CN116332947A - Pyrimidine-2 (1H) -keto bicyclic compounds with MAT2A inhibitory activity and application thereof - Google Patents

Abstract

The present invention relates to pyrimidine-2(1H)-ketobicyclic compounds with MAT2A inhibitory activity and uses thereof, in particular to compounds of formula (I) with MAT2A inhibitory activity, stereoisomers, pharmaceutical compositions and uses thereof. The invention relates in particular to compounds of formula (I), and also to pharmaceutical compositions comprising these compounds and to the use of these compounds for the prevention or treatment of neoplastic diseases.

Description

Pyrimidine-2(1H)-one bicyclic compounds with MAT2A inhibitory activity and their uses

Technical Field

The present invention relates to a pyrimidine-2(1H)-one bicyclic compound, a stereoisomer or a pharmaceutically acceptable salt thereof, and the use thereof in preparing a MAT2A inhibitor, and treating and/or preventing tumors.

Background Art

Cancer treatment is a huge challenge facing the world today. The biggest problem with existing common therapies, such as chemotherapy and immunotherapy, is that the cell-killing effect is often not only targeted at cancer cells, but also has significant side effects on normal cells and tissues. Therefore, there is an urgent need to develop new treatments to better target cancer cells.

Synthetic lethality is defined as the loss of two or more genes leading to cell death, while the loss of any one of the genes alone has no effect. In recent years, a large number of studies have shown that there are multiple gene mutations in cancer cells that make them more sensitive to synthetic lethal treatments. These tumor-specific gene mutations can enable us to use appropriate targeted therapeutic drugs to kill cancer cells without affecting normal cells.

Methionine adenosyltransferase 2A (MAT2A) is an enzyme that catalyzes the reaction of methionine (Met) with ATP to generate S-adenosylmethionine (S-Adenosyl-L-methionine, SAM). SAM is the main methyl donor in the body and can regulate gene expression through transmethylation reactions of DNA, RNA and proteins, thereby having an important impact on cell differentiation, growth and death. Arginine N-methyltransferase 5 (PRMT5) is a methylase that uses SAM as a methyl donor. SAM is essential for the activity of PRMT5, and methylthioadenosine (5'methylthioadenosine, MTA) can inhibit the activity of PRMT5. MTA is a product of the methionine compensation pathway and is maintained at a low level in cells through the catalysis of methylthioadenosine phosphorylase (MTAP) to produce 5-methylthioribose-1-phosphate and adenine.

The MTAP gene is located on chromosome 9, which is deleted in the cells of patients with various cancers, including pancreatic cancer, esophageal cancer, bladder cancer, and lung cancer (cBioPortal database). The loss of MTAP leads to the enrichment of MTA in cells, which makes these cells more dependent on the production of SAM and the activity of MAT2A than normal cells. Studies have shown that inhibiting the expression of MAT2A in MTAP-deficient cancer cells can selectively inhibit cell activity compared with cancer cells with normal MTAP (McDonald et.al., 2017 Cell 170, 577-592). At the same time, reducing the expression of MAT2A can selectively inhibit tumor growth in a mouse xenograft tumor model of MTAP-deficient tumor cells (Marjon et.al., 2016 Cell Reports 15 (3), 574-587). These results indicate that MAT2A inhibitors can provide a new and effective treatment for cancer patients, especially those whose tumors contain MTAP deletions.

Summary of the invention

The present invention provides a compound of formula (I), a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a pharmaceutical composition thereof and a combination thereof. The inventors unexpectedly found that the compound of formula (I) is a good MAT2A inhibitor.

According to the present invention, there is provided a compound as represented by formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof,

in,

_X1 is ^CR3 or N, _X2 is ^CR4 or N, _X3 is ^CR5 or N, _X4 is ^CR6 or N; and at most two of _X1 , _X2 , _X3 , and _X4 are N at the same time;

R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from H, D, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₁ -C ₆ alkylsulfonyl, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, mercapto, C ₁ -C ₆ alkylamino, amino, nitro, carboxyl, and NHCOR ^a , wherein the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen, and R ^a is selected from C ₁ -C ₁₀ alkyl and C ₃ -C ₁₀ cycloalkyl;

Preferably, R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from H, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, and amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen;

Further preferably, R ³ , R ⁵ , and R ⁶ are each independently selected from H and halogen, and R ⁴ is selected from H, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, and amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen;

Further preferably, R ³ , R ⁵ , and R ⁶ are all H, and R ⁴ is selected from H, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, and amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen;

For example, _X1 is CH or N, _X2 is ^CR4 or N, _X3 is CH or N, and _X4 is CH or N; and at most two of _X1 , _X2 , _X3 , and _X4 are N at the same time; and ^R4 is selected from F, Cl, Br, _CH3 , _CD3 , _CF3 , _CF2H , _CF2D , OH, SH, NH2, CN _{, OCH3} , _CH3CH2 , _-CH2CH2CH3 , cyclopropyl, _{-CH2 ( CH3 ) CH3 ,} and _NO2 .

Preferably, at most one of X ₁ , X ₂ , X ₃ , and X ₄ is N.

More preferably, _X1 is ^CR3 , _X2 is ^CR4 , _X3 is ^CR5 , _X4 is ^CR6 , and ^R3 , ^R4 , ^R5 and ^R6 are as defined above.

W is selected from O, NR ^b , S, CHR ^b ; R ^b is selected from H, D, C ₁ -C ₄ alkyl, C ₃ -C ₆ cycloalkyl, the alkyl and cycloalkyl are unsubstituted or substituted by one or more substituents selected from D, halogen, -OH; preferably, R ^b is selected from H, D, C ₁ -C ₃ alkyl, C ₃ -C ₆ cycloalkyl, the alkyl and cycloalkyl are unsubstituted or substituted by one or more substituents selected from halogen, -OH; more preferably, R ^b is selected from H, D, methyl, ethyl, n-propyl, isopropyl, cyclopropyl, cyclobutyl, wherein methyl, ethyl, n-propyl, isopropyl, cyclopropyl, cyclobutyl are unsubstituted or substituted by one or more substituents selected from D, halogen, -OH;

Preferably, W is selected from O, NH, NCH ₃ , NC ₂ H ₄ OH, NCH(CH ₃ ) ₂ , NC ₂ H ₅ , S, CH ₂ ;

R ¹ is selected from unsubstituted or substituted C ₃ -C ₁₀ cycloalkyl, unsubstituted or substituted C ₆ -C ₁₀ aryl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, wherein the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C 1 -C 4 alkoxy, C ₁ -C 4 haloalkyl (such as CF ₃ , CF ₂ H), COOH, CONHR ^c , NHCOR ^c , NHSO 2 R ^c , R ^c is selected from H, C ₁ -C 4 alkyl, C 3 -C ₁₀ cycloalkyl, C 1 -C ₁₀ alkoxy or C 6 -C 10 aryl (such as phenyl, pyridine), wherein the C ₁ -C ₄ alkyl in R c is ^C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy _or C ₆ -C ₁₀ aryl (such as phenyl, pyridine), wherein the C ₁ -C _C4-C10 alkyl, _C3 - _C10 cycloalkyl, _C1 - _C10 alkoxy, _C6 - _C10 aryl are unsubstituted or substituted by one or more selected from halogen, hydroxyl, cyano, and the heteroaryl and heterocycloalkyl contain one or more heteroatoms selected from N, O, S;

Preferably, R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ and the definitions of heteroaryl and heterocycloalkyl are as described above respectively;

Preferably, R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, and the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl (such as CF ₃ , CF ₂ H), and the heteroaryl and heterocycloalkyl contain one or more heteroatoms selected from N, O, and S, respectively;

Preferably, R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl; the 5-10 membered heteroaryl is selected from the following groups:

Wherein, the substitution in R ¹ refers to substitution by one or more substituents selected from Group A; the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl (such as CF ₃ , CF ₂ H), COOH, CONHR ^c , NHCOR ^c , NHSO ₂ R ^c , R ^c is selected from H, C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy or C ₆ -C ₁₀ aryl (such as phenyl, pyridine), wherein the C ₁ -C ₄ alkyl, C ₃ -C 10 cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl in R ^c is _{unsubstituted} or substituted by one or more substituents selected from halogen, hydroxyl, cyano;

Preferably, the R ¹ is selected from

wherein R ¹⁰ is selected from halogen, CN, OH, SH, NH ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl, COOH, CONH ₂ , CONHR ^c , NHCOR ^c , and R ^c is selected from C ₁ -C ₄ alkyl, C ₃ -C ₆ cycloalkyl, C ₁ -C ₄ alkoxy or phenyl which is unsubstituted or substituted with one or more groups selected from halogen, hydroxyl, cyano.

Preferably, the R ¹ is selected from

R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ;

R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, D, unsubstituted or substituted C ₁ -C ₆ alkyl, unsubstituted or substituted C ₁ -C ₆ alkoxy, unsubstituted or substituted C ₂ -C ₆ alkenyl, unsubstituted or substituted C ₂ -C ₆ alkynyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted C ₅ -C ₁₀ aryl, unsubstituted or substituted 3-7 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₆ alkyl), -NHSO ₂ -(C ₁ -C ₆ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution by one or more substituents selected from the following: D, halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C ₃ alkyl which is unsubstituted or substituted by one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted by one or more substituents selected from Group B, or 5-10 membered heteroaryl which is unsubstituted or substituted by one or more substituents selected from Group B; R ^d is H, COOH, Boc, C ₁ -C ₃ alkyl which is unsubstituted or substituted by one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted by one or more substituents selected from Group B, or 5-10 membered heteroaryl which is unsubstituted or substituted by one or more substituents selected from Group B; the substituents in Group B include: C ₁ -C ₆ alkyl, halogen, OH, and amino;

or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 4-6 membered heterocycloalkyl group which is unsubstituted or substituted with one or more substituents selected from the following: OH, halogen, C ₁ -C ₁₀ alkyl, C ₃ -C ₁₀ cycloalkyl;

The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively.

Preferably, wherein R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ;

R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, unsubstituted or substituted C ₁ -C ₃ alkyl, unsubstituted or substituted C ₁ -C ₃ alkoxy, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted phenyl, unsubstituted or substituted 3-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₃ alkyl), and -NHSO ₂ -(C ₁ -C ₃ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution with one or more substituents selected from the following: halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C ₃ alkyl unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C 7 alkyl unsubstituted or substituted with one or more substituents selected from Group B, R ^d is H, COOH, Boc, C 1 -C 3 alkyl which is unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted with one or more substituents selected from Group B, or 5-10 _membered heteroaryl which is unsubstituted or substituted with one _{or more substituents selected from Group B; the substituents in Group B include: C 1 -C 3 alkyl , halogen} , OH, and amino;

or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 5-6 membered heterocycloalkyl group which is unsubstituted or substituted by one or more substituents selected from the following: OH, halogen, C ₁ -C ₃ alkyl, C ₃ -C ₆ cycloalkyl;

The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively.

More preferably, R ² is selected from NH ₂ ,

For example, in one embodiment, in the compound represented by formula (I),

_X1 is ^CR3 or N, _X2 is ^CR4 or N, _X3 is ^CR5 or N, _X4 is ^CR6 or N, at most one of _X1 , _X2 , _X3 , and _X4 is N, ^R3 , ^R5 , and ^R6 are each independently selected from H and halogen, and ^R4 is selected from H, halogen, _C1 - _C6 alkyl, _C2 - _C6 alkenyl, _C2 - _C6 alkynyl, _C1 - _C6 alkoxy, _C3 - _C7 cycloalkyl, cyano, hydroxyl, and amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen;

W is selected from O, NH, NCH ₃ , NC ₂ H ₄ OH, NCH(CH ₃ ) ₂ , NC ₂ H ₅ , S, CH ₂ ;

^R1 is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C3- _{C7 cycloalkyl} , unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl; the 5-10 membered heteroaryl is selected from the following groups:

Wherein, the substitution in R ¹ refers to substitution by one or more substituents selected from Group A; the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl (such as CF ₃ , CF ₂ H), COOH, CONHR ^c , NHCOR ^c , NHSO ₂ R ^c , R ^c is selected from H, C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy or C ₆ -C ₁₀ aryl, wherein the C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl in R ^c is unsubstituted or substituted by one or more substituents selected from halogen, hydroxyl, cyano;

R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ;

R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, unsubstituted or substituted C ₁ -C ₃ alkyl, unsubstituted or substituted C ₁ -C ₃ alkoxy, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted phenyl, unsubstituted or substituted 3-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₃ alkyl), and -NHSO ₂ -(C ₁ -C ₃ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution with one or more substituents selected from the following: halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C ₃ alkyl unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C 7 alkyl unsubstituted or substituted with one or more substituents selected from Group B, R ^d is H, COOH, Boc, C 1 -C 3 alkyl which is unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted with one or more substituents selected from Group B, or 5-10 _membered heteroaryl which is unsubstituted or substituted with one _{or more substituents selected from Group B; the substituents in Group B include: C 1 -C 3 alkyl , halogen} , OH, and amino;

or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 5-6 membered heterocycloalkyl group which is unsubstituted or substituted by one or more substituents selected from the following: OH, halogen, C ₁ -C ₃ alkyl, C ₃ -C ₆ cycloalkyl;

The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively.

For example, in one embodiment, the compound represented by formula (I) is represented by the following formula I-A:

wherein W, R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as described above, respectively.

For example, in one embodiment, the compound represented by formula (I) is represented by the following formula I-A:

Wherein, W is selected from O, NR ^b , S, CHR ^b ; R ^b is selected from H, D, C ₁ -C ₄ alkyl, C ₃ -C ₆ cycloalkyl, and the alkyl and cycloalkyl are unsubstituted or substituted with one or more substituents selected from D, halogen, -OH;

R ³ , R ⁵ , and R ⁶ are each independently selected from H and halogen, R ⁴ is selected from H, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, and amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen;

R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, and Group A substituents include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl;

R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ;

R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, unsubstituted or substituted C ₁ -C ₃ alkyl, unsubstituted or substituted C ₁ -C ₃ alkoxy, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted phenyl, unsubstituted or substituted 3-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₃ alkyl), and -NHSO ₂ -(C ₁ -C ₃ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution with one or more substituents selected from the following: halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C ₃ alkyl unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C 7 alkyl unsubstituted or substituted with one or more substituents selected from Group B, R ^d is H, COOH, Boc, C 1 -C 3 alkyl which is unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted with one or more substituents selected from Group B, or 5-10 _membered heteroaryl which is unsubstituted or substituted with one _{or more substituents selected from Group B; the substituents in Group B include: C 1 -C 3 alkyl , halogen} , OH, and amino;

or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 5-6 membered heterocycloalkyl group which is unsubstituted or substituted by one or more substituents selected from the following: OH, halogen, C ₁ -C ₃ alkyl, C ₃ -C ₆ cycloalkyl;

The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively.

