Substituted pyridinone compounds and uses

Abstract

The present application provides compounds of general formula (A) or pharmaceutically acceptable salts thereof, pharmaceutical compositions and methods of preparation, and uses as MAT2A inhibitors.

Description

[0001] This invention claims priority to the following earlier applications: Patent Application No. 202011105645.3, filed with the China National Intellectual Property Administration (CNIPA) on October 15, 2020, entitled "Substituted Pyridone Compounds and Their Applications"; Patent Application No. 202110182410.2, filed with the CNIPA on February 10, 2021, entitled "Substituted Pyridone Compounds and Their Applications"; and Patent Application No. 202110361446.7, filed with the CNIPA on April 2, 2021, entitled "Substituted Pyridone Compounds and Their Applications". The entire contents of these earlier applications are incorporated herein by reference. Technical Field

[0002] This invention belongs to the pharmaceutical field and relates to a novel pyridone compound or a pharmaceutically acceptable salt thereof, pharmaceutical compositions containing them, and their use as MAT2A inhibitors. Background Technology

[0003] Methionine adenosyltransferase (MAT), also known as S-adenosylmethionine synthase, is a class of enzymes that catalyze the reaction of methionine (Met) with ATP to produce S-adenosyl-L-methionine (SAM). SAM is a major methyl donor in the body and regulates gene expression, transcription, and translation through transmethylation reactions, thus significantly impacting cell growth, death, and differentiation. Furthermore, SAM also participates in the biosynthesis of polyamines and glutathione.

[0004] There are three main subtypes of MAT enzymes: MAT1A, MAT2A, and MAT2B. MAT1A is mainly found in normal hepatocytes, while MAT2A is widely distributed in extrahepatic cells. These two subtypes differ in catalytic efficiency and regulatory mechanisms. MAT2B does not have the ability to catalyze the synthesis of SAM; instead, it acts as a regulatory subunit of MAT2A, forming a complex with MAT2A to regulate its catalytic activity.

[0005] Studies have shown that in liver cancer cells, downregulation of MAT1A expression and upregulation of MAT2A expression promote the proliferation of liver cancer cells. In addition, abnormally high MAT2A expression levels are also observed in many other tumors, and silencing the gene encoding MAT2A can lead to cancer cell death. Furthermore, Marjon et al. (MTAP deletions in cancer create vulnerability to targeting of the MAT2A / PRMT5 / RIOK1axis. Marjon K, et al. Cell Reports. 2016, 15(3), 574–587) found that cancer cell lines lacking MTAP are sensitive to MAT2A inhibition. MTAP, also known as methionine phosphorylase, is widely expressed in normal tissue cells. This enzyme catalyzes the conversion of methionine (MTA) into 5-methylthioribose-1-phosphate and adenine. This process is also an important step in the methionine compensation pathway in the human body. When MTAP is lost, the metabolic pathway of MTA is inhibited, leading to a large accumulation of MTA in the body, which ultimately increases the sensitivity of cancer cells to MAT2A inhibition.

[0006] The gene encoding human MTAP is located in the 9p21 region of chromosome 9 (chr9p21). Its homozygous deletion frequency is approximately 15% across all tumors, and varies among different tumor types. Tumors with higher deletion frequencies include glioma, mesothelioma, melanoma, gastric cancer, esophageal cancer, bladder cancer, pancreatic cancer, non-small cell lung cancer, astrocytoma, osteosarcoma, head and neck cancer, myxoid chondrosarcoma, ovarian cancer, endometrial cancer, breast cancer, soft tissue sarcoma, and non-Hodgkin's lymphoma.

[0007] The 9p21 region of human chromosome contains not only the gene encoding MTAP, but also the tumor suppressor genes p16INK4A (also known as CDKN2A) and p15INK4B. MTAP is also absent in 80%-90% of tumors with CDKN2A deletion.

[0008] Given that MAT2A expression levels are abnormally elevated in various tumor types, including gastric cancer, colon cancer, liver cancer, and pancreatic cancer, and that selective inhibition of MAT2A can reduce the proliferative activity of MTAP-deficient cancer cells, selective inhibition of MAT2A could be considered an effective cancer treatment.

[0009] WO2018039972, WO2018045071 and WO2019191470 disclose heterocyclic compounds that are MAT2A inhibitors for the treatment of tumors. Summary of the Invention

[0010] This invention provides a compound of formula (A) or a pharmaceutically acceptable salt thereof:

[0011]

[0012] Among them, ring Q is a 5-6 member heterocyclic group, C6-C 10 Aryl or 5-10 membered heteroaryl, wherein the 5-6 membered heterocyclic group, C6-C 10 Aryl or 5-10 heteroaryl groups are selectively coated with R. a Instead, the R a Selected from F, Cl, Br, I, OH, CN, C2-C3 alkynyl groups, C1-C 10 Alkyl, C3-C 10 cycloalkyl or C1-C 10 Alkoxy, the C1-C 10 Alkyl, C3-C 10 cycloalkyl or C1-C 10 Alkoxy groups are optionally replaced by R b replace;

[0013] R b Selected from F, Cl, Br, I, OH or CN;

[0014] The ring W can be phenyl, pyridinyl, pyrimidinyl, pyridazinyl, pyridinoneyl, or pyridazinoneyl. Wherein ring M is a 5-10 membered heteroaryl or a 4-10 membered heterocyclic group, and the phenyl, pyridyl, pyrimidinyl, pyridinoneyl, pyridinoneyl or ring M is optionally replaced by R. c Instead, the R c Selected from OH, =O, or arbitrarily subjected to R c1 The following groups are substituted: C1-C 10 Alkyl, C1-C 10 Alkoxy, C3-C 10 Cycloalkyl, P(O)(C1-C3 alkyl)2, 4-6 membered heterocycloalkyl, NHC(O)(C1-C6 alkyl); X1 is selected from C or N, when X1 is selected from C, This indicates a double bond, where X1 is selected from N. Indicates a single key;

[0015] R c1 Selected from deuterium, F, Cl, Br, I, CN, OH, NH2, or optionally R c2 The following groups are substituted: C1-C3 alkyl, C1-C3 alkoxy, NHC(O)O (C1-C6 alkyl), (C1-C6 alkyl)NHC(O)O, NH (C1-C3 alkyl), S(O)2 (C1-C3 alkyl), 4-10 membered heterocyclic groups, 4-10 membered heterocyclic oxy groups, (C3-C10 Cycloalkyl)CH2O, 5-6-membered heteroaryl, 5-6-membered heteroaryloxy;

[0016] R c2 Selected from 4-6 membered heterocyclic alkyl, =O, C1-C3 alkyl, halo-C1-C3 alkyl, F, Cl, Br, I, CN, OH, CH2OH or NH2;

[0017] X is selected from O, S, or NR. 2 ;

[0018] L is selected from O, NH, or chemical bonds;

[0019] R 1 R 2 Independently selected from H, C1-C 10 Alkyl, C3-C 10 Cycloalkyl or 4-10 membered heterocyclic groups, wherein the C1-C 10 Alkyl, C3-C 10 Cycloalkyl or 4-10 membered heterocyclic groups are optionally R d Instead, the R d Selected from F, Cl, Br, I, OH, CN or C1-C3 alkyl groups.

[0020] In some implementations, ring Q is a 5-6 membered heterocyclic group, C6-C 10 Aryl or 5-10 membered heteroaryl, wherein the 5-6 membered heterocyclic group, C6-C 10 Aryl or 5-10 heteroaryl groups are selectively coated with R. a Instead, the R a Selected from F, Cl, Br, I, OH, CN, or optionally R b The following groups are substituted: C1-C 10 Alkyl, C3-C 10 cycloalkyl, C1-C 10 Alkoxy;

[0021] R b Selected from F, Cl, Br, I, OH or CN;

[0022] The ring W can be phenyl, pyridinyl, pyridinone, pyridazinone, or... Wherein ring M is a 5-6 membered heteroaryl or a 4-10 membered heterocyclic group, and the phenyl, pyridyl, pyridone, pyridazinone, or ring M is optionally replaced by R. c Instead, the R c Selected from OH, =O, or arbitrarily subjected to R c1 The following groups are substituted: C1-C 10 Alkyl, C1-C 10 Alkoxy, C3-C 10Cycloalkyl, P(O)(C1-C3 alkyl)2, 4-6 membered heterocycloalkyl, NHC(O)(C1-C6 alkyl); X1 is selected from C or N, when X1 is selected from C, This indicates a double bond, where X1 is selected from N. Indicates a single key;

[0023] R c1 Selected from deuterium, F, Cl, Br, I, CN, OH, NH2, or optionally R c2 The following groups are substituted: C1-C3 alkyl, C1-C3 alkoxy, NHC(O)O (C1-C6 alkyl), (C1-C6 alkyl)NHC(O)O, NH (C1-C3 alkyl), S(O)2 (C1-C3 alkyl), 4-10 membered heterocyclic groups, 4-10 membered heterocyclic oxy groups, (C3-C 10 Cycloalkyl)CH2O, 5-6-membered heteroaryl, 5-6-membered heteroaryloxy;

[0024] R c2 Selected from 4-6 membered heterocyclic alkyl, =O, C1-C3 alkyl, halo-C1-C3 alkyl, F, Cl, Br, I, CN, OH, CH2OH or NH2;

[0025] X is selected from O, S, or NR. 2 ;

[0026] L is selected from O or NH;

[0027] R 1 R 2 Independently selected from H, C1-C 10 Alkyl or C3-C 10 cycloalkyl, the C1-C 10 Alkyl or C3-C 10 cycloalkyl optional R d Instead, the R d Selected from F, Cl, Br, I, OH, CN or C1-C3 alkyl groups.

[0028] In some implementations, ring Q is a 5-6 membered heterocyclic group, C6-C 10 Aryl or 5-10 membered heteroaryl, wherein the 5-6 membered heterocyclic group, C6-C 10 Aryl or 5-10 heteroaryl groups are selectively coated with R. a Instead, the R a Selected from F, Cl, Br, I, OH, CN, or optionally R b The following groups are substituted: C1-C 10 Alkyl, C3-C 10 cycloalkyl, C1-C 10 Alkoxy;

[0029] R b Selected from F, Cl, Br, I, OH or CN;

[0030] Ring W is phenyl, pyridyl, pyridoneyl, or a 9-10 heteroaryl group, wherein the phenyl, pyridyl, pyridoneyl, or 9-10 heteroaryl group is optionally replaced by R. c Instead, the R c Selected from OH or optionally R c1 The following groups are substituted: C1-C 10 Alkyl, C1-C 10 Alkoxy, C3-C 10 Cycloalkyl, P(O)(C1-C3 alkyl) 2, 4-6 membered heterocyclic alkyl;

[0031] R c1 Selected from F, Cl, Br, I, CN, OH, NH2, or optionally R c2 The following groups are substituted: C1-C3 alkyl, C1-C3 alkoxy, NHC(O)O (C1-C6 alkyl), (C1-C6 alkyl)NHC(O)O, NH (C1-C3 alkyl), S(O)2 (C1-C3 alkyl), 4-10 membered heterocyclic groups, 4-10 membered heterocyclic oxy groups, (C3-C 10 Cycloalkyl)CH2O, 5-6-membered heteroaryl, 5-6-membered heteroaryloxy;

[0032] R c2 Selected from 4-6 membered heterocyclic alkyl, =O, C1-C3 alkyl, halo-C1-C3 alkyl, F, Cl, Br, I, CN, OH, CH2OH or NH2;

[0033] X is selected from O, S, or NR. 2 ;

[0034] L is selected from O or NH;

[0035] R 1 R 2 Independently selected from H, C1-C 10 Alkyl or C3-C 10 cycloalkyl, the C1-C 10 Alkyl or C3-C 10 cycloalkyl optional R d Instead, the R d Selected from F, Cl, Br, I, OH, CN or C1-C3 alkyl groups.

[0036] In some embodiments, ring Q is a 5-6 membered heterocyclic group, a phenyl group, or a 5-6 membered heteroaryl group, wherein the 5-6 membered heterocyclic group, phenyl group, or 5-6 membered heteroaryl group is optionally replaced by R. a replace.

[0037] In some embodiments, ring Q is a phenyl or a 5-6-membered heteroaryl group, wherein the phenyl or 5-6-membered heteroaryl group is optionally replaced by R. a replace.

[0038] In some embodiments, ring Q is piperidinyl, phenyl, or pyridinyl, wherein the piperidinyl, phenyl, or pyridinyl group is optionally replaced by R. a replace.

[0039] In some embodiments, ring Q is phenyl or pyridyl, said phenyl or pyridyl group optionally being R a replace.

[0040] In some implementations, ring Q is arbitrarily controlled by R. a Substituted phenyl groups.

[0041] In some implementation schemes, R a It can be F, Cl, Br, I, CN, C2-C3 alkynyl, C1-C6 alkyl, C3-C6 cycloalkyl or C1-C6 alkoxy optionally substituted with F.

[0042] In some implementation schemes, R a It is F, Cl, Br, I, C1-C6 alkyl, C3-C6 cycloalkyl, or optionally F-substituted C1-C6 alkoxy. In some embodiments, R a It can be F, Cl, Br, I, CN, methyl, cyclopropyl, ethynyl, prop-1-ynyl, or a methoxy group optionally substituted with F.

[0043] In some implementation schemes, R a It can be F, Cl, Br, I, methyl, cyclopropyl, or methoxy group optionally substituted with F.

[0044] In some implementation schemes, R a The methoxy group can be optionally replaced by F.

[0045] In some implementation schemes, R a It can be F, Cl, Br, I, CN, methyl, cyclopropyl, ethynyl, prop-1-ynyl, difluoromethoxy, trifluoromethoxy, or methoxy.

[0046] In some implementation schemes, R a It is difluoromethoxy.

[0047] In some implementations, ring Q is selected from...

[0048]

[0049] In some implementations, ring Q is selected from...

[0050] In some implementations, ring Q is selected from...

[0051] In some embodiments, ring W is phenyl, pyridyl, pyrimidinyl, pyridazinyl, 2-pyridoneyl, Wherein ring M is a 5-10 membered heteroaryl or a 5-10 membered heterocyclic group, wherein the phenyl, pyridyl, pyrimidinyl, pyridazinyl, 2-pyridoneyl, Or ring M can be arbitrarily selected by R c Substitution. In some embodiments, ring W is phenyl, pyridinyl, 2-pyridone, or... Wherein ring M is a 5-6 membered heteroaryl or a 5-10 membered heterocyclic group, wherein the phenyl, 2-pyridone, pyridyl, Or ring M can be arbitrarily selected by R c replace.

[0052] In some embodiments, ring W is phenyl, pyridinyl, 2-pyridoneyl, or a 9-10 heteroaryl group, wherein the phenyl, 2-pyridoneyl, pyridinyl, or 9-10 heteroaryl group is optionally replaced by R. c replace.

[0053] In some implementations, ring W is arbitrarily R c The following groups are substituted: Phenyl, 9-10 heteroaryl.

[0054] In some embodiments, ring M is a 5-6 membered heteroaryl, a 9-10 membered heteroaryl, or a 5-9 membered heterocyclic group, wherein ring M is optionally R c replace.

[0055] In some embodiments, ring M is a 5-6 membered heteroaryl or a 6-8 membered heterocyclic group, wherein ring M is optionally R c replace.

[0056] In some implementation schemes, ring W is Wherein ring M is a 5-6 membered heteroaryl, a 9-10 membered heteroaryl, or a 5-9 membered heterocyclic group, and ring M is optionally replaced by R. c replace.

[0057] In some implementation schemes, ring W is Wherein ring M is a 5-6 membered heteroaryl or a 6-8 membered heterocyclic group, and ring M is optionally replaced by R. c replace.

[0058] In some implementation schemes, ring W is Wherein ring M is a 5-6 member heteroaryl group, and ring M is optionally replaced by R c replace.

[0059] In some implementations, ring W is arbitrarily R c The following groups are substituted: Phenyl,

[0060] In some implementations, ring W is arbitrarily R c The following groups are substituted: Phenyl,

[0061] In some implementations, ring W is arbitrarily R c The following groups are substituted: Phenyl,

[0062] In some implementations, ring W is arbitrarily R c The following groups are substituted: Phenyl,

[0063] In some implementation schemes, R c Selected from =O or arbitrarily selected by R c1 The following groups are substituted: C1-C6 alkyl, C1-C6 alkoxy, NHC(O)(C1-C6 alkyl).

[0064] In some implementation schemes, R c Selected from =O or arbitrarily selected by R c1 The following groups are substituted: methyl, methoxy, NHC(O)CH3.

[0065] In some implementation schemes, R c Selected from =O or arbitrarily selected by R c1 The following groups are substituted: methyl, methoxy, NHC(O)CH3.

[0066] In some implementation schemes, R c Selected from R c1 The following groups are substituted: C1-C6 alkyl, C1-C6 alkoxy.

[0067] In some implementation schemes, R c Selected from R c1 The following groups are substituted: methyl, methoxy,

[0068] In some implementation schemes, R c Selected from Rc1 Substituted methyl or optionally R c1 Substituted methoxy group.

[0069] In some implementation schemes, R c1 It is selected from deuterium, F, Cl, Br, I, CN, OH, NH2 or ethoxy.

[0070] In some implementation schemes, R c1 Selected from deuterium, F, Cl, Br, I, CN, and OH.

[0071] In some implementation schemes, R c1 Selected from F, Cl, Br, I, CN, and OH.

[0072] In some implementation schemes, R c Selected from CH2OH, methyl, methoxy, CHF2, CD3, NHC(=O)CH3, =O

[0073] In some implementation schemes, R c Selected from CH2OH, methyl, methoxy, CHF2, CD3, NHC(=O)CH3 or =O.

[0074] In some implementation schemes, R c Selected from CH2OH, methyl, methoxy, CHF2 or

[0075] In some implementation schemes, R c Selected from methyl groups optionally substituted with OH or methoxy groups optionally substituted with OH.

[0076] In some implementation schemes, ring W is selected from In some implementation schemes, ring W is selected from In some implementation schemes, ring W is selected from

[0077] In some implementation schemes, ring W is selected from

[0078] In some implementation schemes, ring W is selected from

[0079] In some implementations, X is selected from O, S, or NH.

[0080] In some implementations, X is selected from S or O.

[0081] In some implementations, X is selected from S.

[0082] In some implementation schemes, R 1 It is selected from H, 4-6 membered heterocyclic groups, C3-C6 cycloalkyl groups optionally substituted with C1-C3 alkyl groups, or C1-C6 alkyl groups optionally substituted with F.

[0083] In some implementation schemes, R 1 Selected from H, C3-C6 cycloalkyl substituted with C1-C3 alkyl, or C1-C6 alkyl substituted with F.

[0084] In some implementation schemes, R 1 Selected from 4-6 membered heterocyclic groups or optional C1-C6 alkyl groups substituted with F.

[0085] In some implementation schemes, R 1 It is selected from 4-membered heterocyclic groups or C1-C3 alkyl groups optionally substituted with F.

[0086] In some implementation schemes, R 1 Selected from C1-C6 alkyl groups.

[0087] In some implementation schemes, R 1 Selected from methyl, ethyl, n-propyl, Or trifluoromethyl.

[0088] In some implementation schemes, R 1 Selected from ethyl.

[0089] In some implementations, L is selected from O or a chemical bond.

[0090] In some implementations, L is selected from O.

[0091] In some implementation schemes, Selected from methoxy, ethoxy, n-propyloxy, Or trifluoromethyl.

[0092] In some embodiments, the compound represented by general formula (A) or a pharmaceutically acceptable salt thereof is selected from the compound of formula (B) or a pharmaceutically acceptable salt thereof:

[0093]

[0094] Among them, rings W and R a R 1 As defined above, n is selected from 0, 1, 2, 3, 4 or 5.

[0095] In some embodiments, the compound represented by general formula (A) or a pharmaceutically acceptable salt thereof is selected from the compound of formula (C) or a pharmaceutically acceptable salt thereof:

[0096]

[0097] Among them, rings W and R a R 1 As defined above.

[0098] In some embodiments, the compound represented by general formula (A) or a pharmaceutically acceptable salt thereof is selected from the compound of formula (D) or a pharmaceutically acceptable salt thereof:

[0099]

[0100] The ring W is defined as above.

[0101] In some embodiments, the compound or pharmaceutically acceptable salt represented by general formula (A) is selected from the following compounds or pharmaceutically acceptable salts:

[0102]

[0103]

[0104]

[0105] In some embodiments, the compound or pharmaceutically acceptable salt represented by general formula (A) is selected from the following compounds or pharmaceutically acceptable salts:

[0106]

[0107] The present invention also provides pharmaceutical compositions comprising a compound of general formula (A) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

[0108] This invention relates to the use of compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, in the preparation of medicaments for the prevention or treatment of tumors.

[0109] Furthermore, the present invention relates to the use of compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, in the preparation of medicaments for the prevention or treatment of tumors with reduced or absent MTAP activity.

