0003] A series of studies in recent years have found that the WRN protein (WernerProtein) in the RNA / DNA helicase RecQ family has a strong correlation with the occurrence and development of MSI-H tumors. Knocking out or knocking down WRN in MSI-H tumor cells can significantly inhibit the proliferation, invasion and migration of tumor cells and promote the apoptosis of tumor cells. These research results indicate that WRN and MSI-H may have a synthetic lethal relationship between tumors. At present, the mechanism by which knocking down WRN promotes apoptosis of MSI-H tumors has also been clearly explained: TA-dinucleotide repeats are highly unstable in MSI cells and will undergo large-scale amplification. These large increases in TArepeat will form non-B DNA secondary structures, stop the replication fork, and activate the ATR checkpoint kinase. In this process, the participation of WRN helicase is required to reactivate the DNA replication fork to allow DNA replication to continue. When there is no WRN, the amplified TA-dinucleotide repeat sequence is easily cut and broken by the MUS81 nuclease, resulting in a large number of chromosome fragments and inducing cell apoptosis.

[0004] Therefore, the development of small molecule inhibitors of WRN is expected to provide a new means of anti-tumor treatment and enable MSI-H tumor patients to obtain clinical benefits. Summary of the invention

[0005] The present invention provides a compound represented by general formula (I) or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof,

[0006]

[0007] in,

[0008] Ring A is selected from cycloalkyl, heterocyclyl, aryl and heteroaryl;

[0009] R 1Selected from hydrogen, halogen, cyano, amino, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclic group, heteroalkenyl, alkoxy, haloalkoxy, hydroxyalkoxy, monoalkylamino, dialkylamino, alkylacylamino, alkylacyl, aminoacyl, alkylaminoacyl, dialkylamino, heterocyclic group, aryl and heteroaryl, and the alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclic group, heteroalkenyl, alkoxy, haloalkoxy, hydroxyalkoxy, monoalkylamino, dialkylamino, alkylacylamino, alkylacyl, aminoacyl, alkylaminoacyl, dialkylamino, aryl and heteroaryl are optionally substituted by one or more groups selected from halogen, hydroxy, alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, amino, monoalkylamino, alkylacylamino, alkylacyl, alkylsulfonyl, aminoacyl, alkylaminoacyl, dialkylamino, alkenyl, alkynyl, haloalkylacyl, hydroxyalkylacyl, cycloalkyl, heterocyclic group, aryl, heteroaryl, oxo group;

[0010] R 2 Selected from aryl, heteroaryl, cycloalkyl and heterocyclic group, and the aryl, heteroaryl, cycloalkyl and heterocyclic group are optionally substituted by one or more groups selected from halogen, hydroxy, cyano, amino, alkyl, haloalkyl, alkoxy, haloalkoxy, cycloalkyl and heterocyclic group;

[0011] R 3 Selected from alkyl, haloalkyl, cycloalkyl, heterocyclic group and hydroxyalkyl;

[0012] Each R 4 Independently selected from halogen, hydroxy, alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, nitro, carboxyl, cyano, amino, monoalkylamino, alkylacylamino, alkylacyl, alkylsulfonyl, aminoacyl, alkylaminoacyl, dialkylamino, alkenyl, alkynyl, haloalkylacyl, hydroxyalkylacyl, cycloalkylacyl, heterocyclic group acyl, cycloalkyl, halocycloalkyl, heterocyclic group, aryl, heteroaryl and oxo group, and the alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, monoalkylamino, alkylacylamino, alkylacyl, alkylsulfonyl, aminoacyl, alkylaminoacyl, dialkylamino, alkenyl, alkynyl, haloalkylacyl, hydroxyalkylacyl, cycloalkylacyl, heterocyclic group acyl, cycloalkyl, halocycloalkyl, heterocyclic group, aryl, heteroaryl are optionally substituted by one or more groups selected from halogen, hydroxy, alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, amino, cycloalkyl, halocycloalkyl, heterocyclic group, aryl, heteroaryl and oxo group, and m is selected from 0, 1, 2, 3 and 4; and

[0013] L is selected from -CH 2 -, -CH 2 CH2 -, -CH 2 NH-, -CH 2 C(O)NH- and -C(O)NH-.

[0014] In some preferred embodiments, the compounds of the present invention are compounds of general formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0015] Ring A is selected from C 6-12 cycloalkyl, 3- to 13-membered heterocyclic group, C 6-12 aryl and 5- to 13-membered heteroaryl;

[0016] More preferably, ring A is selected from C 6-10 cycloalkyl, 3- to 13-membered heterocyclic group, C 6-10 aryl and 5- to 10-membered heteroaryl;

[0017] Even more preferably, ring A is selected from

[0018] In some preferred embodiments, the compounds of the present invention are compounds of general formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0019] R 1 is selected from hydrogen, halogen, cyano, amino, C 1-6 alkyl, C 3-10 cycloalkyl, C 2-10 alkenyl, C 2-10 alkynyl, 3- to 10-membered heterocyclic group, 3- to 10-membered heterocyclic enyl, C 1-6 alkoxy, halo C 1-6 alkoxy, hydroxy C 1-6 alkoxy, mono C 1-6 alkylamino, di C 1-6 alkylamino, C 1-6 alkylacylamino, C 1-6 alkylacyl, aminoacyl, C 1-6 alkylaminoacyl, di C 1-6 alkylamino, C 6-10 aryl and 5- to 10-membered heteroaryl, wherein the C 1-6 alkyl, C 3-10 cycloalkyl, C 2-10 alkenyl, C 2-10 alkynyl, 3- to 10-membered heterocyclic group, 3- to 10-membered heterocyclic enyl, C 1-6 alkoxy, halo C 1-6 alkoxy, hydroxy C 1-6 alkoxy, mono C 1-6 alkylamino, di C1-6 alkylamino, C 1-6 alkylacylamino, C 1-6 alkylacyl, aminoacyl, C 1-6 alkylaminoacyl, di-C 1-6 alkylamino, C 6-10 aryl and 5-10 membered heteroaryl optionally substituted by one or more groups selected from halogen, hydroxy, C 1-6 alkyl, halo-C 1-6 alkyl, hydroxy-C 1-6 alkyl, C 1-6 alkoxy, halo-C 1-6 alkoxy, hydroxy-C 1-6 alkoxy, amino, mono-C 1-6 alkylamino, C 1-6 alkylacylamino, C 1-6 alkylacyl, C 1-6 alkylsulfonyl, aminoacyl, C 1-6 alkylaminoacyl, di-C 1-6 alkylamino, alkenyl, alkynyl, halo-C 1-6 alkylacyl, hydroxy-C 1-6 alkylacyl, C 3-10 cycloalkyl, 3-10 membered heterocyclic group, C 6-10 aryl, 5-10 membered heteroaryl, oxo group substituted;

[0020] Further preferably, R 1 selected from hydrogen, halogen, cyano, amino, C 1-6 alkyl, C 3-8 cycloalkyl, C 2-6 alkenyl, C 2-6 alkynyl, 3-10 membered heterocyclic group, 3-10 membered heterocyclic enyl, C 1-6 alkoxy, halo-C 1-6 alkoxy, hydroxy-C 1-6 alkoxy, mono-C 1-6 alkylamino, di-C 1-6 alkylamino, C 1-6 alkylacylamino, C 1-6 alkylacyl, aminoacyl, C 1-6 alkylaminoacyl, di-C 1-6 alkylamino, C 6-10 aryl and 5-10 membered heteroaryl, the C 1-6 alkyl, C 3-8 cycloalkyl, C 2-6 alkenyl, C 2-6 alkynyl, 3-10 membered heterocyclic group, 3-10 membered heterocyclic enyl, C 1-6 alkoxy, halo-C 1-6 alkoxy, hydroxy-C 1-6Alkoxy, mono-C 1-6 Alkylamino, di-C 1-6 Alkylamino, C 1-6 Alkylacylamino, C 1-6 Alkylacyl, aminoacyl, C 1-6 Alkylaminoacyl, di-C 1-6 Alkylamino, C 6-10 Aryl and 5-10 membered heteroaryl are optionally substituted by one or more groups selected from halogen, hydroxy, C 1-3 Alkyl, halo-C 1-3 Alkyl, hydroxy-C 1-6 Alkyl, C 1-6 Alkoxy, halo-C 1-6 Alkoxy, hydroxy-C 1-6 Alkoxy, amino, mono-C 1-6 Alkylamino, C 1-6 Alkylacylamino, C 1-6 Alkylacyl, C 1-6 Alkylsulfonyl, aminoacyl, C 1-6 Alkylaminoacyl, di-C 1-6 Alkylamino, C 2-6 Alkenyl, C 2-6 Alkynyl, halo-C 1-6 Alkylacyl, hydroxy-C 1-6 Alkylacyl, C 3-10 Cycloalkyl, 3-10 membered heterocyclic group, C 6-10 membered aryl, 5-10 membered heteroaryl, oxo group.

[0021] In some preferred embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0022] R 2 is selected from C 6-10 aryl, 5-13 membered heteroaryl, C 3-13 cycloalkyl and 3-13 membered heterocyclic group, the C 6-10 aryl, 5-13 membered heteroaryl, C 3-13 cycloalkyl and 3-13 membered heterocyclic group are optionally substituted by one or more groups selected from halogen, hydroxy, cyano, amino, C 1-6 alkyl, halo-C 1-6 alkyl, C 1-6 alkoxy, halo-C 1-6 alkoxy, C 3-8 cycloalkyl and 3-10 membered heterocyclic group;

[0023] More preferably, R 2 is selected from C 6-8 aryl, 5-10 membered heteroaryl, C 3-8Cycloalkyl and 3- to 13-membered heterocyclic group, said C 6-8 Aryl, 5- to 10-membered heteroaryl, C 3-8 Cycloalkyl and 3- to 13-membered heterocyclic group are optionally substituted by one or more groups selected from halogen, hydroxy, cyano, amino, C 1-6 Alkyl, halo-C 1-6 Alkyl, C 1-6 Alkoxy, halo-C 1-6 Alkoxy, C 3-8 Cycloalkyl and 3- to 8-membered heterocyclic group;

[0024] Even more preferably, R 2 Is selected from C 6-8 Aryl, 5- to 10-membered heteroaryl, C 3-8 Cycloalkyl and 3- to 13-membered heterocyclic group, said C 6-8 Aryl, 5- to 10-membered heteroaryl, C 3-8 Cycloalkyl and 3- to 13-membered heterocyclic group are optionally substituted by one or more groups selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo-C 1-3 Alkyl, C 1-3 Alkoxy, halo-C 1-3 Alkoxy, C 3-8 Cycloalkyl and 3- to 8-membered heterocyclic group.