The compound represented by formula (I) of the present invention can be specifically selected from the following structures:

The present invention also provides a composition comprising at least one compound represented by formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof.

The present invention also provides a pharmaceutical preparation comprising a therapeutically effective amount of a compound represented by formula (I), a stereoisomer thereof or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.

In another embodiment, the present invention provides a combination, in particular a pharmaceutical combination, comprising a therapeutically effective amount of a compound of the present invention, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, and one or more additional therapeutic agents.

The compounds according to the invention can be used alone, in combination with other compounds according to the invention or in combination with one or more, preferably one or two, other substances simultaneously or sequentially.

The present invention also provides the use of the compound described in formula (I), its stereoisomer or its pharmaceutically acceptable salt, or a composition containing the same, the pharmaceutical preparation, or the pharmaceutical combination in preparing a drug for treating and/or preventing tumors.

Preferably, the tumor includes: MTAP-deficient tumors; MTAP-lowly expressed tumors; MAT2A-abnormally expressed tumors; and other MAT2A-dependent tumors.

Specifically, the tumors include: breast cancer, lung cancer, glioblastoma, brain cancer and spinal cancer, head and neck cancer, skin cancer, reproductive system cancer, gastrointestinal system cancer, esophageal cancer, nasopharyngeal cancer, pancreatic cancer, rectal cancer, hepatocellular carcinoma, bile duct cancer, gallbladder cancer, colon cancer, multiple myeloma, kidney and bladder cancer, bone cancer, malignant mesothelioma, sarcoma, lymphoma, adenocarcinoma, thyroid cancer, heart tumors, germ cell tumors, malignant neuroendocrine tumors, malignant rhabdoid tumors, soft tissue sarcomas, midline tract cancer and unknown primary cancer.

In one embodiment of the present invention, the present invention provides a method for treating or preventing MAT2A-related tumors, the method comprising administering to a patient in need thereof an effective amount of a first therapeutic agent and optionally a second therapeutic agent, wherein the first therapeutic agent is a compound of the present invention, a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, and the second therapeutic agent is one or more other therapeutic agents.

In addition, the present invention provides a product or kit comprising a combined preparation of a compound of the present invention or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof, as defined above and one or more other active agents for simultaneous, separate or sequential use in anticancer therapy.

Terminology

In the present invention, unless otherwise explicitly stated, the terms used in the present invention have the meanings defined below. Terms not explicitly defined in the present invention have the general meanings generally understood by those skilled in the art.

The terms “a,” “an,” “the” and similar terms used in the context of the present invention (especially in the context of the claims) are to be construed to cover both the singular and the plural, unless otherwise specifically indicated herein or clearly contradicted by context.

A hyphen ("-") that is not between two letters or symbols indicates the point of attachment of a substituent. For example, -O(C ₁ -C ₃ alkyl) indicates that the group is attached to the rest of the molecule through an oxygen atom. However, when the point of attachment of a substituent is obvious to one skilled in the art, such as for halogen, hydroxyl, etc., the "-" may be omitted.

时，波浪线表示该基团与分子其余部分的连接位置。When the group has a wavy line

, the wavy line shows where the group is attached to the rest of the molecule.

As used herein, "heteroatom" refers to nitrogen (N), oxygen (O) or sulfur (S) atoms, particularly nitrogen or oxygen, each of which may be substituted or unsubstituted, including oxidized forms thereof. Examples of heteroatoms include, but are not limited to, -O-, -N=, -NR-, -S-, -S(O)-, and -S(O) ₂ -, where R is hydrogen, C ₁ -C ₄ alkyl, or a nitrogen protecting group (e.g., benzyloxycarbonyl, p-methoxybenzylcarbonyl, tert-butoxycarbonyl, acetyl, benzoyl, benzyl, p-methoxy-benzyl, p-methoxy-phenyl, 3,4-dimethoxybenzyl, etc.). Any heteroatom with unsatisfied valences is considered to have hydrogen atoms sufficient to satisfy the valences, unless otherwise indicated.

As used herein, "halogen" or "halo" refers to fluorine, chlorine, bromine and iodine. Preferred halogens as substituents are fluorine and chlorine.

As used herein, "alkyl" refers to a completely saturated straight or branched monovalent hydrocarbon group. The alkyl group preferably contains 1-20 carbon atoms, more preferably 1-16 carbon atoms, 1-10 carbon atoms, 1-8 carbon atoms, 1-6 carbon atoms, 1-4 carbon atoms or 1-3 carbon atoms. The number before the alkyl group represents the number of carbon atoms. For example, "C ₁ -C ₆ alkyl" represents an alkyl group having 1-6 carbon atoms, "C ₁ -C ₄ alkyl" represents an alkyl group having 1-4 carbon atoms, and "C ₁ -C ₃ alkyl" represents an alkyl group having 1-3 carbon atoms. Representative examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, n-heptyl, n-octyl, n-nonyl, n-decyl, etc. This definition applies regardless of whether the term "alkyl" appears alone or as part of another group such as haloalkyl, alkoxy, and the like.

As used herein, "alkenyl" refers to a linear or branched monovalent hydrocarbon group containing at least one double bond. The alkenyl group preferably contains 2-20 carbon atoms, more preferably 2-10 carbon atoms, 2-8 carbon atoms, 2-6 carbon atoms or 2-4 carbon atoms. The number before the alkenyl group represents the number of carbon atoms. Representative examples of alkenyl include, but are not limited to, ethenyl, propenyl, isopropenyl, butenyl, isobutenyl, pentenyl, isopentenyl, hexenyl, heptenyl, octenyl, etc.

As used herein, "alkynyl" refers to a straight or branched monovalent hydrocarbon group containing at least one triple bond. The alkynyl group preferably contains 2-20 carbon atoms, more preferably 2-10 carbon atoms, 2-8 carbon atoms, 2-6 carbon atoms or 2-4 carbon atoms. The number before the alkynyl group represents the number of carbon atoms. Representative examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, isopropynyl, butynyl, isobutynyl, pentynyl, isopentenyl, hexynyl, heptynyl, octynyl, etc.

As used herein, "alkoxy" refers to an alkyl group as defined herein connected by an oxygen bridge, i.e., an alkyl-O- group, and the number before the alkoxy group represents the number of carbon atoms. For example, "C _1- C ₆ alkoxy" represents an alkoxy group having 1-6 carbon atoms, i.e., -OC _1-6 alkyl; "C _1- C ₄ alkoxy" represents an alkoxy group having 1-4 carbon atoms, i.e., -OC _1-4 alkyl; "C _1- C ₃ alkoxy" represents an alkoxy group having 1-3 carbon atoms, i.e., -OC _1-3 alkyl. Representative examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy, 2-propoxy, butoxy, tert-butoxy, pentoxy, hexoxy, and the like. Preferably, the alkoxy group contains about 1-6 or about 1-4 carbon atoms, etc.

As used herein, "cycloalkyl" refers to a saturated or partially saturated non-aromatic carbocyclic ring, including mono-, bi- or tricyclic rings, preferably having 3-12 ring carbon atoms, more preferably 3-10 ring carbon atoms, such _as 3-8, 3-7, 3-6, 4-10, or _4-8 ring carbon atoms. "C3- _C8 cycloalkyl" is intended to include _C3 , _C4 , _C5 , _C6 , _C7 and C8 cycloalkyl groups; " _C3 - _C6 cycloalkyl" is intended to include _C3 , _C4 , _C5 and _C6 cycloalkyl groups; and so on. Exemplary monocyclic cycloalkyls include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl and cyclohexenyl, etc. Exemplary bicyclic cycloalkyl groups include bornyl, tetrahydronaphthyl, decahydronaphthyl, bicyclo[2.1.1]hexyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.1]heptenyl, 6,6-dimethylbicyclo[3.1.1]heptyl, 2,6,6-trimethylbicyclo[3.1.1]heptyl, bicyclo[2.2.2]octyl, etc. Exemplary tricyclic cycloalkyl groups include adamantyl, etc.

As used herein, "haloalkyl" refers to an alkyl group as defined herein in which one or more hydrogen atoms, such as 1, 2, 3, 4, 5, 6 or 7 hydrogen atoms, such as 1, 2 or 3 hydrogen atoms, are replaced by halogen, and when more than one hydrogen atom is replaced by a halogen atom, the halogen atoms may be the same or different from each other. For example, "C ₁ -C ₄ haloalkyl" is intended to include C ₁ , C ₂ , C ₃ and C ₄ haloalkyl groups, and "C ₁ -C ₃ haloalkyl" is intended to include C ₁ , C ₂ and C ₃ haloalkyl groups. Examples of haloalkyl groups include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, trichloromethyl, 1,1-difluoroethyl, 1,1,-difluoropropyl and 1,1,1-trifluoropropyl. Examples of haloalkyl groups also include "fluoroalkyl", which is intended to include alkyl groups as defined herein in which one or more hydrogen atoms are replaced by fluorine atoms. "Haloalkyl" herein is preferably an alkyl group in which up to three hydrogen atoms are replaced by halogen.

As used herein, "haloalkoxy" means a haloalkyl group as defined above with a specified number of carbon atoms attached via an oxygen bridge, wherein one or more hydrogen atoms, such as 1, 2, 3, 4, 5, 6 or 7 hydrogen atoms, such as 1, 2 or 3 hydrogen atoms, are replaced by halogen. For example, "C ₁ -C ₆ haloalkoxy" or "C ₁ to C ₆ haloalkoxy" is intended to include C ₁ , C ₂ , C ₃ , C ₄ , C ₅ and C ₆ haloalkoxy groups. Examples of haloalkoxy include, but are not limited to, fluoromethoxy, difluoromethoxy, trifluoromethoxy, 2-fluoroethoxy, 2,2,2-trifluoroethoxy. Examples of haloalkoxy also include "fluoroalkoxy".

As used herein, "aryl" is a monocyclic, bicyclic or tricyclic carbocyclic hydrocarbon group consisting of one or more rings fused together, having 6-20, preferably 6-14, more preferably 6-12, most preferably 6-10, for example 6-9 ring carbon atoms, wherein at least one ring is aromatic, and the other rings (if present) may be aromatic or non-aromatic. Preferred aryl groups are aryl groups having 6-10 ring carbon atoms, i.e. _C6 - _C10 aryl groups, which include: monocyclic aryl groups (e.g. phenyl); or fused bicyclic systems, wherein one ring is aromatic, and the other ring is aromatic (e.g. in naphthalene, biphenyl) or non-aromatic (e.g. in dihydroindene, tetrahydronaphthalene). Non-limiting examples of aryl groups include phenyl, biphenyl, naphthyl, tetrahydronaphthyl, indenyl, dihydroindene or anthracenyl, etc.

As used herein, "heteroaryl" refers to a 5-14-membered, preferably 5-10-membered, more preferably 5-7-membered or 5-6-membered aromatic ring system containing 1-8, preferably 1-4, also preferably 1-3, more preferably 1 or 2 ring heteroatoms selected from N, O or S, including monocyclic or bicyclic or fused polycyclic rings, and the remaining ring atoms are carbon atoms. Heteroaryl is preferably a 5-10-membered heteroaryl, more preferably a 5-7-membered heteroaryl or a 5-6-membered heteroaryl, each containing 1, 2 or 3 ring heteroatoms selected from N, O or S. Examples of heteroaryl groups include, but are not limited to, pyrrolyl, furanyl, thienyl, pyrazolyl, imidazolyl, triazolyl, thiazolyl, isothiazolyl, oxazolyl, pyridinyl, pyranyl, pyrazinyl, pyridazinyl, pyrimidinyl, oxazinyl, oxadiazinyl, quinolyl, isoquinolyl, cinnolinyl, quinazolinyl, quinoxalinyl, benzoxazinyl, 2H-chromene, benzopyranyl, benzothienyl, indolyl, indazolyl, benzimidazolyl, benzoxazolyl, benzothiazolyl, 7-azaindolyl, 6-azaindolyl, 5-azaindolyl, 4-azaindolyl, 1H-benzo[d][1,2,3]triazolyl, and the like.

As used herein, "heterocycloalkyl" refers to a cycloalkyl group as defined in the present application, provided that one or more ring carbons are replaced by a heteroatom selected from N, O or S, such as -O-, -N=, -NR-, -S-, -S(=O)- and -S(=O) _2- , wherein R is hydrogen, _C1-4 alkyl or a nitrogen protecting group (e.g., benzyloxycarbonyl, p-methoxybenzylcarbonyl, tert-butoxycarbonyl, acetyl, benzoyl, benzyl, p-methoxy-benzyl, p-methoxy-phenyl, 3,4-dimethoxybenzyl, etc.). Preferably, heterocycloalkyl is a monocyclic, bicyclic or tricyclic saturated and partially unsaturated non-aromatic ring having 3-20 ring atoms, such as 3-12 ring atoms, such as 3-8 ring atoms, such as 3-6 ring atoms. More preferably, the heterocycloalkyl group is preferably a 4- to 12-membered heterocycloalkyl group, preferably a 4- to 8-membered heterocycloalkyl group, more preferably a 4- to 7-membered, 4- to 6-membered or 5- to 6-membered heterocycloalkyl group, preferably containing 1, 2 or 3 heteroatoms selected from N, O or S, wherein the heteroatoms are substituted or unsubstituted, for example substituted by C ₁ -C ₄ alkyl. For example, examples of heterocycloalkyl include, but are not limited to, oxiranyl, aziridinyl, azetidinyl, oxetanyl, azopentyl (pyrrolidinyl), tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydrothiophenyl 1,1-dioxide, pyrazolidinyl, imidazolidinyl, oxazolidinyl, thiazolidinyl, isothiazolidinyl, pyrrolidin-2-one, imidazolonyl, piperidinyl (hexahydropyridine), N-methylpiperidinyl, tetrahydropyranyl, oxazinanyl, 1,3-oxazinanyl, hexahydropyrimidinyl, piperazinyl, piperidinylone, 1,4-dioxa-8-aza-spiro[4.5]decan-8-yl, morpholino, thiomorpholino, thiomorpholino-S-monooxide (sulfanomorpholino), thiomorpholino-S,S-dioxide (sulfonomorpholino), octahydropyrrolo[3,2-b]pyrrolyl, and the like.