[0110] This invention relates to the use of compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, for the prevention or treatment of tumors.

[0111] Furthermore, the present invention relates to the use of compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, in the prevention or treatment of tumors with reduced or absent MTAP activity.

[0112] This invention relates to compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, for the prevention or treatment of tumors.

[0113] Furthermore, the present invention relates to compounds of general formula (A) or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof, for the prevention or treatment of tumors with reduced or absent MTAP activity.

[0114] The present invention also relates to a method of treating tumors, the method comprising administering to a patient a therapeutically effective dose of a pharmaceutical preparation comprising a compound of general formula (A) as described herein or a pharmaceutically acceptable salt thereof.

[0115] Furthermore, the present invention also relates to a method for treating tumors with reduced or absent MTAP activity, the method comprising administering to a patient a therapeutically effective dose of a pharmaceutical preparation comprising a compound of general formula (A) as described in the present invention or a pharmaceutically acceptable salt thereof.

[0116] In a preferred embodiment of the present invention, the tumor or tumor with reduced or absent MTAP activity includes, but is not limited to, glioma, mesothelioma, melanoma, gastric cancer, esophageal cancer, bladder cancer, pancreatic cancer, non-small cell lung cancer, astrocytoma, osteosarcoma, head and neck cancer, myxoid chondrosarcoma, ovarian cancer, endometrial cancer, breast cancer, soft tissue sarcoma, non-Hodgkin lymphoma, etc.

[0117] Terminology Definitions and Explanations

[0118] Unless otherwise stated, the definitions of groups and terms recorded in this application specification and claims, including definitions as examples, exemplary definitions, preferred definitions, definitions recorded in tables, and definitions of specific compounds in the examples, can be arbitrarily combined and combined with each other. Such combinations and combinations of group definitions and compound structures should fall within the scope of this application specification. A specific term, without a specific definition, should not be considered uncertain or unclear, but should be understood according to its ordinary meaning in the art. When trade names appear herein, they are intended to refer to the corresponding product or its active ingredient.

[0119] In this article Indicates the connection site.

[0120] The term "pharmaceutically acceptable salt" refers to a pharmaceutically acceptable, non-toxic salt of an acid or base, including salts of inorganic acids and bases, and salts of organic acids and bases.

[0121] The diagrammatic representation of racemic or enantiomerically pure compounds in this article is derived from Maehr, J. Chem. Ed. 1985, 62:114-120. Unless otherwise specified, wedge bonds and virtual wedge bonds are used. The absolute configuration of a solid center is represented by black solid bonds and imaginary bonds. The relative configurations of a stereocenter (e.g., the cis-trans configurations of alicyclic compounds) are indicated. When the compounds described herein contain an alkene double bond or other geometrically asymmetric centers, they include E and Z geometric isomers unless otherwise specified. Similarly, all tautomers are included within the scope of this invention.

[0122] The term "tautomer" refers to a functional group isomer resulting from the rapid movement of an atom between two positions within a molecule. The compounds of this invention can exhibit tautomerism. Tautomers can exist in two or more interconvertible forms. Tautomers generally exist in equilibrium; attempts to isolate a single tautomer typically produce a mixture whose physicochemical properties are consistent with those of the mixture of compounds. The equilibrium position depends on the intramolecular chemical characteristics. For example, in many aliphatic aldehydes and ketones such as acetaldehyde, the ketone form is dominant; while in phenols, the enol form is dominant. This invention encompasses all tautomeric forms of the compounds.

[0123] The term "stereoisomer" refers to isomers that are produced by different spatial arrangements of atoms in a molecule, including cis-trans isomers, enantiomers, non-corresponding isomers, and conformational isomers.

[0124] The compounds of this invention may have asymmetric atoms such as carbon atoms, sulfur atoms, nitrogen atoms, phosphorus atoms, or asymmetric double bonds, and therefore the compounds of this invention may exist in specific geometric or stereoisomer forms. Specific geometric or stereoisomer forms may be cis and trans isomers, E-type and Z-type geometric isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures thereof or other mixtures, such as mixtures enriched with enantiomers or diastereomers. All of these isomers and mixtures thereof are within the scope of the definition of compounds in this application. Alkyl groups and other substituents may contain additional asymmetric carbon atoms, asymmetric sulfur atoms, asymmetric nitrogen atoms, or asymmetric phosphorus atoms. All such isomers involved in all substituents and mixtures thereof are also included within the scope of the definition of compounds in this application. The compounds containing asymmetric atoms of this application can be isolated in optically active pure form or in racemic form. The optically active pure form can be separated from racemic mixtures or synthesized using chiral starting materials or chiral reagents.

[0125] The term "pharmaceutical composition" refers to a mixture of one or more compounds described herein, or physiologically / pharmaceutical acceptable salts or prodrugs thereof, with other chemical components, such as physiologically / pharmaceutical acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate the administration of the compound to a living organism.

[0126] The term "substituted" refers to the substitution of one or more hydrogen atoms on a specific atom by a substituent, provided that the valence state of the specific atom is normal and the resulting compound is stable. When the substituent is oxo (i.e., =O), it means that two hydrogen atoms are substituted; oxo substitution does not occur on aromatic groups.

[0127] The terms “optional” or “optionally” mean that the event or condition subsequently described may or may not occur, including both the occurrence and non-occurrence of said event or condition. For example, the ethyl group being “optionally” substituted with a halogen means that the ethyl group can be unsubstituted (CH2CH3), monosubstituted (e.g., CH2CH2F, CH2CH2Cl, etc.), polysubstituted (e.g., CHFCH2F, CH2CHF2, CHFCH2Cl, CH2CHCl2, etc.), or fully substituted (CF2CF3, CF2CCl3, CCl2CCl3, etc.). Those skilled in the art will understand that for any group containing one or more substituents, no substitution or substitution pattern that is spatially impossible and / or cannot be synthesized is introduced.

[0128] When any variable (e.g., R) a R b When a group appears more than once in the composition or structure of a compound, its definition is independent in each case. For example, if a group is surrounded by two R... b Replaced, then each R b Each has its own independent options.

[0129] When the number of a linking group is 0, such as -(CH2)0-, it indicates that the linking group is a bond.

[0130] When one of the variables is selected as a chemical bond or does not exist, it means that the two groups it is connected to are directly connected. For example, when L in ALZ represents a bond, it means that the structure is actually AZ.

[0131] When a substituent is cross-bonded to two atoms on a ring, it can bond to any atom on that ring. For example, structural units. R represents 5 Substitution can occur at any position on the benzene ring.

[0132] C in this article m -C n This refers to having an integer number of carbon atoms in the range mn. For example, "C1-C 10 "" means that the group can have 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms.

[0133] The term "alkyl" refers to a compound with the general formula C10. n H 2n+1 The alkyl group. This alkyl group can be straight-chain or branched. For example, the term "C1-C..." 10 "Alkyl" should be understood to mean a straight-chain or branched saturated monovalent hydrocarbon group having 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. The alkyl group includes, but is not limited to, methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, 2-methylbutyl, 1-methylbutyl, 1-ethylpropyl, 1,2-dimethylpropyl, neopentyl, 1,1-dimethylpropyl, 4-methylpentyl, 3-methylpentyl, 2 -Methylpentyl, 1-methylpentyl, 2-ethylbutyl, 1-ethylbutyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbutyl, 2,3-dimethylbutyl, 1,3-dimethylbutyl or 1,2-dimethylbutyl, etc.; "C1-C6 alkyl" should be understood as representing straight-chain or branched saturated monovalent hydrocarbon groups with 1, 2, 3, 4, 5, or 6 carbon atoms; "C1-C3 alkyl" refers to methyl, ethyl, propyl, and isopropyl.

[0134] The “C1-C” mentioned in this article 10 "alkyl" may include "C1-C6 alkyl" or "C1-C3 alkyl", and the "C1-C6 alkyl" may further include "C1-C3 alkyl".

[0135] The term "alkoxy" can be understood as "alkyloxy" or "alkyl-O-", referring to a monovalent group formed by the loss of a hydrogen atom from a hydroxyl group in straight-chain or branched alcohols, such as the term "C1-C". 10 "Alkoxy" can be understood as "C1-C" 10 "alkyloxy" or "C1-C" 10 "alkyl-O-", the term "C1-C6 alkoxy" can be understood as "C1-C6 alkyloxy" or "C1-C6 alkyl-O-", preferably, "C1-C 10 The term "alkoxy" can include ranges such as "C1-C6 alkoxy" and "C1-C3 alkoxy".

[0136] The term “halogenated C1-C3 alkyl” includes monohalogenated or polyhalogenated C1-C3 alkyl, examples of which include, but are not limited to, trifluoromethyl, 2,2,2-trichloroethyl or 3-fluoropropyl.

[0137] The term "alkynyl" refers to a straight-chain or branched unsaturated aliphatic hydrocarbon group consisting of carbon and hydrogen atoms and having at least one triple bond. The term "C2-C"... 10 "Alkyne" can be understood as representing a straight-chain or branched unsaturated hydrocarbon group containing one or more triple bonds and having 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. "C2-C"10 Examples of "alkynyl" include, but are not limited to, ethynyl (-C≡CH), propynyl (-C≡CCH3, -CH2C≡CH), buty-1-alkynyl, buty-2-alkynyl, or buty-3-alkynyl. "C2-C 10 "Alynyl" can include "C2-C3 alkynyl", and examples of "C2-C3 alkynyl" include ethynyl (-C≡CH), propynyl-1-alkynyl (-C≡CCH3), and propynyl-2-alkynyl (-CH2C≡CH).

[0138] The term "C3-C" 10 "Cycloalkyl" should be understood as referring to a saturated monovalent monocyclic or bicyclic hydrocarbon ring having 3 to 10 carbon atoms. Examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, or cyclodecyl, or bicyclic hydrocarbon rings such as decahydronaphthalene ring. The term "C3-C6 cycloalkyl" should be understood as referring to a saturated monovalent monocyclic or bicyclic hydrocarbon ring having 3 to 6 carbon atoms.

[0139] The term “cycloalkoxy” can be understood as “cycloalkyloxy” or “cycloalkyl-O-”.

[0140] The term "heterocyclic group" refers to a fully saturated or partially saturated (not aromatic as a whole) monocyclic, fused, spirocyclic, or bridged ring group containing 1 to 5 heteroatoms or heteroatomic groups (i.e., groups containing heteroatoms). The "heteroatoms or heteroatomic groups" include, but are not limited to, nitrogen (N), oxygen (O), sulfur (S), phosphorus (P), boron (B), -S(=O)2-, -S(=O)-, -P(=O)2-, -P(=O)-, -NH-, -S(=O)(=NH)-, -C(=O)NH-, or -NHC(=O)NH-, etc.

[0141] The term "4-10 membered heterocyclic group" refers to a heterocyclic group with 4, 5, 6, 7, 8, 9, or 10 ring atoms, and whose ring atoms contain 1-5 independently selected heteroatoms or heteroatomic groups as described above. "4-10 membered heterocyclic group" includes "4-7 membered heterocyclic group", wherein specific examples of 4 membered heterocyclic groups include, but are not limited to, azirrocyclobutane or oxacyclobutane; specific examples of 5 membered heterocyclic groups include, but are not limited to, tetrahydrofuranyl, dioxacyclopentenyl, pyrrolyl, imidazoalkyl, pyrazolyl, pyrrolinyl, 4,5-dihydrooxazolyl, or 2,5-dihydro-1H-pyrrolyl; specific examples of 6 membered heterocyclic groups include, but are not limited to, tetrahydropyranyl, piperidinyl, morpholinyl, dithiaalkyl, thiomorpholinyl, piperazine, trithiaalkyl, tetrahydropyridinyl, or 4H-[1,3,4]thiadiazinyl; and specific examples of 7 membered heterocyclic groups include, but are not limited to, diazacycloheptane. The heterocyclic group can also be a bicyclic group, wherein specific examples of 5,5-membered bicyclic groups include, but are not limited to, hexahydrocyclopentano[c]pyrrolo-2(1H)-yl; specific examples of 5,6-membered bicyclic groups include, but are not limited to, hexahydropyrrolo[1,2-a]pyrazin-2(1H)-yl, 5,6,7,8-tetrahydro-[1,2,4]triazolo[4,3-a]pyrazinyl, or 5,6,7,8-tetrahydroimidazo[1,5-a]pyrazinyl. Optionally, the heterocyclic group can be a benzofused cyclic group of the above-mentioned 4-7-membered heterocyclic groups, specific examples of which include, but are not limited to, dihydroisoquinolinyl, etc. Preferably, "4-10 membered heterocyclic group" can include the ranges of "5-10 membered heterocyclic group", "5-9 membered heterocyclic group", "4-6 membered heterocyclic group", "5-6 membered heterocyclic group", "6-9 membered heterocyclic group", "6-8 membered heterocyclic group", "5-10 membered heterocyclic alkyl group", "5-9 membered heterocyclic alkyl group", "4-6 membered heterocyclic alkyl group", "5-6 membered heterocyclic alkyl group", and "6-8 membered heterocyclic alkyl group". According to the present invention, although some bicyclic heterocyclic groups partially contain a benzene ring or a heteroaromatic ring, the heterocyclic group as a whole is still non-aromatic.

[0142] The term "heterocyclic oxy group" can be understood as "heterocyclic group -O-".

[0143] The term "heterocyclic alkyl" refers to a fully saturated 4- to 10-membered cyclic group that may exist as a monocyclic, fused, bridged, or spirocyclic ring. Unless otherwise indicated, the ring atom of the heterocycle contains 1 to 5 heteroatoms or heterogroups (i.e., groups containing heteroatoms), including but not limited to nitrogen (N), oxygen (O), sulfur (S), phosphorus (P), boron (B), -S(=O)2-, -S(=O)-, -NH-, -S(=O)(=NH)-, -C(=O)NH-, or -NHC(=O)NH-, etc. The term "4-6 membered heterocyclic alkyl" refers to a heterocyclic alkyl group having 4, 5, or 6 ring atoms, and containing 1-3 independent heteroatoms or heteroatomic groups selected from those described above. Non-limiting examples of 4-membered heterocyclic alkyl groups include, but are not limited to, acridine, oxadiazolyl, and thiobutyl. Examples of 5-membered heterocyclic alkyl groups include, but are not limited to, tetrahydrofuranyl, tetrahydrothiophenyl, pyrrolidinyl, isoxazolyl, oxazolyl, isothiazolyl, thiazolyl, imidazolyl, and tetrahydropyrazolyl. Examples of 6-membered heterocyclic alkyl groups include, but are not limited to, piperidinyl, tetrahydropyranyl, tetrahydrothiaranyl, morpholinyl, piperazine, 1,4-thiaoxalyl, 1,4-dioxane, thiomorpholinyl, 1,3-dithiaalkyl, and 1,4-dithiaalkyl.

[0144] The term "aryl" refers to an aromatic ring group consisting of an all-carbon monocyclic or fused polycyclic aromatic cyclic group with a conjugated π-electron system. For example, aryl groups can have 6-20 carbon atoms, 6-14 carbon atoms, or 6-12 carbon atoms. The term "C6-C"... 10 "Aryl" should be understood to preferably represent an aromatic or partially aromatic monocyclic or bicyclic group with 6 to 10 carbon atoms, particularly a ring with 6 carbon atoms ("C6 aryl"), such as phenyl; or a ring with 9 carbon atoms ("C9 aryl"), such as indenyl or indenyl; or a ring with 10 carbon atoms ("C9 aryl"). 10 Aryl), such as tetrahydronaphthyl, dihydronaphthyl or naphthyl.

[0145] The term "heteroaryl" refers to an aromatic monocyclic or fused polycyclic system containing at least one ring atom selected from N, O, or S, with the remaining ring atoms being C aromatic cyclic groups. The term "5-10-membered heteroaryl" should be understood to include monovalent monocyclic or bicyclic aromatic ring systems having 5, 6, 7, 8, 9, or 10 ring atoms, particularly 5, 6, 9, or 10 ring atoms, and containing 1-5, preferably 1-3, heteroatoms independently selected from N, O, and S. Furthermore, in each case, it may be benzofused. Specifically, the heteroaryl group is selected from thienyl, furanyl, pyrroleyl, oxazolyl, thiazolyl, imidazoleyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazolyl, and their benzo[derivatives], such as benzofuranyl, benzothienyl, benzothiazolyl, benzooxazolyl, benzoisooxazolyl, benzoimidazolyl, benzotriazolyl, indazoleyl, indolyl, isoindolyl, etc.; or pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, etc., and their benzo[derivatives], such as quinolinyl, quinazolinyl, isoquinolinyl, etc.; or acrylinyl, inazinyl, purinyl, etc., and their benzo[derivatives]; or cyclolinyl, phthalazinyl, quinazolinyl, quinoxolinyl, naphthinyl, pteridinyl, carbazolyl, acrylinyl, phenazinyl, phenthiazinyl, phenoxazinyl, etc. The term "5-6-membered heteroaryl" refers to an aromatic ring system having 5 or 6 ring atoms, and containing 1 to 3, preferably 1 to 2, heteroatoms independently selected from N, O, and S. The term "9-10-membered heteroaryl" refers to an aromatic ring system having 9 or 10 ring atoms, and containing 1 to 5, preferably 1 to 3, heteroatoms independently selected from N, O, and S.

[0146] The term "heteroaryloxy" can be understood as "heteroaryl-O-".

[0147] The term "excipient" refers to pharmaceutically acceptable inert ingredients.

[0148] Examples of the types of excipients, without limitation, include binders, disintegrants, lubricants, flow aids, stabilizers, fillers, and diluents. Excipients enhance the handling properties of pharmaceutical formulations, i.e., by increasing flowability and / or adhesion, making the formulation more suitable for direct compression. Typical examples of pharmaceutically acceptable carriers suitable for the above formulations include sugars, starches, cellulose, and their derivatives, which are commonly used excipients in pharmaceutical formulations.

[0149] The term "treatment" means administering the compound or preparation described in this application to prevent, improve, or eliminate a disease or one or more symptoms related to said disease, and includes:

[0150] (i) To prevent the occurrence of disease or disease state in mammals, especially when such mammals are susceptible to the disease state but have not yet been diagnosed with the disease state;

[0151] (ii) To suppress the disease or disease state, that is, to curb its development;

[0152] (iii) Alleviate the disease or disease state, even if the disease or disease state subsides.

[0153] The term "therapeutic effective amount" means the amount of the compound of the present invention used to treat or prevent a particular disease, condition, or disorder; (ii) to reduce, improve, or eliminate one or more symptoms of a particular disease, condition, or disorder; or (iii) to prevent or delay the onset of one or more symptoms of a particular disease, condition, or disorder described herein. The amount of the compound of the present invention constituting a "therapeutic effective amount" varies depending on the compound, the disease state and its severity, the route of administration, and the age of the mammal to be treated, but may routinely be determined by those skilled in the art based on their own knowledge and the content of this disclosure.

[0154] The term "pharmaceuticalally acceptable excipient" refers to excipients that do not cause significant irritation to the organism and do not impair the biological activity and properties of the active compound. Suitable excipients are well known to those skilled in the art, such as carbohydrates, waxes, water-soluble and / or water-swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, etc.

[0155] The words “comprise,” “comprise,” or “comprise,” and their English variations such as comprises or comprising, should be understood in an open, non-exclusive sense, meaning “including but not limited to.”

[0156] The compounds of the present invention can be prepared by a variety of synthetic methods known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthetic methods, and equivalent substitutions known to those skilled in the art. Preferred embodiments include, but are not limited to, the embodiments of the present invention.

[0157] This application also includes compounds of this application that are identical to those described herein, but with one or more atoms replaced by isotopes whose atomic weights or mass numbers differ from those commonly found in nature. Examples of isotopes that can be incorporated into compounds of this application include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as... 2 H, 3 H, 11 C 13 C 14 C 13 N、 15 N、 15 O、 17 O、 18 O、31 P, 32 P, 35 S, 18 F, 123 I, 125 I and 36 Cl, etc.

[0158] Certain isotope-labeled compounds of this application (e.g., using...) 3 H and 14 Those labeled with C can be used in the analysis of compound and / or substrate tissue distribution. Tritiumization (i.e. 3 H) and carbon-14 (i.e. 14 C) Isotopes are particularly preferred due to their ease of preparation and detectability. Positron-emitting isotopes, such as... 15 O、 13 N、 11 C and 18 F can be used in positron emission tomography (PET) studies to determine substrate occupancy. The isotopically labeled compounds of this application can typically be prepared by replacing the unlabeled reagent with an isotopically labeled reagent using a procedure similar to those disclosed in the schemes and / or examples below.

[0159] The pharmaceutical compositions of this application can be prepared by combining the compounds of this application with suitable pharmaceutically acceptable excipients, for example, in solid, semi-solid, liquid or gaseous formulations, such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, suppositories, injections, inhalers, gels, microspheres and aerosols.