[0025] In some preferred embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0026] R 3 Is selected from C 1-6 Alkyl, halo-C 1-6 Alkyl, C 3-8 Cycloalkyl, 3- to 8-membered heterocyclic group and hydroxy-C 1-6 Alkyl;

[0027] Even more preferably, R 3 Is selected from C 1-3 Alkyl, halo-C 1-3 Alkyl, C 3-8 Cycloalkyl, 3- to 6-membered heterocyclic group and hydroxy-C 1-3 Alkyl.

[0028] In some preferred embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0029] Each R 4 Independently is selected from halogen, hydroxy, C 1-6 Alkyl, halo-C 1-6 Alkyl, hydroxy-C1-6 Alkyl, C 1-6 Alkoxy, halo C 1-6 Alkoxy, hydroxy C 1-6 Alkoxy, nitro, carboxy, cyano, amino, mono C 1-6 Alkylamino, C 1-6 Alkylacylamino, C 1-6 Alkylacyl, C 1-6 Alkylsulfonyl, aminoacyl, C 1-6 Alkylaminoacyl, bis C 1-6 Alkylamino, C 2-10 Alkenyl, C 2-10 Alkynyl, halo C 1-6 Alkylacyl, hydroxy C 1-6 Alkylacyl, C 3-12 Cycloalkylacyl, 3-12 membered heterocyclic acyl, C 3-12 Cycloalkyl, halo C 3-12 Cycloalkyl, 3-12 membered heterocyclic, C 6-12 Aryl, 5-12 membered heteroaryl and oxo group, the C 1-6 Alkyl, halo C 1-6 Alkyl, hydroxy C 1-6 Alkyl, C 1-6 Alkoxy, halo C 1-6 Alkoxy, hydroxy C 1-6 Alkoxy, mono C 1-6 Alkylamino, C 1-6 Alkylacylamino, alkylacyl, C 1-6 Alkylsulfonyl, aminoacyl, C 1-6 Alkylaminoacyl, bis C 1-6 Alkylamino, C 2-10 Alkenyl, C 2-10 Alkynyl, halo C 1-6 Alkylacyl, hydroxy C 1-6 Alkylacyl, C 3-12 Cycloalkylacyl, 3-12 membered heterocyclic acyl, C 3-12 Cycloalkyl, halo C 3-12 Cycloalkyl, 3-12 membered heterocyclic, C 6-12 Aryl, 5-12 membered heteroaryl optionally substituted with one or more selected from halogen, hydroxy, C 1-6 Alkyl, halo C 1-6 Alkyl, hydroxy C 1-6 Alkyl, C 1-6 Alkoxy, halo C 1-6 Alkoxy, hydroxy C 1-6 Alkoxy, amino, C 3-12 Cycloalkyl, halo C 3-12Substituted by cycloalkyl, 3- to 12-membered heterocyclic group, 6- to 12-membered aryl group, 5- to 12-membered heteroaryl group and oxo group, m is selected from 0, 1, 2, 3 and 4;

[0030] Further preferably, each R 4 is independently selected from halogen, hydroxy, C 1-3 alkyl, halo C 1-3 alkyl, hydroxy C 1-3 alkyl, C 1-3 alkoxy, halo C 1-3 alkoxy, hydroxy C 1-3 alkoxy, nitro, carboxyl, cyano, amino, mono-C 1-3 alkylamino, C 1-3 alkylacylamino, C 1-3 alkylacyl, C 1-3 alkylsulfonyl, aminoacyl, C 1-3 alkylaminoacyl, di-C 1-3 alkylamino, C 2-6 alkenyl, C 2-6 alkynyl, halo C 1-3 alkylacyl, hydroxy C 1-3 alkylacyl, C 3-8 cycloalkylacyl, 3- to 8-membered heterocyclic acyl group, C 3-8 cycloalkyl, halo C 3-8 cycloalkyl, 3- to 8-membered heterocyclic group, C 6-8 aryl, 5- to 8-membered heteroaryl group and oxo group, the C 1-3 alkyl, halo C 1-3 alkyl, hydroxy C 1-3 alkyl, C 1-3 alkoxy, halo C 1-3 alkoxy, hydroxy C 1-3 alkoxy, mono-C 1-3 alkylamino, C 1-3 alkylacylamino, alkylacyl, C 1-3 alkylsulfonyl, aminoacyl, C 1-3 alkylaminoacyl, di-C 1-3 alkylamino, C 2-6 alkenyl, C 2-6 alkynyl, halo C 1-3 alkylacyl, hydroxy C 1-3 alkylacyl, C 3-8 cycloalkylacyl, 3- to 8-membered heterocyclic acyl group, C 3-8 cycloalkyl, halo C 3-8 cycloalkyl, 3- to 8-membered heterocyclic group, C 6-8 aryl, 5- to 8-membered heteroaryl group is optionally substituted by one or more selected from halogen, hydroxy, C 1-3 alkyl, halo C 1-3Alkyl, hydroxy C 1-3 Alkyl, C 1-3 Alkoxy, halo C 1-3 Alkoxy, hydroxy C 1-3 Alkoxy, amino, C 3-8 Cycloalkyl, halo C 3-8 Cycloalkyl, 3- to 8-membered heterocyclic group, C 6-8 Substituted with a group selected from aryl, 5- to 8-membered heteroaryl and oxo group, and m is selected from 0, 1, 2, 3, and 4.

[0031] In some embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0032] R 1 Selected from And Optionally substituted with one or more groups selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo C 1-3 Alkyl, C 1-3 Alkoxy, halo C 1-3 Alkoxy, C 3-8 Cycloalkyl, 3- to 8-membered heterocyclic group and oxo group; further optionally substituted with one or more groups selected from fluorine, chlorine, bromine, iodine, hydroxy, cyano, amino, methyl, ethyl, propyl, isopropyl, halo C 1-3 Alkyl such as trifluoromethyl, C 1-3 Alkoxy, halo C 1-3 Alkoxy, C 3-8 Cycloalkyl, 3- to 8-membered heterocyclic group and oxo group.

[0033] In some embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein:

[0034] R 2 Selected from And Optionally substituted with one or more groups selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo C 1-3 Alkyl, C 1-3 Alkoxy, halo C 1-3 Alkoxy, C 3-8 Cycloalkyl, 3- to 8-membered heterocyclic group and oxo group; further optionally substituted with one or more groups selected from fluorine, chlorine, bromine, iodine, hydroxy, cyano, amino, methyl, ethyl, propyl, isopropyl, halo C 1-3 Alkyl such as trifluoromethyl, C 1-3 Alkoxy, halo C1-3 Alkoxy, C 3-8 Substituted by a cycloalkyl group, a 3-8 membered heterocyclic group and an oxo group.

[0035] In some embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein R 3 Is selected from methyl, ethyl and propyl.

[0036] In some embodiments, the compounds of the present invention are compounds of formula (I) or pharmaceutically acceptable salts, isomers, solvates, crystals or prodrugs thereof, wherein each R 4 Is independently selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo C 1-3 Alkyl, C 1-3 Alkoxy, halo C 1-3 Alkoxy, C 3-8 Cycloalkyl and 3-8 membered heterocyclic group, which is optionally substituted by one or more groups selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo C 1-3 Alkyl and oxo group, and m is selected from 0, 1, 2, 3 and 4.

[0037] In some embodiments, the present invention provides a compound of formula (Ia) or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof,

[0038]

[0039] R 1 、R 2 、R 3 、R 4 、m and L have the definitions in the above formula (I), X 1 、X 2 And X 3 Are each independently selected from N and -C(R 5 )-, R 5 Is selected from hydrogen, halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halo C 1-3 Alkyl, hydroxy C 1-3 Alkyl, C 1-3 Alkoxy and halo C 1-3 Alkoxy; Ring B is selected from C 3-8 Cycloalkyl, 3-10 membered heterocyclic group, C 6-8Aryl and 5- to 8-membered heteroaryl, preferably, ring B is selected from phenyl, imidazolyl, pyrrolyl, furyl, thienyl, pyrazolyl, oxazolyl, thiazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, indolyl, pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, piperidinyl, piperazinyl, and tetrahydroindazolyl.

[0040] In some embodiments, the present invention provides a compound of formula (Ia) or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof, wherein

[0041] Ring B is selected from R 5 is selected from fluorine, chlorine, bromine, iodine, hydroxyl, cyano, amino, methyl, ethyl, propyl, isopropyl, halo-C 1-3 alkyl such as trifluoromethyl, hydroxy-C 1-3 alkyl, C 1-3 alkoxy, and halo-C 1-3 alkoxy.

[0042] In some embodiments, the present invention provides a compound of formula (Ia) or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof, wherein

[0043] is selected from Each R 4 is independently selected from halogen, hydroxyl, cyano, amino, C 1-3 alkyl, halo-C 1-3 alkyl, C 1-3 alkoxy, halo-C 1-3 alkoxy, C 3-8 cycloalkyl, and 3- to 8-membered heterocyclic group, which is optionally substituted by one or more groups selected from halogen, hydroxyl, cyano, amino, C 1-3 alkyl, halo-C 1-3 alkyl, and oxo group, and m is selected from 0, 1, 2, 3, and 4.

[0044] In some embodiments, the present invention provides a compound of formula (Ib) or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof,

[0045]

[0046] wherein ring A, R 1 , R 3 , R 4 , m have the definitions in the above general formula (I), and R 6 , R 7 are each independently selected from halogen, hydroxyl, cyano, amino, C 1-3 alkyl, halo-C1-3 alkyl, C 1-3 alkoxy and halo C 1-3 alkoxy; n and p are each independently selected from 0, 1, 2, 3, and 4.

[0047] In some embodiments, a compound represented by formula (I) or (Ib) according to the present invention, or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof, wherein

[0048] selected from

[0049] The present invention provides the following specific compounds or pharmaceutically acceptable salts, isomers, solvates, crystals, or prodrugs thereof:

[0050]

[0051]

[0052] On the other hand, the present invention provides a pharmaceutical composition comprising a compound of the present invention or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof.

[0053] In some embodiments, the present invention provides a compound of the present invention or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof, and a pharmaceutical composition comprising the compound of the present invention or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof, wherein the compound or pharmaceutical composition is used for treating diseases related to WRN.