As used herein, "partially or fully saturated heterocycle" refers to a partially or fully hydrogenated non-aromatic ring that can exist as a monocycle, a bicyclic ring (including fused rings) or a spirocycle. Unless otherwise specified, a heterocycle is typically a 3- to 12-membered ring, preferably a 5- to 10-membered ring, more preferably a 9-10-membered monocyclic or bicyclic ring system containing 1 to 3, such as 1 to 3, preferably 1 or 2 heteroatoms independently selected from S, O and/or N, wherein the heteroatoms are, for example, -O-, -N＝, -NR- or -S-, wherein R is hydrogen, C _1-4 alkyl or a nitrogen protecting group. When the term "partially or fully saturated heterocycle" is used, it is intended to include "heterocycloalkyl" and "partially saturated heterocycle". Examples of spirocycles include 2,6-diazaspiro[3.3]heptanyl, 3-azaspiro[5.5]undecyl, 3,9-diazaspiro[5.5]undecyl, and the like. Partially saturated heterocycles include groups such as dihydropyrrolyl, dihydrofuranyl, dihydrooxazolyl, dihydropyridinyl, imidazolinyl, 1H-dihydroimidazolyl, 2H-pyranyl, 2H-chromenyl, dihydrooxazinyl, etc. Partially saturated heterocycles also include heterocycles with fused aryl or heteroaryl rings, preferably with 9-10 ring members (e.g., dihydrobenzofuranyl, dihydroisobenzofuranyl, dihydroindole (or 2,3-dihydroindole), dihydrobenzothienyl, dihydrobenzothiazolyl, dihydrobenzopyranyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, tetrahydropyrido[3,4-b]pyrazinyl, etc.).

As used herein, the term "-C(=O)" is a carbonyl group, "-S(=O)" is a sulfoxide group, and "-S(=O) ₂ " is a sulfone group. "=O" means an oxo group, that is, an oxygen atom is connected to other atoms through a double bond.

As used herein, "optional", "optional" or "optionally" means that the subsequently described event may or may not occur, and the description includes instances where the event occurs as well as instances where the event does not occur. For example, "optionally substituted alkyl" includes "unsubstituted alkyl" and "substituted alkyl" as defined herein. "Optionally substituted with halogen" includes instances of "substituted with halogen" and instances of "not substituted with halogen", such as substituted with 0-3 halogens. It should be understood by those skilled in the art that for any group containing one or more substituents, the group does not include any sterically impractical, chemically incorrect, synthetically infeasible and/or inherently unstable substitution patterns.

When a dashed ring is used in a ring structure, this indicates that the ring structure may be saturated, partially saturated, or unsaturated.

The term "substituted", "substituted" or "substituted by..." as used herein means that one or more hydrogen atoms on a given atom or group are replaced by one or more substituents selected from a given substituent group, provided that the normal valence of the given atom is not exceeded. When the substituent is oxo (i.e., =O), two hydrogen atoms on a single atom are replaced by oxygen. There is no oxo substituent on the aromatic part. When a ring system (e.g., a carbocycle or a heterocycle) is substituted by a carbonyl group or a double bond, it is intended that the carbonyl group or the double bond is part of the ring (i.e., within the ring). Such combinations are allowed only when the combination of substituents and/or variables leads to chemically correct and stable compounds. Chemically correct and stable compounds mean that the compound is stable enough to be isolated from a reaction mixture and the chemical structure of the compound can be determined, and then it can be formulated into a formulation that at least has practical utility. For example, in the absence of a clear listing of substituents, the term "substituted", "substituted" or "substituted by..." as used herein means that one or more hydrogen atoms on a given atom or group are independently replaced by one or more, such as 1, 2, 3 or 4 substituents. When an atom or group is substituted with multiple substituents, the substituents may be the same or different.

Unless otherwise indicated, the term "compound of the present invention" or "compound of the present invention" refers to one or more compounds of formula (I) or its subformula defined herein, such as formula (I-1), (I-2), or a pharmaceutically acceptable salt thereof, and all isomers such as stereoisomers (including diastereomers, enantiomers and racemates), geometric isomers, conformational isomers (including rotational isomers and atropisomers), tautomers, internal addition products of isomers, prodrugs, and isotopically labeled compounds (including deuterium substitution) and inherently formed parts (e.g., polymorphs, solvates and/or hydrates). When there is a part capable of forming a salt, it also includes salts, especially pharmaceutically acceptable salts. The presence of tautomers or internal addition products of isomers can be identified by those skilled in the art using tools such as NMR. The compound of formula (I) of the present invention can easily form tautomers and internal addition products of isomers as depicted herein.

Those skilled in the art will recognize that the compounds of the present invention may contain chiral centers and as such may exist in different isomeric forms. "Isomers" as used herein refer to different compounds that have the same molecular formula but differ in arrangement and configuration of the atoms.

As used herein, "enantiomers" are a pair of stereoisomers that are non-superimposable mirror images of each other. A 1:1 mixture of a pair of enantiomers is a "racemic" mixture. Where appropriate, the term is used to refer to a racemic mixture. When indicating the stereochemistry of the compounds of the invention, the conventional RS system is used to specify single stereoisomers with known relative and absolute configurations of two chiral centers (e.g., (1S, 2S)); single stereoisomers with known relative configurations but unknown absolute configurations are marked with an asterisk (e.g., (1R*, 2R*)); racemates with two letters (e.g., (1RS, 2RS) is a racemic mixture of (1R, 2R) and (1S, 2S); (1RS, 2SR) is a racemic mixture of (1R, 2S) and (1S, 2R)). "Diastereomers" are stereoisomers that have at least two asymmetric atoms but are not mirror images of each other. Absolute stereochemistry is indicated according to the Cahn-lngold-Prelog R-S system. When the compound is a pure enantiomer, the stereochemistry at each chiral carbon can be described by R or S. The unknown resolved compound of absolute configuration can be designated as (+) or (-) according to the direction (right-handed or left-handed) of rotating plane polarized light at the wavelength of the sodium D line. Alternatively, the resolved compound can be defined by the respective retention time of the corresponding enantiomer/diastereomer via chiral HPLC.

Some of the compounds described herein contain one or more asymmetric centers or axes and may thus give rise to enantiomers, diastereomers, and other stereoisomers that may be defined in terms of absolute stereochemistry as (R)- or (S)-.

Geometric isomers can occur when the compound contains a double bond or some other feature that imparts a certain amount of structural rigidity to the molecule. If the compound contains a double bond, the substituents can be in the E or Z conformation. If the compound contains a disubstituted cycloalkyl, the cycloalkyl substituent can have a cis or trans configuration.

Conformational isomers are isomers that differ by rotation about one or more valence bonds. Rotamers are conformational isomers that differ by rotation about only a single valence bond.

"Atropisomers" refer to structural isomers based on axial or planar chirality resulting from restricted rotation in the molecule.

Unless otherwise indicated, the compounds of the invention are intended to include all such possible isomers, including racemic mixtures, optically active forms and intermediate mixtures. Optically active (R)- and (S)-isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques.

The compounds of the present invention may be separated into optically active forms or racemic forms. Optically active forms may be prepared by resolution of the racemic form or by synthesis from optically active starting materials. All methods for preparing the compounds of the present invention and intermediates prepared therein are considered to be part of the present invention. When enantiomers or diastereoisomer products are prepared, they may be separated by conventional methods such as by chromatography or fractional crystallization.

Depending on the process conditions, the end products of the present invention are obtained in free (neutral) or salt form. The free and salt forms of these end products are within the scope of the present invention. If desired, one form of the compound can be converted into other forms. Free bases or acids can be converted into salts; salts can be converted into free compounds or other salts; mixtures of isomeric compounds of the present invention can be separated into individual isomers.

As used herein, "pharmaceutically acceptable salts" refer to salts that retain the biological effects and properties of the compounds of the present invention, and the salts are not biologically or otherwise undesirable. Non-limiting examples of the salts include non-toxic, inorganic or organic base or acid addition salts of the compounds of the present invention. In many cases, due to the presence of amino and/or carboxyl groups or groups similar thereto, the compounds of the present invention are able to form acid salts and/or base salts. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like; ammonium, potassium, sodium, calcium, and magnesium salts are particularly preferred. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines (including naturally occurring substituted amines), cyclic amines, basic ion exchange resins, and the like, particularly, for example, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. Pharmaceutically acceptable salts of the present invention can be synthesized from parent compounds (basic or acidic moieties) by conventional chemical methods. Generally speaking, the salts can be prepared as follows: reacting the free acid form of the compound with a stoichiometric amount of an appropriate base (e.g., hydroxide, carbonate, bicarbonate, etc. of Na, Ca, Mg, or K) or reacting the free base form of the compound with a stoichiometric amount of an appropriate acid. Such reactions are usually carried out in water or an organic solvent or a mixed solvent of the two. In general, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred where practicable. Other suitable salts may be found in Remington's Pharmaceutical Sciences, 20th edition, Mack Publishing Company, Easton, Pa., (1985), which is incorporated herein by reference.

As used herein, "pharmaceutically acceptable excipients" include any and all solvents, dispersion media, coatings, surfactants, antioxidants, preservatives (e.g., antibacterial agents, antifungal agents), isotonic agents, absorption delaying agents, salts, preservatives, drugs, drug stabilizers, binders, excipients, disintegrants, lubricants, sweeteners, flavoring agents, dyes, the like, and combinations thereof, which are well known to those of ordinary skill in the art (see, e.g., Remington's Pharmaceutical Sciences, 18th edition, Mack Printing Company, 1990, pp. 1289-1329, incorporated herein by reference). Unless any conventional carrier is incompatible with the active ingredient, it may be considered for use in the therapeutic or pharmaceutical composition.

Any formula given herein is also intended to represent the unlabeled form of the compound as well as the isotope-labeled form. In addition to one or more atoms being replaced by atoms with a selected atomic mass or mass number, isotope-labeled compounds have the structure described by the formula given herein. Examples of isotopes that can be incorporated into the compounds of the present invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine and chlorine, such as ² H (i.e. D), ³ H (i.e. T), ¹¹ C, ¹³ C, ¹⁴ C, ¹⁵ N, ¹⁸ F ³¹ P, ³² P, ³⁵ S, ³⁶ Cl, ¹²⁵ I, respectively. The present invention includes different isotope-labeled compounds as defined herein, such as those in which radioactive isotopes such as ³ H, ¹³ C and ¹⁴ C are present. Such isotopically labeled compounds are useful in metabolic studies (with ¹⁴ C), reaction kinetic studies (e.g., with ² H or ³ H), detection or imaging techniques, such as positron emission tomography (PET) or single photon emission computed tomography (SPECT), including drug or substrate tissue distribution assays, or in radiotherapy of patients. In particular, ¹⁸ F or labeled compounds may be particularly desirable for PET or SPECT studies. Isotopically labeled compounds of the invention can generally be prepared by carrying out the procedures described in the schemes hereinafter or those described in the Examples and Preparations, substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent.

Moreover, substitution by heavier isotopes, particularly deuterium (i.e. ² H or D), can also obtain certain therapeutic benefits caused by greater metabolic stability, such as extending the half-life in vivo or reducing dosage requirements or improving the therapeutic index. It is understood that deuterium in this context can be regarded as a substituent of the compounds of the present invention. The concentration of such heavier isotopes, particularly deuterium, can be defined by an isotopic enrichment factor. "Isotopic enrichment factor" represents the ratio between the isotopic abundance and the natural abundance of a specified isotope.

As used herein, a "therapeutically effective amount" of a compound of the invention refers to an amount of a compound of the invention that can elicit a biological or medical response in an individual or improve symptoms, slow or delay progression of a disease, or prevent a disease, etc. A "therapeutically effective amount" can be determined by the attending physician or veterinary practitioner and will vary with factors such as the compound, the disease state being treated, the severity of the disease being treated, the age and relevant health conditions of the individual, the route and form of administration, the judgment of the attending physician or veterinary practitioner, and the like.

As used herein, "subject" refers to an animal. Preferably, the animal is a mammal. Subject also refers to, for example, primates (e.g., humans), cows, sheep, goats, horses, dogs, cats, rabbits, rats, mice, fish, birds, etc. In a preferred embodiment, the subject is a human.

As used herein, "inhibit" refers to a reduction or suppression of a particular condition, symptom or disorder or disease, or a significant decrease in the baseline activity of a biological activity or process.

As used herein, the term "treating" any disease or condition refers in one embodiment to ameliorating the disease or condition (i.e., arresting or slowing the development of the disease or at least one of its clinical symptoms). In another embodiment, "treating" refers to improving at least one physical parameter, which may not be perceived by the patient. In another embodiment, "treating" refers to modulating the disease or condition physically (e.g., stabilizing a perceptible symptom) or physiologically (e.g., stabilizing a physical parameter), or both.

As used herein, "prevention" refers to the administration of one or more pharmaceutical substances, particularly compounds of the present invention and/or pharmaceutically acceptable salts thereof, to an individual with a predisposition to the disease in question, in order to prevent the individual from developing the disease.

In general, the term "about" is used herein to modify a given numerical value by 20%, such as 10%, such as 5% above or below that value.

Technical and scientific terms used herein without specific definition have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

DETAILED DESCRIPTION

The following examples illustrate the invention described above, which however do not limit the scope of the invention in any way.The beneficial effects of the combination of the invention can also be determined by other test models known to those skilled in the art.

In the present invention, the sources and trade names of the reagents and equipment used are indicated when they first appear. The same reagents used thereafter are the same as those indicated for the first time unless otherwise specified. Conventional unlabeled reagents were purchased from Sinopharm Chemical Reagent Co., Ltd.

Reaction Formula A

Compound 1-a is dissolved in an inert solvent and reacted with methyl chloroacetate under alkaline conditions to obtain intermediate 2-a; intermediate compound 2-a is subjected to coupling reaction with boric acid connected with an ^R1 group under alkaline conditions to obtain intermediate 3-a; intermediate compound 3-a is reacted with chlorosulfonyl isocyanate in an inert solvent under low temperature conditions to obtain intermediate compound 4-a; compound 4-a is cyclized in a polar solvent under alkaline conditions to obtain compound 5-a.

Reaction Formula B

Compound 5-a reacts with a nucleophilic reagent containing R ² in a polar solvent such as acetonitrile in the presence of an organic base and a condensation reagent such as 1H-benzotriazol-1-yloxytripyrrolidino hexafluorophosphate to obtain a compound of formula I.

Reaction Formula C

Compound 5-a is dissolved in an organic solvent and reacted with phosphorus oxychloride in the presence of an organic base such as triethylamine under the protection of an inert gas to obtain a chlorinated intermediate 6-a; intermediate 6-a reacts with a nucleophilic reagent containing an ^R2 group such as methylamine to obtain a compound of the general formula I.