[0160] Typical routes of administration for the compounds of this application or their pharmaceutically acceptable salts or pharmaceutical compositions thereof include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, vaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, and intravenous administration.

[0161] The pharmaceutical composition of this application can be manufactured using methods well known in the art, such as conventional mixing, dissolving, granulation, sugar-coated pill making, grinding, emulsification, freeze drying, etc.

[0162] In some embodiments, the pharmaceutical composition is in oral form. For oral administration, the pharmaceutical composition can be formulated by mixing the active compound with pharmaceutically acceptable excipients well known in the art. These excipients enable the compounds of this application to be formulated into tablets, pills, lozenges, sugar-coated tablets, capsules, liquids, gels, pastes, suspensions, etc., for oral administration to patients.

[0163] Solid oral compositions can be prepared using conventional mixing, filling, or tableting methods. For example, they can be obtained by mixing the active compound with solid excipients, optionally milling the resulting mixture, adding other suitable excipients if necessary, and then processing the mixture into granules to obtain the core of a tablet or sugar-coated formulation. Suitable excipients include, but are not limited to, binders, diluents, disintegrants, lubricants, glidants, sweeteners, or flavoring agents.

[0164] The pharmaceutical composition may also be suitable for parenteral administration, such as in suitable unit dosage forms of sterile solutions, suspensions or lyophilized products.

[0165] In all methods of administration of the compound of general formula I described herein, the daily dose is from 0.01 to 100 mg / kg body weight, preferably from 0.05 to 50 mg / kg body weight, more preferably from 0.1 to 30 mg / kg body weight, in the form of single or separate doses.

[0166] The chemical reactions in the specific embodiments of this invention are carried out in a suitable solvent, which must be suitable for the chemical changes of this invention and the reagents and materials required therefor. To obtain the compounds of this invention, it is sometimes necessary for those skilled in the art to modify or select the synthesis steps or reaction flow based on existing embodiments. Detailed Implementation

[0167] The following embodiments describe the technical solutions of the invention in detail, but the scope of protection of the invention includes, but is not limited to, these embodiments.

[0168] Unless otherwise stated, the proportions of mixed solvents are volume-based.

[0169] Unless otherwise stated, % refers to weight percentage (wt%).

[0170] The eluent or mobile phase can be a mixture of two or more solvents, with the ratio being the volume ratio of each solvent. For example, "0-10% methanol / dichloromethane" indicates that the volume ratio of methanol to dichloromethane in the mixed eluent or mobile phase is 0:100 to 10:100.

[0171] Compounds are processed manually or Software naming conventions are used; commercially available compounds use supplier catalog names.

[0172] The structure of the compound was determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts are measured in units of 10⁻⁶. -6 (ppm). The solvents used for NMR determination included deuterated dimethyl sulfoxide, deuterated chloroform, and deuterated methanol, with tetramethylsilane (TMS) as the internal standard. "IC 50"Hardest effective concentration" refers to the concentration at which half of the maximum inhibitory effect is achieved. "DCM" refers to dichloromethane, "PE" refers to petroleum ether, and "EA" refers to ethyl acetate.

[0173] Example 1: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(hydroxymethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 1)

[0174]

[0175] Step 1: Synthesis of N-((5-bromo-6-methoxypyridin-2-yl)aminomeththioyl)benzamide (intermediates 1-2)

[0176] Starting material 1-1 (1 g, 4.93 mmol) was dissolved in tetrahydrofuran (15 mL), and benzoyl isothiocyanate (763.59 mg, 4.68 mmol) was added at 0 °C. The mixture was stirred at 0 °C for 1 hour, and then stirred at 25 °C for 16 hours. LCMS analysis showed that the reaction was complete. The solvent was removed by concentration under reduced pressure to obtain the title compound (1.8 g), which was directly used in the next reaction.

[0177] MS m / z (ESI): 365.9 [M+H] + .

[0178] Step 2: Synthesis of 1-(5-bromo-6-methoxypyridin-2-yl)thiourea (intermediates 1-3)

[0179] Intermediate 1-2 (1.8 g, 4.91 mmol) was dissolved in water (1.5 mL) and tetrahydrofuran (8 mL), and sodium hydroxide (5 M, 1.18 mL) was added. The mixture was heated to 80 °C and stirred for 3 hours. LCMS analysis confirmed the reaction was complete. The solvent was removed by concentration under reduced pressure. Water (50 mL) and ethyl acetate (50 mL) were added, and the pH was adjusted to 1 with hydrochloric acid. The mixture was then extracted with a 1:3 (120 mL) mixture of isopropanol and dichloromethane. The organic phases were combined, dried over sodium sulfate, filtered, concentrated under reduced pressure, and the residue was purified by rapid column chromatography. 20g The title compound (1.1 g) was obtained by rapid silica column chromatography with a mobile phase of 5-25% tetrahydrofuran / petroleum ether (5% DCM) gradient at 30 mL / min.

[0180] MS m / z(ESI): 261.8 [M+H] + .

[0181] Step 3: Synthesis of 6-bromo-5-methoxythiazo[4,5-b]pyridine-2-amine (intermediates 1-4)

[0182] Intermediate 1-3 (1 g, 3.81 mmol) was dissolved in chloroform (20 mL) and cooled to 0 °C. Liquid bromine (621.86 mg, 3.89 mmol) was dissolved in chloroform (5 mL) and added dropwise to the reaction solution. The reaction solution was reacted at 0 °C for 30 minutes, then heated to 65 °C and reacted for 2.5 hours. LCMS analysis confirmed complete reaction of the starting material. The reaction solution was cooled to -20 °C, filtered, and the filter cake was washed with n-pentane. The title compound (1.1 g) was obtained and used directly in the next reaction step.

[0183] MS m / z(ESI): 260.0 [M+H] + .

[0184] Step 4: Synthesis of 2-amino-6-bromothiazo[4,5-b]pyridin-5(4H)-one (intermediates 1-5)

[0185] Intermediate 1-4 (600 mg, 2.31 mmol) was dissolved in 1,2-dichloroethane (15 mL), and boron tribromide (2.23 g, 8.90 mmol) was added. The reaction mixture was stirred at 50 °C for 16 hours. LC-MS analysis confirmed complete reaction of the starting material. The reaction mixture was diluted with 1,2-dichloroethane, and methanol (15 mL) was added dropwise under nitrogen protection at 0 °C. The reaction mixture was concentrated to obtain a solid, which was then added with methanol (15 mL), and the pH was adjusted to 9 with ammonia. The mixture was filtered to obtain the title compound (100 mg).

[0186] MS m / z (ESI): 247.9 [M+H] + .

[0187] Step 5: Synthesis of 6-bromo-2-chlorothiazo[4,5-b]pyridin-5(4H)-one (intermediates 1-6)

[0188] Copper chloride (327.82 mg, 2.44 mmol) and tert-butyl nitrite (399.00 mg, 3.87 mmol) were dissolved in acetonitrile (6 mL), followed by intermediate 1-5 (300 mg, 1.22 mmol) dissolved in acetonitrile (4 mL). The solutions were added dropwise to the reaction mixture at 0 °C. The reaction mixture was stirred at 25 °C for 16 hours. LC-MS analysis confirmed complete reaction of the starting materials. 6 M HCl (2 mL) was added dropwise to the reaction mixture, and the mixture was stirred for 30 minutes. The pH was adjusted to 8 with sodium bicarbonate. The mixture was extracted with a mixed solvent of methanol and dichloromethane (10:1). The organic phases were combined and concentrated to give the title compound (310 mg).

[0189] MS m / z(ESI): 264.9 [M+H] + .

[0190] Step 6: Synthesis of 6-bromo-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (intermediates 1-7)

[0191] Intermediate 1-6 (400 mg, 1.51 mmol) was dissolved in ethanol (15 mL), followed by the addition of sodium ethoxide (512.59 mg, 7.53 mmol). The mixture was stirred at 25 °C for 16 hours. Saturated ammonium chloride was then added to the reaction mixture, and the mixture was stirred for 15 minutes. The mixture was extracted with a mixed organic solvent of ethanol and dichloromethane (10:1, 100 mL * 5), dried over sodium sulfate, and concentrated under reduced pressure to remove the solvent. The residue was purified by preparative thin-layer chromatography (silica, petroleum ether: tetrahydrofuran = 3:2) to give the title compound (130 mg).

[0192] MS m / z(ESI): 275 [M+H] + .

[0193] Step 7: Synthesis of 6-bromo-4-(4-(difluoromethoxy)phenyl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (intermediates 1-8)

[0194] Intermediate 1-7 (120 mg, 436.17 μmol) was dissolved in dichloromethane (5 mL), followed by the addition of (4-(difluoromethoxy)phenyl)boronic acid (163.94 mg, 872.34 μmol), copper acetate (87.15 mg, 479.79 μmol), and pyridine (103.50 mg, 1.31 mmol). The mixture was stirred at 40 °C for 16 hours under an oxygen atmosphere. Water (5 mL), ammonia (0.5 mL), and ethyl acetate (5 mL) were then added, followed by extraction. The organic phases were combined and concentrated. The residue was purified by preparative thin-layer chromatography (silica, petroleum ether:tetrahydrofuran = 1:1). The title compound (110 mg) was then extracted.

[0195] MS m / z(ESI): 417[M+H] + .

[0196] Step 8: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(hydroxymethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 1)

[0197] Intermediates 1-8 (110 mg, 263.65 μmol) and 1-9 (120 mg, 416.45 μmol) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (171.80 mg, 527.29 μmol) was added to the reaction system, and Pd(dtbpf)Cl2 (17.18 mg, 26.36 μmol) was added under a nitrogen atmosphere. The reaction mixture was stirred at 90 °C for 16 hours. Water (15 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (10 mL). The organic phase was dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative high performance liquid chromatography (alkaline conditions, column: Phenomenex Luna C18 100*30 mm*5 μm; mobile phase: [water (0.05% ammonia v / v)-acetonitrile]; B%: 43%-63%, 9 min). The title compound was obtained (12 mg).

[0198] MS m / z(ESI): 499.1 [M+H] + .

[0199] 1 H NMR (400MHz, METHANOL-d4) δ8.18(s,1H),7.89(s,1H),7.65(d,J=8.4Hz,1H),7.54(d,J=8.4Hz,1H),7.45(d,J=8.9Hz,2H ), 7.33 (d, J = 8.8Hz, 2H), 6.96 (t, J = 73.8Hz, 1H), 4.81 (s, 2H), 4.39 (q, J = 7.1Hz, 2H), 3.92 (s, 3H), 1.36 (t, J = 7.1Hz, 3H).

[0200] Example 2, 4-[4-(difluoromethoxy)phenyl]-2-ethoxy-6-(4-methoxyphenyl)thiazo[4,5-b]pyridine-5(4H)-one (Compound 2)

[0201]

[0202] Step 1: Synthesis of 4-[4-(difluoromethoxy)phenyl]-2-ethoxy-6-(4-methoxyphenyl)thiazo[4,5-b]pyridine-5(4H)-one (compound 2)

[0203] Intermediate 1-8 (110 mg, 263.65 μmol) and reactant 2-1 (92.58 mg, 395.48 μmol) were dissolved in dioxane (3 mL) and water (0.75 mL). Cesium carbonate (171.80 mg, 527.30 μmol) and Pd(dtbpf)Cl2 (17.18 mg, 26.37 μmol) were added. The mixture was then heated to 90 °C under a nitrogen atmosphere and reacted for 16 hours. The reaction was confirmed by LCMS. The reaction solution was diluted with water (5 mL), extracted twice with ethyl acetate (10 mL), and the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by thin-layer chromatography (silica, tetrahydrofuran / petroleum ether = 2 / 1) and then by high-performance liquid chromatography (alkaline conditions, column: Boston Prime C18 150*30mm*5μm; mobile phase: [A: water (0.05% ammonia v / v), B: acetonitrile]; B%: 50%-80%, 9 min). The title compound was obtained (2.6 mg).

[0204] MS m / z (ESI): 445.1 [M+H] + .

[0205] 1 H NMR (400MHz, Methanol-d4) δ8.05 (s, 1H), 7.60 (d, J = 8.8Hz, 2H), 7.44-7.40 (m, 2H), 7.35-7.29 (m, 2H), 6.96 (t, J = 73Hz, 1H), 6.94 (m, 2H), 4.38 (q, J = 7.3Hz, 2H), 3.82 (s, 3H), 1.35 (t, J = 7.1Hz, 3H).

[0206] Example 3: 4-[4-(difluoromethoxy)phenyl]-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 3)

[0207]

[0208] Intermediate 1-8 (60 mg, 143.81 μmol) and intermediate 3-1 (50.71 mg, 215.71 μmol) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (93.71 mg, 287.61 μmol) and Pd(dtbpf)Cl2 (9.37 mg, 14.38 μmol) were added, and the mixture was heated to 90 °C under a nitrogen atmosphere for 16 hours. The reaction was completed by LCMS. The reaction solution was diluted with water (5 mL), extracted twice with ethyl acetate (10 mL), and the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by thin-layer chromatography (silica, tetrahydrofuran / petroleum ether = 2 / 1) and then by high-performance liquid chromatography (alkaline conditions, column: Boston Prime C18 150*30mm*5μm; mobile phase: [water (0.05% ammonia v / v)-acetonitrile]; B%: 42%-72%, 9 min). The title compound was obtained (3.2 mg).

[0209] MS m / z(ESI): 446.0 [M+H] + .

[0210] 1 H NMR (400MHz, Methanol-d4) δ8.20(d,J=2.4Hz,1H),8.14(s,1H),7.90(dd,J=2.6,9.4Hz,1H),7.46-7.38(m,2H),7.36- 7.28(m,2H),6.95(t,J=73.7Hz,1H),6.60(d,J=9.4Hz,1H),4.38(q,J=7.1Hz,2H),3.62(s,3H),1.35(t,J=7.1Hz,3H).

[0211] Example 4: 4-(4-(difluoromethoxyphenyl)-2-ethoxy-6-(1-isobutyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 4)

[0212]

[0213] Intermediate 4-1 (53 mg, 191.74 μmol, synthesized according to the method reported on page 48 of patent WO2019102256) and intermediate 1-8 (40 mg, 95.87 μmol) were dissolved in dioxane (1 mL) and water (0.25 mL). Cesium carbonate (62.47 mg, 191.74 μmol) and Pd(dtbpf)Cl2 (6.25 mg, 9.59 μmol) were added to the solution. The reaction mixture was stirred at 100 °C for 2 h under nitrogen protection. The reaction was completed by LC-MS. The reaction solution was filtered, concentrated to dryness under reduced pressure, and purified by preparative high performance liquid chromatography (column: Gemini NX C18 5μm*10*150mm; mobile phase: A: water (0.225% ammonium bicarbonate v / v), B: acetonitrile; B%: 30%-50%, 11 min) to obtain the title compound (10.0 mg).

[0214] MS m / z(ESI): 488.1 [M+H] + .

[0215] 1 H NMR (400MHz, DMSO-d6) δ8.25–8.14(m,2H),7.75–7.67(m,1H),7.42–7.35(m,2H),7.30(t,J=72Hz 1H),7.29–7.22(m,2H),6.37(d,J=9.5Hz,1H),4.31–4.19(m,2H),3.67(d,J= 7.4Hz, 2H), 2.11–1.90 (m, 1H), 1.24 (t, J = 7.0Hz, 3H), 0.80 (d, J = 6.7Hz, 6H).

[0216] Example 5: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 5)

[0217]

[0218] Intermediate 1-8 (40 mg, 95.87 μmol) and intermediate 5-1 (35 mg, 134.22 μmol) were dissolved in dioxane (1 mL) and water (0.2 mL). Cesium carbonate (62.47 mg, 191.74 μmol) and Pd(dtbpf)Cl2 (6.25 mg, 9.59 μmol) were added to the solution. The reaction mixture was stirred at 90 °C for 2 h under nitrogen protection. The reaction was confirmed by LC-MS. The reaction mixture was filtered, concentrated to dryness under reduced pressure, and purified by preparative high-performance liquid chromatography (column: Gemini NX C18 5 μm*10*150 mm; mobile phase: A: water (0.225% trifluoroacetic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to obtain the title compound (23.0 mg).

[0219] MS m / z (ESI): 469.1 [M+H] + .

[0220] 1 H NMR(400MHz,DMSO-d6)δ9.23(s,1H),8.31(s,1H),8.19(s,1H),7.91–7.79(m,2H),7.47–7.40(m,2H ),7.37–7.30(m,2H),7.20(t,J=72.0Hz,1H),4.38–4.25(m,2H),4.01(s,3H),1.31(t,J=7.0Hz,3H).

[0221] Example 6: 4-(4-(difluoromethoxy)phenyl)-6-(1-(difluoromethyl)-6-oxo-1,6-dihydropyridin-3-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (Compound 6)

[0222]

[0223] Step 1: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborpentane-2-yl)thiazo[4,5-b]pyridine-5(4H)-one (intermediate 6-1)

[0224] Compounds 1-8 (4.2 g, 10 mmol) and bis-pinacolborate (3.8 g, 15 mmol) were dissolved in anhydrous dioxane (42 mL). Potassium acetate (1.9 g, 20 mmol) and Pd(dppf)Cl2 (1.1 g, 1.5 mmol) were added to the solution. The reaction mixture was stirred at 100 °C for 15 h under nitrogen protection. The reaction was confirmed by LC-MS. After cooling to room temperature, the solvent was removed by rotary evaporation. Water (40 mL) and ethyl acetate (50 mL) were added sequentially. The organic phase was washed with water (15 mL x 2) and dried over an appropriate amount of anhydrous sodium sulfate. The title compound (3.2 g) was obtained by preparative thin-layer chromatography (silica, petroleum ether: ethyl acetate = 4:1).

[0225] MS m / z (ESI): 465.1 [M+H] + .

[0226] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(1-(difluoromethyl)-6-oxo-1,6-dihydropyridin-3-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (compound 6)

[0227] Intermediate 6-2 (50 mg, 0.22 mmol) and intermediate 6-1 (124.5 mg, 0.268 mmol) were dissolved in anhydrous dioxane solution (0.5 mL). Water (0.1 mL), cesium carbonate (145.5 mg, 0.446 mmol), and Pd(dtbpf)Cl2 (14.6 mg, 0.022 mmol) were added. The reaction mixture was stirred at 100 °C for 2 h under nitrogen protection. The reaction was confirmed by LC-MS. The reaction mixture was filtered, concentrated to dryness under reduced pressure, and purified by high pressure [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile in decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 12 min] to obtain the title compound (15 mg).

[0228] MS m / z(ESI): 482.0 [M+H] + .

[0229] 1 H NMR(400MHz,DMSO-d6)δ8.43(s,1H),8.36(s,1H),7.99-7.70(m,2H),7.49-7 .12(m,5H),6.55(d,J=9.8Hz,1H),4.29-4.23(m,2H),1.24(t,J=7.0Hz,3H).

[0230] Example 7: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)thiazo[4,5-b]pyridin-5-(4H)-one (Compound 7)

[0231]

[0232] Step 1: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)thiazo[4,5-b]pyridin-5-(4H)-one (compound)

[0233] Intermediate 1-8 (40 mg, 95.87 μmol) and intermediate 7-1 (48.7 mg, 191.74 μmol) were dissolved in dioxane (2 mL). Potassium acetate (28.2 mg, 287.61 μmol) and Pd(dppf)Cl2 (7.0 mg, 9.59 μmol) were added to the solution. The reaction mixture was stirred at 90 °C for 1 h under nitrogen protection. The reaction was confirmed by LC-MS. Then, intermediate 7-2 (36 mg, 191.74 μmol), cesium carbonate (62.6 mg, 191.74 μmol), Pd(dtbpf)Cl2 (6.3 mg, 9.5 μmol), and water (0.5 mL) were added to the reaction mixture. The mixture was stirred at 90 °C for 1 h under nitrogen protection. The reaction was confirmed by LC-MS. The reaction solution was filtered, concentrated to dryness under reduced pressure, and purified by preparative high performance liquid chromatography (column: Gemini NX C18 5μm*10*150mm; mobile phase: A: water (0.225% ammonium bicarbonate v / v), B: acetonitrile; B%: 30%-50%, 11min) to obtain the title compound (10.6mg).

[0234] MS m / z (ESI): 447.1 [M+H] + .

[0235] 1 H NMR (400MHz, DMSO-d6) δ8.51(s,1H),8.01(d,J=9.7Hz,1H),7.48(d,J=8.9Hz,2H),7.37(t,J=76.0Hz,1 H),7.33(d,J=8.8Hz,2H),6.93(d,J=9.7Hz,1H),4.47–4.25(m,2H),3.72(s,3H),1.31(t,J=7.0Hz,3H).