[0054] In some embodiments, the present invention provides a pharmaceutical composition comprising a compound of the present invention or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof and a pharmaceutically acceptable carrier.

[0055] The compound of the present invention or a pharmaceutically acceptable salt, isomer, solvate, crystal, or prodrug thereof can be mixed with a pharmaceutically acceptable carrier, diluent, or excipient to prepare a pharmaceutical preparation suitable for oral or parenteral administration. The administration methods include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, and oral routes. The preparation can be administered by any route, such as by infusion or bolus injection, and by a route of absorption through epithelial or skin mucosa (such as oral mucosa or rectum, etc.). The administration can be systemic or local. Examples of oral administration preparations include solid or liquid dosage forms, specifically, including tablets, pills, granules, powders, capsules, syrups, emulsions, suspensions, etc. The preparation can be prepared by methods known in the art and contains carriers, diluents, or excipients commonly used in the field of pharmaceutical preparations.

[0056] Thirdly, the present invention provides a method for treating a WRN-related disease by using a compound represented by formula (I), (Ia) or (Ib) of the present invention, or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, or a pharmaceutical composition containing the same, and the use thereof in the preparation of a drug for treating a WRN-related disease.

[0057] In some preferred embodiments, the present invention provides a method for treating a WRN-related disease by using a compound represented by formula (I), (Ia) or (Ib) of the present invention, or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, or a pharmaceutical composition containing the same, wherein the WRN-related diseases include but are not limited to: cancer, proliferative diseases, blood diseases or metabolic diseases. In some embodiments, the WRN-related disease of the present invention is cancer.

[0058] In some embodiments, the WRN-related diseases of the present invention include but are not limited to: pancreatic cancer, lung cancer, colorectal cancer, cholangiocarcinoma, multiple myeloma, melanoma, uterine cancer, endometrial cancer, thyroid cancer, acute myeloid leukemia, bladder cancer, urothelial cancer, gastric cancer, cervical cancer, head and neck squamous cell carcinoma, diffuse large B-cell lymphoma, esophageal cancer, chronic lymphocytic leukemia, hepatocellular carcinoma, breast cancer, ovarian cancer, prostate cancer, glioblastoma, renal cancer and sarcoma.

[0059] In some embodiments, the present invention provides the use of a compound represented by formula (I), (Ia) or (Ib) of the present invention, or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, or a pharmaceutical composition containing the same in the preparation of a drug for treating and / or preventing microsatellite instability (MSI) or DNA mismatch repair (dMMR) cancer.

[0060] In some embodiments, the microsatellite instability (MSI) or DNA mismatch repair (dMMR) cancers of the present invention include but are not limited to: colorectal cancer, gastric cancer, prostate cancer, endometrial cancer, adrenocortical cancer, uterine cancer, cervical cancer, esophageal cancer, breast cancer, renal cancer and ovarian cancer.

[0061] Term Definition

[0062] Unless otherwise stated, the terms used in the specification and claims have the following meanings.

[0063] The "hydrogen", "carbon", and "oxygen" in the compounds of the present invention include all their isotopes. Isotopes are to be understood as including those atoms having the same atomic number but different mass numbers. For example, the isotopes of hydrogen include protium, tritium, and deuterium, and the isotopes of carbon include 12 C, 13 C, and 14 C, and the isotopes of oxygen include 16 O and 18 O, etc.

[0064] The "isomers" of the present invention refer to molecules having the same atomic composition and mode of connection but different three-dimensional spatial arrangements, including but not limited to diastereomers, enantiomers, cis-trans isomers, and mixtures thereof, such as racemic mixtures. Many organic compounds exist in optically active forms, i.e., they have the ability to rotate the plane of plane-polarized light. When describing optically active compounds, the prefixes D, L or R, S are used to denote the absolute configuration of the chiral centers of the molecule. The prefixes D, L or (+), (-) are used to name the sign of the rotation of plane-polarized light by the compound, (-) or L means that the compound is levorotatory, and the prefix (+) or D means that the compound is dextrorotatory. The chemical structures of these stereoisomers are the same, but their stereostructures are different. A specific stereoisomer may be an enantiomer, and a mixture of isomers is usually called a mixture of enantiomers. A 50:50 mixture of enantiomers is called a racemic mixture or racemate, which may result in no stereoselectivity or stereospecificity during a chemical reaction. The terms "racemic mixture" and "racemate" refer to a mixture of equimolar amounts of two enantiomers, lacking optical activity.

[0065] Depending on the choice of starting materials and methods, the compounds of the present invention may exist in the form of one or a mixture of possible isomers, such as racemates and mixtures of diastereoisomers (depending on the number of asymmetric carbon atoms). Optically active (R)- or (S)-isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques.

[0066] Any mixture of stereoisomers obtained can be separated into pure or substantially pure geometric isomers, enantiomers, diastereoisomers based on differences in the physicochemical properties of the components, for example, by chromatography and / or fractional crystallization.

[0067] The "halogen" of the present invention refers to fluorine, chlorine, bromine, and iodine. The "halogenated" of the present invention means being substituted by fluorine, chlorine, bromine, or iodine.

[0068] "Alkyl" in the present invention refers to a straight-chain or branched-chain saturated aliphatic hydrocarbon group, preferably a straight-chain or branched-chain group containing 1 to 6 carbon atoms, more preferably a straight-chain or branched-chain group containing 1 to 3 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, etc. The alkyl group can be substituted or unsubstituted. When substituted, the substituent can be at any available attachment point.

[0069] "Carbonyl" and "acyl" in the present invention both refer to -C(O)-.

[0070] "Sulfonyl" in the present invention refers to -S(O) 2 -.

[0071] "Sulfonamido" in the present invention refers to -S(O) 2 NH-.

[0072] "Haloalkyl" in the present invention refers to an alkyl group that is at least substituted by one halogen.

[0073] "Hydroxyalkyl" in the present invention refers to an alkyl group that is at least substituted by one hydroxyl group.

[0074] "Alkoxy" in the present invention refers to -O-alkyl. Non-limiting examples of alkoxy include: methoxy, ethoxy, propoxy, n-propoxy, isopropoxy, isobutoxy, sec-butoxy, etc. The alkoxy group can be optionally substituted or unsubstituted. When substituted, the substituent can be at any available attachment point.

[0075] "Cycloalkyl" in the present invention refers to a cyclic saturated hydrocarbon group. Suitable cycloalkyl groups can be substituted or unsubstituted monocyclic, bicyclic or tricyclic saturated hydrocarbon groups having 3 to 12 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl.

[0076] The "heterocyclic group" of the present invention refers to a group of a 3- to 15-membered non-aromatic ring system having 1 to 4 ring heteroatoms (where each heteroatom is independently selected from nitrogen, oxygen, sulfur, boron, phosphorus, and silicon) ("3-15 membered heterocyclic group"). In a heterocyclic group containing one or more nitrogen atoms, the point of attachment can be a carbon or nitrogen atom, provided that the valence allows. The heterocyclic group can either be monocyclic ("monocyclic heterocyclic group") or a fused, bridged, or spiro ring system (such as a bicyclic system (also known as "bicyclic heterocyclic group")) and can be saturated or can be partially unsaturated, where the bicyclic heterocyclic group includes, but is not limited to, benzazacyclic group, benzoxacyclic group, benzothiacyclic group, benzodiazacyclic group, benzodioxacyclic group, benzodithiacyclic group, benzoxazacyclic group, benzothiazacyclic group. Suitable heterocyclic groups include, but are not limited to, piperidinyl, azetidinyl, aziridinyl, pyrrolidinyl, piperazinyl, dihydroquinazolinyl, oxiranyl, oxetanyl, tetrahydrofuryl, tetrahydropyranyl, dihydrobenzoxacyclohexenyl, dihydroquinolinyl, tetrahydroquinolinyl, dihydroisoquinolinyl, and tetrahydroisoquinolinyl, etc. Each instance of the heterocyclic group can be optionally substituted or unsubstituted, and when substituted, the substituents can be at any available point of attachment.

[0077] The "aryl" of the present invention refers to an aromatic system that can include a monocyclic or fused polycyclic ring, preferably a monocyclic or fused bicyclic aromatic system, which contains 6 to 12 carbon atoms, preferably about 6 to about 10 carbon atoms. Suitable aryl groups include, but are not limited to, phenyl, naphthyl, anthryl, fluorenyl, indanyl. The aryl can be optionally substituted or unsubstituted, and when substituted, the substituents can be at any available point of attachment.

[0078] The "heteroaryl" of the present invention refers to an aryl in which at least one carbon atom is replaced by a heteroatom, preferably composed of 5 - 12 atoms (5-12 membered heteroaryl), more preferably composed of 5 - 10 atoms (5-10 membered heteroaryl), and the heteroatoms are O, S, N. The heteroaryl includes, but is not limited to, imidazolyl, pyrrolyl, furyl, thienyl, pyrazolyl, oxazolyl, thiazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, tetrazolyl, indolyl, pyridyl, pyrimidinyl, pyridazinyl, pyrazinyl, triazinyl, isoindolyl, benzopyrazolyl, benzimidazolyl, benzofuryl, benzopyranyl, benzothienyl, benzoxazolyl, benzothiazolyl, benzisoxazolyl, benzisothiazolyl, quinolinyl, isoquinolinyl, quinazolinyl, cinnolinyl, quinoxalinyl, benzoxazinyl, benzothiazinyl, imidazopyridyl, pyrimidinopyrazolyl, pyrimidinobenzimidazolyl, pyridazinopyrrolyl, pyridazinobenzimidazolyl, pyridinopyrrolyl, pyridinopyrazolyl, etc. The heteroaryl can be optionally substituted or unsubstituted, and when substituted, the substituents can be at any available point of attachment.

[0079] The "cycloalkylalkoxy" of the present invention refers to an alkoxy group in which at least one hydrogen atom of the alkoxy group has been replaced by a cycloalkyl group as defined herein.

[0080] The "heterocyclicalkoxy" of the present invention refers to an alkoxy group in which at least one hydrogen atom of the alkoxy group has been replaced by a heterocyclic group as defined herein.

[0081] The "pharmaceutically acceptable salt" of the present invention refers to a salt of a compound of the present invention, which has safety and effectiveness when used in mammals and has the due biological activity.

[0082] The "solvate" of the present invention conventionally refers to a complex formed by the combination of a solute (such as an active compound, a salt of an active compound) and a solvent (such as water). The solvent refers to a solvent known to those skilled in the art or easily determined. If it is water, the solvate is usually called a hydrate, such as a hemihydrate, a monohydrate, a dihydrate, a trihydrate or an alternative amount thereof, etc.