Reaction Formula D

Compound 1-b is dissolved in a polar aprotic solvent such as dimethyl sulfoxide, and reacts with an ester containing a W portion such as glycine methyl ester under heating conditions to obtain an intermediate 2-a; compound 2-a and a halogenated ^R1 reagent are reacted with a catalyst containing a metal such as Pd and a ligand containing a P element to obtain an intermediate compound 3-a through a coupling reaction; compound 3-a reacts with chlorosulfonyl isocyanate under low temperature conditions to obtain an intermediate compound 4-a; compound 4-a is cyclized under alkaline conditions to obtain compound 5-a.

In the above general formula, each substituent of W, R ¹ , R ² , R ³ , R ⁴ , R ⁵ , and R ⁶ has the same meaning as described above.

Example 1. Synthesis of Compound 1-A

Step 1: Synthesis of intermediate compound 2-1

The reaction was carried out in a 100 mL three-necked flask. 5-Chloro-2-hydroxybenzonitrile 1-1 (5.0 g, 32.47 mmol), methyl chloroacetate (3.5 g, 32.47 mmol), and potassium carbonate (9.0 g, 64.94 mmol) were dissolved in DMF (15 mL) and stirred at 80 ° C for 6 h under nitrogen protection until the reaction was complete. The reaction solution was poured into 200 mL of water, and a large amount of solid was precipitated. The filter cake was washed with a small amount of water, then dissolved with EA, and the remaining small amount of water was separated with a separatory funnel. The organic phase was dried with anhydrous sodium sulfate, filtered, and the filtrate was spin-dried to obtain compound 2-1, a white solid (8.5 g, yield> 100%), which can be directly used in the next step without purification. ¹ H NMR (400MHz, DMSO) δ8.08 (d, J = 1.8 Hz, 1H), 7.57–7.47 (m, 2H), 6.42 (s, 2H), 3.82 (s, 3H). LCMS (ESI) m/z = 226.0 [M + H] ⁺ .

Step 2: Synthesis of intermediate compound 3-1

The reaction was carried out in a 100 mL three-necked flask. Compound 2-1 (2.0 g, 8.87 mmol), m-methylphenylboronic acid (2.4 g, 17.74 mmol), copper acetate (1.9 g, 8.87 mmol), and 4A molecular sieve (0.25 g) obtained in the previous step were dissolved in DCM (30 mL), and then 2.5 mL of triethylamine was added, and an oxygen ball was inserted, and the atmosphere was replaced three times to form an oxygen atmosphere. Stir at room temperature for 12 hours until the reaction was complete. The reaction solution was filtered through diatomaceous earth, and the filter cake was rinsed with dichloromethane several times. The filtrate was combined, silica gel was added to mix the sample, and it was spin-dried, and then petroleum ether: ethyl acetate (50:1) was passed through a column to obtain compound 3-1 as an orange-red oil (2.12 g, yield: 76%). ¹ H NMR (400MHz, DMSO) δ8.25 (s, 1H), 7.70 (d, J = 8.9Hz, 1H), 7.54 (dd, J ₁ = 8.9Hz, J ₂ = 2.2Hz, 1H), 7.20 (t, J = 7.8Hz, 1H), 7.13 (d, J = 2.1Hz, 1H), 6.95 (s, 1H), 6 .92–6.87(m,2H),3.85(s,3H),2.27(s,3H).

Step 3: Synthesis of intermediate compound 4-1

The reaction was carried out in a 250 mL three-necked flask. Intermediate 3-1 (2.0 g, 6.35 mmol) was added to THF (30 ml), the temperature was lowered to -15 ° C under nitrogen protection, and compound chlorosulfonyl isocyanate (1.34 g, 9.52 mmol) was added and stirred for 1 h. Ammonium chloride aqueous solution (20 ml) was added to quench, water (200 ml) was added, and EA (200 ml) was used for extraction twice. The organic phase was concentrated to obtain compound 4-1 (2.2 g, yield: 97.9%) as a white solid. LCMS (ESI) m/z = 359.0 [M + H] ⁺ .

Step 4: Synthesis of Compound 1-A

The reaction was carried out in a 100 mL three-necked flask. Intermediate 4-1 (2.2 g, 6.13 mmol) was dissolved in methanol (30 ml), sodium hydroxide (490 mg, 12.26 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. The pH was adjusted to 3-4 with 1 mol/L HCl, and a white solid 1-A (1.2 g, yield: 60.0%) was obtained by filtration. LCMS (ESI) m/z = 327.0 [M + H] ⁺ .

Compound 1-B: was prepared by replacing the starting material 5-chloro-2-hydroxybenzonitrile 1-1 in the first step of the synthesis step in Example 1 with 4-chloro-2-hydroxybenzonitrile, and referring to the method of Example 1.

¹ H NMR (400MHz, DMSO) δ11.88 (s, 1H), 8.02 (d, J = 1.5Hz, 1H), 7.53 (d, J = 7.6Hz, 1H), 7.47 (d, J = 7.7Hz, 1H), 7.45–7.38 (m, 2H), 7.27 (dd, J ₁ = 8.7Hz, J ₂ = 1.6Hz, 1H), 6.09 (d, J = 8.7Hz, 1H), 2.41 (s, 3H).

Referring to the method of Example 1, the starting material 5-chloro-2-hydroxybenzonitrile 1-1 in the first step of the synthesis step was replaced by 5-fluoro-2-hydroxybenzonitrile and 2-chloro-6-hydroxybenzonitrile to prepare compounds 1-C and 1-D, respectively.

Compound 1-C

¹ H NMR (400MHz, DMSO) δ11.90 (s, 1H), 7.86 (dd, J ₁ = 9.2Hz, J ₂ = 4.0Hz, 1H), 7.55 (t, J = 7.6Hz, 1H), 7.51– 7.39 (m, 4H), 5.68 (dd, J ₁ = 8.4Hz, J ₂ = 2.4Hz, 1H), 2.42 (s, 3H).

Compound 1-D

LCMS (ESI) m/z = 327.0 [M + H] ⁺ .

Referring to the method of Example 1, the m-methylphenylboric acid in the second step of the synthesis step was replaced by m-chlorophenylboric acid, m-fluorophenylboric acid, o-methylphenylboric acid, and p-methylphenylboric acid to prepare Compound 1-E, Compound 1-F, Compound 1-G, and Compound 1-H, respectively.

Compound 1-E

LCMS(ESI)m/z=347.10[M+H] ⁺ .

Compound 1-F

LCMS(ESI)m/z=331.10[M+H] ⁺ .

Compound 1-G

LCMS (ESI) m/z = 327.0 [M + H] ⁺ .

Compound 1-H

¹ H NMR (400MHz, DMSO) δ11.90 (s, 1H), 7.85 (d, J = 9.0Hz, 1H), 7.61 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H), 7.53– 7.43(m,4H),6.03(d,J＝2.2Hz,1H),2.47(s,3H).LCMS(ESI)m/z＝327.0[M+H] ⁺ .

Example 2. Synthesis of Compound 17

The reaction was carried out in a 10 mL microwave tube. 1-A (150 mg, 0.46 mmol) was dissolved in acetonitrile (2 ml), and PyBOP (358 mg, 0.69 mmol), DBU (140 mg, 0.92 mmol), and 2-aminoethane-1-ol (56 mg, 0.920 mmol) were added. The reaction was carried out at 50° C. for 2 h. Purification by Prep-HPLC (H ₂ O/ACN, 0.1% TFA) gave 9 mg of 17 as a white solid in a yield of 6%. ¹ H NMR (400MHz, DMSO) δ8.82 (s, 1H), 7.79 (d, J = 9.0Hz, 1H), 7.60 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H), 7.53 (t, J = 7.7Hz, 1H), 7.45 (d, J = 7.6Hz, 1H), 7.37–7.26 (m, 2H),6.09(d,J＝2.2Hz,1H),3.63(t,J＝5.6Hz,2H),3.60–3.55(m,2H),2.41(s,3H).LCMS(ESI)m/z＝370.1[M+H] ⁺ .

Referring to the aforementioned method, methylamine, cyclopropylamine, tetrahydropyrrole, 3-hydroxypyrrolidine, N-methylethylamine, isopropylamine, DL-aminopropanol, cyclobutylamine, cis-3-aminocyclobutanol, trans-3-aminocyclobutanol, 3-oxetanamine, 3-aminotetrahydrofuran, 4-aminotetrahydropyran, 1-methyl-1H-pyrazole-5-methylamine, ethylamine, 2-methoxyethylamine, 3-amino-1-propanol, cyclopropylmethylamine, N,N-dimethylethylenediamine, and N-acetylethylenediamine were used to replace 2-aminoethane-1-ol and react with 1-A to prepare compounds 1, 18, 19, 20, 21, 24, 25, 26, 28, 29, 30, 31, 33, 35, 36, 37, 38, 39, 40, and 41, respectively.

Compound 1:

¹ H NMR (400MHz, DMSO) δ8.59(d,J＝4.4Hz,1H),7.77(d,J＝9.2Hz,1H),7.57(dd,J＝9.0,2.2Hz,1H),7.52( t,1H),7.43(d,J＝7.6Hz,1H),7.32–7.25(m,2H),6.10(d,J＝2.4Hz,1H),2.97(d,J＝4.4Hz,3H), 2.41(s,3H).LCMS(ESI)m/z＝340.0[M+1] ⁺ .

Compound 18:

¹ H NMR (400MHz, DMSO) δ8.73 (s, 1H), 7.74 (d, J = 8.9Hz, 1H), 7.57 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H), 7.52 ( t,J＝7.7Hz,1H),7.44(d,J＝7.6Hz,1H),7.34–7.27(m,2H),6.09(d,J＝2.1Hz,1H),3.08(dd,J ₁ = 7.3Hz, J ₂ =3.5Hz, 1H), 2.41 (s, 3H), 0.79 (d, J = 7.1Hz, 2H), 0.72 (t, J = 3.9Hz, 2H). LCMS (ESI) m/z ＝366.1[M+H] ⁺ .

Compound 19:

¹ H NMR (400MHz, DMSO) δ7.80 (d, J = 9.0Hz, 1H), 7.57 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H), 7.52 (t, J = 7.7Hz, 1H),7.44(d,J＝7.6Hz,1H),7.29(s,1H),7.25(d,J＝7.9Hz,1H),6.04(d,J＝2.2Hz,1H),4.06(t, J＝6.7Hz,2H),3.66(t,J＝6.8Hz,2H),2.41(s,3H),2.11–2.02(m,2H),1.98–1.89(m,2H).LCMS(ESI)m /z＝380.1[M+H] ⁺ .

Compound 20:

¹ H NMR (400MHz, DMSO) δ7.81 (dd, J ₁ = 8.9Hz, J ₂ = 5.2Hz, 1H), 7.57 (dd, J ₁ = 9.0Hz, J ₂ =2.1Hz,1H),7.52(t,J＝7.7Hz,1H),7.44(d,J＝7.6Hz,1H),7.28(d,J＝15.7Hz,2H),6.04(d,J＝2.1 Hz,1H),5.15(d,J＝28.3Hz,1H),4.47(d,J＝39.9Hz,1H),4.28–3.95(m,2H),3.85–3.58(m,2H),2.41(s ,3H),2.13(m,1H),2.06–1.87(m,1H).LCMS(ESI)m/z＝396.1[M+1] ⁺ .

Compound 21:

¹ H NMR (400MHz, CDCl ₃ ) δ7.53–7.37(m,4H),7.25–7.15(m,2H),6.19(s,1H),4.04(s,2H),3.50(s,3H), 2.45(s,3H),1.40(s,3H).LCMS(ESI)m/z＝368.1[M+H] ⁺ .

Compound 24:

¹ H NMR (400MHz, DMSO) δ8.50 (d, J = 8.0Hz, 1H), 7.75 (d, J = 8.9Hz, 1H), 7.57 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H),7.52(t,J＝7.6Hz,1H),7.43(d,J＝7.7Hz,1H),7.32(s,1H),7.29(d,J＝7.8Hz,1H),6.09(d, J＝2.1Hz,1H),4.42(m,1H),2.41(s,3H),1.26(d,J＝6.6Hz,6H).LCMS(ESI)m/z＝368.0[M+1] ⁺ .

Compound 25:

¹ H NMR (400MHz, DMSO) δ8.34 (d, J = 8.1Hz, 1H), 7.76 (d, J = 9.0Hz, 1H), 7.58 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H),7.52(t,J＝7.7Hz,1H),7.43(d,J＝7.6Hz,1H),7.32(s,1H),7.29(d,J＝7.7Hz,1H),6.09(d, J＝2.0Hz,1H),4.84(t,J＝5.7Hz,1H),4.40–4.26(m,1H),3.56(dd,J ₁ ＝11.0Hz,J ₂ ＝5.5Hz,1H),3.44( dd,J ₁ =11.1Hz,J ₂ ＝5.6Hz,1H),2.41(s,3H),1.21(d,J＝6.7Hz,3H).LCMS(ESI)m/z＝384.0[M+1] ⁺ .

Compound 26:

¹ H NMR (400MHz, DMSO) δ8.88 (d, J = 7.5Hz, 1H), 7.76 (d, J = 8.9Hz, 1H), 7.58 (dd, J ₁ = 9.0Hz, J ₂ = 2.1Hz, 1H),7.51(t,J＝7.6Hz,1H),7.43(d,J＝7.6Hz,1H),7.33–7.25(m,2H),6.08(d,J＝2.1Hz,1H),4.67( m,1H),2.40(s,3H),2.27–2.29(m,2H),2.22–2.12(m,2H),1.70–1.76(m,2H).LCMS(ESI)m/z＝380.2[M +1] ⁺ .

Compound 28:

¹ H NMR (400MHz, DMSO) δ8.87 (d, J = 7.1Hz, 1H), 7.77 (d, J = 9.0Hz, 1H), 7.58 (dd, J ₁ = 9.0, J ₂ ＝2.1Hz,1H),7.51(t,J＝7.7Hz,1H),7.43(d,J＝7.7Hz,1H),7.31(s,1H),7.28(d,J＝7.9Hz,1H), 6.08(d,J＝2.1Hz,1H),5.17(d,J＝5.6Hz,1H),4.13(d,J＝7.3Hz,1H),3.90(d,J＝6.6Hz,1H),2.64– 2.54(m,2H),2.40(s,3H),2.01(d,J＝8.7Hz,2H).LCMS(ESI)m/z＝396.1[M+1] ⁺ .