[0236] Example 8: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-(methyl-d3)-1Hbenzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 8)

[0237]

[0238] Step 1: Synthesis of 6-bromo-1-(methyl-d3)-1H-benzo[d]imidazole (intermediate 8-2)

[0239] Reactant 8-1 (5.0 g, 25.38 mmol) was dissolved in tetrahydrofuran solution (50 mL), and sodium hydride (60%) (1.12 g, 27.91 mmol) was added at 0-5 °C. The reaction solution was stirred at 5 °C for 0.5 h, and CD3I (4.41 g, 30.45 mmol) was added to the reaction solution. After the addition was complete, the mixture was stirred at room temperature for 12 h. After the reaction was detected by LC-MS, the reaction solution was slowly poured into a saturated ammonium chloride aqueous solution (50 mL), extracted with ethyl acetate (60 mL * 2), the organic phases were combined, washed with saturated brine (100 mL), and the washed organic phase was dried with an appropriate amount of anhydrous sodium sulfate to obtain 5.0 g of crude product. 1.0 g of crude product was prepared by supercritical liquid chromatography (column: DAICL CHIRALPAK IG (250 mm * 50 mm, 10 μm); mobile phase: [A is CO2, B: ethanol solution containing 0.1% ammonia]; B%: 35%) to obtain the title compound (400 mg).

[0240] MS m / z(ESI): 214.0 [M+H] + .

[0241] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-(methyl-d3)-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 8)

[0242] Intermediate 6-1 (150 mg, 0.32 mmol) and intermediate 8-2 (83 mg, 0.39 mmol) were dissolved in anhydrous dioxane solution (2 mL). Water (0.4 mL), cesium carbonate (211 mg, 0.65 mmol), and 1,1-bis(tert-butylphosphine)ferrocene palladium chloride (Pd(dtbpf)Cl2) (21 mg, 0.03 mmol) were added. The reaction mixture was stirred at 100 °C for 2 h under nitrogen protection. The reaction was detected by LC-MS after completion. After the reaction was cooled to room temperature, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The solution was then purified by high performance liquid chromatography (HPLC) [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.05% NH4HCO3) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 55%-80%, elution time 13 min] to obtain the title compound (16 mg).

[0243] MS m / z(ESI): 472.0 [M+H] + .

[0244] 1 H NMR (400MHz, DMSO-d6) δ8.34(s,1H),8.20(s,1H),7.97(d,J=1.6Hz,1H),7.6 6(d,J=8.5Hz,1H),7.57–7.21(m,6H),4.36–4.31(m,2H),1.35–1.31(m,3H).

[0245] Example 9: N-(6-(4-(4-(difluoromethoxy)phenyl)-2-ethoxy-5-oxo-4,5-dihydrothiazo[4,5-b]pyridin-6-yl)-1-methyl-1H-benzo[d]imidazol-2-yl)acetamide (compound 9)

[0246]

[0247] Step 1: Synthesis of N-(6-(4-(4-(difluoromethoxy)phenyl)-2-ethoxy-5-oxo-4,5-dihydrothiazo[4,5-b]pyridin-6-yl)-1-methyl-1H-benzo[d]imidazol-2-yl)acetamide (compound 9)

[0248] Intermediate 9-1 (67 mg, 0.25 mmol) and intermediate 6-1 (115 mg, 0.30 mmol) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (164 mg, 0.50 mmol) and 1,1-bis(tert-butylphosphine)ferrocene palladium chloride (Pd(dtbpf)Cl2) (24 mg, 0.04 mmol) were added to the solution. The reaction mixture was stirred at 80 °C for 16 h under nitrogen protection. The reaction was detected by LC-MS. The reaction solution was filtered, concentrated to dryness under reduced pressure, and purified by preparative high performance liquid chromatography (HPLC) using a YMC-Actus Triart C18 column (5 μm silica, 30 mm diameter, 150 mm length); water (containing 0.05% NH4HCO3) and a mixture of acetonitrile with decreasing polarity were used as the eluent; acetonitrile gradient ratio 30%-50%, elution time 12 min) to give the title compound (11 mg).

[0249] MS m / z(ESI): 526.1 [M+H] + .

[0250] 1 H NMR(400MHz,DMSO-d6)δ10.62(s,1H),8.31(s,1H),7.87(s,1H),7.60–7.43(m,4H),7 .39–7.30(m,3H),4.35–4.30(m,2H),3.30(s,3H),2.16(s,3H),1.32(t,J=7.0Hz,3H).

[0251] Example 10: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(4-oxo-1,2,3,9b-tetrahydro-4λ) 4 -Benzo[c]thieno[2,1-e]isothiazo-8-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 10)

[0252]

[0253] Intermediate 10-1 (200 mg, 0.52 mmol, synthesized according to the method reported in the literature "Tetrahydrobenzo[c]thieno-[2,1-e]isothiazole 4-Oxides: Three-Dimensional Heterocycles as Cross-Coupling Building Blocks") and intermediate 6-1 (201 mg, 0.62 mmol) were dissolved in dioxane (2 mL) and water (0.4 mL). Potassium phosphate (330 mg, 1.56 mmol) and 1,1-bis(tert-butylphosphine)ferrocene palladium chloride (Pd(dtbpf)Cl2) (33 mg, 0.05 mmol) were added to the solution. The reaction mixture was stirred at 100 °C for 16 h under nitrogen protection. The reaction was completed by LC-MS. After the reaction was cooled to room temperature, water (30 mL) and ethyl acetate (40 mL) were added sequentially. The organic phase was washed with water (30 mL * 2) and dried over an appropriate amount of anhydrous sodium sulfate. The crude product (210 mg) was obtained by thin-layer chromatography (silica, dichloromethane: methanol = 5:1), and then purified by preparative high-performance liquid chromatography (YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.05% trifluoroacetic acid) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 40%-60%, elution time 13 min) to obtain the title compound (30 mg).

[0254] MS m / z (ESI): 530.1 [M+H] + .

[0255] 1 H NMR(400MHz, DMSO-d6)δ8.16(s,1H),7.61(d,J=1.9Hz,1H),7.57–7.11(m,6H),6.76(d,J=8.3Hz,1H),4.91–4.84(m,1H),4.36–4.25(m,2 H),3.77–3.67(m,1H),3.62–3.49(m,1H),2.79–2.67(m,1H),2.22–2.13(m,1H),2.14–2.05(m,1H),1.67–1.51(m,1H),1.35–1.21(m,3H).

[0256] Example 11, 4-(4-(difluoromethoxy)phenyl)-6-(2,3-dihydro-1H-benzo[d]pyrrolo[1,2-a]imidazol-7-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (Compound 11)

[0257]

[0258] Intermediate 11-1 (149 mg, 628.03 μmol, synthesized according to the method reported in the literature "Metal-Free Sequential C(sp2)-H / OH and C(sp3)-HAminations of Nitrosoarenes and N-Heterocycles to Ring-Fused Imidazoles") and intermediate 6-1 (200 mg, 523.36 μmol) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (341 mg, 1.05 mmol) and 1,1-bis(tert-butylphosphine)ferrocene palladium chloride (Pd(dtbpf)Cl2) (34 mg, 52.01 μmol) were added to the solution. The reaction mixture was stirred at 90 °C for 2 h under nitrogen protection. The reaction was completed by LC-MS. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (YMC-ActusTriart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.05% formic acid) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 30%-50%, elution time 11 min) to give the title compound (57.0 mg).

[0259] MS m / z(ESI): 495.0 [M+H] + .

[0260] 1 H NMR(400MHz,DMSO-d6)δ8.29(s,1H),7.86(s,1H),7.56–7.19(m,6H),4.35-4.29(m,2H ),4.09(t,J=7.0Hz,2H),2.96–2.94(m,2H),2.67–2.64(m,2H),1.32(t,J=7.0Hz,3H).

[0261] Example 12: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-methyl-2H-indazol-5-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 12)

[0262]

[0263] Intermediate 1-8 (150 mg, 0.36 mmol) and intermediate 12-1 (111 mg, 0.43 mmol) were dissolved in anhydrous dioxane solution (2 mL). Water (0.4 mL), cesium carbonate (234 mg, 0.72 mmol), and 1,1-bis(tert-butylphosphine)ferrocene palladium chloride (Pd(dtbpf)Cl2) (23 mg, 0.03 mmol) were added to the solution. The reaction mixture was stirred at 100 °C for 2 h under nitrogen protection. The reaction was detected by LC-MS after completion. After the reaction was cooled to room temperature, the reaction solution was filtered, concentrated to dryness under reduced pressure, and purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.05% NH4HCO3) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 55%-80%, elution time 14 min] to obtain the title compound (12 mg).

[0264] MS m / z (ESI): 469.1 [M+H] + .

[0265] 1 H NMR (400MHz, DMSO-d6) δ8.36(s,1H),8.27(s,1H),8.05(s,1H),7.59–7.19(m,7H),4.34–4.29(m,2H),4.17(s,3H),1.33–1.30(m,3H).

[0266] Example 13, 4-(6-cyclopropylpyridin-3-yl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 13)

[0267]

[0268] Step 1: Synthesis of 6-bromo-4-(6-cyclopropylpyridin-3-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (intermediate 13-2)

[0269] Under an oxygen atmosphere, intermediate 1-7 (80.0 mg) was dissolved in 2 mL of dichloroethane (DCE) solution. Intermediate 13-1 (56.87 mg), copper acetate (105.6 mg), and pyridine (69.0 mg) were added to the reaction solution. After addition, the mixture was heated to 45 °C and stirred for 24 h. Subsequently, the reaction solution was concentrated to dryness under reduced pressure, and the residue was purified by silica gel column chromatography (PE / EA = 5 / 1-2 / 1). The title compound (40 mg) was obtained.

[0270] MS m / z(ESI): 392 [M+H] + .

[0271] Step 2: Synthesis of 4-(6-cyclopropylpyridin-3-yl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 13)

[0272] Under a nitrogen atmosphere, intermediate 13-2 (40.0 mg) was dissolved in dioxane solution (0.4 mL). Intermediate 3-1 (28.8 mg), potassium phosphate (43.3 mg), water (0.08 mL), and Pd(dtbpf)Cl2 (6.7 mg) were added to the reaction solution. After addition, the mixture was heated to 100 °C and stirred for 2 h. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by high-pressure preparation [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (2 mg).

[0273] MS m / z(ESI): 421[M+H] + .

[0274] 1 H NMR (400MHz, DMSO-d6) δ8.32(s,1H),8.22(d,J=2.6Hz,1H),8.20(s,1H),7.71–7.69(m,1H),7.66–7.63(m,1H),7.41(d,J=8. 3Hz,1H),6.37(d,J=9.5Hz,1H),4.28–4.23(m,2H),3.39(s,3H),2.20–2.10(m,1H),1.24(t,J=7.0Hz,3H),0.99–0.86(m,4H).

[0275] Example 14, 2-(2,2-difluoroethoxy)-4-(4-(difluoromethoxy)phenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 14)

[0276]

[0277] Step 1: Synthesis of 6-bromo-2-(2,2-difluoroethoxy)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 14-2)

[0278] Intermediate 14-1 (150 mg) was dissolved in tetrahydrofuran (THF) (2 mL), and NaH (77 mg, 60% wt) was added under an ice-water bath. After the addition was complete, the mixture was stirred at room temperature for 0.5 hours. 2,2-Difluoroethanol (119 mg) was then added under an ice-water bath. After the addition was complete, the mixture was stirred at room temperature for 12 hours. Subsequently, the reaction mixture was poured into a saturated ammonium chloride aqueous solution (5 mL), extracted with ethyl acetate (5 mL * 3), and the organic phases were combined. The mixture was washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to dryness under reduced pressure to give the title compound (110 mg).

[0279] MS m / z(ESI): 311.0 [M+H] + .

[0280] Step 2: Synthesis of 6-bromo-2-(2,2-difluoroethoxy)-4-(4-(difluoromethoxy)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 14-4)

[0281] Intermediate 14-2 (120 mg) was dissolved in dichloroethane (2 mL). Copper acetate (140 mg), pyridine (92 mg), and intermediate 14-3 (824 mg) were added to the reaction solution. After the addition was complete, air was bubbled into the reaction solution, and the solution was heated to 60 °C and stirred for 12 h. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (DCM / MeOH = 10 / 1) to give the title compound (90 mg).

[0282] MS m / z(ESI): 453.0 [M+H] + .

[0283] Step 3: Synthesis of 2-(2,2-difluoroethoxy)-4-(4-(difluoromethoxy)phenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 14)

[0284] Under a nitrogen atmosphere, intermediate 14-4 (90 mg) was dissolved in dioxane solution (1 mL), and intermediate 3-1 (56 mg), cesium carbonate (129 mg), water (0.2 mL), and Pd(dtbpf)Cl2 (13 mg) were added. After addition, the mixture was heated to 100 °C and stirred for 2 hours. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (HPLC) [YMC-ActusTriart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile in decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (25 mg).

[0285] MS m / z(ESI): 482.1 [M+H] + .

[0286] 1 H NMR (400MHz, DMSO-d6) δ8.24–8.21(m,2H),7.72–7.69(m,1H),7.49–7.12(m,5H),6.43–6.15(m,2H),4.57–4.49(m,2H),3.40(s,3H).

[0287] Example 15, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(6-methoxypyridazin-3-yl)thiazo[4,5-b]pyridine-5(4H)-one (Compound 15)

[0288]

[0289] Step 1: Synthesis of (4-(4-(difluoromethoxy)phenyl)-2-ethoxy-5-oxo-4,5-dihydrothiazo[4,5-b]pyridin-6-yl)boronic acid (intermediate 15-1)

[0290] Intermediate 1-8 (4.2 g) and bis-pinacol boronic acid ester (3.8 g) were dissolved in anhydrous dioxane (42 mL), and potassium acetate (1.9 g) and Pd(dppf)Cl2 (1.1 g) were added. The reaction mixture was stirred at 100 °C for 15 h under nitrogen protection. The reaction was detected by LC-MS. After the reaction was cooled to room temperature, the solvent was removed by rotary evaporation. Water (40 mL) and ethyl acetate (50 mL) were added sequentially. The organic phase was washed with water (15 mL * 2), and the washed organic phase was dried with an appropriate amount of anhydrous sodium sulfate. The mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm * 10 * 150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%) to give the title compound (3.2 g).

[0291] MS m / z (ESI): 383.1 [M+H] + .

[0292] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(6-methoxypyridazin-3-yl)thiazo[4,5-b]pyridine-5(4H)-one (compound 15)

[0293] Intermediate 15-1 (36.6 mg) and intermediate 15-2 (36.24 mg) were dissolved in dioxane (1 mL) and water (0.25 mL). Cesium carbonate (62.48 mg) and Pd(dtbpf)Cl2 (6.25 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 16 h under nitrogen protection. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to obtain the title compound (9.0 mg).

[0294] MS m / z (ESI): 447.1 [M+H] + .

[0295] 1 H NMR (400MHz, DMSO-d6) δ8.86 (s, 1H), 8.32 (d, J = 9.3Hz, 1H), 7.56–7.19 (m, 6H), 4.37-4.31 (m, 2H), 4.06 (s, 3H), 1.32 (t, J = 6.0Hz, 3H).

[0296] Example 16, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-methoxypyrimidin-5-yl)thiazo[4,5-b]pyridine-5(4H)-one (Compound 16)

[0297]

[0298] Intermediate 1-8 (40 mg) and intermediate 16-1 (45 mg) were dissolved in dioxane (1 mL) and water (0.2 mL), and cesium carbonate (62.47 mg) and Pd(dtbpf)Cl2 (6.25 mg) were added. Nitrogen gas was purged, and the mixture was stirred at 90 °C for 1 h. Thiol silica gel (50 mg) was added to the reaction mixture, and the mixture was stirred for 30 min. The mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (HPLC) (column: Phenomenex Luna C18 100*30 mm*5 μm; mobile phase: [water (0.05% CF3COOH, v / v)-acetonitrile]; B%: 43%-63%, 9 min) to obtain the title compound (10 mg).

[0299] MS m / z (ESI): 447.1 [M+H] + .

[0300] 1H NMR (400MHz, DMSO-d6) δ8.93(s,2H),8.42(s,1H),7.56–7.19(m,5H),4.37–4.31(m,2H),3.96(s,3H),1.32(t,J=6.0Hz,3H).

[0301] Example 17, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(2-hydroxypropane-2-yl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 17)

[0302]

[0303] Step 1: Synthesis of 2-(6-bromo-1-methyl-1H-benzo[d]imidazol-2-yl)propane-2-ol (intermediate 17-2)

[0304] Intermediate 17-1 (500 mg) was dissolved in dilute hydrochloric acid (4 mL, 2N), and 2-hydroxy-2-methylpropionic acid (388 mg) was added. The reaction mixture was stirred at 100 °C for 8 h. After the reaction was completed, the mixture was cooled to room temperature, and water (15 mL) and ethyl acetate (20 mL) were added sequentially. The organic phase was washed with water (10 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the mixture was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (dichloromethane:methanol = 10:1) to give the title compound (610 mg).

[0305] MS m / z(ESI): 269.0 [M+H] +

[0306] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(2-hydroxypropane-2-yl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 17)

[0307] Intermediate 6-1 (150 mg) and intermediate 17-2 (104 mg) were dissolved in 1,4-dioxane (1.2 mL) and water (0.3 mL). Pd(dtbpf)Cl2 (32 mg) and Cs2CO3 (211 mg) were added. The reaction mixture was stirred at 100 °C for 4 h under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (dichloromethane:methanol = 10:1) to obtain the crude product, which was then purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.225% formic acid) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 30%-60%, elution time 15 min] to obtain the title compound (3.1 mg).

[0308] MS m / z (ESI): 527.1 [M+H] + .

[0309] 1 H NMR(400MHz,DMSO-d6)δ8.17(s,1H),7.86(s,1H),7.65–7.63(m,1H),7.53–7.51(m,1H),7.46–7.44(m,2H) ,7.34–7.32(m,2H),7.14–6.77(m,1H),4.42–4.36(m,2H),4.10(s,3H),1.74(s,6H),1.36(t,J=7.0Hz,3H).

[0310] Example 18, 4-(4-(difluoromethoxy)phenyl)-6-(2,3-dihydrobenzofuran-5-yl)-2-ethoxythiazo[4,5-b]pyridine-5(4H)-one (Compound 18)

[0311]

[0312] Intermediate 6-1 (80 mg) and intermediate 18-1 (51 mg) were dissolved in 1,4-dioxane (1.2 mL) and water (0.3 mL). Pd(dtbpf)Cl2 (17 mg) and Cs2CO3 (112 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 10 h under nitrogen protection. After the reaction was completed, the reaction solution was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (dichloromethane:methanol = 20:1) to obtain the crude product, which was then purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.225% formic acid) and acetonitrile in decreasing polarity mixture as eluent; acetonitrile gradient ratio 40%-70%, elution time 15 min] to obtain the title compound (9 mg).

[0313] MS m / z (ESI): 457.1 [M+H] + .

[0314] 1 H NMR(400MHz,DMSO-d6)δ8.16(s,1H),7.57(s,1H),7.56–7.17(m,6H),6.77(d,J=8.3Hz,1H ),4.55(t,J=8.7Hz,2H),4.33–4.28(m,2H),3.19(t,J=8.7Hz,2H),1.31(t,J=7.0Hz,3H).

[0315] Example 19, 4-(4-(difluoromethoxy)phenyl)-6-(1-methyl-1H-benzo[d]imidazol-6-yl)-2-(oxacyclobut-3-yloxy)thiazo[4,5-b]pyridin-5(4H)-one (Compound 19)

[0316]

[0317] Step 1: Synthesis of 2,6-dibromo-5-methoxythiazo[4,5-b]pyridine (intermediate 19-1)

[0318] Copper bromide (103 g) and isoamyl nitrite (86 g) were dissolved in acetonitrile (1000 mL) and cooled to 0 °C. Intermediate 1-4 (60 g) was added in portions to the reaction flask, and the reaction mixture was stirred at 25 °C for 16 h. The reaction mixture was then poured into dilute HCl solution and stirred for 30 min. The pH was adjusted to 8 with sodium bicarbonate, and the mixture was extracted with a mixed solvent of dichloromethane:methanol = 10:1 (200 mL * 3). The combined organic phases were dried over anhydrous sodium sulfate. The title compound (12 g) was obtained by preparative thin-layer chromatography (silica, dichloromethane).

[0319] MS m / z(ESI): 323.1 [M+H] + .

[0320] Step 2: Synthesis of 2,6-dibromothiazo[4,5-b]pyridin-5(4H)-one (intermediate 19-2)

[0321] Intermediate 19-1 (8 g) was dissolved in acetic acid (50 mL), and 48% hydrobromic acid (52 mL) was added. The reaction mixture was stirred at 80 °C for 5 h. After the reaction was completed, the solvent was removed by concentration under reduced pressure, and saturated sodium bicarbonate aqueous solution (60 mL) was added. The mixture was filtered, and the filter cake was washed with water (40 mL * 2) to obtain the title compound (9.5 g).

[0322] MS m / z(ESI): 309.1 [M+H] + .