[0083] The in vivo action of the compound having the chemical formula (I) can be partially exerted by one or more metabolites formed in the human body or an animal body after administration of the compound having the chemical formula (I). As described above, the in vivo action of the compound having the chemical formula (I) can also be exerted via the metabolism of a prodrug. The "prodrug" of the present invention refers to a compound that is converted into a compound of the present invention under physiological conditions in a living organism due to reactions with enzymes, gastric acid, etc., that is, a compound that is converted into a compound of the present invention by oxidation, reduction, hydrolysis, etc. of enzymes and / or a compound that is converted into a compound of the present invention by hydrolysis reactions such as gastric acid, etc.

[0084] The "crystal" of the present invention refers to a solid whose internal structure is regularly repeated in three dimensions to form atoms (or their groups), which is different from an amorphous solid that does not have such a regular internal structure.

[0085] The "pharmaceutical composition" of the present invention refers to a mixture comprising any one of the compounds described in the present invention, including corresponding isomers, prodrugs, solvates, pharmaceutically acceptable salts or their chemically protected forms, and one or more pharmaceutically acceptable carriers and / or one or more other drugs. The purpose of the pharmaceutical composition is to facilitate the administration of the compound to a living organism. The composition is usually used for preparing a drug for treating and / or preventing a disease mediated by one or more kinases.

[0086] The "pharmaceutically acceptable carrier" of the present invention refers to a carrier that does not cause significant irritation to the organism and does not interfere with the biological activity and properties of the administered compound, including all solvents, diluents or other excipients, dispersants, surfactants, isotonic agents, thickening agents or emulsifying agents, preservatives, solid binders, lubricants, etc. Unless any conventional carrier medium is incompatible with the compounds of the present invention. Some examples of pharmaceutically acceptable carriers include, but are not limited to, saccharides such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, as well as cellulose and cellulose acetate; malt, gelatin, etc.

[0087] The "excipient" of the present invention refers to an inert substance added to a pharmaceutical composition to further facilitate the administration of the compound. Excipients may include calcium carbonate, calcium phosphate, various saccharides and various types of starches, cellulose derivatives, gelatin, vegetable oils, polyethylene glycol. Detailed implementation mode

[0088] The present invention will be further elaborated in detail below in conjunction with embodiments, but the present invention is not limited to these embodiments. Materials used in the following embodiments are commercially available unless otherwise specified.

[0089] Example 1: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(3,5-difluoro-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0090]

[0091] Step 1: Preparation of methyl 2-chloro-3-oxopentanoate

[0092]

[0093] Dissolve methyl 3-oxopentanoate (5.0 g, 38.4 mmol) in 200 mL of anhydrous ethanol, protect with argon, cool to below 0 °C, slowly add thionyl chloride (6.74 g, 99.6 mmol), and after the addition, slowly raise the temperature to room temperature and stir overnight. After the reaction is completed, quench the reaction with water, extract with dichloromethane, combine the organic phases, dry over anhydrous sodium sulfate, and concentrate to obtain the title compound. ESI-MS m / z: 164.9 [M+H] + . Step 2: Preparation of tert-butyl 4-(1-methoxy-1,3-dioxolan-2-yl)piperazine-1-carboxylate

[0094]

[0095] Dissolve methyl 2-chloro-3-oxopentanoate (6.34 g, 38.42 mmol) in 50 mL of acetonitrile. Sequentially add triethylamine (11.67 g, 115.3 mmol) and N-Boc piperazine (7.1 g, 38.42 mmol). After addition, heat to 60 °C and stir the reaction overnight. After the reaction is completed, quench the reaction with water, extract with ethyl acetate, dry over anhydrous sodium sulfate, and concentrate and purify by column chromatography to obtain the title compound. ESI-MS m / z [M+H] + : 315.2.

[0096] Step 3: Preparation of tert-butyl 4-(2-bromo-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate

[0097]

[0098] Dissolve 3-bromo-1H-1,2,4-triazol-5-amine (1.5 g, 9.2 mmol) in 5 mL of ethanol, then add tert-butyl 4-(1-methoxy-1,3-dioxolan-2-yl)piperazine-1-carboxylate (3.2 g, 10.12 mmol) and phosphoric acid (902 mg, 9.2 mmol). Stir the reaction at 80 °C overnight. After the reaction is completed, concentrate and purify by column chromatography to obtain the title compound. ESI-MS m / z [M+H] + : 427.2.

[0099] Step 4: Preparation of tert-butyl 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate

[0100]

[0101] Dissolve tert-butyl 4-(2-bromo-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (300 mg, 0.70 mmol), 3,6-dihydro-2H-pyran-4-ylboronic acid pinacol ester (221 mg, 1.43 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (51 mg, 0.07 mmol), and sodium carbonate (150 mg, 1.41 mmol) in 30 mL of anhydrous 1,4-dioxane. Then add 10 mL of water. After protecting with argon, heat to 100 °C and stir the reaction for 36 hours. After the reaction is completed, filter through diatomaceous earth, concentrate the filtrate, and purify by column chromatography to obtain the title compound. ESI-MS m / z [M-H] - : 429.1.

[0102] Step 5: Preparation of tert-butyl 4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate

[0103]

[0104] Dissolve tert-butyl 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (270 mg, 0.63 mmol) in 20 mL of N,N-dimethylformamide, then add N,N-diisopropylethylamine (245 mg, 1.89 mmol) and 2-chloro-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide (171 mg, 0.63 mmol), and stir the reaction at 50 °C overnight. After the reaction is completed, concentrate the reaction solution and perform column chromatography to obtain the title compound. ESI-MS m / z + : 666.2.

[0105] Step 6: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate

[0106]

[0107] Dissolve tert-butyl 4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (150 mg, 0.223 mmol) in 10 mL of anhydrous dichloromethane, then add 1.0 mL of trifluoroacetic acid, and stir the reaction at room temperature for 1 hour. After the reaction is completed, concentrate the reaction solution to obtain the title compound. ESI-MS m / z + : 566.1.

[0108] Step 7: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(3,5-difluoro-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0109]

[0110] Dissolve 3,5-difluoro-2-hydroxybenzoic acid (34 mg, 0.19 mmol) in 5 ml of N,N-dimethylformamide. Sequentially add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (37 mg, 0.19 mmol), 1-hydroxybenzotriazole (26 mg, 0.19 mmol). After stirring at room temperature for 30 minutes, add N,N-diisopropylethylamine (0.5 ml) and N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate (120 mg, 0.18 mmol). After the addition is complete, stir the reaction at room temperature for 2 hours. After the reaction is completed, pour the reaction solution into water to quench it, extract with ethyl acetate, combine the organic phases, dry, concentrate, and perform column chromatography to obtain the title compound. ESI-MS m / z: 722.1 [M+H] + 。 1 HNMR(400MHz,CDCl 3 )δ8.99(s,1H),8.48(d,J=8Hz,1H),7.64(s,1H),7.54(d,J=8Hz,1H),6.98-6.94(m,2H),6.84(d,J=8Hz,1H),5.11(s,2H),4.43-4.33(m,4H),3.93-3.76(m,4H),3.27-3.16(m,4H),2.80-2.73(m,4H),1.33(t,J=8.0Hz,3H).

[0111] Example 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(2,4-difluoro-6-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0112]

[0113] The preparation method is the same as that of Example 1, except that 3,5-difluoro-2-hydroxybenzoic acid is replaced with 2,4-difluoro-6-hydroxybenzoic acid to obtain the title compound. ESI-MS m / z: 722.2 [M+H] + 。 11H NMR (400 MHz, DMSO-d6) δ 10.91 (s, 1H), 10.37 (s, 1H), 8.06 (d, J = 8.0 Hz, 1H), 7.96 (s, 1H), 7.72 (d, J = 8.0 Hz, 1H), 6.83 (s, 1H), 6.73 (d, J = 12.0 Hz, 1H), 6.53 (d, J = 8.0 Hz, 1H), 5.31 (s, 2H), 4.53 (d, J = 12.0 Hz, 2H), 4.25 (s, 2H), 3.84 - 3.76 (m, 2H), 3.06 - 2.84 (m, 4H), 2.83 - 2.69 (m, 2H), 2.68 - 2.59 (m, 2H), 2.44 - 2.24 (m, 2H), 1.18 (t, J = 8.0 Hz, 3H).

[0114] Example 3: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(2-hydroxy-3-(trifluoromethyl)benzoyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0115]

[0116] The preparation method is the same as that of Example 1, except that 3,5-difluoro-2-hydroxybenzoic acid is replaced by 2-hydroxy-3-(trifluoromethyl)benzoic acid to obtain the title compound. ESI-MS m / z: 754.4 [M + H] + 。 1 1H NMR (400 MHz, CDCl 3 ) δ 10.39 (s, 1H), 8.99 (s, 1H), 8.48 (d, J = 8.0 Hz, 1H), 7.64 (s, 2H), 7.55 (dd, J = 20, 8.0 Hz, 2H), 7.02 - 6.89 (m, 2H), 5.12 (s, 2H), 4.37 (s, 4H), 3.92 (t, J = 4.0 Hz, 2H), 3.81 (t, J = 12.0 Hz, 2H), 3.32 - 3.09 (m, 4H), 2.85 - 2.65 (m, 4H), 1.33 (t, J = 8.0 Hz, 3H).

[0117] Example 4: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(5-cyano-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0118]

[0119] The preparation method was the same as that of Example 1. Replace 3,5-difluoro-2-hydroxybenzoic acid with 5-cyano-2-hydroxybenzoic acid to obtain the title compound. ESI-MS m / z: 711.7 [M+H] + 。 1 H NMR(400MHz, DMSO-d 6 ) δ 11.21(s, 1H), 10.35(s, 1H), 8.05(d, J = 8.6Hz, 1H), 7.96(s, 1H), 7.76 - 7.60(m, 3H), 6.99(d, J = 8.6Hz, 1H), 6.83(s, 1H), 5.31(s, 2H), 4.58 - 4.47(m, 1H), 4.30 - 4.21(m, 2H), 3.86 - 3.75(m, 2H), 3.52 - 3.41(m, 2H), 3.25 - 3.17(m, 2H), 3.03 - 2.89(m, 3H), 2.81 - 2.75(m, 1H), 2.70 - 2.65(m, 1H), 2.36 - 2.29(m, 2H), 1.26 - 1.13(m, 3H).