Compound 29:

¹ H NMR (400MHz, DMSO) δ9.09(d,J＝6.3Hz,1H),7.78(d,J＝9.0Hz,1H),7.61–7.57(m,1H),7.51(t,J＝7.7 Hz,1H),7.45(s,1H),7.32(s,1H),7.29(d,J＝7.7Hz,1H),6.08(d,J＝2.1Hz,1H),4.69–4.60(m,1H ),4.33–4.38(m,1H),2.54(s,1H),2.41(s,3H),2.37–2.43(m,2H),2.20–2.27(m,2H).LCMS(ESI)m/z ＝396.1[M+1] ⁺ .

Compound 30:

¹ H NMR (400MHz, DMSO) δ9.34 (d, J = 5.1Hz, 1H), 7.80 (d, J = 9.0Hz, 1H), 7.60 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H),7.51(d,J＝7.6Hz,1H),7.44(d,J＝7.6Hz,1H),7.33–7.26(m,2H),6.09(d,J＝2.1Hz,1H),5.18( dd,J ₁ =12.7Hz, J ₂ =6.5Hz, 1H), 4.83 (t, J = 6.8Hz, 2H), 4.69 (t, J = 6.5Hz, 2H), 2.41 (s, 3H).LCMS ( ESI)m/z＝382.0[M+1] ⁺ .

Compound 31:

¹ H NMR (400MHz, DMSO) δ8.82 (d, J = 6.5Hz, 1H), 7.76 (d, J = 9.0Hz, 1H), 7.58 (dd, J ₁ = 8.9Hz, J ₂ = 2.2Hz, 1H),7.52(t,J＝7.7Hz,1H),7.44(d,J＝7.6Hz,1H),7.34–7.27(m,2H),6.09(d,J＝1.9Hz,1H),4.71( d, J＝6.5Hz, 1H), 3.92 (q, J＝8.2Hz, 2H), 3.73 (ddd, J ₁ ＝ 12.9Hz, J ₂ ＝ 11.4Hz, J ₃ ＝6.1Hz,2H),2.41(s,3H),2.23(dq,J＝14.8,7.6Hz,1H),2.06(td,J＝12.5,6.0Hz,1H).LCMS(ESI)m/z＝ 396.0[M+1] ⁺ .

Compound 33:

¹ H NMR (400MHz, DMSO) δ9.09 (s, 1H), 7.80 (d, J = 9.0Hz, 1H), 7.62 (dd, J ₁ = 8.9Hz, J ₂ = 2.1Hz, 1H), 7.54 ( t,J＝7.6Hz,1H),7.47(d,J＝7.8Hz,1H),7.36–7.30(m,2H),6.08(d,J＝2.1Hz,1H),4.35–4.32(m,1H ),3.94(d,J＝8.2Hz,2H),3.42(t,J＝11.2Hz,2H),2.42(s,3H),1.85(d,J＝10.3Hz,2H),1.73(dd,J ₁ =11.0Hz,J ₂ ＝8.0Hz,2H).LCMS(ESI)m/z＝410.2[M+1] ⁺ .

Compound 35:

¹ H NMR (400MHz, DMSO) δ9.29 (t, J = 5.2Hz, 1H), 7.78 (d, J = 9.0Hz, 1H), 7.59 (dd, J ₁ = 9.0Hz, J ₂ = 2.2Hz, 1H),7.52(t,J＝7.8Hz,1H),7.44(d,J＝7.6Hz,1H),7.33(d,J＝1.6Hz,2H),7.30(d,J＝8.0Hz,1H) ,6.25(d,J＝1.6Hz,1H),6.08(d,J＝2.2Hz,1H),4.74(d,J＝5.0Hz,2H),3.90(s,3H),2.40(s,3H) .LCMS(ESI)m/z＝420.1[M+1] ⁺ .

Compound 36:

¹ H NMR (400MHz, DMSO) δ8.65 (s, 1H), 7.76 (d, J = 9.0Hz, 1H), 7.57 (dd, J = 8.9, 2.2Hz, 1H), 7.51 (t, J = 7.7 Hz,1H),7.43(d,J＝7.6Hz,1H),7.31(s,1H),7.28(d,J＝7.9Hz,1H),6.09(d,J＝2.1Hz,1H),3.50( dd,J ₁ =6.4Hz, J ₂ =2.8Hz, 2H), 2.40 (s, 3H), 1.21 (t, J = 7.2Hz, 3H). LCMS (ESI) m/z = 353.95 [M+1] ⁺ .

Compound 37:

¹ H NMR (400MHz, DMSO) δ8.67(t,J＝5.4Hz,1H),7.78(d,J＝8.9Hz,1H),7.58(dd,J＝9.0,1.9Hz,1H),7.52( t,J＝7.6Hz,1H),7.44(d,J＝7.7Hz,1H),7.32(s,1H),7.29(d,J＝7.7Hz,1H),6.09(d,J＝1.7Hz, 1H),3.64(t,J＝5.4Hz,2H),3.58(d,J＝5.3Hz,2H),3.30(s,3H),2.41(s,3H).LCMS(ESI)m/z＝384.0 [M+1] ⁺ .

Compound 38:

¹ H NMR (400MHz, DMSO) δ8.61 (s, 1H), 7.77 (d, J = 9.0Hz, 1H), 7.57 (m, J = 9.0Hz, 1H), 7.52 (t, J = 7.7Hz, 1H),7.43(d,J＝7.6Hz,1H),7.32(s,1H),7.29(d,J＝7.8Hz,1H),6.09(d,J＝2.0Hz,1H),4.60(t, J＝5.1Hz,1H),3.56–3.47(m,4H),2.41(s,3H),1.83–1.74(m,2H).LCMS(ESI)m/z＝384.1[M+1] ⁺ .

Compound 39:

¹ H NMR (400MHz, DMSO) δ9.14–9.05 (m, 1H), 7.79 (d, J = 9.0Hz, 1H), 7.61 (dd, J ₁ = 9.0, J ₂ = 2.1Hz, 1H), 7.52 (d,J＝7.7Hz,1H),7.46(s,1H),7.34(s,1H),7.31(d,J＝7.8Hz,1H),6.09(d,J＝2.0Hz,1H),3.38 (t,J＝6.3Hz,2H),2.41(s,3H),1.25–1.13(m,1H),0.50(d,J＝7.9Hz,2H),0.33(d,J＝4.9Hz,2H) .LCMS(ESI)m/z＝380.1[M+1] ⁺ .

Compound 40:

¹ H NMR (400MHz, DMSO) δ8.47 (t, J = 5.3Hz, 1H), 7.77 (d, J = 9.0Hz, 1H), 7.57 (dd, J ₁ = 8.9Hz, J ₂ = 2.2Hz, ,1H),7.51(t,J＝7.7Hz,1H),7.43(d,J＝7.6Hz,1H),7.31(s,1H),7.28(d,J＝7.9Hz,1H),6.08(d ,J＝2.1Hz,1H),3.57(q,J＝6.3Hz,2H),2.52(q,J＝6.3Hz,2H),2.40(s,3H),2.20(s,6H).LCMS(ESI )m/z＝397.0[M+1] ⁺ .

Compound 41:

¹ H NMR (400MHz, DMSO) δ8.64 (t, J = 5.4Hz, 1H), 8.05 (t, J = 5.2Hz, 1H), 7.78 (d, J = 9.2Hz, 1H), 7.58 (dd, J ₁ =8.8Hz, J ₂ =2.0Hz, 1H), 7.52 (t, J = 7.6Hz, 1H), 7.44 (d, J = 7.6Hz, 1H), 7.32 (s, 1H), 7.28 (d, J＝7.6Hz,1H),6.10(d,J＝1.6Hz,1H),3.52(dd,J ₁ ＝12.4Hz,J ₂ ＝6.4Hz,2H),3.32(dd,J ₁ ＝12.4Hz,J ₂ ＝6.4Hz,2H),2.41(s,3H),1.83(s,3H).LCMS(ESI)m/z＝411.0[M+1] ⁺ .

Referring to Example 2. The synthesis method of compound 17, methylamine was used instead of 2-aminoethane-1-ol in Example 2 to react with 1-B, 1-C, and 1-D, respectively, to prepare compound 6, compound 3, and compound 5, respectively.

Compound 6:

¹ H NMR (400MHz, DMSO) δ = 9.15 (s, 1H), 7.96 (d, J = 1.6Hz, 1H), 7.52 (t, J = 7.6Hz, 1H), 7.43 (d, J = 7.6Hz, 1H),7.34–7.27(m,3H),6.19(d,J＝8.6Hz,1H),3.02(d,J＝3.6Hz,3H),2.40(s,3H).

LCMS (ESI) m/z = 340.1 [M + H] ⁺ .

Compound 3:

¹ H NMR (400MHz, DMSO) δ8.57 (d, J = 4.8Hz, 1H), 7.78 (dd, J ₁ = 9.2Hz, J ₂ = 4.0Hz, 1H), 7.51 (t, J = 7.6Hz, 1H),7.45–7.39(m,2H),7.30(s,1H),7.27(d,J＝7.6Hz,1H),5.79(dd,J ₁ ＝8.4Hz,J ₂ ＝2.8Hz,1H), 2.97(d,J＝4.8Hz,3H),2.41(s,3H).LCMS(ESI)m/z＝324.1[M+H] ⁺ .

Compound 5:

¹ H NMR (400MHz, DMSO) δ7.76 (d, J＝8.4Hz, 1H), 7.54–7.37 (m, 5H), 7.23 (dd, J＝7.8, 0.7Hz, 1H), 6.03 (s, 1H ),2.74(d,J＝4.3Hz,3H),2.39(s,3H).LCMS(ESI)m/z＝339.9[M+H] ⁺ .

Referring to Example 2. The synthesis method of compound 17, cyclopropylamine was used instead of 2-aminoethane-1-ol in Example 2, and reacted with compound 1-E, compound 1-F, compound 1-G, and compound 1-H, respectively, to prepare compound 42, compound 43, compound 45, and compound 46, respectively.

Compound 42:

¹ H NMR (400MHz, DMSO) δ8.79(d,J＝4.3Hz,1H),7.77(d,J＝9.0Hz,1H),7.74–7.69(m,2H),7.69–7.64(m,1H ), 7.60 (dd, J ₁ = 9.0Hz, J ₂ = 2.1Hz, 1H), 7.53 (dd, J ₁ = 7.4Hz, J ₂ = 1.6Hz, 1H), 6.17 (d, J = 2.0Hz, 1H ), 3.08 (d, J = 3.9Hz, 1H), 0.79 (dd, J ₁ = 6.8Hz, J ₂ = 4.7Hz, 2H), 0.72 (t, J = 3.8Hz, 2H).LCMS(ESI)m /z＝386.04[M+1] ⁺ .

Compound 43:

¹ H NMR (400MHz, DMSO) δ8.79 (d, J = 4.2Hz, 1H), 7.77 (d, J = 8.9Hz, 1H), 7.69 (dd, J ₁ = 15.1Hz, J ₂ = 7.8Hz, 1H), 7.60 (dd, J ₁ = 8.9Hz, J ₂ = 2.1Hz, 1H), 7.50 (t, J = 8.1Hz, 2H), 7.40 (d, J = 8.1Hz, 1H), 6.16 (d, J＝2.0Hz,1H),3.08(d,J＝3.8Hz,1H),0.80(d,J＝7.2Hz,2H),0.72(t,J＝3.9Hz,2H).LCMS(ESI)m/ z＝369.9[M+1] ⁺ .

Compound 45:

¹ H NMR (400MHz, DMSO) δ8.75 (s, 1H), 7.75 (d, J = 8.9Hz, 1H), 7.59–7.49 (m, 3H), 7.43 (dd, J ₁ = 17.1, J ₂ = 5.1Hz,2H),5.90(s,1H),3.10(s,1H),2.07(d,J＝3.5Hz,3H),0.80(d,J＝7.2Hz,2H),0.73(s,2H) .LCMS(ESI)m/z＝366.0[M+1] ⁺ .

Compound 46:

¹ H NMR (400MHz, DMSO) δ9.34 (s, 1H), 7.79 (d, J = 9.0Hz, 1H), 7.65–7.58 (m, 1H), 7.47 (d, J = 8.2Hz, 2H), 7.40 (d, J = 8.2Hz, 2H), 6.13 (d, J = 2.0Hz, 1H), 3.02 (td, J ₁ = 6.6Hz, J ₂ = 4.0Hz, 1H), 2.48 (s, 3H), 0.90–0.81(m,2H),0.81–0.72(m,2H).LCMS(ESI)m/z＝366.1[M+1] ⁺ .

Referring to the synthesis method of compound 17 in Example 2, methylamine is used instead of 2-aminoethane-1-ol in Example 2 to react with 1-G to prepare compound 64.

64:

¹ H NMR (400MHz, DMSO) δ8.63 (d, J = 4.6Hz, 1H), 7.78 (d, J = 9.0Hz, 1H), 7.57 (dd, J ₁ = 8.9Hz, J ₂ = 2.2Hz, 1H),7.55–7.51(m,2H),7.48–7.42(m,1H),7.40(d,J＝7.5Hz,1H),5.91(d,J＝2.1Hz,1H),2.98(d,J ＝4.6Hz,3H),2.06(s,3H).LCMS(ESI)m/z＝340.0[M+1] ⁺ .

Example 3. Synthesis of Compound 2

Step 1: Synthesis of intermediate compound 1-A-1

The reaction was carried out in a 25 ml single-necked flask. Compound 1-A (100 mg, 0.3 mmol), Pd/C (10 mg), and methanol (5 ml) were placed in a single-necked flask, replaced with hydrogen three times, stirred at room temperature for overnight reaction, and samples were taken to detect the complete reaction of the raw materials. Post-treatment: The reaction solution was filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain compound 1-A-1 (80.0 mg, yield: 89.5%), a white solid, which was directly subjected to the next step of reaction without purification.

LCMS (ESI) m/z = 293.1 [M + H] ⁺ .

Step 2: Synthesis of Compound 2

The reaction was carried out in a microwave tube. Compound 1-A-1 (80 mg, 0.27 mmol) and PyBOP (213.3 mg, 0.41 mmol) were dissolved in acetonitrile (6 ml), and DBU (83.7 mg, 0.55 mmol) was added dropwise at room temperature. After the addition was complete, the mixture was stirred at room temperature for 0.5 hours, and then a 1M methylamine tetrahydrofuran solution (1.1 ml, 1.1 mmol) was added dropwise. The mixture was reacted at 50°C for 1 hour in a microwave oven and the sample was sent for testing. Post-treatment: The reaction solution was spin-dried, extracted twice with dichloromethane (50 ml), the organic layer was washed twice with purified water and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated to dryness under reduced pressure, purified by Prep-HPLC (0.1% trifluoroacetic acid system), and freeze-dried to obtain 4-(methylamino)1-(m-benzyl)benzofuran[3,2-d]pyrimidine 2(1H)-one (compound 2) as a white solid (5.7 mg, yield: 6.8%). ¹ H NMR (400MHz, MeOD) δ7.71(d,J＝8.8Hz,1H),7.64–7.56(m,2H),7.53(d,J＝7.6Hz,1H),7.40(s,1H),7.36(d,J＝7.6Hz,1H),7.18(t,J＝7.8Hz,1H),6.37(d, J＝8.0Hz,1H),3.23(s,3H),2.50(s,3H).LCMS(ESI)m/z＝306.0[M+H] ⁺ .