[0323] Step 3: Synthesis of 6-bromo-2-(oxetane-3-yloxy)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 19-3)

[0324] 3-oxetanebutanol (358.5 mg, 4.84 mmol) was dissolved in tetrahydrofuran (10 mL), and NaH (193.6 mg, 4.84 mmol) was added at 0 °C. After the addition was complete, the mixture was stirred for 10 minutes, and then intermediate 19-2 (500 mg, 1.61 mmol) was added. The mixture was stirred at room temperature for 3 hours. The reaction was quenched by adding water (10 mL), and the mixture was extracted with ethyl acetate (20 mL * 3). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (DCM:CH3OH = 10:1) to give the title compound (52.0 mg).

[0325] MS m / z (ESI): 303.1 [M+H] + .

[0326] Step 4: Synthesis of 6-bromo-4-(4-(difluoromethoxy)phenyl)-2-(oxetane-3-yloxy)thiazo[4,5-b]pyridine-5(4H)-one (intermediate 19-4)

[0327] Intermediate 19-3 (50 mg) and 4-(difluoromethoxy)phenylboronic acid (62 mg) were dissolved in dichloroethane (2 ml), and then Cu(OAc)2 (44.9 mg) and pyridine (39.14 mg) were added. The mixture was reacted at 45 °C for 12 h. The reaction solution was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (DCM:CH3OH = 10:1) to give the title compound (57.9 mg).

[0328] MS m / z (ESI): 445.1 [M+H] +

[0329] Step 5: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(1-methyl-1H-benzo[d]imidazol-6-yl)-2-(oxetane-3-yloxy)thiazo[4,5-b]pyridin-5(4H)-one (compound 19)

[0330] Intermediate 5-1 (57.97 mg) and intermediate 19-1 (50 mg) were dissolved in dioxane (1 mL) and water (0.2 mL). Cesium carbonate (73.17 mg) and Pd(dtbpf)Cl2 (7.32 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 16 h under nitrogen protection. The reaction mixture was then filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to give the title compound (9.8 mg).

[0331] MS m / z(ESI): 497.1 [M+H] + .

[0332] 1 H NMR (400MHz, DMSO-d6) δ8.27(s,1H),8.13(s,1H),7.88(s,1H),7.59-7.49(m,1H),7. 45-7.12(m,6H),5.43-5.40(m,1H)4.63-4.60(m,2H)4.55-4.52(m,2H),3.76(s,3H).

[0333] Example 20: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(imidazo[1,2-a]pyridin-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 20)

[0334]

[0335] Intermediate 15-1 (40 mg) and intermediate 20-1 (24.75 mg) were dissolved in dioxane (1 mL) and water (0.25 mL). Cesium carbonate (68.2 mg) and Pd(dtbpf)Cl2 (6.82 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 16 h under nitrogen protection. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C185 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to obtain the title compound (18.0 mg).

[0336] MS m / z (ESI): 455.1 [M+H] + .

[0337] 1 H NMR (400MHz, DMSO-d6) δ9.14(s,1H),8.45(s,1H),7.98(s,1H),7.60-7.19(m,8H),4.36-4.31(m,2H),1.31(t,J=7.1Hz,3H).

[0338] Example 21: 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-methyl-2,3-dihydro-1H-benzo[d]imidazol[1,2-a]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 21)

[0339]

[0340] Compound 28 (57 mg) was dissolved in anhydrous tetrahydrofuran (0.8 mL), and a solution of bis(trimethylsilylamine) lithium (1 mol / L THF solution, 0.15 mL) was added at 0 °C. After 30 min, iodomethane (33 mg) was added, and the reaction mixture was stirred at 30 °C for 4 h. After the reaction was completed, the solvent was removed by concentration under reduced pressure, and the residue was purified by preparative high performance liquid chromatography [YMC-ActusTriart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.225% formic acid) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 40%-70%, elution time 14 min] to obtain the title compound (16 mg).

[0341] MS m / z (ESI): 510.1 [M+H] + .

[0342] 1H NMR(400MHz,DMSO-d6)δ8.20–8.16(m,1H),7.59–7.09(m,8H),4.33–4.28(m,2 H),4.14–4.08(m,2H),3.94–3.85(m,2H),2.93(s,3H),1.31(t,J=7.0Hz,3H).

[0343] Example 22, 6-(benzo[4,5]imidazo[1,2-a]pyrimidin-7-yl)-4-(4-(difluoromethoxy)phenyl)-2-ethoxythiazo[4,5-b]pyridine-5(4H)-one (Compound 22)

[0344]

[0345] Intermediate 22-1 (48.69 mg) and intermediate 15-1 (50 mg) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (85.26 mg) and Pd(dtbpf)Cl2 (8.53 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 16 h under nitrogen protection. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to obtain the title compound (4.5 mg).

[0346] MS m / z(ESI): 506.0 [M+H] + .

[0347] 1 H NMR (400MHz, DMSO-d6) δ9.56–9.54(m,1H),8.84–8.82(m,1H),8.74(s,1H),8.38( s,1H),7.88(s,2H),7.56–7.14(m,6H),4.37-4.31(m,2H),1.32(t,J=8.0Hz,3H).

[0348] Example 23, 4-(4-(difluoromethoxy)phenyl)-6-(1-(methyl-d3)-1H-benzo[d]imidazol-6-yl)-2-(trifluoromethyl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 23)

[0349]

[0350] Step 1: Synthesis of 1-(methyl-d3)-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborone-2-yl)-1H-benzi[d]imidazole (intermediate 23-1)

[0351] Under a nitrogen atmosphere, intermediate 8-2 (1.0 g) was dissolved in 10 mL of dioxane solution, and potassium acetate (917 mg), bis-pinacolborate (1.42 g), and Pd(dppf)Cl2 (342 mg) were added. The reaction mixture was heated to 100 °C and stirred for 2 h. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (PE / EA = 5 / 1-3 / 1) to give the title compound (500 mg).

[0352] MS m / z(ESI): 262.0 [M+H] + .

[0353] Step 2: Synthesis of 2,6-dibromo-4-(4-(difluoromethoxy)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 23-3)

[0354] Intermediate 23-2 (450 mg) was dissolved in 1,2-dichloroethane (5 mL), and copper acetate (318 mg), pyridine (0.4 mL), and intermediate 14-3 (409 mg) were added. After the addition was complete, air was bubbled through the reaction mixture, and the reaction solution was heated to 60 °C and stirred for 48 h. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (DCM:MeOH = 20:1) to give the title compound (450 mg).

[0355] MS m / z(ESI): 451.0 [M+H] + .

[0356] Step 3: Synthesis of 6-bromo-4-(4-(difluoromethoxy)phenyl)-2-(trifluoromethyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 23-4)

[0357] Under a nitrogen atmosphere, intermediate 23-3 (400 mg) was dissolved in DMF (5 mL), and cuprous iodide (202 mg) and methyl 2,2-difluoro-2-(fluorosulfonyl)acetate (204 mg) were added. The reaction mixture was heated to 100 °C and stirred for 12 h. The reaction mixture was then filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (DCM:MeOH = 20 / 1-10 / 1) to give the title compound (150 mg).

[0358] MS m / z(ESI): 441.0 [M+H] + .

[0359] Step 4: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(1-(methyl-d3)-1H-benzo[d]imidazol-6-yl)-2-(trifluoromethyl)thiazo[4,5-b]pyridin-5(4H)-one (compound 23)

[0360] Under a nitrogen atmosphere, intermediate 23-4 (120.0 mg) was dissolved in dioxane solution (2 mL), and intermediate 23-1 (71 mg), cesium carbonate (177 mg), water (0.2 mL), and Pd(dtbpf)Cl2 (18 mg) were added. After the addition was complete, the mixture was heated to 100 °C and stirred for 2 h. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (HPLC) [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (25 mg).

[0361] MS m / z (ESI): 496.1 [M+H] + .

[0362] 1 H NMR (400MHz, DMSO-d6) δ8.66(s,1H),8.27(s,1H),8.03(s,1H),7.73(d,J=8.4Hz,1H),7.60–7.24(m,6H).

[0363] Example 24, 4-(4-(difluoromethoxy)phenyl)-2-methoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 24)

[0364]

[0365] Step 1: Synthesis of 6-bromo-4-(4-(difluoromethoxy)phenyl)-2-methoxythiazo[4,5-b]pyridin-5(4H)-one (intermediate 24-1)

[0366] Intermediate 23-3 (250 mg) was dissolved in methanol (5 mL), sodium methoxide (120 mg) was added, and the reaction was carried out at 25 °C for 5 h. Subsequently, the reaction solution was concentrated under reduced pressure to remove the solvent, the residue was washed with water (10 mL), extracted with ethyl acetate (10 mL * 2), and the organic phase was dried with an appropriate amount of anhydrous sodium sulfate. The title compound (172 mg) was obtained by preparative thin-layer chromatography (petroleum ether: ethyl acetate = 2:1).

[0367] MS m / z(ESI): 402.9 [M+H] + .

[0368] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-methoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 24)

[0369] Intermediate 24-1 (80 mg) and intermediate 3-1 (70 mg) were dissolved in 1,4-dioxane (1.2 mL) and water (0.3 mL). Pd(dtbpf)Cl2 (19 mg) and cesium carbonate (196 mg) were added to the solution, and the mixture was reacted at 100 °C for 4 h under a nitrogen atmosphere. The reaction solution was then filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NXC 185 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 17 min) to give the title compound (13 mg).

[0370] MS m / z(ESI): 432.0 [M+H] + .

[0371] 1 H NMR (400MHz, DMSO-d6) δ8.30(d,J=2.5Hz,1H),8.26(s,1H),7.79–7.76(m,1H),7.62–7.14(m,5H),6.44(d,J=9.5Hz,1H),3.97(s,3H),3.46(s,3H).

[0372] Example 25, 2-ethoxy-4-(4-fluorophenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 25)

[0373]

[0374] Step 1: Synthesis of 6-bromo-2-ethoxy-4-(4-fluorophenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 25-2)

[0375] Intermediate 1-7 (100 mg) and intermediate 25-1 (156 mg) were dissolved in anhydrous 1,2-dichloroethane (5 mL), and pyridine (86 mg) and copper acetate (79 mg) were added. After the addition was complete, the reaction mixture was stirred at 40 °C for 4 hours. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to remove the solvent. The residue was purified by residual column chromatography (dichloromethane:methanol = 20:1) to give the title compound (100 mg).

[0376] MS m / z(ESI): 368.0 [M+H] + .

[0377] Step 2: Synthesis of 2-ethoxy-4-(4-fluorophenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 25)

[0378] Intermediate 25-2 (55 mg) was dissolved in 1,4-dioxane solution (5 mL) and water (0.5 mL). Intermediate 3-1 (73 mg), Pd(dppf)Cl2 (11 mg), and cesium carbonate (151 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (5 mL * 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100 * 25 mm * 5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (38 mg).

[0379] MS m / z(ESI): 398.0 [M+H] + .

[0380] 1 H NMR (400MHz, DMSO-d6) δ8.32(d,J=2.6Hz,1H),8.27(s,1H),7.80(dd,J=9.5,2.7Hz,1H),7.48–7 .44(m,2H),7.42–7.34(m,2H),6.46(d,J=9.5Hz,1H),4.36–4.30(m,2H),1.32(t,J=7.0Hz,3H).

[0381] Example 26, 4-(3-chlorophenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 26)

[0382]

[0383] Step 1: Synthesis of 6-bromo-4-(3-chlorophenyl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (intermediate 26-2)

[0384] Intermediates 1-7 (100 mg) and 26-1 (156 mg) were dissolved in anhydrous 1,2-dichloroethane (5 mL), and pyridine (86 mg) and copper acetate (79 mg) were added. After the addition was complete, the reaction mixture was stirred at 40 °C for 4 hours. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (dichloromethane:methanol = 20:1) to give the title compound (170 mg).

[0385] MS m / z(ESI): 385.0 [M+H] + .

[0386] Step 2: Synthesis of 4-(3-chlorophenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 26)

[0387] Intermediate 26-2 (60 mg) was dissolved in 1,4-dioxane solution (5 mL) and water (0.5 mL). Intermediate 3-1 (73 mg), Pd(dppf)Cl2 (11 mg), and cesium carbonate (151 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (5 mL x 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100 x 25 mm x 5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (38 mg).

[0388] MS m / z(ESI): 414.0 [M+H] + .

[0389] 1H NMR (400MHz, DMSO-d6) δ8.31(d,J=2.6Hz,1H),8.27(s,1H),7.80(dd,J=9.5,2.6Hz,1H),7.62–7.51(m ,3H),7.42–7.38(m,1H),6.45(d,J=9.5Hz,1H),4.44–4.18(m,2H),3.48(s,3H),1.32(t,J=7.1Hz,3H).

[0390] Example 27, 4-(4-(difluoromethoxy)phenyl)-6-(3,4-dihydro-2H-benzo[4,5]imidazo[2,1-b][1,3]oxazin-7-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (compound 27)

[0391]

[0392] Step 1: Synthesis of 1-(3-hydroxypropyl)-3-(4-iodophenyl)thiourea (intermediate 27-3)

[0393] Intermediate 27-1 (1.0 g) and intermediate 27-2 (261.5 mg) were dissolved in anhydrous tetrahydrofuran (10 mL) and stirred at room temperature for 2 h. The reaction solution was concentrated under reduced pressure to give the title compound (1.1 g).

[0394] MS m / z (ESI): 337.1 [M+H] + .

[0395] Step 2: Synthesis of N-(4-iodophenyl)-5,6-dihydro-4H-1,3-oxazine-2-amine (intermediate 27-4)

[0396] Intermediate 27-3 (1.1 g) was dissolved in tetrahydrofuran (20 mL), and a solution of sodium hydroxide (327.19 mg) in water (5 mL) was added. The mixture was stirred at room temperature for 10 min. Then, p-toluenesulfonyl chloride (TsCl, 748.56 mg) was added, and the mixture was stirred at room temperature for 3 h. Subsequently, the reaction solution was concentrated under reduced pressure, extracted with ethyl acetate (40 mL), washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (EA:Et3N = 100:1) to give the title compound (1.1 g).

[0397] MS m / z (ESI): 303.1 [M+H] + .

[0398] Step 3: Synthesis of 7-iodo-3,4-dihydro-2H-benzo[4,5]imidazo[2,1-b][1,3]oxazine (intermediate 27-6)

[0399] Intermediate 27-4 (200 mg) was dissolved in acetonitrile (10 mL), and intermediate 27-5 was added at 0 °C. The mixture was stirred at room temperature for 2 h. Subsequently, the reaction solution was washed with saturated sodium bicarbonate solution (10 mL) and extracted with dichloromethane (20 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (EA:Et3N = 100:1) to give the title compound (110 mg).

[0400] MS m / z (ESI): 301.1 [M+H] + .

[0401] Step 4: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(3,4-dihydro-2H-benzo[4,5]imidazo[2,1-b][1,3]oxazin-7-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (compound 27)

[0402] Intermediate 27-6 (94.23 mg) and intermediate 15-1 (60 mg) were dissolved in dioxane (2 mL) and water (0.5 mL). Cesium carbonate (102.31 mg) and Pd(dtbpf)Cl2 (10.22 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 16 h under nitrogen protection. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm*10*150 mm; mobile phase: A: water (0.225% formic acid v / v), B: acetonitrile; B%: 30%-50%, 11 min) to give the title compound (14 mg).

[0403] MS m / z (ESI): 511.1 [M+H] + .

[0404] 1 H NMR(400MHz,Methanol-d4)δ8.26(s,1H),7.73(s,1H),7.56-7.19(m,7H),4.53-4.50( m,2H),4.34-4.29(m,2H),4.11-4.08(m,2H),2.26-2.23(m,2H),1.31(t,J=7.0Hz,3H)

[0405] Example 28, 4-(4-(difluoromethoxy)phenyl)-6-(2,3-dihydro-1H-benzo[d]imidazo[1,2-a]imidazo-6-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (Compound 28)

[0406]

[0407] Step 1: Synthesis of 2-((6-bromo-1H-benzo[d]imidazol-2-yl)amino)ethanol-1-ol (intermediate 28-2)

[0408] Intermediate 28-1 (4.6 g) was dissolved in ethanolamine (3.6 mL), and the reaction mixture was stirred at 140 °C for 14 h. After the reaction was completed, the mixture was cooled to room temperature, and saturated sodium bicarbonate solution (25 mL) was added. A solid precipitated, the reaction mixture was filtered, and the filter cake was washed with water (40 mL) and dried to obtain the crude product of the title compound (4.1 g).

[0409] MS m / z (ESI): 255.9 [M+H] +

[0410] Step 2: Synthesis of 6-bromo-2,3-dihydro-1H-benzo[d]imidazo[1,2-a]imidazolium (intermediate 28-3)

[0411] Intermediate 28-2 (2.6 g) was dissolved in 1,2-dichloroethane (20 mL), and thionyl chloride (0.9 mL) was added. The reaction mixture was stirred at 80 °C for 50 min. After the reaction was complete, the reaction mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, and saturated sodium bicarbonate aqueous solution (30 mL) was added to the residue. The mixture was extracted with a dichloromethane / methanol co-solvent (25 mL * 2). The organic phase was dried over an appropriate amount of anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and xylene (20 mL) was added to the residue. The reaction mixture was stirred at 110 °C for 20 h. After the reaction mixture was cooled to room temperature, the precipitate was collected by filtration, washed with petroleum ether, and dried to give the title compound (2.2 g).

[0412] MS m / z(ESI): 238.0 [M+H] +

[0413] Step 3: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(2,3-dihydro-1H-benzo[d]imidazo[1,2-a]imidazo-6-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (compound 28)

[0414] Intermediate 6-1 (180 mg) and intermediate 28-3 (138 mg) were dissolved in dioxane (1.2 mL) and water (0.3 mL). Cesium carbonate (253 mg) and Pd(dtbpf)Cl2 (38 mg) were added. The reaction mixture was stirred at 100 °C for 4 h under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.225% formic acid) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 40%-70%, elution time 12 min] to obtain the title compound (88.4 mg).

[0415] MS m / z(ESI): 496.0 [M+H] + .

[0416] 1 H NMR(400MHz,Chloroform-d)δ8.21–8.18(m,1H),7.55(s,1H),7.52–7.08(m,7H),6.96(d,J= 17.1Hz,1H),4.33–4.28(m,2H),4.04–4.08(m,2H),3.98–3.95(m,2H),1.31(t,J=7.0Hz,3H).

[0417] Example 29, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-oxo-2,3-dihydro-1H-benzo[d]pyrrolo[1,2-a]imidazol-7-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 29)

[0418]

[0419] Step 1: Synthesis of 1-(5-bromo-2-nitrophenyl)pyrrolidine-2,5-dione (intermediate 29-1)

[0420] 4-Bromonitrobenzene (1 g) and succinimide (588 mg) were dissolved in anhydrous N,N-dimethylformamide (20 mL), and potassium carbonate (2.38 g) was added. The mixture was stirred at 25 °C for 12 hours. After the reaction was complete, water (50 mL) and ethyl acetate (50 mL x 3) were added sequentially. The organic phase was washed with saturated brine (25 mL x 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 1:1) to give the title compound (1 g).

[0421] MS m / z(ESI): 299.0 [M+H] +

[0422] Step 2: Synthesis of 1-(2-amino-5-bromophenyl)pyrrolidine-2,5-dione (intermediate 29-2)

[0423] Intermediate 29-1 (1 g) was dissolved in acetic acid (10 mL). Iron powder (1.87 g) was added to the reaction solution, and the reaction solution was stirred at 25 °C for 12 hours. After the reaction was completed, water (100 mL) and ethyl acetate (100 mL) were added sequentially. The organic phase was washed with water (50 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 1:1) to give the title compound (0.6 g).

[0424] MS m / z(ESI): 269.0 [M+H] +

[0425] Step 3: Synthesis of 7-bromo-2,3-dihydro-1H-benzo[d]pyrrole[1,2-a]imidazol-1-one (intermediate 29-3)

[0426] Intermediate 29-2 (1 g) was dissolved in acetic acid (10 mL), and the reaction mixture was stirred at 40 °C for 12 hours. After the reaction was complete, water (100 mL) and ethyl acetate (100 mL) were added sequentially. The organic phase was washed with water (50 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent, and the residue was purified by column chromatography (petroleum ether: ethyl acetate = 1:1) to give the title compound (0.2 g).

[0427] MS m / z(ESI): 251.0 [M+H] +

[0428] Step 4: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(trimethyltinyl)thiazo[4,5-b]pyridine-5(4H)-one (intermediate 29-4)

[0429] Under nitrogen protection, intermediate 1-8 (50 mg) was added to anhydrous dioxane (6 mL), followed by the addition of tetra(triphenylphosphine)palladium (13.8 mg) and hexamethyldistin (58 mg). The reaction mixture was then stirred at 120 °C for 16 hours under nitrogen protection. After the reaction was complete, the reactants were filtered and concentrated to dryness under reduced pressure. The residue was analyzed by preparative thin-layer chromatography (petroleum ether:ethyl acetate = 10:1) to give the title compound (40 mg).