[0120] Example 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(4-cyano-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0121]

[0122] The preparation method was the same as that of Example 1. Replace 3,5-difluoro-2-hydroxybenzoic acid with 4-cyano-2-hydroxybenzoic acid to obtain the title compound. ESI-MS m / z: 711.2 [M+H] + 。 1 H NMR(400MHz, DMSO-d 6)δ 10.75 (s, 1H), 10.36 (s, 1H), 8.06 (d, J = 8.8 Hz, 1H), 7.96 (s, 1H), 7.71 (d, J = 8.8 Hz, 1H), 7.41 - 7.27 (m, 2H), 7.20 (s, 1H), 6.83 (s, 1H), 5.31 (s, 2H), 4.58 - 4.47 (m, 1H), 4.25 (s, 2H), 3.86 - 3.75 (m, 2H), 3.53 - 3.40 (m, 2H), 3.24 - 3.14 (m, 2H), 3.04 - 2.88 (m, 3H), 2.83 - 2.74 (m, 1H), 2.69 - 2.59 (m, 2H), 2.35 - 2.31 (m, 1H), 1.24 - 1.10 (m, 3H).

[0123] Example 6: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(2-hydroxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0124]

[0125] Step 1: Preparation of methyl 4-cyano-2-methoxybenzoate

[0126]

[0127] Dissolve 4-cyano-2-methoxybenzoic acid (1.0 g, 5.64 mmol) in methanol (30 mL), add p-toluenesulfonic acid (525.7 mg, 1.467 mmol), and reflux with stirring overnight after addition. After completion of the reaction, cool to room temperature, quench with saturated aqueous sodium carbonate solution, extract with ethyl acetate, dry the organic phase with anhydrous sodium sulfate, and concentrate to obtain the title product. ESI-MS m / z: 192.1 [M + H] + .

[0128] Step 2: Preparation of methyl (Z)-4-(N'-hydroxycarbamimidoyl)amino-2-methoxybenzoate

[0129]

[0130] Hydroxylamine hydrochloride (1.47 g, 21.18 mmol) was dissolved in dimethyl sulfoxide (18 mL), sodium bicarbonate (2114.0 mg, 26.36 mmol) was added, and after reacting at 40 °C for 0.5 h, methyl 4-cyano-2-methoxybenzoate (900.0 mg, 4.70 mmol) was added. After the addition was complete, the mixture was stirred at 90 °C overnight. After the reaction was completed, it was cooled to room temperature and quenched with water, extracted with ethyl acetate, the organic phase was dried over anhydrous sodium sulfate, and concentrated to obtain the title product. ESI-MS m / z: 225.1 [M+H] + 。

[0131] Step 3: Preparation of methyl 2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoate

[0132]

[0133] (Z)-Methyl 4-(N'-hydroxycarbamoylamino)-2-methoxybenzoate (900.0 mg, 3.69 mmol) was dissolved in tetrahydrofuran (30 mL), N,N'-carbonyldiimidazole (239.0 mg, 1.47 mmol) was added, and 1,8-diazabicyclo[5.4.0]undec-7-ene (207.0 mg, 1.34 mmol) was added. After the addition was complete, the mixture was stirred at room temperature for 3 h. After the reaction was completed, it was quenched with a saturated aqueous solution of potassium bisulfate, extracted with ethyl acetate, the organic phase was dried over anhydrous sodium sulfate, and the organic phase was concentrated to obtain the title product. ESI-MS m / z: 251.1 [M+H] + 。

[0134] Step 4: Preparation of 2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoic acid

[0135]

[0136] Methyl 2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoate (700.0 mg, 2.80 mmol) was dissolved in 1,4-dioxane (16 mL), lithium hydroxide (224.8 mg, 5.60 mmol) was added, and then water (4 mL) was added. After the addition was complete, the mixture was stirred at room temperature overnight. After the reaction was completed, 10 mL of water was added, concentrated, and the pH was adjusted to 4-5 with 2N dilute hydrochloric acid, filtered by suction, the filter cake was washed with water, and the filter cake was dried to obtain the title compound. ESI-MS m / z: 237.1 [M+H] + 。

[0137] Step 5: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0138]

[0139] Dissolve N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate (120.0 mg, 0.17 mmol) in N,N-dimethylformamide (1 mL), add 2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoic acid (54.2 mg, 0.23 mmol), then sequentially add 2-(7-azabenzotriazol)-N,N,N',N'-tetramethyluronium hexafluorophosphate (73.9 mg, 0.194 mmol) and N,N-diisopropylethylamine (68.4 mg, 0.53 mmol), and stir at room temperature overnight. After the reaction is completed, quench with saturated ammonium chloride solution, extract with ethyl acetate, dry and concentrate the organic phase, and purify by reverse phase chromatography to obtain the title compound. ESI-MS m / z: 784.1 [M+H] + 。

[0140] Step 6: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(2-hydroxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0141]

[0142] N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(2-methoxy-4-(5-oxo-4,5-dihydro-1,2,4-oxadiazol-3-yl)benzoyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (90.0 mg, 0.11 mmol) was dissolved in dichloromethane (4 mL). Boron tribromide (0.5 mL) was added at -78 °C. After addition, the mixture was stirred at -78 °C for 1 hour. After completion of the reaction, water was added to quench the reaction, and the mixture was extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated, and then purified by reverse-phase chromatography to obtain the title compound. ESI-MS m / z: 770.2 [M+H] + 。 1 H NMR (400 MHz, DMSO-d 6 ): δ 12.99 (s, 1H), 10.48 (s, 1H), 10.36 (s, 1H), 8.06 (d, J = 8.0 Hz, 1H), 7.96 (s, 1H), 7.72 (d, J = 8.0 Hz, 1H), 7.42 - 7.32 (m, 2H), 7.29 (d, J = 7.7 Hz, 1H), 6.83 (s, 1H), 5.32 (s, 2H), 4.61 - 4.47 (m, 1H), 4.25 (s, 2H), 3.87 - 3.74 (m, 2H), 3.53 - 3.41 (m, 2H), 3.26 - 3.11 (m, 2H), 3.07 -.87 (m, 3H), 2.84 - 2.73 (m, 1H), 2.70 - 2.60 (m, 1H), 2.44 - 2.30 (m, 2H), 1.24 - 1.11 (m, 3H).

[0143] Example 7: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1.5-a]pyrimidin-4(7H)-yl)acetamide

[0144]

[0145] Step 1: Preparation of ethyl 4-hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carboxylate

[0146]

[0147] Ethyl 4-methylpyrazole-3-carboxylate (770.0 mg, 5.0 mmol) was dissolved in dimethyl sulfoxide (10 mL). Ethyl 4-bromocrotonate (1930 mg, 10.0 mmol) and potassium carbonate (1380 mg, 10.0 mmol) were added successively. After addition, the mixture was stirred overnight at 150 °C in an oil bath. After the reaction was completed, it was cooled to room temperature, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate and concentrated by column chromatography to obtain the title compound. ESI-MS m / z: 221.0 [M+H] + 。

[0148] Step 2: Preparation of 4-hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carboxylic acid

[0149]

[0150] Ethyl 4-hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carboxylate (800.0 mg, 3.636 mmol) was dissolved in tetrahydrofuran / water (15 / 5 mL). Lithium hydroxide (436.0 mg, 18.181 mmol) was added, and the mixture was heated to 80 °C in an oil bath and stirred overnight. After the reaction was completed, it was cooled to room temperature, 5 mL of water was added and concentrated. After concentration, the pH was adjusted to 4-5 with 2N dilute hydrochloric acid, filtered by suction, and the filter cake was washed with water. After drying the filter cake, the title compound was obtained. ESI-MS m / z: 193.1 [M+H] + 。

[0151] Step 3: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1.5-a]pyrimidin-4(7H)-yl)acetamide

[0152]

[0153] N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate (112.0 mg, 0.16 mmol) was dissolved in N,N-dimethylformamide (1 mL). 4-Hydroxy-3-methylpyrazolo[1,5-a]pyridine-5-carboxylic acid (46 mg, 0.24 mmol) was added, followed by 2-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (66.5 mg, 0.17 mmol) and N,N-diisopropylethylamine (61.5 mg, 0.48 mmol). The mixture was stirred at room temperature overnight. After completion of the reaction, the reaction was quenched with saturated ammonium chloride solution, extracted with ethyl acetate, and the organic phase was dried, concentrated and purified by reverse phase chromatography to obtain the title compound. ESI-MS m / z: 740.1 [M+H] + 。1H NMR (400 MHz, DMSO-d 6 ): δ 10.64 (s, 1H), 10.35 (s, 1H), 8.13 (d, J = 7.1 Hz, 1H), 8.06 (d, J = 8.7 Hz, 1H), 7.96 (s, 1H), 7.72 (d, J = 8.7 Hz, 2H), 6.83 (s, 1H), 6.59 (d, J = 7.1 Hz, 1H), 5.32 (s, 2H), 4.25 (s, 2H), 4.16 - 4.02 (m, 2H), 3.86 - 3.75 (m, 2H), 3.55 - 3.45 (m, 2H), 3.23 - 3.11 (m, 2H), 3.06 - 2.94 (m, 2H), 2.79 - 2.69 (m, 2H), 2.69 - 2.63 (m, 1H), 2.44 (s, 3H), 2.35 - 2.31 (m, 1H), 1.27 - 1.13 (m, 3H).

[0154] Example 8: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(3-Chloro-4-hydroxypyrazolo[1,5-a]pyridine-5-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1.5-a]pyrimidin-4(7H)-yl)acetamide

[0155]

[0156] The preparation method was the same as that of Example 7. Ethyl 4-methylpyrazole-3-carboxylate in step 1 was replaced with ethyl 4-chloropyrazole-3-carboxylate to obtain the title compound. ESI-MS m / z: 760.1 [M+H] +。 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.75 (s, 1H), 10.36 (s, 1H), 8.25 (d, J = 6.5 Hz, 1H), 8.11 - 8.01 (m, 2H), 7.97 (s, 1H), 7.72 (d, J = 7.0 Hz, 1H), 6.83 (s, 1H), 6.73 (d, J = 7.0 Hz, 1H), 5.32 (s, 2H), 4.25 (s, 2H), 4.14 - 4.00 (m, 2H), 3.86 - 3.74 (m, 2H), 3.55 - 3.43 (m, 2H), 3.24 - 3.08 (m, 3H), 3.05 - 2.92 (m, 2H), 2.79 - 2.69 (m, 2H), 2.37 - 2.29 (m, 1H), 1.24 - 1.13 (m, 3H).