Example 4. Synthesis of Compound 4

Step 1: Synthesis of intermediate compound 2-4

The reaction was carried out in a 100 mL eggplant flask. 2-Hydroxy-5-trifluoromethylbenzonitrile 1-4 (500 mg, 2.673 mmol) was dissolved in DMF (8 mL), and methyl chloroacetate (318 mg, 2.941 mmol) and potassium carbonate (553 mg, 4.010 mmol) were added respectively, and stirred at 60°C for 12 h until the reaction was complete. The reaction mixture was cooled to room temperature, and water (20 ml) was added and filtered. The filter cake was dried to obtain compound 2-4 (400 mg, yield: 57.7%) as a yellow solid. LCMS (ESI) m/z = 260.0 [M + H] ⁺ .

Step 2: Synthesis of intermediate compound 3-4

The reaction was carried out in a 100 mL three-necked flask. The intermediate 2-4 (400 mg, 1.538 mmol) obtained in the previous step was dissolved in 6 mL THF solution, potassium tert-butoxide (190 mg, 1.698 mmol) was added, and stirred in an ice bath for 2 h. 20 mL of saturated aqueous ammonium chloride solution was added to the system, and extracted three times with 20 mL of ethyl acetate. After the organic phase was dried over anhydrous sodium sulfate, the reaction mixture was concentrated in vacuo to obtain compound 3-4 (240 mg, yield: 60.0%) as a yellow solid. ¹ H NMR (400 MHz, DMSO) δ8.50 (s, 1H), 7.82 (dd, J ₁ ＝8.8 Hz, J 2 ＝1.8 Hz, 1H) _, 7.72 (d, J＝8.8 Hz, 1H), 6.56 (s, 2H), 3.84 (s, 3H).

Step 3: Synthesis of intermediate compound 4-4

The reaction was carried out in a 100 mL three-necked flask. The intermediate compound 3-4 (240 mg, 0.926 mmol), 3-methylphenylboronic acid (250 mg, 1.853 mmol), copper acetate (84 mg, 0.463 mmol), and triethylamine (94 mg, 0.926 mmol) were dissolved in 5 mL DCM, replaced with an oxygen balloon three times, and stirred at room temperature for 12 h. The reaction solution was filtered, and the filtrate was concentrated to obtain a crude product. The crude product was purified by silica gel flash chromatography (PE: EA = 10: 1) to obtain compound 4-4 (200 mg, yield: 61.8%) as a yellow solid. LCMS (ESI) m/z = 350.0 [M + H] ⁺ .

Step 4: Synthesis of intermediate compound 5-4

The reaction was carried out in a 100 mL three-necked flask. Intermediate 4-4 (200 mg, 0.573 mmol) was added to THF (5 ml), the temperature was lowered to -15 ° C under nitrogen protection, and compound chlorosulfonyl isocyanate (122 mg, 0.865 mmol) was added and stirred for 1 hour. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction twice. The organic phase was concentrated to obtain compound 5-4 (220 mg, yield: 97.9%) as a white solid. LCMS (ESI) m/z = 393.0 [M + H] ⁺ .

Step 5: Synthesis of intermediate compound 6-4

The reaction was carried out in a 100 mL three-necked flask. Intermediate 5-4 (220 mg, 0.559 mmol) was dissolved in methanol (3 ml), sodium hydroxide (45 mg, 1.125 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. The pH was adjusted to 3-4 with 1 mol/L HCl, and compound 6-4 (50 mg, yield: 24.8%) was obtained by filtration. LCMS (ESI) m/z = 361.0 [M + H] ⁺ .

Step 6: Synthesis of compound 4

The reaction was carried out in a 10 mL microwave tube. Intermediate 6-4 (50 mg, 0.138 mmol) was dissolved in acetonitrile (1 ml), PyBOP (108 mg, 0.207 mmol) and DBU (42 mg, 0.276 mmol) were added, and the mixture was stirred at room temperature for 30 min. 0.3 ml of 2M methylamine tetrahydrofuran solution was added, and the mixture was reacted at 50°C for 1 hour in a microwave oven. Prep-TLC purification gave 80 mg of crude product, which was purified by Prep-HPLC (H ₂ O/ACN, 0.1% TFA) to give 6 mg of white solid 4. ¹ H NMR (400MHz, DMSO) δ8.84(s,1H),7.97(d,J＝8.8Hz,1H),7.92–7.86(m,1H),7.53(t,J＝7.6Hz,1H),7.46(d,J＝7.7Hz,1H),7.36–7.30(m,2H),6.45(s,1H ),3.01(d,J＝4.6Hz,3H),2.40(s,3H).LCMS(ESI)m/z＝374.2[M+H] ⁺ .

Referring to the synthesis method of Example 4 (Compound 4), the 2-hydroxy-5-trifluoromethylbenzonitrile in the first step of the synthesis step is replaced by 2-cyano-3-hydroxypyridine, and Compound 7 can be prepared accordingly.

Compound 7:

¹ H NMR (400MHz, DMSO) δ9.19 (s, 1H), 8.45 (dd, J ₁ = 4.6Hz, J ₂ = 1.1Hz, 1H), 8.21 (dd, J ₁ = 8.6Hz, J ₂ = 1.1 Hz, 1H), 7.56 (dd, J ₁ = 8.6Hz, J ₂ = 4.6Hz, 1H), 7.39 (t, J = 7.7Hz, 1H), 7.29 (d, J = 7.6Hz, 1H), 7.25– 7.20(m,2H),3.04(d,J＝3.6Hz,3H),2.37(s,3H).LCMS(ESI)m/z＝307.1[M+1] ⁺ .

Example 5. Synthesis of Compound 10

Step 1: Synthesis of intermediate compound 2-10

The reaction was carried out in a 250 mL three-necked flask. 3-cyano-2-fluoropyridine 1-10 (2.0 g, 16.4 mmol) was dissolved in tetrahydrofuran (20 mL). Methyl glycolate (1.62 g, 18.0 mmol) was dissolved in 10 mL THF under nitrogen protection and then added to the reaction solution. The reaction solution was cooled to 0°C, potassium tert-butoxide (5.5 g, 49.0 mmol) was slowly added to the reaction system, and stirred in an ice bath for 60 min. The reaction solution changed from colorless to yellow. After the reaction was complete, 100 mL of saturated ammonium chloride was added to the reaction solution and extracted three times with 100 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and then spin-dried and mixed. Purification was performed by silica gel flash chromatography (mobile phase: petroleum ether/ethyl acetate = 3:1). After spin-dried, compound 2-10 (0.7 g, yield: 23%) was obtained as a yellow solid. ¹ H NMR (400MHz, DMSO) δ8.47 (dd, J ₁ = 4.8Hz, J ₂ = 1.7Hz, 1H), 8.42 (dd, J ₁ = 7.8Hz, J ₂ = 1.8Hz, 1H), 7.37 (dd, J ₁ = 7.8Hz, J ₂ = 4.8Hz, 1H), 6.56 (s, 2H), 3.83 (s,3H).

Step 2: Synthesis of intermediate compound 3-10

The reaction was carried out in a 100 mL eggplant flask. The intermediate 2-10 (700 mg, 3.6 mmol) obtained in the previous step was dissolved in 7 mL of dichloromethane, and copper acetate (326 mg, 1.8 mmol), m-methylphenylboronic acid (493 mg, 3.6 mmol) and triethylamine (367 mg, 3.6 mmol) were added. The reaction was stirred for 24 h in an oxygen (14 psi) environment. The reaction solution was filtered, the mother liquor was dried by spin drying, and the crude product was purified by silica gel flash chromatography (20% ethyl acetate/petroleum ether) to obtain compound 3-10 (600 mg, yield: 58%) as a yellow solid. LCMS (ESI) m/z = 283.1 [M + H] ⁺ .

Step 3: Synthesis of intermediate compound 4-10

The reaction was carried out in a 100mL three-necked flask. Intermediate 3-10 (600mg, 2.1mmol) was added to 5ml tetrahydrofuran solution, replaced with nitrogen three times, then cooled to -15°C, chlorosulfonyl isocyanate (448mg, 3.1mmol) solution was added dropwise, and the reaction was kept at -15°C for 1 hour after the addition was completed. 2ml of purified water was added dropwise to the reaction solution, the pH value was adjusted to 13 with sodium hydroxide, and the temperature was raised to room temperature and stirred for 2 hours. Post-treatment: The reaction solution was poured into ice water (15ml), stirred for 0.5 hours, filtered to obtain the product, washed once with purified water and then directly freeze-dried to obtain 500mg of compound 4-10 as a white solid (purity 85%, yield 75%). LCMS (ESI) m/z = 312.1 [M + H] ⁺ .

Step 4: Synthesis of intermediate compound 5-10

The reaction was carried out in a 100 mL eggplant flask. Intermediate 4-10 (400 mg, 1.3 mmol) was weighed, sodium acetate (2.19 g, 15.0 mmol) was added, 4 mL of acetic anhydride was added, and the mixture was stirred at 80°C until the reaction was complete. Post-treatment: The reaction solution was directly spin-dried, 20 mL of dichloromethane was added to the reaction solution to dilute and filter, the filtrate was mixed with silica gel, and purified by Flash column (mobile phase: 0-53% ethyl acetate/petroleum ether) to obtain compound 5-10 (150 mg, yield: 39%) as a white solid. LCMS (ESI) m/z = 294.1 [M + H] ⁺ .

Step 5: Synthesis of compound 10

The reaction was carried out in a 10 mL microwave tube. Compound 5-10 (150 mg, 0.5 mmol), methylamine (1 ml, 2.0 mmol, 2 mol/L tetrahydrofuran solution), PyBOP (397 mg, 0.76 mmol) were dissolved in 3 ml of acetonitrile, and DBU (190 mg, 0.96 mmol) was slowly added dropwise at room temperature, purged with nitrogen three times, and the tube was sealed. The reaction temperature was raised to 50°C for 2 hours. Post-treatment: The reaction solution was spin-dried, extracted three times with ethyl acetate (50 ml×3), the organic layer was washed twice with saturated brine, dried over anhydrous sodium sulfate, filtered and concentrated to dryness, and prep-HPLC (H ₂ O:CAN, 0.1% NH ₃ .H ₂ O) was used to prepare 20.17 mg of 10 as a yellow solid (purity 99.0%, yield 6.5%). ¹ H NMR (400MHz, DMSO) δ8.67 (s, 1H), 8.50 (dd, J ₁ = 4.8Hz, J ₂ = 1.6Hz, 1H), 7.50 (t, J = 7.7Hz, 1H), 7.40 (d, J = 7.7Hz, 1H), 7.31 (dt, J ₁ = 11.2Hz, J ₂ = 6.6Hz, 3 H), 6.70 (dd, J ₁ = 7.9Hz, J ₂ = 1.6Hz, 1H), 2.98 (s, 3H), 2.40 (s, 3H). LCMS (ESI) m/z = 307.1 [M+H] ⁺ .

Referring to the synthesis method of Example 5 (Compound 10), 3-cyano-2-fluoropyridine in the first step of the synthesis step was replaced by 4-fluoronicotinonitrile and 3-fluoro-4-cyanopyridine, respectively, to prepare Compound 8 and Compound 9 accordingly.

Compound 8:

¹ H NMR (400MHz, DMSO) δ8.65(d,J＝4.6Hz,1H),8.61(d,J＝5.9Hz,1H),7.81(d,J＝5.9Hz,1H),7.53(t, J＝7.7Hz,1H),7.44(d,J＝10.3Hz,2H),7.35(s,1H),7.32(d,J＝8.2Hz,1H),2.98(d,J＝4.6Hz,3H) ,2.41(s,3H).LCMS(ESI)m/z＝307.0[M+1] ⁺ .

Compound 9:

¹ H NMR (400MHz, DMSO) δ9.17 (s, 1H), 9.13 (s, 1H), 8.36 (d, J = 5.4Hz, 1H), 7.52 (t, J = 7.6Hz, 1H), 7.44 ( d,J＝7.6Hz,1H),7.35–7.26(m,2H),6.24(d,J＝5.4Hz,1H),3.03(d,J＝3.4Hz,3H),2.41(s,3H). LCMS(ESI)m/z=307.1[M+H] ⁺ .

Example 6. Synthesis of Compound 14

Step 1: Synthesis of intermediate compound 2-14

Compound 2-1 (2.98 g, 13.2 mmol), 3-bromopyridine (2.09 g, 13.2 mmol), Pd ₂ (dba) ₃ (1.21 mg, 1.32 mmol), Xantphos (1.53 mg, 2.6 mmol), Cs ₂ CO ₃ (5.65 mg, 18.5 mmol) were dissolved in 30 mL of 1,4-dioxane and stirred at 110° C. overnight under nitrogen protection. The reaction solution was cooled to room temperature and filtered, and the filtrate was concentrated to obtain a crude product, which was purified by Flash column (mobile phase: 98% DCM + 2% MeOH) to obtain compound 2-14 (1.98 g, yield: 66%) as a white solid. ¹ H NMR (400MHz, DMSO) δ8.60(s,1H),8.37(d,J=2.6Hz,1H),8.20(dd, _J1 =4.6Hz, _J2 =1.2Hz,1H),7.73(d,J=8.9Hz,1H),7.57(dd, _J1 =8.9Hz, _J2 =2.2Hz,1H) ,7.38(m,1H),7.32–7.26(m,2H),3.82(s,3H).LCMS(ESI)m/z＝303.1[M+H] ⁺ .

Step 2: Synthesis of intermediate compound 3-14

Intermediate 2-14 (1.98 g, 6.53 mmol) was added to THF (20 mL), the temperature was lowered to -15°C under nitrogen protection, chlorosulfonyl isocyanate (1.38 g, 9.80 mmol) was added and stirred for 1 hour. Saturated NH ₄ Cl aqueous solution was added to quench the reaction, and the mixture was extracted with DCM:MeOH=10:1 (30 mL×2). The organic phase was washed with saturated NaCl, and the organic phase was concentrated to obtain a crude product of compound 3-14 (3.94 g) as a yellow solid. LCMS (ESI) m/z=346.0[M+H] ⁺ .