[0430] MS m / z(ESI): 503.0 [M+H] +

[0431] Step 5: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-oxo-2,3-dihydro-1H-benzo[d]pyrrolo[1,2-a]imidazol-7-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 29)

[0432] Under nitrogen protection, intermediates 29-4 (40 mg) and 29-3 (10 mg) were added to anhydrous dioxane (6 mL), and tetra(triphenylphosphine)palladium (9.2 mg) was added to the reaction mixture. The reaction mixture was then stirred at 120 °C for 16 hours under nitrogen protection. After the reaction was complete, the reactants were filtered and concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high-performance liquid chromatography (column: Waters Xbridge BEH C18 100*25mm*5μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 27%-47%, 14 min) to give the title compound (1.5 mg).

[0433] MS m / z(ESI): 509.0 [M+H] +

[0434] 1 H NMR(400MHz,Chloroform-d)δ8.01(d,J=1.6Hz,1H),7.93(d,J=8.4Hz,1H),7.85(s,1H),7.79–7.77(m,1H),7.42(s,1H),7.40 (s,1H),7.28(s,2H),6.58(t,J=73.6Hz,1H),4.41–4.36(m,2H),3.35–3.32(m,2H),3.26–3.23(m,2H),1.39(t,J=7.1Hz,3H).

[0435] Example 30, 4-(4-(difluoromethoxy)phenyl)-6-(1-(methyl-d3)-1H-benzo[d]imidazol-6-yl)-2-propoxythiazo[4,5-b]pyridin-5(4H)-one (Compound 30)

[0436]

[0437] Step 1: Synthesis of 6-bromo-4-(4-(difluoromethoxy)phenyl)-2-propoxythiazo[4,5-b]pyridin-5(4H)-one (intermediate 30-1)

[0438] Sodium hydride (133 mg) was dissolved in anhydrous tetrahydrofuran (2 mL), and n-propanol (198 mg) was added at 0 °C. After 30 min, intermediate 23-3 (150 mg) was added, and the reaction mixture was stirred at 25 °C for 4 hours. After the reaction was complete, the pH was adjusted to 7 with dilute hydrochloric acid (2N), the solvent was removed by concentration under reduced pressure, water (10 mL) was added, and the mixture was extracted with ethyl acetate (10 mL * 2). The organic phase was dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated to dryness under reduced pressure, and the residue was purified by column chromatography (petroleum ether: ethyl acetate = 2:1) to give the title compound (71 mg).

[0439] MS m / z (ESI): 431.8 [M+H] +

[0440] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-6-(1-(methyl-d3)-1H-benzo[d]imidazol-6-yl)-2-propoxythiazo[4,5-b]pyridin-5(4H)-one (compound 30)

[0441] Intermediate 30-1 (68 mg) and intermediate 23-1 (62 mg) were dissolved in dioxane (1.2 mL) and water (0.3 mL). Cesium carbonate (103 mg) and Pd(dtbpf)Cl2 (15 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 10 h under nitrogen protection. After the reaction was completed, the reaction solution was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure and purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent of a mixture of water (containing 0.225% formic acid) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 40%-70%, elution time 14 min] to obtain the title compound (17 mg).

[0442] MS m / z (ESI): 486.1 [M+H] + .

[0443] 1 H NMR (400MHz, DMSO-d6) δ8.32–8.27(m,1H),8.19–8.18(m,1H),7.96(s,1H),7.70–7.36( m,5H),7.36–7.31(m,2H),4.25–4.22(m,2H),1.77–1.68(m,2H),0.91(t,J=7.4Hz,3H).

[0444] Example 31, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(methyl-d3)-2H-indazol-5-yl)thiazo[4,5-b]pyridine-5(4H)-one (Compound 31)

[0445]

[0446] Under a nitrogen atmosphere, intermediate 6-1 (100 mg) was dissolved in dioxane solution (1 mL), and intermediate 31-1 (55 mg), cesium carbonate (140 mg), water (0.2 mL), and Pd(dtbpf)Cl2 (14 mg) were added. The reaction solution was heated to 100 °C and stirred for 2 h. Subsequently, the reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (HPLC) [YMC-ActusTriart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (15 mg).

[0447] MS m / z(ESI): 472.1 [M+H] +

[0448] 1 H NMR (400MHz, DMSO-d6) δ8.35(s,1H),8.28(s,1H),8.05(s,1H),7.59–7.19(m,7H),4.35–4.29(m,2H),1.32(t,J=7.0Hz,3H).

[0449] Example 32, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(ethoxymethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 32)

[0450]

[0451] Step 1: Synthesis of 6-bromo-2-(ethoxymethyl)-1-methyl-1H-benzi[d]imidazole (intermediate 32-2)

[0452] Intermediate 32-1 (200 mg) was dissolved in THF (2 mL), and NaH (66 mg, 60% wt) was added under an ice-water bath. After the addition was complete, the mixture was stirred at room temperature for 0.5 hours. Iodoethane (388 mg) was added under an ice-water bath, followed by stirring at room temperature for 12 hours. TLC analysis showed that the reaction was complete. The reaction mixture was poured into a saturated ammonium chloride aqueous solution (5 mL), extracted with ethyl acetate (5 mL * 3), and the organic phases were combined. The mixture was washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to give the title compound (140 mg).

[0453] MS m / z(ESI): 269.0 [M+H] +

[0454] Step 2: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(2-(ethoxymethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 32)

[0455] Under a nitrogen atmosphere, intermediate 32-2 (75 mg) was dissolved in dioxane (1 mL), and intermediate 6-1 (129 mg), cesium carbonate (182 mg), water (0.2 mL), and Pd(dtbpf)Cl2 (18 mg) were added. The reaction mixture was heated to 100 °C and stirred for 2 h. Subsequently, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high-performance liquid chromatography (HPLC) [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile in decreasing polarity; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (25 mg).

[0456] MS m / z (ESI): 527.1 [M+H] +

[0457] 1 H NMR (400MHz, DMSO-d6) δ8.34(s,1H),7.95(s,1H),7.63–7.20(m,7H),4.75(s,2H),4.33(t,J= 7.0Hz,2H),3.82(s,3H),3.53(t,J=7.0Hz,2H),1.33(t,J=7.0Hz,3H),1.16(t,J=7.0Hz,3H).

[0458] Example 33, 6-(2-(aminomethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)-4-(4-(difluoromethoxy)phenyl)-2-ethoxythiazo[4,5-b]pyridine-5(4H)-one (compound 33)

[0459]

[0460] Step 1: Synthesis of 2-(azidomethyl)-6-bromo-1-methyl-1H-benzi[d]imidazole (intermediate 33-1)

[0461] Intermediate 32-1 (500 mg) was dissolved in THF (5 mL), and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (474 ​​mg) and diphenyl azidophosphate (DPPA) (685 mg) were added. The mixture was stirred at room temperature for 12 h. Subsequently, the reaction solution was poured into water (5 mL), extracted with ethyl acetate (5 mL * 3), and the organic phases were combined and washed with saturated brine (10 mL). The organic phases were dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure. The residue was purified by column chromatography (PE / EA = 5 / 1-2 / 1) to give the title compound (450 mg).

[0462] MS m / z(ESI): 266.0 [M+H] +

[0463] Step 2: Synthesis of (6-bromo-1-methyl-1H-benzo[d]imidazol-2-yl)methylamine (intermediate 33-2)

[0464] Intermediate 33-1 (500 mg) was dissolved in THF (5 mL), and water (1 mL) and triphenylphosphine (591 mg) were added. After the addition was complete, the mixture was stirred at room temperature for 12 h. Subsequently, the reaction solution was poured into water (5 mL), extracted with ethyl acetate (5 mL * 3), the organic phases were combined, washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, the solvent was removed under reduced pressure, and the concentrated residue was purified by column chromatography (DCM / MeOH = 20 / 1) to give the title compound (160 mg).

[0465] MS m / z(ESI): 240.0 [M+H] +

[0466] Step 3: Synthesis of 6-(2-(aminomethyl)-1-methyl-1H-benzo[d]imidazol-6-yl)-4-(4-(difluoromethoxy)phenyl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (compound 33)

[0467] Under a nitrogen atmosphere, intermediate 6-1 (100 mg) was dissolved in dioxane solution (1 mL), and intermediate 33-2 (52 mg), cesium carbonate (140 mg), water (0.2 mL), and Pd(dtbpf)Cl2 (14 mg) were added. The reaction solution was heated to 100 °C and stirred for 2 h. LCMS monitoring showed that the reaction was complete. The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by high-performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.05% NH4HCO3) and a mixture of acetonitrile with decreasing polarity as eluent; acetonitrile gradient ratio 55%-80%, elution time 13 min], to obtain the title compound (13 mg).

[0468] MS m / z (ESI): 498.1 [M+H] +

[0469] 1 H NMR(400MHz,DMSO-d6)δ8.31(s,1H),7.89(s,1H),7.57–7.55(m,1H),7.50–7.19 (m,6H),4.32(d,J=7.1Hz,2H),3.98(s,2H),3.77(s,3H),1.32(t,J=7.0Hz,3H).

[0470] Example 34, 2-ethoxy-4-(4-ethynylphenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 34)

[0471]

[0472] Step 1: Synthesis of 6-bromo-2-ethoxy-4-(4-iodophenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 34-2)

[0473] Intermediate 1-7 (100 mg) and intermediate 34-1 (135 mg) were dissolved in anhydrous 1,2-dichloroethane (5 mL). Pyridine (86 mg) and copper acetate (79 mg) were added dropwise. After the addition was complete, the mixture was stirred at 40 °C for 4 hours. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100*25mm*5μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (170 mg).

[0474] MS m / z(ESI): 477.0 [M+H] +

[0475] Step 2: Synthesis of 6-bromo-2-ethoxy-4-(4-((trimethylsilyl)ethynyl)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 34-3)

[0476] Intermediate 34-2 (100 mg) and 1-(trimethylsilyl)acetylene (25 mg) were dissolved in anhydrous tetrahydrofuran solution (5 mL), and cuprous iodide (12 mg), triethylamine (21 mg), and tetra(triphenylphosphine)palladium (12.10 mg) were added. The reaction mixture was stirred at 25 °C for 12 hours under nitrogen protection. After the reaction mixture cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (50 mL x 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent, and the residue was purified by column chromatography (dichloromethane:methanol = 20:1) to give the title compound (70 mg).

[0477] MS m / z(ESI): 447.0 [M+H] +

[0478] Step 3: Synthesis of 2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-4-(4-((trimethylsilyl)ethynyl)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 34-4)

[0479] Intermediate 34-3 (70 mg) was dissolved in 1,4-dioxane (5 mL) and water (0.5 mL). Intermediate 3-1 (73.56 mg), Pd(dppf)Cl2 (11 mg), and potassium phosphate (99 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (5 mL x 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100 x 25 mm x 5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (40 mg).

[0480] MS m / z(ESI): 476.0 [M+H] +

[0481] Step 4: Synthesis of 2-ethoxy-4-(4-ethynylphenyl)-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 34)

[0482] Intermediate 34-4 (40 mg) was dissolved in anhydrous ethanol (2 mL), and potassium carbonate (35 mg) was added. The reaction mixture was stirred at 25 °C for 2 hours. After the reaction was complete, the mixture was filtered, and the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100*25mm*5μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (15 mg).

[0483] MS m / z(ESI): 404.0 [M+H] +

[0484] 1 H NMR (400MHz, DMSO-d6) δ8.30(d,J=2.6Hz,1H),8.25(s,1H),7.78(dd,J=9.5,2.7Hz,1H),7.66–7.62(m,2 H),7.45–7.35(m,2H),6.44(d,J=9.5Hz,1H),4.31(d,J=6.8Hz,3H),3.46(s,3H),1.30(t,J=7.0Hz,3H).

[0485] Example 35, 2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-4-(4-(prop-1-yn-1-yl)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 35)

[0486]

[0487] Step 1: Synthesis of 6-bromo-2-ethoxy-4-(4-(prop-1-yn-1-yl)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 35-1)

[0488] Intermediate 34-2 (100 mg) and 1-(trimethylsilyl)propyne (25 mg) were dissolved in anhydrous tetrahydrofuran solution (5 mL). Cuprous iodide (12 mg), triethylamine (21 mg), tetrabutylammonium fluoride (58 mg), and tetra-triphenylphosphine palladium (12.10 mg) were added. The reaction mixture was stirred at 25 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (50 mL * 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (dichloromethane:methanol = 20:1) to give the title compound (60 mg).

[0489] MS m / z(ESI): 389.0 [M+H] +

[0490] Step 2: Synthesis of 2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-4-(4-(prop-1-yn-1-yl)phenyl)thiazo[4,5-b]pyridin-5(4H)-one (compound 35)

[0491] Intermediate 35-1 (60 mg) was dissolved in 1,4-dioxane (5 mL) and water (0.5 mL), and intermediate 3-1 (73 mg), Pd(dppf)Cl2 (11 mg), and potassium phosphate (99 mg) were added. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, water (10 mL) and ethyl acetate (10 mL) were added sequentially. The organic phase was washed with water (5 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100 * 25 mm * 5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (40 mg).

[0492] MS m / z(ESI): 418.0 [M+H] +

[0493] 1H NMR (400MHz, DMSO-d6) δ8.31(d,J=2.6Hz,1H),8.26(s,1H),7.79(dd,J=9.5,2.7Hz,1H),7.58 -7.51(m,2H),7.41 -7.32(m,2H),6.45(d,J=9.6Hz,1H),4.32(q,J=7.0Hz,2H),3.47(s,3H),2.10(s,3H),1.31(t,J=7.0Hz,3H).

[0494] Example 36, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(1,2,3,4-tetrahydrobenzo[4,5]imidazo[1,2-a]pyridin-8-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 36)

[0495]

[0496] Intermediate 6-1 (80 mg) and intermediate 36-1 (65 mg) were dissolved in 1,4-dioxane (1.2 mL) and water (0.3 mL). Pd(dtbpf)Cl2 (21 mg) and Cs2CO3 (143 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 12 h under nitrogen protection. After the reaction was completed, the reaction solution was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (dichloromethane:methanol = 20:1) to obtain the crude product, which was then purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; water (containing 0.225% formic acid) and a mixture of acetonitrile with decreasing polarity as the eluent; acetonitrile gradient ratio 40%-70%, elution time 15 min] to obtain the title compound (25 mg).

[0497] MS m / z (ESI): 509.1 [M+H] + .

[0498] 1 H NMR(400MHz,DMSO-d6)δ8.21(s,1H),7.77(s,1H),7.51–7.08(m,7H),4.29–4.20(m,2H),4.00(t, J=5.8Hz,2H),2.89(t,J=6.1Hz,2H),2.02–1.94(m,2H),1.89–1.85(m,2H),1.25(t,J=7.0Hz,3H).

[0499] Example 37, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(3-methyl-4-oxo-3,4-dihydroquinazolin-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 37)

[0500]

[0501] Step 1: Synthesis of 6-bromo-3-methylquinazolin-4(3H)-one (intermediate 37-2)

[0502] Intermediate 37-1 (600 mg) was dissolved in acetonitrile (6 mL), and potassium carbonate (1.1 g) and iodomethane (3.8 g) were added. The reaction mixture was stirred at 30 °C for 10 h. After the reaction was complete, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 3:1) to give the title compound (305 mg).

[0503] MS m / z(ESI): 238.9 [M+H] +

[0504] Step 2: Synthesis of 3-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-yl)quinazolin-4(3H)-one (intermediate 37-3)

[0505] Intermediate 37-2 (100 mg) and bis-pinacolborate (170 mg) were dissolved in anhydrous dioxane (2 mL), and potassium acetate (82 mg) and Pd(dppf)Cl2 (46 mg) were added. The reaction mixture was stirred at 100 °C for 4 h under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, and water (8 mL) and ethyl acetate (10 mL * 2) were added sequentially. The organic phase was washed with water (10 mL * 2), dried with an appropriate amount of anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the residue was purified by column chromatography (silica, petroleum ether: ethyl acetate = 4:1) to give the title compound (105 mg).

[0506] MS m / z(ESI): 287.1 [M+H] +

[0507] Step 3: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(3-methyl-4-oxo-3,4-dihydroquinazolin-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 37)

[0508] Intermediate 37-3 (98 mg) and intermediate 1-8 (110 mg) were dissolved in dioxane (1.2 mL) and water (0.3 mL). Cesium carbonate (172 mg) and Pd(dtbpf)Cl2 (26 mg) were added to the solution. The reaction mixture was stirred at 80 °C for 10 h under nitrogen protection. After the reaction was completed, the reaction solution was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure and purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent of a mixture of water (containing 0.225% formic acid) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 40%-70%, elution time 15 min] to obtain the title compound (21 mg).

[0509] MS m / z(ESI): 497.0 [M+H] + .

[0510] 1 H NMR (400MHz, DMSO-d6) δ8.55(s,1H),8.44(s,1H),8.37(s,1H),8.15(d,J=8.6Hz,1H),7.68 (d,J=8.6Hz,1H),7.58–7.15(m,5H),4.36–4.30(m,2H),3.51(s,3H),1.32(t,J=7.0Hz,3H).

[0511] Example 38, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(4-oxo-4H-pyrido[1,2-a]pyrimidin-7-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 38)

[0512]

[0513] Intermediate 15-1 (79 mg) and intermediate 38-1 (70 mg) were dissolved in dioxane (1.2 mL) and water (0.3 mL). Cesium carbonate (136 mg) and Pd(dtbpf)Cl2 (25 mg) were added to the solution. The reaction mixture was stirred at 90 °C for 7 h under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure. The residue was purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent of a mixture of water (containing 0.225% formic acid) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 40%-70%, elution time 18 min] to obtain the title compound (28 mg).

[0514] MS m / z(ESI): 483.0 [M+H] + .

[0515] 1 H NMR (400MHz, DMSO-d6) δ9.66(d,J=1.8Hz,1H),8.65(s,1H),8.40–8.26(m,2H),7.75(d,J=9. 3Hz,1H),7.60–7.11(m,5H),6.40(d,J=6.3Hz,1H),4.38–4.32(m,2H),1.32(t,J=7.0Hz,3H).

[0516] Example 39, 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(4-methyl-3-oxy-3,4-dihydroquinoxalin-6-yl)thiazo[4,5-b]pyridine-5(4H)-one (Compound 39)

[0517]

[0518] Step 1: Synthesis of 7-bromo-1-methylquinoxalin-2(1H)-one (intermediate 39-2)

[0519] Intermediate 39-1 (400 mg) was dissolved in acetonitrile (4 mL), and potassium carbonate (736 mg) and iodomethane (2.5 g) were added. The reaction mixture was stirred at 30 °C for 10 h. After the reaction was completed, the reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 2:1) to give the title compound (160 mg).

[0520] MS m / z(ESI): 238.9 [M+H] +

[0521] Step 2: Synthesis of 1-methyl-7-(4,4,5,5-tetramethyl-1,3,2-dioxaborpentan-2-yl)quinoxalin-2(1H)-one (intermediate 39-3)

[0522] Intermediate 39-2 (100 mg) and bis-pinacolborate (170 mg) were dissolved in anhydrous dioxane (2 mL), and potassium acetate (82 mg) and Pd(dppf)Cl2 (46 mg) were added. The reaction mixture was stirred at 100 °C for 4 h under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, and water (8 mL) and ethyl acetate (10 mL * 2) were added sequentially. The organic phase was washed with water (10 mL * 2), dried with an appropriate amount of anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the residue was purified by column chromatography (silica, petroleum ether: ethyl acetate = 4:1) to give the title compound (98 mg).

[0523] MS m / z(ESI): 287.1 [M+H] +

[0524] Step 3: Synthesis of 4-(4-(difluoromethoxy)phenyl)-2-ethoxy-6-(4-methyl-3-oxo-3,4-dihydroquinoxalin-6-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 39)

[0525] Intermediate 39-3 (98 mg) and intermediate 1-8 (110 mg) were dissolved in dioxane (1.2 mL) and water (0.3 mL). Cesium carbonate (172 mg) and Pd(dtbpf)Cl2 (26 mg) were added to the solution. The reaction mixture was stirred at 80 °C for 10 h under nitrogen protection. After the reaction was completed, the reaction solution was cooled to room temperature and filtered. The filtrate was concentrated to dryness under reduced pressure and purified by preparative high performance liquid chromatography [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent of a mixture of water (containing 0.225% formic acid) and acetonitrile with decreasing polarity; acetonitrile gradient ratio 40%-70%, elution time 15 min] to obtain the title compound (23 mg).

[0526] MS m / z(ESI): 497.0 [M+H] + .