[0157] Example 9: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxypyrazolo[1,5-a]pyridine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0158]

[0159] The preparation method is the same as that of Example 7. Replace ethyl 4-methylpyrazole-3-carboxylate in step 1 with ethyl 1H-pyrazole-5-carboxylate to obtain the title compound. ESI-MS m / z: 726.2 [M + H] + 。 1 H NMR (400 MHz, DMSO-d 6 ) δ 10.69 (s, 1H), 10.36 (s, 1H), 8.27 (s, 1H), 8.10 - 7.85 (m, 2H), 7.72 (s, 1H), 6.87 (d, J = 6.5 Hz, 1H), 6.71 (s, 1H), 5.46 - 5.02 (m, 2H), 4.39 - 4.13 (m, 2H), 4.13 - 3.94 (m, 1H), 3.89 - 3.63 (m, 2H), 3.61 - 3.37 (m, 2H), 3.38 - 3.30 (m, 3H), 3.27 - 3.08 (m, 2H), 3.06 - 2.85 (m, 2H), 2.81 - 2.60 (m, 2H), 1.19 (t, J = 5.3 Hz, 3H).

[0160] Example 10: 2-(6-(4-(3,5-Difluoro-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(5,6,7,8-tetrahydronaphthalen-2-yl)acetamide

[0161]

[0162] Step 1: Preparation of tert-butyl 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4-(2-oxo-2-((5,6,7,8-tetrahydronaphthalen-2-yl)amino)ethyl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate

[0163]

[0164] Dissolve tert-butyl 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (1.5 g, 3.49 mmol) and 2-chloro-N-(5,6,7,8-tetrahydronaphthalen-2-yl)acetamide (0.93 g, 4.19 mmol) in 11 ml of N,N-dimethylformamide, slowly add N,N-diisopropylethylamine (1.4 g, 10.5 mmol), and after addition, heat to 100 °C and react for 6 hours. After the reaction is completed, cool to room temperature, add saturated ammonium chloride solution, extract with ethyl acetate, dry the organic phase, and concentrate and purify by column chromatography to obtain the title compound. ESI-MS m / z: 618.7 [M+H] + .

[0165] Step 2: Preparation of 2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(5,6,7,8-tetrahydronaphthalen-2-yl)acetamide trifluoroacetate

[0166]

[0167] tert-Butyl 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4-(2-oxo-2-((5,6,7,8-tetrahydronaphthalen-2-yl)amino)ethyl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate (730 mg, 1.18 mmol) was dissolved in 14.6 ml of anhydrous dichloromethane, and then 1.46 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 1 hour. After the reaction was completed, it was concentrated to obtain the title compound. ESI-MS m / z: 518.1 [M - 114 + H] + 。

[0168] Step 3: Preparation of 2-(6-(4-(3,5-difluoro-2-hydroxybenzoyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(5,6,7,8-tetrahydronaphthalen-2-yl)acetamide

[0169]

[0170] 3,5-Difluoro-2-hydroxybenzoic acid (226 mg, 1.3 mmol) was dissolved in 5 ml of N,N-dimethylformamide. 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide (249 mg, 1.3 mmol) and 1-hydroxybenzotriazole (176 mg, 1.3 mmol) were added successively. After stirring at room temperature for 30 minutes, N,N-diisopropylethylamine (0.62 ml) and 2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(5,6,7,8-tetrahydronaphthalen-2-yl)acetamide trifluoroacetate (758 mg, 1.2 mmol) were added. After the addition, the mixture was stirred at room temperature for 2 hours. After the reaction was completed, the reaction solution was poured into water for quenching, extracted with ethyl acetate, and the combined organic phases were dried and concentrated. Column chromatography was used to obtain the title compound. ESI-MS m / z: 674.3 [M + H] + 。 1 H NMR(400MHz,CDCl 3)δ8.63(s,1H),8.45(s,1H),7.21(s,1H),7.09(d,J=4.0Hz,1H),7.04 - 6.88(m,3H),6.84(d,J=4.0Hz,1H),5.15 - 4.86(m,2H),4.48 - 4.17(m,4H),3.98 - 3.85(m,2H),3.83 - 3.64(m,2H),3.36 - 3.01(m,4H),2.82 - 2.55(m,8H),1.83 - 1.68(m,4H),1.38 - 1.18(m,3H).

[0171] Example 11: N-(6-(4-(4-(2-(2-Chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carbonyl)-5-hydroxypyridazin-3-yl)cyclopropanecarboxamide

[0172]

[0173] Step 1: Preparation of methyl 6-chloro-4-methoxypyridazine-3-carboxylate

[0174]

[0175] Dissolve methyl 4,6-dichloropyridazine-3-carboxylate (3.0 g, 14.49 mmol) in 50 mL of tetrahydrofuran, cool to -10 °C, add sodium methoxide in methanol solution (2.6 mL, 14.49 mmol), and stir the reaction at room temperature for 5.0 hours. After the reaction is completed, quench with 1N HCl aqueous solution, extract with ethyl acetate, dry the organic phase with anhydrous sodium sulfate and concentrate by column chromatography to obtain the title compound. ESI-MS m / z: 203.1 [M + H] + .

[0176] Step 2: Preparation of methyl 6-(cyclopropanecarboxamide)-4-methoxypyridazine-3-carboxylate

[0177]

[0178] Methyl 6-chloro-4-methoxypyridazine-3-carboxylate (2.28 g, 11.23 mmol), cyclopropanecarboxamide (1.43 g, 16.85 mmol), (2'-amino-1,1'-biphenyl-2-yl)palladium(II) (2-dicyclohexylphosphino)-3,6-dimethoxy-2',4',6'-triisopropyl-1,1'-biphenyl] (2.04 g, 2.25 mmol), 2-(dicyclohexylphosphino)-3,6-dimethoxy-2'-4'-6'-tri-I-propyl-11'-biphenyl (1.21 g, 2.25 mmol), and cesium carbonate (7.28 g, 22.4 mmol) were dissolved in 100 mL of anhydrous dioxane. After protection with argon, the mixture was stirred at 100 °C in an oil bath for 5 h. After the reaction was completed, the mixture was concentrated, water and ethyl acetate were added for extraction, the organic phase was dried over anhydrous sodium sulfate, and the title compound was obtained by column chromatography after concentration. ESI-MS m / z: 252.1 [M+H] + 。

[0179] Step 3: Preparation of 6-(cyclopropanecarboxamido)-4-methoxypyridazine-3-carboxylic acid

[0180]

[0181] Methyl 6-(cyclopropanecarboxamido)-4-methoxypyridazine-3-carboxylate (1.0 g, 3.98 mmol) was dissolved in 30 mL of tetrahydrofuran, and then an aqueous lithium hydroxide solution (382 mg in 30 mL H 2 O) was added. After the addition, the mixture was stirred at room temperature for 1 h. After the reaction was completed, the reaction solution was concentrated, and the pH was adjusted to 3-4 with 2N dilute hydrochloric acid. The title compound was obtained by reverse-phase column chromatography. ESI-MS m / z: 238.1 [M+H] + 。

[0182] Step 4: Preparation of N-(6-(4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carbonyl)-5-methoxypyridazin-3-yl)cyclopropanecarboxamide

[0183]

[0184] N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate (126 mg, 0.18 mmol) was dissolved in N,N-dimethylformamide (5 mL). 6-(Cyclopropanecarboxamido)-4-methoxypyridazine-3-carboxylic acid (43 mg, 0.18 mmol) was added, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (68 mg, 0.18 mmol) and N,N-diisopropylethylamine (70 mg, 0.54 mmol). The reaction mixture was stirred at room temperature for 1 h. After completion of the reaction, the reaction was quenched with saturated ammonium chloride solution, extracted with ethyl acetate, the organic phase was dried over anhydrous sodium sulfate and concentrated. The residue was purified by reverse-phase chromatography to obtain the title compound. ESI-MS m / z: 785.1 [M+H] + 。

[0185] Step 5: Preparation of N-(6-(4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carbonyl)-5-hydroxypyridazin-3-yl)cyclopropanecarboxamide

[0186]

[0187] N-(6-(4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carbonyl)-5-methoxypyridazin-3-yl)cyclopropanecarboxamide (80 mg, 0.10 mmol) was dissolved in 5 mL of N,N-dimethylformamide. Trimethylsilyl iodide (27 mg, 0.13 mmol) was added and the reaction mixture was stirred at 85 °C in an oil bath for 8 h. After completion of the reaction, the reaction solution was cooled to room temperature, poured into ice water, extracted with ethyl acetate, the organic phase was dried over anhydrous sodium sulfate, concentrated and purified by column chromatography to obtain the title compound. ESI-MS m / z: 771.1 [M+H] + 。

[0188] 1 H NMR(400MHz,DMSO-d 6)δ11.21(s,1H),10.34(s,1H),8.05(d,J=8.6Hz,1H),7.95(s,1H),7.71(d,J=7.7Hz,1H),6.83(s,1H),5.31(s,2H),4.50(s,1H),4.25(d,J=2.3Hz,2H),3.80(t,J=5.4Hz,2H),3.51 - 3.39(m,2H),3.36(d,J=12.1Hz,1H),3.24(t,J=11.4Hz,1H),3.02 - 2.90(m,3H),2.80(d,J=9.9Hz,1H),2.63(d,J=9.7Hz,1H),2.02 - 1.90(m,1H),1.26 - 1.16(m,4H),0.94 - 0.83(m,5H).

[0189] Example 12: N-(2-Chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(6-(6-(2-Chloro-4-fluorophenyl)-4-hydroxypyridazin-3-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0190]

[0191] Step 1: Preparation of methyl 6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carboxylate

[0192]

[0193] Methyl 6-chloro-4-methoxypyridazine-3-carboxylate (800 mg, 3.96 mmol), 2-(2-chloro-4-fluorophenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (1.0 g, 3.96 mmol), tetrakis(triphenylphosphine)palladium (229 mg, 0.2 mmol), and sodium carbonate (839 mg, 7.92 mmol) were dissolved in a mixed solvent of 40 ml of 1,4-dioxane and 4 ml of water. After argon protection, the mixture was stirred at 80 °C in an oil bath for 2.5 hours. After the reaction was completed, the mixture was concentrated, water and ethyl acetate were added for extraction, the organic phase was dried over anhydrous sodium sulfate, and after drying, it was concentrated and purified by column chromatography to obtain the title compound. ESI-MS m / z: 297.1 [M + H] + .