Step 3: Synthesis of intermediate compound 4-14

The crude intermediate 3-14 (3.94 g, 12.54 mmol) was dissolved in methanol (30 mL), sodium hydroxide (1.0 g, 25.10 mmol) was added, and the mixture was stirred at 60°C for 2 h under nitrogen protection. Methanol was removed by rotary evaporation, water was added, and the pH was adjusted to 3-4 with dilute hydrochloric acid, and the crude product was filtered and purified by Flash column (mobile phase: 98% DCM + 2% MeOH) to obtain compound 4-14 (280 mg, two-step yield: 14%) as a white solid. ¹ H NMR (400MHz, DMSO) δ12.08 (s, 1H), 8.92–8.82 (m, 2H), 8.18–8.12 (m, 1H), 7.89 (d, J = 9.0Hz, 1H), 7.76 (dd, J ₁ = 8.1Hz, J ₂ = 4.8Hz, 1H), 7.64 (dd, J ₁ = 9.0 Hz,J ₂ =2.2Hz,1H),6.02(d,J=2.1Hz,1H).LCMS(ESI)m/z=314.0[M+H] ⁺ .

Step 4: Synthesis of compound 14

The reaction was carried out in a 10 mL microwave tube. Compound 4-14 (30 mg, 0.096 mmol) was dissolved in acetonitrile (5 mL), and DBU (29 mg, 0.191 mmol), PyBOP (75 mg, 0.144 mmol), and 0.25 mL (2 M) of methylamine tetrahydrofuran solution were added, and the mixture was reacted at 50°C for 1 h. Purification by Prep-HPLC (H ₂ O:ACN, 0.1% FA) gave 4.5 mg of white solid 14, with a yield of 14.3%. ¹ HNMR (400MHz, MeOD) δ8.82 (d, J = 4.8Hz, 1H), 8.77 (d, J = 2.2Hz, 1H), 8.12–8.04 (m, 1H), 7.76 (dd, J ₁ = 8.1Hz, J ₂ = 4.9Hz, 1H), 7.68 (d, J = 9.0Hz, 1H), 7.54 (dd, J ₁ =9.0, J ₂ =2.1Hz, 1H), 6.31 (d, J = 2.1Hz, 1H), 3.14 (s, 3H). LCMS (ESI) m/z = 326.9 [M+H] ⁺ .

Referring to the method of Example 6 (Compound 14), the methylamine tetrahydrofuran solution in the fourth step of the synthesis step was replaced by dimethylamine tetrahydrofuran solution, methylethylamine, cyclopropylamine, 2,2-difluoroethylamine, 2,2,2-trifluoroethylamine, and (1S,2R)-2-fluorocyclopropylamine, and 15, 16, 23, 52, 53, and 68 were prepared accordingly.

Compound 15:

¹ H NMR (400MHz, DMSO) δ8.82 (dd, J ₁ = 4.8Hz, J ₂ = 1.2Hz, 1H), 8.73 (d, J = 2.4Hz, 1H), 8.04–8.00 (m, 1H), 7.85(d,J＝9.2Hz,1H),7.71(dd, _J1 ＝8.2Hz, _J2 ＝4.8Hz,1H) _, 7.62(dd,J1＝9.0Hz, _J2 ＝2.4Hz,1H), 6.05(d,J＝2.0Hz,1H),3.61(s,3H),3.24(s,3H).LCMS(ESI)m/z＝341.0[M+H] ⁺ .

Compound 16:

¹ H NMR (400MHz, DMSO) δ8.83 (d, J = 3.6Hz, 1H), 8.76 (d, J = 1.6Hz, 1H), 8.05 (dd, J ₁ = 6.0Hz, J ₂ = 2.0Hz, 1H), 7.86 (d, J = 9.2Hz, 1H), 7.73 (dd, J ₁ = 8.0Hz, J ₂ = 4.8Hz, 1H), 7.62 (dd, J ₁ = 8.8Hz, J ₂ ＝2.4Hz,1H),6.07(d,J＝2.0Hz,1H),4.03(s,1H),3.77(s,1H),3.59(s,1H),3.24(s,2H),1.28(d ,J＝36.8Hz,3H).LCMS(ESI)m/z＝355.1[M+H] ⁺ .

Compound 23:

¹ H NMR (400MHz, MeOD) δ8.93 (d, J = 4.7Hz, 1H), 8.89 (s, 1H), 8.21 (d, J = 8.3Hz, 1H), 7.87 (dd, J ₁ = 8.1Hz , J ₂ =5.0Hz, 1H), 7.77 (d, J = 9.0Hz, 1H), 7.67 (dd, J ₁ = 9.1Hz, J ₂ = 2.0Hz, 1H), 6.35 (d, J = 1.9Hz, 1H),3.07(m,1H),1.11-1.14(m,2H),1.01–0.92(m,2H).LCMS(ESI)m/z＝353.0[M+H] ⁺ .

Compound 52:

¹ H NMR (400MHz, DMSO) δ9.14 (s, 1H), 8.85–8.80 (m, 1H), 8.79 (d, J = 2.3Hz, 1H), 8.10–8.04 (m, 1H), 7.85 (d ,J＝9.0Hz,1H),7.73(dd,J ₁ ＝8.1Hz,J ₂ ＝4.8Hz,1H),7.65(dd,J ₁ ＝9.0,J ₂ ＝2.2Hz,1H),6.26(tt, J ₁ =55.6Hz, J ₂ =3.9Hz, 1H), 6.16 (d, J = 2.0Hz, 1H), 3.92 (t, J = 15.1Hz, 2H). LCMS (ESI) m/z = 377.0[M +1] ⁺ .

Compound 53:

¹ H NMR (400MHz, DMSO) δ9.37 (t, J = 6.4Hz, 1H), 8.82 (d, J = 4.4Hz, 1H), 8.79 (s, 1H), 8.07 (d, J = 8.0Hz, 1H), 7.85 (d, J = 9.0Hz, 1H), 7.72 (dd, J ₁ = 7.6Hz, J ₂ = 5.2Hz, 1H), 7.66 (d, J = 9.2Hz, 1H), 6.15 (s, 1H),4.42–4.31(m,2H).LCMS(ESI)m/z＝395.0[M+1] ⁺ .

Compound 68:

Referring to the method of Example 6 for compound 14, the 3-bromopyridine in the first step of the synthesis step was replaced by 2-methyl-3-bromopyridine, 5-bromo-3-methylpyridine, and 5-bromo-2-methylpyridine, and the methylamine tetrahydrofuran solution in the fourth step was replaced by cyclopropylamine, and compounds 47, 49, and 50 were prepared accordingly.

Compound 47:

¹ H NMR (400MHz, DMSO) δ8.86 (s, 1H), 8.71 (dd, J ₁ = 4.8Hz, J ₂ = 1.3Hz, 1H), 7.92 (dd, J ₁ = 7.9Hz, J ₂ = 1.3 Hz, 1H), 7.78 (d, J ₁ = 9.0Hz, 1H), 7.60 (dd, J ₁ = 9.0Hz, J ₂ = 2.1Hz, 1H), 7.53 (dd, J ₁ = 7.8Hz, J ₂ = 4.8Hz,1H),6.00(d,J＝2.0Hz,1H),3.23–3.01(m,1H),2.29(s,3H),0.81(d,J＝7.6Hz,2H),0.74(dd, J ₁ =7.6Hz,J ₂ ＝3.7Hz,2H).LCMS(ESI)m/z＝367.10[M+1] ⁺ .

Compound 49:

¹ H NMR (400MHz, DMSO) δ8.81 (d, J = 4.4Hz, 1H), 8.58 (d, J = 2.4Hz, 1H), 7.88 (dd, J ₁ = 8.4Hz, J ₂ = 2.4Hz, 1H), 7.78 (d, J = 8.8Hz, 1H), 7.61 (dd, J ₁ = 9.2Hz, J ₂ = 2.0Hz, 1H), 7.54 (d, J = 8.0Hz, 1H), 6.23 (d, J＝1.6Hz,1H),3.08(m,1H),2.64(s,3H),0.86–0.77(m,2H),0.74–0.64(m,2H).LCMS(ESI)m/z＝367.0[ M+1] ⁺ .

Compound 50:

¹ H NMR (400MHz, DMSO) δ8.83 (d, J = 4.4 Hz, 1H), 8.65 (s, 1H), 8.54 (s, 1H), 7.86 (s, 1H), 7.78 (d, J = 9.2 Hz,1H),7.60(d,J＝9.2Hz,1H),6.17(s,1H),3.09(d,J＝4.4Hz,1H),2.43(s,3H),0.80(m,2H), 0.75–0.68(m,2H).LCMS(ESI)m/z＝367.2[M+1] ⁺ .

Example 7. Synthesis of Compound 11

Step 1: Synthesis of intermediate compound 2-11

The reaction was carried out in a 250 mL eggplant flask. 5-Chloro-2-fluorobenzonitrile 1-11 (10 g, 65.7 mmol) was dissolved in DMSO (100 mL), and glycine methyl ester hydrochloride (16 g, 98 mmol) and potassium carbonate (27.2 g, 196 mmol) were added, and stirred at 100 ° C for 12 h until the reaction was complete. Water (90 ml) was added and filtered, and the filter cake was dried to obtain the product compound 2-11 (4.3 g, yield: 43%) as a white solid. ¹ H NMR (400 MHz, DMSO) δ10.63 (s, 1H), 7.87 (d, J＝1.7 Hz, 1H), 7.25–7.15 (m, 2H), 5.74 (s, 2H), 3.82 (s, 3H).

Step 2: Synthesis of intermediate compound 3-11

The reaction was carried out in a 250 mL single-mouth bottle. Compound 2-11 (4.3 g, 19.1 mmol), meta-iodotoluene (4.97 g, 22.9 mmol), palladium acetate (0.42 g, 1.87 mmol), BINAP (1.18 g, 1.89 mmol), cesium carbonate (8.7 g, 26.7 mmol) were dissolved in 50 mL toluene, replaced with nitrogen three times, and reacted at 110 ° C for 16 hours. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by Flash column (PE/EA=2:1) to obtain the product compound 3-11 (0.9 g, yield: 20.9%) as a yellow solid. ¹ H NMR (400MHz, DMSO) δ11.64(s,1H),7.67(s,1H),7.42(d,J＝8.8Hz,1H),7.29–7.24(m,1H),7.23(d,J＝2.0Hz,1H),7.04(s,1H),6.68(s,1H),6.61(d,J＝ 7.8Hz,2H),3.83(s,3H),2.20(s,3H).

Step 3: Synthesis of Compound 4-11

The reaction was carried out in a 100 mL three-necked flask. Intermediate 3-11 (300 mg, 0.952 mmol) was added to THF (5 ml), the temperature was lowered to -15 ° C under nitrogen protection, and compound chlorosulfonyl isocyanate (268 mg, 1.90 mmol) was added and stirred for 1 hour. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and the mixture was extracted twice with EA (20 ml). After drying over anhydrous sodium sulfate, the organic phase was concentrated to obtain compound 4-11 (200 mg, yield: 66.6%) as a white solid. LCMS (ESI) m/z = 358.0 [M + H] ⁺ .

Step 4: Synthesis of Compound 5-11

The reaction was carried out in a 100 mL single-mouth bottle. Intermediate 4-11 (200 mg, 0.559 mmol) was dissolved in methanol (3 ml), sodium hydroxide (45 mg, 1.125 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction twice. The organic phase was dried over anhydrous sodium sulfate and concentrated to obtain a white solid compound 5-11 (150 mg, yield: 68%). LCMS (ESI) m/z = 326.1 [M + H] ⁺ .

Step 5: Synthesis of compound 11

The reaction was carried out in a 50 mL single-mouth bottle. Compound 5-11 (150 mg, 0.460 mmol) was dissolved in acetonitrile (3 ml), and PyBOP (358 mg, 0.688 mmol), DBU (140 mg, 0.915 mmol), and 0.5 ml (2 M) of methylamine tetrahydrofuran solution were added. The reaction was carried out at room temperature for 2 h. Purification by Prep-HPLC (H ₂ O:ACN, 0.1% TFA) gave 14.51 mg of white solid 11, with a yield of 9.6%. ¹ H NMR (400MHz, DMSO) δ11.42(s,1H),8.82–8.47(m,1H),7.66(d,J＝8.9Hz,1H),7.54(t,J＝7.7Hz,1H),7.46(d,J＝7.8Hz,1H),7.35(m,J＝8.9,1H),7.30(s, 1H),7.27(d,J＝7.7Hz,1H),5.94(d,J＝1.8Hz,1H),3.12(d,J＝4.1Hz,3H),2.42(s,3H).LCMS(ESI)m/z＝339.0[M+H] ⁺ .

Referring to the method of Example 7 for compound 11, the m-iodotoluene in the second step of the synthesis process is replaced by 2-methyl-3-bromopyridine, and compound 57 can be prepared accordingly.

Compound 57:

¹ H NMR (400MHz, DMSO) δ11.20(s,1H),8.70–8.65(m,1H),7.92(s,1H),7.81(d,J＝6.8Hz,1H),7.62(d,J =8.9Hz, 1H), 7.51 (dd, J ₁ = 7.8, J ₂ = 4.8Hz, 1H), 7.27 (m, J = 8.8Hz, 1H), 5.85 (d, J = 1.9Hz, 1H), 3.04 (s,3H),2.20(s,3H).LCMS(ESI)m/z＝340.0[M+1] ⁺ .

Example 8. Synthesis of Compound 27

Step 1: Synthesis of intermediate compound 2-27

The reaction was carried out in a 10mL microwave tube. Compound 1-A (327mg, 1mmol) was dissolved in acetonitrile (10mL), DBU (304mg, 2mmol) and PyBOP (780mg, 1.5mmol) were added and stirred for 15min, then 1-tert-butyloxycarbonyl-3-aminocyclobutylamine (344mg, 2mmol) was added to the reaction solution, and the reaction was carried out at 50°C for 1h. Post-treatment: The reaction solution was slowly poured into water (25ml), extracted twice with ethyl acetate, the organic phases were combined, the organic layer was washed with water and saturated brine again, dried over anhydrous sodium sulfate, filtered, mixed with silica gel, passed through a Flash column, and the eluent (DCM:MeOH=20:1) to obtain compound 2-27 (125mg, yield: 26.0%) as a white solid. LCMS (ESI) m/z=481.1[M+H] ⁺ .