[0527] 1 H NMR(400MHz, DMSO-d6)δ8.53(s,1H),8.21(s,1H),7.98(d,J=1.4Hz,1H),7.84–7.82(m,1H), 7.77–7.75(m,1H),7.57–7.18(m,5H),4.37–4.32(m,2H),3.62(s,3H),1.32(t,J=7.0Hz,3H).

[0528] Example 40: Synthesis of 4-(4-(difluoromethoxy)-3-fluorophenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 40)

[0529]

[0530] Step 1: Synthesis of 6-bromo-2-ethoxy-5-oxothiazolo[4,5-b]pyridine-4(5H)-carboxylic acid tert-butyl ester (intermediate 40-1)

[0531] Intermediate 1-7 (2 g) was dissolved in anhydrous tetrahydrofuran (20 mL), followed by the addition of tert-butyl carbonate (2.38 g), triethylamine (2.21 g), and 4-dimethylaminopyridine (266.43 mg). The mixture was stirred at 25 °C for 12 hours after the addition was complete. After the reaction was complete, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was then purified using flash filtration. 20g The title compound (1.5 g) was obtained by collecting and concentrating the fractions using a Silica Flash column with a gradient of 0-5% methanol / dichloromethane at 80 mL / min.

[0532] MS m / z(ESI): 375.0 [M+H] +

[0533] Step 2: Synthesis of tert-butyl 2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-5-oxothiazolo[4,5-b]pyridine-4(5H)-carboxylic acid (intermediate 40-2)

[0534] Intermediate 40-1 (1.5 g) was dissolved in anhydrous 1,4-dioxane solution (20 mL) and water (5 mL). Intermediate 3-1 (1.13 g), Pd(dppf)Cl2 (260 mg), and potassium phosphate (2.54 g, 11.99 mmol) were added. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was completed, the reaction mixture was cooled to room temperature, and water (100 mL) and ethyl acetate (100 mL) were added sequentially. The organic phase was washed with water (50 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (petroleum ether: ethyl acetate = 1:1) to give the title compound (1 g).

[0535] MS m / z(ESI): 404.0 [M+H] +

[0536] Step 3: Synthesis of 2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (intermediate 40-3)

[0537] Intermediate 40-2 (1 g) was dissolved in anhydrous dichloromethane (10 mL), and hydrochloric acid / dioxane (6 mL, 4 N) was added dropwise. After the addition was complete, the mixture was stirred at 25 °C for 4 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure to remove the solvent, and saturated sodium bicarbonate (50 mL) and ethyl acetate (50 mL * 3) were added sequentially. The organic phase was washed with water (20 mL * 2), and the washed organic phase was dried with an appropriate amount of anhydrous sodium sulfate, filtered, and concentrated to obtain the title compound (450 mg).

[0538] MS m / z(ESI): 304.0 [M+H] +

[0539] Step 4: Synthesis of (4-(difluoromethoxy)-3-fluorophenyl)boronic acid (intermediate 40-4)

[0540] 300 mg of 4-bromo-1-(difluoromethoxy)-2-fluorobenzene and 375 mg of triisopropyl borate were dissolved in 3 mL of anhydrous tetrahydrofuran solution. A tetrahydrofuran solution of n-BuLi (1.6 M, 0.895 mL) was added at 0 °C, and the reaction mixture was stirred at 0 °C for 4 h under nitrogen protection. Subsequently, the reaction mixture was concentrated to dryness under reduced pressure, and the residue was purified by column chromatography (PE / EA = 10 / 1-3 / 1) to give the title compound (120 mg).

[0541] Step 5: Synthesis of 4-(4-(difluoromethoxy)-3-fluorophenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 40)

[0542] Intermediate 40-4 (36 mg) and intermediate 40-3 (35 mg) were dissolved in 1,2-dichloroethane (1 mL), and anhydrous copper acetate (25 mg) and pyridine (27 mg) were added. The reaction mixture was stirred at 50 °C for 12 h under nitrogen protection. Subsequently, after the reaction was cooled to room temperature, the reaction mixture was filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by high-pressure preparation [YMC-Actus Triart C18 column, 5 μm silica, 30 mm diameter, 150 mm length; eluent: a mixture of water (containing 0.05% NH4HCO3) and acetonitrile in decreasing polarity; acetonitrile gradient ratio 55%-75%, elution time 12 min] to obtain the title compound (12 mg).

[0543] MS m / z(ESI): 464.0 [M+H]+ .

[0544] 1 H NMR(400MHz,DMSO-d6)δ8.30(d,J=2.7Hz,1H),8.27(s,1H),7.80–7.76(m,1H),7.65–7.61(m,1H) ,7.56–7.20(m,3H),6.44(d,J=9.5Hz,1H),4.36–4.31(m,2H),3.46(s,3H),1.32(t,J=7.0Hz,3H).

[0545] Example 41, 4-(6-(difluoromethoxy)pyridin-3-yl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 41)

[0546]

[0547] Step 1: Synthesis of 2-(difluoromethoxy)-5-iodopyridine (intermediate 41-2)

[0548] Intermediate 41-1 (1.11 g) and sodium difluorochloroacetate (1.54 g) were dissolved in anhydrous N,N-dimethylformamide (20 mL), and cesium carbonate (4.92 g) was added. After the addition was complete, the mixture was stirred at 100 °C for 12 hours. After the reaction was complete, saturated ammonium chloride (100 mL) and ethyl acetate (100 mL x 3) were added sequentially. The organic phase was washed with water (50 mL x 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography (dichloromethane:methanol = 20:1) to give the title compound (600 mg).

[0549] MS m / z(ESI): 272.0 [M+H] +

[0550] Step 2: Synthesis of (6-(difluoromethoxy)pyridin-3-yl)boronic acid (intermediate 41-3)

[0551] Intermediate 41-2 (550 mg) and trimethyl borate (314 mg) were dissolved in anhydrous tetrahydrofuran (10 mL). Butyllithium (2.5 mL, 1.6 M) was added dropwise at -78 °C. After the addition was complete, the mixture was stirred at -78 °C for 4 hours. After the reaction was complete, hydrochloric acid (3 mL, 1 N) was added sequentially, and the tetrahydrofuran was removed by concentration under reduced pressure. The residue was purified by high-performance liquid chromatography (HPLC) (column: Waters Xbridge BEH C18 100*25 mm*5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (80.0 mg).

[0552] MS m / z(ESI): 189.0 [M+H] +

[0553] Step 3: Synthesis of 6-bromo-4-(6-(difluoromethoxy)pyridin-3-yl)-2-ethoxythiazo[4,5-b]pyridin-5(4H)-one (intermediate 41-4)

[0554] Intermediate 41-3 (53 mg) and intermediate 1-7 (51 mg) were dissolved in anhydrous 1,2-dichloroethane (10 mL), and pyridine (66 mg) and copper acetate (51 mg) were added. After the addition was complete, the mixture was stirred in air at 40 °C for 4 hours. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18 100*25mm*5μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (40 mg).

[0555] MS m / z (ESI): 418.1 [M+H] +

[0556] Step 4: Synthesis of 4-(6-(difluoromethoxy)pyridin-3-yl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (compound 41)

[0557] Intermediate 41-4 (40 mg) was dissolved in 2 mL of 1,4-dioxane solution and 0.5 mL of water. Intermediate 3-1 (45 mg), Pd(dppf)Cl2 (7 mg), and cesium carbonate (93 mg) were added to the solution. The reaction mixture was stirred at 100 °C for 12 hours under nitrogen protection. After the reaction was cooled to room temperature, 10 mL of water and 10 mL of ethyl acetate were added sequentially. The organic phase was washed with water (50 mL x 2) and dried over an appropriate amount of anhydrous sodium sulfate. After filtration, the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by high performance liquid chromatography (HPLC) (column: Waters Xbridge BEH C18 100 x 25 mm x 5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (10 mg).

[0558] MS m / z(ESI): 447.0 [M+H] +

[0559] 1 H NMR (400MHz, DMSO-d6) δ8.37(d,J=2.6Hz,1H),8.29(d,J=3.7Hz,2H),8.06(dd,J=8.7,2.6Hz,1H),7.99–7.63 (m,2H),7.30(d,J=8.7Hz,1H),6.46(d,J=9.5Hz,1H),4.37–4.32(m,2H),3.48(s,3H),1.33(t,J=7.0Hz,3H).

[0560] Example 42, 4-(4-(difluoromethoxy)-3-methylphenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5-(4H)-one (Compound 42)

[0561]

[0562] Step 1: Synthesis of (4-(difluoromethoxy)-3-methylphenyl)boronic anhydride (intermediate 42-2)

[0563] Intermediate 42-1 (500 mg) and triisopropyl borate (800 mg) were dissolved in tetrahydrofuran (5 mL), and n-butyllithium (4.26 mL) was added dropwise at -78 °C. The mixture was stirred at 0 °C for 2 h. Subsequently, H2O (20 mL) and ethyl acetate (20 mL * 3) were added to the reaction solution in sequence. The organic phase was washed with saturated brine (30 mL * 2), and the washed organic phase was dried with an appropriate amount of anhydrous sodium sulfate and concentrated to obtain the title compound (350 mg).

[0564] MS m / z(ESI): 247.2 [M+H] + .

[0565] Step 2: Synthesis of 4-(4-(difluoromethoxy)-3-methylphenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5-(4H)-one (compound 42)

[0566] Intermediate 42-2 (48 mg) and intermediate 40-2 (25 mg) were dissolved in dichloromethane (2 mL), and pyridine (10 mg) and copper acetate (4 mg) were added. The reaction mixture was stirred at 45 °C for 16 hours under nitrogen protection. After the reaction was cooled to room temperature, water (30 mL) and dichloromethane (40 mL) were added sequentially. The organic phase was washed with saturated brine (30 mL * 2), and the washed organic phase was dried over an appropriate amount of anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Gemini NX C18 5 μm * 10 * 150 mm; mobile phase: A: 0.05% TFA v / v, B: acetonitrile; B%: 40%-45%, 12 min) to give the title compound (26 mg).

[0567] MS m / z (ESI): 460.1 [M+H] + .

[0568] 1 H NMR(400MHz, DMSO-d6)δ8.31(d,J=2.6Hz,1H),8.26(s,1H),7.84–7.75(m,1H),7.54–7.08(m ,4H),6.45(d,J=9.5Hz,1H),4.38–4.28(m,2H),3.48(s,3H),2.29(s,3H),1.37–1.28(m,3H).

[0569] Example 43: 2-(difluoromethoxy)-5-(2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-5-oxothiazo[4,5-b]pyridin-4(5H)-yl)benzonitrile (compound 43)

[0570]

[0571] Step 1: Synthesis of 5-bromo-2-(difluoromethoxy)benzonitrile (intermediate 43-2)

[0572] Intermediate 43-1 (1 g) and sodium difluorochloroacetate (1.54 g) were dissolved in anhydrous N,N-dimethylformamide (20 mL), and cesium carbonate (4.92 g) was added. The mixture was stirred at 100 °C for 12 hours after the addition was complete. After the reaction was complete, saturated ammonium chloride (100 mL) and ethyl acetate (100 mL x 3) were added sequentially. The organic phase was washed with water (50 mL x 2), dried over an appropriate amount of anhydrous sodium sulfate, filtered, and the organic phase was concentrated under reduced pressure to remove the solvent. The residue was purified by column chromatography. 20g The title compound (500 mg) was obtained by using a Silica Flash column with a gradient of 0-5% methanol / dichloromethane at 80 mL / min.

[0573] MS m / z(ESI): 248.0 [M+H] +

[0574] Step 2: Synthesis of (3-cyano-4-(difluoromethoxy)phenyl)boronic acid (intermediate 43-3)

[0575] Intermediate 43-2 (500 mg) and trimethyl borate (314 mg) were dissolved in anhydrous tetrahydrofuran (10 mL). Butyllithium (2.5 mL, 1.6 M) was added dropwise at -78 °C, and the mixture was stirred at -78 °C for 4 hours. After the reaction was complete, hydrochloric acid (3 mL, 1 N) was added sequentially. The tetrahydrofuran was removed by concentration under reduced pressure. The residue was purified by high performance liquid chromatography (HPLC) (column: Waters Xbridge BEH C18 100*25 mm*5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (80.0 mg).

[0576] Step 3: Synthesis of 2-(difluoromethoxy)-5-(2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-5-oxothiazo[4,5-b]pyridin-4(5H)-yl)benzonitrile (compound 43)

[0577] Intermediate 43-3 (60 mg) and intermediate 40-2 (51.28 mg) were dissolved in anhydrous 1,2-dichloroethane (10 mL), and pyridine (66.86 mg) and copper acetate (51 mg) were added. After the addition was complete, the mixture was stirred in air at 40 °C for 4 hours. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The residue was purified by high performance liquid chromatography (column: Waters Xbridge BEH C18 100*25mm*5μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (10.4 mg).

[0578] MS m / z(ESI): 471.0 [M+H] +

[0579] 1 H NMR (400MHz, DMSO-d6) δ8.34–8.27(m,2H),8.17(d,J=2.5Hz,1H),7.93–7.90(m,1H),7.82– 7.40(m,2H),6.47(d,J=9.5Hz,1H),4.38–4.33(m,2H),3.49(s,3H),1.34(t,J=7.0Hz,3H).

[0580] Example 44: 3-(2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)-5-oxothiazo[4,5-b]pyridin-4(5H)-yl)benzonitrile (compound 44)

[0581]

[0582] Intermediate 40-2 (20 mg) and 3-cyanobenzonic acid (14.53 mg) were dissolved in anhydrous 1,2-dichloroethane (5 mL), and pyridine (15.65 mg) and copper acetate (11.93 mg) were added. After the addition was complete, the mixture was stirred at 40 °C for 4 hours. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The residue was purified by preparative high performance liquid chromatography (column: Waters Xbridge BEH C18100*25 mm*5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (2.8 mg).

[0583] MS m / z(ESI): 405.0 [M+H] +

[0584] 1 H NMR(400MHz,Chloroform-d)δ8.34(d,J=2.7Hz,1H),7.84–7.74(m,2H),7.71–7.58( m,4H),6.64(d,J=9.6Hz,1H),4.38–4.33(m,2H),3.58(s,3H),1.39(t,J=7.1Hz,3H).

[0585] Example 45, 4-(3-(difluoromethoxy)phenyl)-2-ethoxy-6-(1-methyl-6-oxo-1,6-dihydropyridin-3-yl)thiazo[4,5-b]pyridin-5(4H)-one (Compound 45)

[0586]

[0587] Intermediate 40-3 (15 mg) and intermediate 45-1 (13.94 mg) were dissolved in anhydrous 1,2-dichloroethane (5 mL), and pyridine (15.65 mg) and copper acetate (11.93 mg) were added. After the addition was complete, the mixture was stirred at 40 °C for 4 hours. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The residue was purified by high performance liquid chromatography (column: Waters Xbridge BEH C18100*25 mm*5 μm; mobile phase: [A: water (0.225% formic acid), B: acetonitrile]; B%: 23%-50%, 12 min) to give the title compound (5.4 mg).

[0588] MS m / z(ESI): 446.0 [M+H] +

[0589] 1 H NMR(400MHz,DMSO-d6)δ8.30(d,J=2.6Hz,1H),8.26(s,1H),7.80–7.77(m,1H),7.68–7.53(m,1H) ,7.50–7.10(m,4H),6.44(d,J=9.5Hz,1H),4.34–4.29(m,2H),3.46(s,3H),1.30(t,J=7.0Hz,3H).

[0590] Biological activity and related property test examples

[0591] Test Example 1: Biochemical Test

[0592] Brief introduction to the experimental principle: L-methionine and ATP can be converted into SAM, inorganic phosphate, and inorganic diphosphate under the catalysis of MAT2A enzyme. By adding a colorimetric reagent, such as ammonium molybdate, to the enzyme reaction mixture, the content of inorganic phosphate in the sample can be quantitatively detected, thereby reflecting the enzyme activity of MAT2A.

[0593] Materials: MAT2A screening kit was purchased from BPS Bioscience (USA); 384-well plates were purchased from Corning (USA).

[0594] 1. MAT2a protein (Pharmaron (Beijing) New Drug Technology Co., Ltd.);

[0595] 2. L-Methionine (Sigma#M9625-5G)

[0596] 3. ATP (Sigma#A7699-1G)

[0597] 4. KCl (Sigma#60142-500ML-F)

[0598] 5. Tris (Sigma#T2663-1L)

[0599] 6. MgCl2 (Sigma#M1028)

[0600] 7. EDTA (Invitrogen #AM9260G)

[0601] 8.BSA(Sangon Biotech#A500023-0100)

[0602] 9. PiColorLock(abcam#ab270004)

[0603] Detection method:

[0604] The compound was dissolved in DMSO, diluted to a final concentration of 10 μM using Echo, diluted 3-fold, and transferred 80 nL to a 384-well plate.

[0605] Prepare the experimental buffer (50 mM Tris, 50 mM KCl, 15 mM MgCl2, 100 μM EDTA, 0.005% BSA). Dilute the MAT2a protein with the experimental buffer (final concentration 4 μg / mL). Add 40 μL of 2X MAT2a solution to a 384-well plate, centrifuge at 1000 rpm for 1 minute, and incubate at room temperature for 120 minutes.

[0606] Dilute L-methionine and ATP with experimental buffer (final L-methionine concentration: 200 μM, final ATP concentration: 400 μM). Add 40 μL of 2X L-methionine and ATP solution to start the reaction, centrifuge at 1000 rpm for 1 minute, and incubate at room temperature for 90 minutes.

[0607] Install PiColorLock according to the instructions. TM Reaction catalyst and PiColorLock TM Mix buffer solution 1:100, add 20 μL to each well and shake for 30 seconds. Add 8 μL of stabilizing reagent and shake for 30 seconds. Incubate at room temperature for 30 minutes and then detect the signal value.

[0608] Data Analysis:

[0609] Calculate %Compound inhibition and fit the IC50 of the compound. 50 .

[0610] Compound inhibition=(100-100*(Signal-Bottom) / (Top-Bottom))%

[0611] Experimental results:

[0612] Under the experimental conditions, the inhibitory effect of the test compound on MAT2A can be expressed by the IC50 value, which measures the level of inhibition of phosphate production during the enzymatic reaction. 50 Values ​​are used to represent the MAT2A inhibitory activity of the compounds of this invention. See Table 1 for details.

[0613] Table 1

[0614]

[0615]

[0616] Test Example 2: Detection of Intracellular SAM Levels

[0617] Brief Introduction to the Experimental Principle: SAM, as a catalytic product of MAT2A, can be used to reflect the inhibitory effect of the test compound on MAT2A by measuring the level of SAM production in cells. After co-incubating the test MAT2A inhibitor with cancer cells for a period of time, the cells were lysed with a termination reagent to quench the MAT2A enzyme activity. The MAT2A catalytic product SAM in the cell lysate was quantitatively measured by LC-MS / MS, thereby reflecting the intracellular MAT2A activity.

[0618] Materials and Cells: HCT116 MTAP - / - Cells were purchased from Kangyuan Bochuang; fetal bovine serum, McCoy's 5a medium and penicillin-streptomycin were purchased from Gibco (USA); 96-well plates were purchased from Corning (USA); and PBS was purchased from Cytiva (USA).

[0619] Cell culture: HCT116 MTAP - / - Cells were cultured in McCoy's 5a medium containing 10% fetal bovine serum and 1% penicillin-streptomycin at 37°C and 5% CO2. Only cells in the logarithmic growth phase were used for experiments.

[0620] LC-MS / MS detection: LC-MS / MS was used to detect the effect of the compound on HCT116 MTAP. - / -Effect of SAM production levels in cell lines. Cells were seeded at a concentration of 50,000 per well in 96-well plates and incubated overnight at 37°C and 5% CO2. The compound was dissolved in DMSO, then diluted sequentially with DMSO and culture medium and transferred to the cell culture plates to a final concentration of 10 μM (3-fold dilution). Cells were incubated at 37°C and 5% CO2 for another 6 hours. The supernatant was removed, and cells were washed once with PBS and then lysed with glacial acetic acid. The lysate was processed and analyzed by LC-MS / MS to determine the SAM concentration.

[0621] Data Analysis:

[0622] Calculate %Compound inhibition and fit the IC50 of the compound. 50 .

[0623] %Compound inhibition=1-100%*(Signal-Bottom) / (Top-Bottom)

[0624] Experimental results:

[0625] Table 2 shows that the compound of the present invention inhibits intracellular SAM production at the nanomolar level, and the compound of the present invention exhibits strong MAT2A inhibitory activity.

[0626] Table 2

[0627]

[0628] Test Example 3: Human colon cancer HCT116 cell proliferation inhibition assay

[0629] Brief introduction to the experimental principle: After co-incubating the MAT2A inhibitor to be tested with cancer cells for a period of time, the effect of the test compound on cell proliferation is measured by a cell proliferation counting method based on ATP content.