[0194] Step 2: Preparation of 6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carboxylic acid

[0195]

[0196] Dissolve methyl 6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carboxylate (190 mg, 0.64 mmol) in a mixed solution of 6 mL of tetrahydrofuran and 2 mL of water. Add lithium hydroxide (38 mg, 1.28 mmol). After addition, stir the reaction at room temperature for 0.5 hour. After the reaction is completed, concentrate the reaction solution. After concentration, adjust the pH to 3 - 4 with 2N dilute hydrochloric acid, extract with ethyl acetate, dry the organic phase with anhydrous sodium sulfate, and concentrate after drying to obtain the title compound. ESI-MS m / z: 283.1 [M + H] + 。

[0197] Step 3: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(6-(6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0198]

[0199] Dissolve N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide trifluoroacetate (160 mg, 0.23 mmol) in N,N-dimethylformamide (5 mL). Add 6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carboxylic acid (70 mg, 0.25 mmol), then successively add 2-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (140 mg, 0.37 mmol) and N,N-diisopropylethylamine (89 mg, 0.69 mmol). Stir the reaction at room temperature for 0.5 hour. After the reaction is completed, quench with saturated ammonium chloride solution, extract with ethyl acetate, dry the organic phase with anhydrous sodium sulfate and concentrate. After concentration, purify by reverse-phase chromatography to obtain the title compound. ESI-MS m / z: 830.8 [M + H] + 。

[0200] Step 4: Preparation of N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(6-(6-(2-chloro-4-fluorophenyl)-4-hydroxypyridazine-3-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0201]

[0202] Dissolve N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(6-(4-(6-(6-(2-chloro-4-fluorophenyl)-4-methoxypyridazine-3-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide (105 mg, 0.13 mmol) in 5 ml of N,N-dimethylformamide, add trimethylsilyl iodide (33 mg, 0.16 mmol), and stir the reaction at 85 °C in an oil bath for 12 hours. After the reaction is completed, cool the reaction solution to room temperature, pour it into ice water, extract with ethyl acetate, dry the organic phase with anhydrous sodium sulfate, concentrate, and perform column chromatography to obtain the title compound. ESI-MS m / z: 816.1 [M+H] + 。 1 H NMR (400 MHz, DMSO-d 6 ) δ 13.66 (s, 1H), 10.35 (s, 1H), 8.06 (d, J = 8.6 Hz, 1H), 7.96 (s, 1H), 7.73 (dd, J = 12.4, 6.9 Hz, 3H), 7.45 (dd, J = 9.5, 7.4 Hz, 1H), 6.83 (s, 1H), 6.49 (s, 1H), 5.32 (s, 2H), 4.50 (d, J = 12.4 Hz, 1H), 4.26 (s, 2H), 3.79 - 3.81 (m, 2H), 3.50 - 3.45 (m, 3H), 3.32 - 3.30 (m, 1H), 3.01 - 2.96 (m, 3H), 2.81 (d, J = 10.3 Hz, 1H), 2.69 (d, J = 10.6 Hz, 1H), 1.24 - 1.17 (m, 3H).

[0203] The compounds of Examples 13 - 25 in Table 1 were prepared by referring to the synthesis method of Example 1 of the present invention using different raw materials.

[0204] Table 1

[0205]

[0206]

[0207]

[0208] Experimental Example 1: Evaluation of WRN Enzyme Activity

[0209] 1 Experimental Materials

[0210] 1.1 Experimental Reagents and Instruments

[0211] Experimental Instruments: 655 SYSTEM (LABCYTE, 655); Centrifuge (Cence, TDZ5-WS); Microplate Reader (BMG, PHERAstar FSX); Optiplate-384 (PE, 6057480); 96-Well Plates (Nunc, 249944).

[0212] Experimental reagents: ADP-Glo TM Kinase Assay (Promega, V9102); dsDNA (Genscript, / ); WRN (517-1238) (ICE, S2212T-H56HZ); NSC617145 (MCE, HY-110185).

[0213] 1.2 Preparation of compound solutions

[0214] All compounds were prepared as 10 mM stock solutions in DMSO. Subsequently, starting from the highest initial concentration of 20 μM, 10 concentration gradients were serially diluted 3-fold with DMSO, and centrifuged at 1000 g for 1 min for subsequent experiments.

[0215] 2 Experimental methods

[0216] 2.1. Prepare 500× compounds in a 96-well plate, then transfer 40 μL to a new 96-well plate for 2-fold dilution. Then transfer 50 μL of the compound or DMSO to a 384-well dilution plate per well;

[0217] 2.2. In the 384-well dilution plate, the compounds were serially diluted 10 concentration gradients at a ratio of 1:3 in DMSO;

[0218] 2.3. Use Echo to transfer 0.1 μL of the diluted compound solution per row to 384 assay plates, with 2 replicates;

[0219] 2.4. Add 5 μL of the enzyme working solution to the 384-well assay plates and centrifuge at 1000 rpm for 1 min;

[0220] 2.5. Incubate at 25 °C for 10 min;

[0221] 2.6. Add 5 μL of the substrate (ATP and dsDNA) working solution to initiate the reaction;

[0222] 2.7. Incubate at 25 °C for 60 min;

[0223] 2.8. Add 5 μL of the ADP-Glo working solution to initiate the reaction;

[0224] 2.9. Incubate at 25 °C for 40 minutes;

[0225] 2.10. Add 10 μL of the detection working solution to initiate the reaction;

[0226] 2.11. Incubate at 25 °C for 40 minutes;

[0227] 2.12. Read the fluorescence signal with a BMG. Calculate using the formula: inh% = 100 * (ave High control - cpdwell) / (ave High control - ave Low control). After fitting the data, the IC50 value is obtained. The experimental results are shown in Table 2.

[0228] Table 2

[0229]

[0230]

[0231] The experimental results show that the compounds of the present invention have good inhibitory activity against the WRN enzyme.

[0232] Experimental Example 2: Evaluation of the in vitro cell activity of the compound

[0233] 1. Experimental materials

[0234] Experimental cells: Human colorectal cancer cells HCT116 were purchased from Wuhan Procell Life Science & Technology Co., Ltd.

[0235] Test compounds: The compounds of the present invention prepared in the above examples. Each compound was formulated into a 20 mM stock solution with DMSO, and the drug was diluted with complete medium. The final drug concentration started from the highest concentration of 5 μM and was serially diluted to 9 concentration gradients at a 1:3 ratio: 5 μM, 1.25 μM, 1.11 μM, 0.313 μM, 0.0195 μM, 0.00488 μM, 0.00122 μM, 0.000305 μM, 0.000076 μM.

[0236] Reagents: McCoy’s 5A (Gibco, 16600 - 082); Fetal Bovine Serum (Gibco, 10099 - 141); Penicillin - Streptomycin (Gibco, 15140 - 122); CellTiter - Glo Luminescent Cell Viability Assay (Promega, G7571).

[0237] 2. Experimental method

[0238] 2.1 Cell resuscitation:

[0239] Take out the HCT116 cell cryopreservation tube from the liquid nitrogen tank and place it in a 37°C water bath. Gently shake it to thaw as soon as possible. After thawing, take out the cryopreservation tube, disinfect it with an alcohol cotton ball, unscrew the cap, suck out the cell suspension and inject it into a centrifuge tube, and add 1 mL of McCoy's 5A medium containing 10% FBS. Mix well and place it in a centrifuge, centrifuge at 1000 rpm for 5 min. Then discard the supernatant, add complete medium to resuspend the cells. Inoculate the cells in a culture dish at an appropriate concentration. Incubate at 37°C, 5% CO 2 Incubate in an incubator.

[0240] 2.2 Cell culture and passage:

[0241] When the cells grow to a confluence of about 80 - 90%, aspirate and discard the original culture medium (McCoy's 5A medium + 10% FBS + 1% penicillin - streptomycin), add 1 mL of PBS to wash the residual medium and then aspirate and discard it. Add 1 mL of trypsin digestion solution to digest for 2 - 3 min. Observe under the microscope that the cell pseudopods retract and become round but the cells have not yet detached in sheets. At this time, aspirate the trypsin and use 1 - 2 mL of complete medium to terminate the digestion. Gently pipette and collect the cell suspension, centrifuge at 1000 rpm for 5 min. Remove the supernatant, resuspend the cells with complete medium, inoculate the cells in a culture dish at the required density, and place it in an incubator at 37°C, 5% CO 2 Incubate in an incubator. Change the culture medium or passage every 2 - 3 days according to the cell growth situation.

[0242] 2.3 Experimental procedures:

[0243] Day 1 of the experiment:

[0244] After sub - culturing, resuspend the HCT116 cells in complete medium at the corresponding density and inoculate them into a 96 - well culture plate: Fill 36 outer - most wells of the 96 - well plate with 200 μL of PBS to prevent the culture conditions in the internal wells from being too different due to faster evaporation of the medium at the edges; Plate the remaining 60 wells; Each well contains 100 μL of complete medium with 750 cells, and set 2 replicates for each concentration. Place it in a 5% CO 2 Incubate at 37°C in an incubator for 24 h.

[0245] Day 2 of the experiment:

[0246] Based on the original medium (100 μL) of HCT116 cells, add 100 μL of (2x) drug, set two replicates for each concentration group, and continue to place it in a 5% CO 2 Incubate in an incubator for 4 days.

[0247] Day 5 of the experiment:

[0248] After 4 days of treating HCT116 cells, take out the CellTiter-Glo Luminescent Cell Viability Assay 30 min in advance and equilibrate it to room temperature. Aspirate the culture medium in the wells, dilute the Celltiter-Glo reagent with complete medium at a ratio of 1:1 (equal volume), add 150 μL of the diluted Celltiter-Glo reagent to each well, and shake at room temperature for 2 min. After continuing to incubate at room temperature for 10 min, take 100 μL from each well to detect the chemiluminescence signal, shake, and the injection detection condition for Read is 500 ms. According to the A.U. value exported by the microplate reader, calculate the inhibition rate of each well relative to the solvent control well: Inhibition (%) = 100 - (A.U. experimental well - A.U. blank well) / (A.U. solvent control well - A.U. blank well) * 100. According to different drug concentrations and their corresponding inhibition rates, use GraghPad 6.0 software to fit a four-parameter logistic S-shaped curve, analyze the data and obtain the IC 50 value. The experimental results are shown in Table 3.