[0630] Materials and Cells: HCT116 WT Cells and HCT116 MTAP - / - Cells were purchased from Kangyuan Bochuang; fetal bovine serum, McCoy's 5a medium and penicillin-streptomycin were purchased from Gibco (USA); 96-well plates were purchased from Corning (USA); and Cell-Titer Glo reagent was purchased from Prometheus (USA).

[0631] Cell culture: HCT116 WT cells and HCT116 MTAP cells - / - Cells were cultured in McCoy's 5a medium containing 10% fetal bovine serum and 1% penicillin-streptomycin at 37°C and 5% CO2. Only cells in the logarithmic growth phase were used for experiments.

[0632] Cell proliferation activity assay: Cell-Titer Glo reagent was used to detect the effects of compounds on HCT116 WT and HCT116MTAP. - / - Inhibitory activity of the two cell lines on proliferation. The cell concentration was adjusted to 400 cells per well, seeded in 96-well plates, and incubated overnight at 37°C and 5% CO2.

[0633] The compound was dissolved in DMSO, then diluted sequentially with DMSO and culture medium, and transferred to cell plates to a final concentration of 10 μM, followed by a 3-fold dilution. Cells were incubated at 37°C and 5% CO2 for 6 days. Cell-Titer Glo reagent was added to assess cell viability.

[0634] Data Analysis:

[0635] Calculate %Compound inhibition and fit the IC50 of the compound. 50 .

[0636] %Compound inhibition=1-100%*(Signal-Bottom) / (Top-Bottom)

[0637] HCT116 MTAP - / - The cells were obtained by site-specific knockout of wild-type HCT116 (HCT116 WT) cells using gene knockout techniques, resulting in cells that do not express MTAP protein. Since MAT2A inhibition and MTAP deficiency can produce synthetic lethality, leading to tumor cell death, this study tested the effect of MAT2A inhibitors on HCT116 MTAP. - / - The antiproliferative activity of cells and HCT116 WT can be used to evaluate the activity and selectivity of MAT2A inhibitors.

[0638] Experimental results:

[0639] Under the experimental conditions, the test compound showed a response to HCT116 MTAP. - / - The cells exhibited strong inhibitory activity against proliferation, and compared to HCT116 MTAP... + / + The cells exhibited good selectivity. The specific anti-cell proliferation activities of the test compounds are shown in Table 3.

[0640] Table 3

[0641]

[0642]

[0643] Test Example 4: Determination of the metabolic stability of the compound of the present invention in liver microsomes

[0644] The metabolic stability of the compounds of this invention in liver microsomes was determined using the following experimental method.

[0645] I. Experimental Materials and Instruments

[0646] 1. Human liver microsomes (Corning 452117), beagle liver microsomes (XENOTECH D1000), SD rat liver microsomes (XENOTECH R1000) and CD-1 mouse liver microsomes (XENOTECH M1000)

[0647] 2. Na2HPO4 (Tianjin Guangfu Fine Chemical Research Institute 20180130)

[0648] 3. KH2PO4 (Tianjin Guangfu Fine Chemical Research Institute, 20180920)

[0649] 4. MgCl2 (Tianjin Guangfu Fine Chemical Research Institute, 20191216)

[0650] 5. NADPH (Solarbio 1216C022)

[0651] 6. Positive control compound verapamil (Sigma MKBV4993V)

[0652] 7. AB Sciex API4000 Liquid Chromatography-Mass Spectrometry System

[0653] II. Experimental Procedure

[0654] 1. Preparation of 100mM Phosphate Buffered Solution (PBS): Weigh 7.098g Na₂HPO₄ and dissolve it in 500mL of pure water by sonication to obtain solution A. Weigh 3.400g KH₂PO₄ and dissolve it in 250mL of pure water by sonication to obtain solution B. Slowly add solution B to solution A on a stirrer until the pH reaches 7.4 to prepare 100mM PBS buffer.

[0655] 2. Preparation of the reaction system

[0656] Prepare the reaction system according to the table below:

[0657]

[0658] 3. Incubate the reaction system in a 37°C water bath for 10 minutes. Add 40 μL of 10 mM NADPH solution (NADPH dissolved in 100 mM phosphate buffer) to the reaction system, bringing the final NADPH concentration to 1 mM. Use 40 μL of phosphate buffer instead of NADPH solution as a negative control. The negative control is used to eliminate the influence of the compound's own chemical stability.

[0659] 4. Add 4 μL of the 100 μM compound of this invention and the positive control compound verapamil to the reaction system to start the reaction. The final concentration of the compound is 1 μM.

[0660] 5. After thorough mixing on a vortex mixer, 50 μL of the incubated sample was taken out at 0.5, 15, 30, 45, and 60 minutes, respectively, and the reaction was terminated with 200 μL of ice-cold acetonitrile containing the internal standard. The sample was centrifuged at 3220 g for 45 minutes. After centrifugation, 90 μL of the supernatant was transferred to a sample plate, and 90 μL of ultrapure water was added and mixed well for LC-MS / MS analysis.

[0661] All data were calculated using Microsoft Excel software. Peak areas were detected by extracting ion spectra, and the in vitro half-life (t0.05) of the compound was determined by linearly fitting the natural logarithm of the elimination percentage of the compound against time. 1 / 2 ).

[0662] In vitro half-life (t) 1 / 2 ) Calculated by slope:

[0663] in vitro t 1 / 2 =0.693 / k

[0664] In vitro intrinsic clearance rate (unit: μL / min / mg protein) is calculated using the following formula:

[0665] in vitro CL int = k × volume of incubation (μL) / amount of proteins (mg)

[0666] Table 4

[0667]

[0668] The experimental results in Table 4 show that all compounds of the present invention exhibit good metabolic stability in liver microsomes.

[0669] Test Example 5: Determination of membrane permeability and transport properties of the compounds of the present invention

[0670] The membrane permeability and transport properties of the compounds of this invention were determined using the following experimental methods.

[0671] I. Experimental Materials and Instruments

[0672] 1. Caco-2 cells (ATCC)

[0673] 2. HEPES (Solarbio 804D049), penicillin / streptomycin (Solarbio 20200109), PBS (Solarbio20200620), and kanamycin (MP Biomedicals 194531)

[0674] 3. Fetal bovine serum (FBS) (Sigma WXBD0055V), fluorescein (Sigma MKCJ3738), and NaHCO3 (Sigma SLBZ4647)

[0675] 4. Hank's balanced salt solution (HBSS) (Gibco 2085528), non-essential amino acids (NEAA) (Gibco 2211548), and Trypsin / EDTA (Gibco 2120732).

[0676] 5. High-sugar DMEM (Corning 20319014)

[0677] 6.HTS Transwell-96Well Permeable(Corning,3391)

[0678] 7. Resistance meter (Millipore, ERS-2)

[0679] 8. Vision (Nexcelom Bioscience)

[0680] 9. Infinite 200PRO microplate reader (Tecan, Infinite M200PRO)

[0681] 10. Positive control compounds: Metoprolol (Sinopharm 100084-201403), Erythromycin (MCE84550), and Cimetidine (Sinopharm 100158-201406).

[0682] 11. ABIQTrap 5500 Liquid Chromatography-Mass Spectrometry (LC-MS) System

[0683] II. Experimental Procedure

[0684] 1. Caco-2 cell culture

[0685] 1) Preparation of transport buffer (HBSS containing 25mM HEPES, pH 7.4): Accurately weigh 5.958g HEPES and 0.35g NaHCO3, add 900mL of pure water to dissolve them, then add 100mL of 10×HBSS and stir well. Adjust the pH to 7.4 and filter.

[0686] 2) Preparation of Caco-2 cell culture medium: FBS, penicillin / streptomycin, kanamycin and NEAA were added to high glucose DMEM (containing L-glutamine) medium to prepare a cell culture medium containing 10% FBS, 100 units of penicillin / 0.1 mg / mL streptomycin, 0.6 μg / mL kanamycin and 1×NEAA.

[0687] 3) Culture the cells in a T-75 culture flask at 37℃ and 5% CO2. When the cells reach 80-90% confluence, discard the culture medium. Wash the cells with 5mL PBS, add 1.5mL Trypsin / EDTA, and then incubate at 37℃ for 5-10 minutes until the cells detach in a quicksand-like manner. Finally, neutralize the Trypsin / EDTA with FBS-containing medium.

[0688] 4) Centrifuge the cell suspension at 120g for 10 minutes and discard the supernatant.

[0689] 5) Resuspend the cells in cell culture medium and adjust the density to 6.86 × 10⁻⁶. 5 Cell suspension with cells / mL.

[0690] 2. Caco-2 cell inoculation

[0691] 1) Add 50 μL of culture medium to each well of the Transwell chamber, add 25 mL of culture medium to the bottom layer, and preheat in a 37°C, 5% CO2 incubator for 1 hour.

[0692] 2) Add 50 μL of cell suspension to each well of the preheated Transwell chamber, resulting in a final seeding density of 2.4 × 10⁻⁶ cells / well. 5 cells / cm 2 .

[0693] 3) Incubate for 14-18 days, changing the culture medium every other day, and changing the culture medium within 48 hours after the initial inoculation. The culture medium must be changed the day before the experiment.

[0694] 3. Assess the integrity of the monolayer cell membrane

[0695] 1) After 14 days of cell culture, the cells fused and differentiated, ready for transport experiments.

[0696] 2) Measure the resistance of the single-layer film using a resistance meter and record the resistance of each hole.

[0697] 3) After the measurement is completed, the Transwell culture plate is incubated again.

[0698] 4) Calculate the TEER value:

[0699] TEER value = TEER measurement (Ω) × membrane area (cm²) 2 )

[0700] The electrical resistance of a single cell membrane is <230 Ω·cm 2 This indicates that the single-layer cell membrane has poor density and cannot be used for experiments.

[0701] 4. Transport experiment

[0702] 1) Dilute 10 mM of the stock solution of the compound of the present invention or the positive control compound with DMSO to obtain a 2 mM stock solution, and then dilute the 2 mM stock solution with transfer buffer to obtain a 10 μM working solution of the compound of the present invention or the positive control compound.

[0703] 2) Remove the Caco-2 cell plate from the incubator, then wash the Transwell culture plate twice with preheated transport buffer, and then incubate it in a 37°C incubator for 30 minutes.

[0704] 3) To determine the transport rate of the compound from the top to the base (A→B), 108 μL of the compound's working solution was added to the Transwell chamber (top). Simultaneously, 8 μL of sample was immediately removed from the top and transferred to 72 μL of transport buffer. 240 μL of stop solution containing an internal standard was added to terminate the transport and serve as the initial top sample. At the same time, 300 μL of transport buffer was added to the receiving end (base). The experiment used a two-sample setup.

[0705] 4) To determine the transport rate of the compound from the base end to the top end (B→A), add 308 μL of the compound's working solution to the base end, and immediately remove 8 μL of the sample from the base end into 72 μL of transport buffer. Add 240 μL of stop solution containing internal standard to terminate the transport and use it as the initial base end sample. Simultaneously, add 100 μL of transport buffer to the Transwell chamber (top end). The experiment uses a two-sample setup.

[0706] 5) Incubate the cell culture plate in a 37°C CO2 incubator for 2 hours.

[0707] 6) After the transport experiment, take 8 μL of sample from the dosing end (i.e., the top end in the A→B direction and the base end in the B→A direction) and add it to 72 μL of transport buffer. Then, add 240 μL of stop solution containing internal standard to terminate the transport. Take 80 μL of sample from the receiving end (i.e., the base end in the A→B direction and the top end in the B→A direction) and add it to 240 μL of stop solution containing internal standard. Vortex at 1000 rpm for 10 minutes and centrifuge at 3220 g for 30 minutes. Take 100 μL of supernatant into the sample plate, add 100 μL of ultrapure water and mix well. Use for LC-MS / MS analysis.

[0708] 7) After the transport experiment, measure the fluorescence value. Prepare a 10mM fluorescein stock solution with water, then dilute it to 100μM with transport buffer. Add 100μL of fluorescein solution to the top of the Transwell chamber and 300μL of transport buffer to the bottom. Incubate at 37℃ in a CO2 incubator for 30 minutes. Transfer 80μL of the solution from the top and bottom to a 96-well plate and measure the cell fluorescence value using a microplate reader at an excitation wavelength of 485nm and an emission wavelength of 530nm (to check membrane integrity).

[0709] The fluorescence value of the Caco-2 cell monolayer membrane is calculated using the following formula:

[0710] LY Leakage={I acceptor ×0.3 / (I acceptor ×0.3+I donor ×0.1)}×100%I acceptor The fluorescence density on the receiving side (0.3 mL), I donor The fluorescence density refers to the fluorescence density on the administration side (0.1 mL). LY > 1.0% indicates poor compactness of the monolayer cell membrane, and the corresponding result will be excluded from the evaluation.

[0711] The peak areas of the compound on the administration and receptive sides were determined, and the apparent permeability coefficient (P0) of the compound was calculated. app (Unit: cm / s) and Efflux ratio:

[0712] P app ={V A ×[drug] acceptor / (Area×incubation time×[drug] initial donor}

[0713] V A The volume of the receiving solution is 0.3 mL for A→B and 0.1 mL for B→A. Area is the membrane area of ​​the Transwell-96-well plate (0.143 cm²). 2); incubation time is the incubation period (unit: s).

[0714]

[0715] P app(B-A) P represents the apparent permeability from the basal end to the apex. app(A-B) The apparent permeability coefficient is measured from the top to the base.

[0716] Table 5

[0717] compound <![CDATA[P app (A-B)(10 -6 cm / s)]]> <![CDATA[P app (B-A)(10 -6 cm / s)]]> Efflux Ratio Compound 5 3.44 4.70 1.37 Compound 8 2.66 4.29 1.36 Compound 9 3.65 3.95 1.08 Compound 10 1.78 6.32 3.54 Compound 11 2.29 3.87 1.70

[0718] Caco-2 is a human clonal colon adenocarcinoma cell line with a structure and function similar to differentiated small intestinal epithelial cells, and can be used to conduct experiments simulating in vivo intestinal transport. A higher Papp(AB) indicates better permeability of the compound, suggesting potentially higher bioavailability after oral absorption. The Efflux Ratio indicates that the compound is affected by transporters, with a greater likelihood of being excreted by transporters, suggesting that the membrane permeability of the compound is more significantly affected by transporters, which is detrimental to oral absorption and the ability to cross target cell membranes to exert therapeutic effects. Table 5 shows that the compounds of this invention have good membrane permeability, indicating a strong ability to be absorbed orally.

[0719] Test Example 6: Inhibitory effect of the compounds of the present invention on the activities of CYP2C9, CYP2D6, and CYP3A4 enzymes.

[0720] The inhibition of CYP2C9, CYP2D6, and CYP3A4 enzyme activities by the compounds of this invention was determined using the following experimental method.

[0721] I. Experimental Materials and Instruments

[0722] 1. Human liver microsomes (Corning 452117)

[0723] 2. Na₂HPO₄ (Sigma SLBZ 6180)

[0724] 3. KH2PO4 (Sigma SLBT6559)

[0725] 4. NADPH (Solarbio 705Y021)

[0726] 5. Positive substrates: diclofenac (Sigma SLBV3438), dextromethorphan (TRC 3-EDO-175-1), and midazolam (Cerilliant FE01161704).

[0727] 6. Positive inhibitors: sulfamethoxazole (D. Ehrenstorfer GmbH 109012), quinidine (TCI WEODL-RE), and ketoconazole (Sigma 100M1091V)

[0728] 7. AB Sciex Triple Quad 5500 Liquid Chromatography-Mass Spectrometry (LC-MS) System

[0729] II. Experimental Procedure

[0730] 1. Preparation of 100mM Phosphate Buffered Solution (PBS): Weigh 7.098g Na₂HPO₄ and dissolve it in 500mL of pure water by sonication to obtain solution A. Weigh 3.400g KH₂PO₄ and dissolve it in 250mL of pure water by sonication to obtain solution B. Slowly add solution B to solution A on a stirrer until the pH reaches 7.4 to prepare 100mM PBS buffer.

[0731] 2. Prepare a 10 mM NADPH solution using 100 mM PBS buffer. Dilute 10 mM of the stock solution of the compound of this invention with DMSO to obtain 200× concentration working solutions of the compound (6000, 2000, 600, 200, 60, 20, 0 μM). Dilute the positive inhibitor stock solution with DMSO to obtain 200× concentration working solutions of the positive inhibitor (sulfamethoxazole, 1000, 300, 100, 30, 10, 3, 0 μM; quinidine / ketoconazole, 100, 30, 10, 3, 1, 0.3, 0 μM). Prepare 200× concentration substrate working solutions (120 μM diclofenac, 400 μM dextromethorphan, and 200 μM midazolam) with water, acetonitrile, or acetonitrile / methanol.

[0732] 3. Take 2 μL of 20 mg / ml liver microsomal solution, 1 μL of substrate working solution, 1 μL of compound working solution, and 176 μL of PBS buffer, mix well, and pre-incubate in a 37°C water bath for 15 minutes. For the positive control group, add 1 μL of sulfadiazine, quinidine, or ketoconazole working solution instead of the compound working solution. Simultaneously, pre-incubate 10 mM NADPH solution in a 37°C water bath for 15 minutes. After 15 minutes, add 20 μL of NADPH to each well to start the reaction, incubating at 37°C for 5 minutes (CYP2C9), 20 minutes (CYP2D6), or 5 minutes (CYP3A4). All incubated samples should be in duplicate. After the appropriate incubation time, add 400 μL of ice-cold methanol containing the internal standard to all samples to terminate the reaction. Vortex to mix, and centrifuge at 3220 g, 4°C for 40 minutes. After centrifugation, transfer 100 μL of supernatant to the sample plate, add 100 μL of ultrapure water and mix well for LC-MS / MS analysis.

[0733] Table 6

[0734]

[0735] Drug-drug interaction (DDI) refers to the physical or chemical changes that occur between two or more drugs, and the resulting alterations in drug efficacy. Understanding drug interactions can provide better pharmaceutical care for patients, promote rational drug use, and minimize the occurrence of adverse reactions. Drug interactions are primarily metabolic interactions, which are mainly related to CYP450 enzymes involved in drug metabolism. The experimental results in Table 6 show that the compounds of this invention have weak inhibitory ability on CYP450, indicating a low potential risk of DDI.

Claims

1. A compound of general formula (A) or a pharmaceutically acceptable salt thereof: in, Ring Q is a phenyl or a 5-6-membered heteroaryl group, wherein the phenyl or 5-6-membered heteroaryl group is optionally replaced by R. a Instead, the R a Selected from F, Cl, Br, I, CN, C2-C3 alkynyl, C1-C6 alkyl, C3-C6 cycloalkyl, or C1-C6 alkoxy, wherein the C1-C6 alkyl, C3-C6 cycloalkyl, or C1-C6 alkoxy is optionally converted by R. b replace; R b Selected from F, Cl, Br, I or CN; Ring W is arbitrarily selected by R c The following groups are substituted: phenyl, pyridyl, pyrimidinyl, pyridazinyl, pyridinoneyl, pyridazinoneyl. The R c Selected from OH, =O, or arbitrarily subjected to R c1 The following groups are substituted: C1-C6 alkyl, C1-C6 alkoxy, NHC(O)(C1-C6 alkyl); R c1 Selected from deuterium, F, Cl, Br, I, CN, OH, NH2, or ethoxy; X is selected from S; L is selected from O or chemical bonds; R 1 Selected from H, C1-C6 alkyl, C3-C6 cycloalkyl, or 4-6 membered heterocyclic groups, wherein the C1-C6 alkyl, C3-C6 cycloalkyl, or 4-6 membered heterocyclic group is optionally R d Instead, the R d Selected from F, Cl, Br, I, OH, CN or C1-C3 alkyl groups.

2. The compound of general formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, R c Selected from =O or arbitrarily selected by R c1 The following groups are substituted: C1-C6 alkyl, C1-C6 alkoxy, NHC(O) (C1-C6 alkyl), wherein R c1 It is selected from deuterium, F, Cl, Br, I, CN, OH, NH2 or ethoxy.

3. The compound of general formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, Ring W is selected from 4. The compound of formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, Ring Q is selected from 5. The compound of general formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, The compound represented by formula (A) or a pharmaceutically acceptable salt thereof is selected from the compound represented by formula (B) or a pharmaceutically acceptable salt thereof: Among them, rings W and R a R 1 As defined in claim 1, n is selected from 0, 1, 2, 3, 4 or 5.

6. The compound of general formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, The compound represented by general formula (A) or a pharmaceutically acceptable salt thereof is selected from the compound of formula (C) or a pharmaceutically acceptable salt thereof: Among them, rings W and R a R 1 As defined in claim 1.

7. The compound of general formula (A) according to claim 1, or a pharmaceutically acceptable salt thereof, characterized in that, The compound is selected from one of the following structures:

8. A pharmaceutical composition comprising a compound of any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

9. Use of the compound of any one of claims 1 to 7 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 8, in the preparation of a medicament for the prevention or treatment of tumors with reduced or absent MTAP activity.

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