[0249] Table 3

[0250]

[0251] The experimental results show that the compounds of the present invention have good proliferation inhibitory activity against human colorectal cancer cell line HCT116.

[0252] Although the present invention has been described in detail above, those skilled in the art understand that various modifications and changes can be made to the present invention without departing from the spirit and scope of the present invention. The scope of the rights of the present invention is not limited to the detailed description above, but should be attributed to the claims.

Claims

1. A compound represented by general formula (I) or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, in, Ring A is selected from cycloalkyl, heterocyclyl, aryl and heteroaryl; R 1 is selected from the group consisting of hydrogen, halogen, cyano, amino, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclyl, heterocycloalkenyl, alkoxy, haloalkoxy, hydroxyalkoxy, monoalkylamino, dialkylamino, alkylacylamino, alkylacyl, aminoacyl, alkylaminoacyl, dialkylamino, heterocyclyl, aryl and heteroaryl, the alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclyl, heterocycloalkenyl, alkoxy, haloalkoxy, hydroxyalkoxy, monoalkylamino, dialkylamino, alkylacylamino, Alkylacyl, aminoacyl, alkylaminoacyl, dialkylamino, aryl and heteroaryl are optionally substituted with one or more groups selected from halogen, hydroxy, alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, amino, monoalkylamino, alkylacylamino, alkylacyl, alkylsulfonyl, aminoacyl, alkylaminoacyl, dialkylamino, alkenyl, alkynyl, haloalkylacyl, hydroxyalkylacyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, oxo; R 2 is selected from aryl, heteroaryl, cycloalkyl and heterocyclyl, wherein the aryl, heteroaryl, cycloalkyl and heterocyclyl are optionally substituted with one or more groups selected from halogen, hydroxy, cyano, amino, alkyl, haloalkyl, alkoxy, haloalkoxy, cycloalkyl and heterocyclyl; R 3 is selected from the group consisting of alkyl, haloalkyl, cycloalkyl, heterocyclyl and hydroxyalkyl; Each R 4 The alkyl, alkyl, alkyl halide, alkyl hydroxy, alkyl alkoxy, alkyl halide, alkyl hydroxy, alkyl sulfonyl, alkyl amino, alkyl amino, alkyl acyl, alkyl acyl, alkyl sulfonyl, alkyl amino ... alkylamino, alkylacylamino, alkylacyl, alkylsulfonyl, aminoacyl, alkylaminoacyl, dialkylamino, alkenyl, alkynyl, haloalkylacyl, hydroxyalkylacyl, cycloalkylacyl, heterocyclylacyl, cycloalkyl, halocycloalkyl, heterocyclyl, aryl, heteroaryl, and oxo are optionally substituted with one or more groups selected from halogen, hydroxy, alkyl, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, hydroxyalkoxy, amino, cycloalkyl, halocycloalkyl, heterocyclyl, aryl, heteroaryl, and oxo, and m is selected from 0, 1, 2, 3, and 4; and L is selected from -CH2-, -CH2CH2-, -CH2NH-, -CH2C(O)NH- and -C(O)NH-.

2. The compound according to claim 1 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein ring A is selected from C 6-12 Cycloalkyl, 3-13 membered heterocyclic group, C 6-12 Aryl and 5-13 membered heteroaryl.

3. The compound according to claim 1 or 2, or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein R 1 Selected from hydrogen, halogen, cyano, amino, C 1-6 Alkyl, C 3-10 Cycloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, 3-10 membered heterocyclic group, 3-10 membered heterocyclic alkenyl group, C 1-6 Alkoxy, halogenated C 1-6 Alkoxy, hydroxyl C 1-6 Alkoxy, single C 1-6 Alkylamino, di-C 1-6 Alkylamino, C 1-6 Alkyl acylamino, C 1-6 Alkyl acyl, amino acyl, C 1-6 Alkylaminoacyl, di-C 1-6 Alkylamino, C 6-10 Aryl and 5-10 membered heteroaryl, the C 1-6 Alkyl, C 3-10 Cycloalkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, 3-10 membered heterocyclic group, 3-10 membered heterocyclic alkenyl group, C 1-6 Alkoxy, halogenated C 1-6 Alkoxy, hydroxyl C 1-6 Alkoxy, single C 1-6 Alkylamino, di-C 1-6 Alkylamino, C 1-6 Alkyl acylamino, C 1-6 Alkyl acyl, amino acyl, C 1-6 Alkylaminoacyl, di-C 1-6 Alkylamino, C 6-10 The aryl and 5-10 membered heteroaryl groups may be optionally substituted by one or more selected from halogen, hydroxyl, C 1-6 Alkyl, halogenated C 1-6 Alkyl, hydroxyl C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkoxy, hydroxyl C 1-6 Alkoxy, amino, mono C 1-6 Alkylamino, C 1-6 Alkyl acylamino, C 1-6 Alkyl acyl, C 1-6 Alkylsulfonyl, aminoacyl, C 1-6 Alkylaminoacyl, di-C 1-6 Alkylamino, alkenyl, alkynyl, halo C 1-6 Alkyl acyl, hydroxyl C 1-6 Alkyl acyl, C 3-10 Cycloalkyl, 3-10 membered heterocyclic group, C 6-10 substituted with 5- to 10-membered aryl, 5- to 10-membered heteroaryl, or oxo; 2 Selected from C 6-10 Aryl, 5-13 membered heteroaryl, C 3-13 Cycloalkyl and 3-13 membered heterocyclic group, the C 6-10 Aryl, 5-13 membered heteroaryl, C 3-13 The cycloalkyl and 3-13 membered heterocyclic groups are optionally substituted by one or more selected from halogen, hydroxy, cyano, amino, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkoxy, C 3-8 substituted with cycloalkyl and 3-10 heterocyclic groups; and R 3 Selected from C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 3-8 Cycloalkyl, 3-8 membered heterocyclic group and hydroxyl C 1-6 alkyl.

4. The compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein Ring A is selected from Each R 4 are independently selected from halogen, hydroxyl, C 1-3 Alkyl, halogenated C 1-3 Alkyl, hydroxyl C 1-3 Alkyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, hydroxyl C 1-3 Alkoxy, nitro, carboxyl, cyano, amino, mono C 1-3 Alkylamino, C 1-3 Alkyl acylamino, C 1-3 Alkyl acyl, C 1-3 Alkylsulfonyl, aminoacyl, C 1-3 Alkylaminoacyl, di-C 1-3 Alkylamino, C 2-6 Alkenyl, C 2-6 Alkynyl, halo C 1-3 Alkyl acyl, hydroxyl C 1-3 Alkyl acyl, C 3-8 Cycloalkyl acyl, 3-8 membered heterocyclic acyl, C 3-8 Cycloalkyl, halogenated C 3-8 Cycloalkyl, 3-8 membered heterocyclic group, C 6-8 Aryl, 5-8 membered heteroaryl and oxo group, the C 1-3 Alkyl, halogenated C 1-3 Alkyl, hydroxyl C 1-3 Alkyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, hydroxyl C 1-3 Alkoxy, single C 1-3 Alkylamino, C 1-3 Alkyl acylamino, alkyl acyl, C 1-3 Alkylsulfonyl, aminoacyl, C 1-3 Alkylaminoacyl, di-C 1-3 Alkylamino, C 2-6 Alkenyl, C 2-6 Alkynyl, halo C 1-3 Alkyl acyl, hydroxyl C 1-3 Alkyl acyl, C 3-8 Cycloalkyl acyl, 3-8 membered heterocyclic acyl, C 3-8 Cycloalkyl, halogenated C 3-8 Cycloalkyl, 3-8 membered heterocyclic group, C 6-8 Aryl, 5-8 membered heteroaryl, optionally substituted with one or more selected from halogen, hydroxyl, C 1-3 Alkyl, halogenated C 1-3 Alkyl, hydroxyl C 1-3 Alkyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, hydroxyl C 1-3 Alkoxy, amino, C 3-8 Cycloalkyl, halogenated C 3-8 Cycloalkyl, 3-8 membered heterocyclic group, C 6-8 The aryl, 5-8 membered heteroaryl and oxo groups are substituted, and m is selected from 0, 1, 2, 3 and 4.

5. The compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein R 2 Selected from It is optionally substituted by one or more selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halogenated C 1-3 Alkyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, C 3-8 substituted by a cycloalkyl, a 3-8 membered heterocyclic group and an oxo group; further, it is optionally substituted by one or more selected from fluorine, chlorine, bromine, iodine, hydroxyl, cyano, amino, methyl, ethyl, propyl, isopropyl, halogenated C 1-3 Alkyl groups such as trifluoromethyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, C 3-8 The cycloalkyl group, the 3- to 8-membered heterocyclic group and the oxo group are substituted.

6. The compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein R 1 Selected from It is optionally substituted by one or more selected from halogen, hydroxy, cyano, amino, C 1-3 Alkyl, halogenated C 1-3 Alkyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, C 3-8 substituted by a cycloalkyl, a 3-8 membered heterocyclic group and an oxo group; further, it is optionally substituted by one or more selected from fluorine, chlorine, bromine, iodine, hydroxyl, cyano, amino, methyl, ethyl, propyl, isopropyl, halogenated C 1-3 Alkyl groups such as trifluoromethyl, C 1-3 Alkoxy, halogenated C 1-3 Alkoxy, C 3-8 The cycloalkyl group, the 3- to 8-membered heterocyclic group and the oxo group are substituted.

7. The compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein the general formula (I) has the structure of the following general formula (Ia), in, R 1 , R 2 , R 3 , R 4 , m and L have the definitions in the general formula (I) in claims 1 to 6, X 1 , X 2 and X 3 Each independently selected from N and -C(R 5 )-, R 5 Selected from hydrogen, halogen, hydroxyl, cyano, amino, C 1-3 Alkyl, halogenated C 1-3 Alkyl, hydroxyl C 1-3 Alkyl, C 1-3 Alkoxy and halogenated C 1-3 Alkoxy; Ring B is selected from C 3-8 Cycloalkyl, 3-10 membered heterocyclic group, C 6-8 Aryl and 5-8 membered heteroaryl.

8. The compound according to claim 1 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof, wherein the compound is selected from the following compounds:

9. A pharmaceutical composition comprising the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof and a pharmaceutically acceptable carrier.

10. Use of the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt, isomer, solvate, crystal or prodrug thereof or the pharmaceutical composition according to claim 9 in the preparation of a medicament for treating a disease associated with WRN, preferably the disease is cancer.

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