Step 2: Synthesis of compound 27

The reaction was carried out in a 25 mL single-mouth bottle. Compound 2-27 (60 mg, 0.12 mmol) was added to a 1,4-dioxane hydrochloride solution (8 mL) and stirred at room temperature overnight. Post-treatment: The reaction solution was concentrated to dryness under reduced pressure. Prep-HPLC (H ₂ O:ACN, 0.1% NH ₄ OH) purification was performed to obtain 27 (11.46 mg, yield: 24.1%) as a white solid. ¹ H NMR (400MHz, DMSO) δ9.14 (s, 1H), 7.78 (d, J = 8.8Hz, 1H), 7.59 (dd, J ₁ = 8.8Hz, J ₂ = 2.0Hz, 1H), 7.52 (t, J = 7.6Hz, 1H), 7.44 (d, J = 8.0Hz, 1H), 7.33-7.25 (m, 2H), 6.08 (d, J＝2.0Hz, 1H), 4.99–4.88 (m, 1H), 3.74 (d, J＝7.2Hz, 4H), 2.41 (s, 3H). LCMS (ESI) m/z＝381.0[M+H] ⁺ .

Referring to the synthesis method of compound 27 in Example 8, the 1-tert-butoxycarbonyl-3-aminocyclobutylamine in the first step of the synthesis step is replaced by 1-tert-butoxycarbonyl-3-aminocyclopentylamine, and the compound 32 can be prepared accordingly.

Compound 32:

¹ H NMR (400MHz, DMSO) δ8.79 (s, 1H), 7.79 (d, J = 8.8Hz, 1H), 7.61 (dd, J ₁ = 8.8Hz, J ₂ = 2.0Hz, 1H), 7.54 ( t,J＝7.6Hz,1H),7.46(d,J＝7.6Hz,1H),7.34–7.27(m,2H),6.10(d,J＝1.6Hz,1H),4.75(s,1H), 3.52–3.40(m,2H),3.27–3.12(m,3H),2.42(s,3H),2.25(dd,J ₁ =12.8Hz,J ₂ ＝6.8Hz,1H),2.07(d,J＝6.8Hz,1H).LCMS(ESI)m/z＝395.0[M+H] ⁺ .

Example 9. Synthesis of Compound 12

Step 1: Synthesis of Compound 2-11

The reaction was carried out in a 250 mL eggplant flask. 5-Chloro-2-fluorobenzonitrile 1-11 (7.0 g, 46.0 mmol) was dissolved in DMSO (70 mL), and glycine methyl ester hydrochloride (11.2 g, 68.7 mmol) and potassium carbonate (19.0 g, 137 mmol) were added, and stirred at 100 ° C for 12 h until the reaction was complete. Water (90 ml) was added and filtered, and the filter cake was dried to obtain the product compound 2-11 (2.6 g, yield: 37%) as a black solid. ¹ H NMR (400 MHz, DMSO) δ10.63 (s, 1H), 7.87 (d, J＝1.7 Hz, 1H), 7.25–7.15 (m, 2H), 5.74 (s, 2H), 3.82 (s, 3H).

Step 2: Synthesis of compound 3-11

The reaction was carried out in a 250 mL single-mouth bottle. Compound 2-11 (2.6 g, 11.5 mmol), meta-iodotoluene (3.7 g, 16.9 mmol), palladium acetate (0.25 g, 1.15 mmol), 1,1'-binaphthyl-2,2'-bis(diphenylphosphine) BINAP (1.43 g, 2.3 mmol), cesium carbonate (5.2 g, 16.1 mmol) were dissolved in 50 mL toluene, replaced with nitrogen three times, and reacted at 110 ° C for 16 hours. After the reaction was completed, the concentrated reaction solution was added with dichloromethane silica gel to mix the sample, and passed through a Flash column (PE/EA=2:1) to obtain the product compound 3-11 (0.4 g, yield: 15.3%), a yellow solid. ¹ H NMR (400MHz, DMSO) δ11.64(s,1H),7.67(s,1H),7.42(d,J＝8.8Hz,1H),7.29–7.24(m,1H),7.23(d,J＝2.0Hz,1H),7.04(s,1H),6.68(s,1H),6.61(d,J＝ 7.8Hz,2H),3.83(s,3H),2.20(s,3H).

Step 3: Synthesis of Compound 4-12

The reaction was carried out in a 50 mL single-mouth bottle. Compound 3-11 (400 mg, 1.26 mmol), potassium carbonate (526 mg, 3.8 mmol), iodomethane (268 mg, 1.9 mmol) were dissolved in 5 mL DMF and reacted at room temperature for 2 hours. After the reaction was completed, the concentrated reaction solution was added with water (20 ml), extracted three times with ethyl acetate (60 ml), washed with saturated brine (60 ml), concentrated, dried over anhydrous sodium sulfate, concentrated, added with dichloromethane silica gel, and mixed with the sample. The sample was passed through a Flash column (PE/EA=2:1) to obtain compound 4-12 (0.26 g, yield: 65%) as a yellow solid. ¹ H NMR (400MHz, DMSO) δ7.71(s,1H),7.64(d,J＝9.0Hz,1H),7.35(dd,J＝9.0,2.0Hz,1H),7.23(d,J＝1.9Hz,1H),7.04(t,J＝7.7Hz,1H),6.67(s,1H),6.60(t,J =7.8Hz,2H),3.95(s,3H),3.80(s,3H),2.19(s,3H).

Step 4: Synthesis of Compound 5-12

The reaction was carried out in a 50 mL three-necked flask. Intermediate 4-12 (260 mg, 0.79 mmol) was added to THF (5 ml), the temperature was lowered to -15 ° C under nitrogen protection, and chlorosulfonyl isocyanate CSI (222 mg, 1.57 mmol) was added and stirred for 1 hour. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction three times. The organic phase was concentrated to obtain compound 5-12 (228 mg, yield: 66.6%) as a white solid. LCMS (ESI) m/z = 372.1 [M + H] ⁺ .

Step 5: Synthesis of Compound 6-12

The reaction was carried out in a 100 mL single-mouth bottle. Intermediate 5-12 (200 mg, 0.559 mmol) was dissolved in methanol (3 ml), sodium hydroxide (43 mg, 1.07 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction twice. The organic phase was concentrated to obtain compound 6-12 (150 mg, yield: 75%) as a white solid. LCMS (ESI) m/z = 340.0 [M + H] ⁺ .

Step 6: Synthesis of Compound 12

The reaction was carried out in a 50 mL single-mouth bottle. Compound 6-12 (100 mg, 0.29 mmol) was dissolved in acetonitrile (3 ml), and PyBOP (229 mg, 0.44 mmol), DBU (90 mg, 0.58 mmol), and 0.58 ml (2 M) of methylamine tetrahydrofuran solution were added. The reaction was carried out at room temperature for 2 h. Purification by Prep-HPLC (H ₂ O:ACN, 0.1% TFA) gave 14.51 mg of white solid 12, with a yield of 14.5%. ¹ HNMR (400MHz, DMSO) δ7.67 (d, J = 9.1Hz, 1H), 7.51 (t, J = 7.7Hz, 1H), 7.42 (d, J = 6.5Hz, 2H), 7.35 (dd, J ₁ = 9.0, J ₂ = 2.0Hz, 1H), 7.22 (s, 1H), 7.18 (d, J = 7.7Hz ,1H),5.88(d,J＝1.9Hz,1H),4.04(s,3H),3.02(s,3H),2.40(s,3H).LCMS(ESI)m/z＝353.0[M+H] ⁺ .

Referring to the synthesis method of compound 12 in Example 9, the m-iodotoluene in the second step of the synthesis process is replaced by 2-methyl-3-bromopyridine, and compound 71 can be prepared accordingly.

Compound 71:

¹ H NMR (400MHz, DMSO) δ8.78–8.72(m,1H),7.99(s,1H),7.93–7.87(m,1H),7.76(d,J＝9.1Hz,1H),7.59(m ,J＝7.8Hz,1H),7.43(dd,J ₁ ＝9.1,J ₂ ＝2.0Hz,1H),5.80(d,J＝1.8Hz,1H),4.09(s,3H),3.10(s, 3H),2.25(s,3H).LCMS(ESI)m/z＝354.1[M+1] ⁺ .

Referring to the synthesis method of compound 12 in Example 9, the m-iodotoluene in the second step of the synthesis process is replaced by 2-methyl-3-bromopyridine, and the methylamine tetrahydrofuran solution in the sixth step is replaced by cyclopropylamine, and compound 87 can be prepared accordingly.

Compound 87:

¹ H NMR (400MHz, DMSO) δ8.78 (d, J = 3.4Hz, 1H), 8.33 (brs, 1H), 7.96 (d, J = 6.7Hz, 1H), 7.74 (d, J = 7.7Hz, 1H),7.60(m,J=7.8,1H),7.45(s,1H),5.78(s,1H),4.05(s,3H),3.10(s,1H),2.29(s,3H),0.94 (d,J＝6.6Hz,2H),0.82(s,2H).LCMS(ESI)m/z＝380.1[M+1] ⁺ .

Example 10. Synthesis of Compound 44

Step 1: Synthesis of Compound 2-44

The reaction was carried out in a 250 mL single-mouth bottle. Compound 2-1 (2.25 g, 10 mmol), m-bromophenylboronic acid (3 g, 15 mmol), copper acetate (0.9 g, 5 mmol), triethylamine (1.01 g, 10 mmol) were dissolved in 30 mL of dichloromethane, oxygen was replaced three times, and the reaction was carried out at room temperature for 16 hours. After the reaction was completed, the concentrated reaction solution was added with dichloromethane silica gel to mix the sample, and passed through a Flash column (PE/EA=5:1) to obtain the product compound 2-44 (1.8 g, yield: 80%), a yellow solid. ¹ H NMR (400MHz, CDCl ₃ ) δ7.42 (dt, J = 8.9, 5.4Hz, 2H), 7.30–7.27 (m, 2H), 7.26 (s, 1H), 7.21 (t, J = 7.9Hz, 1H), 7.04 (d, J = 7.8Hz, 1H), 4.00 (s, 3H).

Step 2: Synthesis of Compound 3-44

The reaction was carried out in a 100 mL three-necked flask. Intermediate 2-44 (1.8 g, 4.76 mmol) was added to THF (20 ml), the temperature was lowered to -15 ° C under nitrogen protection, CSI (1.34 g, 9.5 mmol) was added and stirred for 1 hour. Ammonium chloride aqueous solution (20 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction three times. The organic phase was concentrated to obtain compound 3-44 (2 g, yield: 95%) as a white solid. ¹ H NMR (400MHz, DMSO) δ7.83(d,J＝8.9Hz,1H),7.63–7.55(m,2H),7.49(d,J＝1.8Hz,1H),7.34(d,J＝3.5Hz,1H),7.24(d,J＝5.8Hz,2H),6.54(s,2H),3.85(s, 3H).LCMS(ESI)m/z＝424.9[M+H] ⁺ .

Step 3: Synthesis of Compound 4-44

The reaction was carried out in a 100 mL single-mouth bottle. Intermediate 3-44 (2 g, 4.75 mmol) was dissolved in methanol (3 ml), sodium hydroxide (0.38 g, 9.5 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. Ammonium chloride aqueous solution (5 ml) was added to quench, water (20 ml) was added, and EA (20 ml) was used for extraction twice. The organic phase was concentrated to obtain a white solid compound 4-44 (1.6 g, yield: 80%). LCMS (ESI) m/z = 392.9 [M + H] ⁺ .

Step 4: Synthesis of Compound 5-44

The reaction was carried out in a 50 mL single-mouth bottle. Compound 4-44 (500 mg, 1.27 mmol), anhydrous potassium ferrocyanide (110 mg, 0.298 mmol), palladium acetate (30 mg, 0.137 mmol), dppf (140 mg, 0.252 mmol), sodium carbonate (134 mg, 0.123 mmol) were dissolved in 5 mL N-methylpyrrolidone and reacted at 120 ° C for 16 hours. After the reaction was completed, water (20 ml) was added, and EA (20 ml) was used for extraction three times. The organic phase was concentrated, and dichloromethane and silica gel were added to the concentrated reaction solution to mix the sample, and the product compound 5-44 (0.2 g, yield: 40%) was obtained as a black solid through a Flash column (PE/EA=1:5). LCMS (ESI) m/z=337.9[M+H] ⁺ .

Step 5: Synthesis of compound 44

The reaction was carried out in a 50 mL single-mouth bottle. Compound 5-44 (100 mg, 0.295 mmol) was dissolved in DMF (1 ml), and PyBOP (230 mg, 0.442 mmol), DBU (90 mg, 0.588 mmol), and cyclopropylamine (33 mg, 0.578 mmol) were added. The reaction was carried out at room temperature for 4 h. Purification by Prep-HPLC (H ₂ O:ACN, 0.1% FA) gave 6.37 mg of white solid 44, with a yield of 6.3%. ¹ H NMR (400MHz, DMSO) δ8.85 (s, 1H), 8.14 (s, 1H), 8.11 (d, J = 7.6Hz, 1H), 7.93 (d, J = 8.1Hz, 1H), 7.86 (t, J = 7.8Hz, 1H), 7.78 (d, J = 8.9Hz, 1H), 7.61 (m, J = 9.0 Hz,1H),6.15(d,J＝1.8Hz,1H),3.09(s,1H),0.84–0.77(m,2H),0.73(t,J＝3.7Hz,2H).LCMS(ESI)m/z＝377.0[M+H] ⁺ .

Example 11. Synthesis of Compound 22

Step 1: Synthesis of Compound 2-22

The reaction was carried out in a 100 mL three-necked flask. 2-Cyano-3-hydroxypyridine 1-22 (5 g, 41.63 mmol) was dissolved in 50 mL ACN:H ₂ O (5:1), NBS (8.15 g, 45.79 mmol) was dissolved in 50 mL ACN:H ₂ O (5:1) solution at 0°C, and slowly added dropwise to the reaction solution, and reacted at low temperature for 6 h. After the reaction was completed, H ₂ O (50 mL × 3) was added to the reaction solution, and then EA (100 mL × 3) was added to extract, the organic phase was dried with saturated NaCl solution and concentrated, and then purified by Flash column (mobile phase: 99% DCE + 1% MeOH) to obtain crude compound 2-22 (3.5 g, yield: 43%) as a white solid. LCMS (ESI) m/z = 199.0 [M + H] ⁺ .

Step 2: Synthesis of compound 3-22

The reaction was carried out in a 100 mL single-mouth bottle. Compound 2-22 (3.5 g, 17.59 mmol), methyl chloroacetate (2.1 g, 19.35 mmol), potassium carbonate (4.86 g, 35.17 mmol) were dissolved in DMF (30 mL), and reacted at 80 ° C for 6 h under nitrogen protection. After the reaction, water (100 mL) was added to precipitate solids, and the solids were filtered to obtain compound 3-22 (2.3 g, yield: 49%) as a white solid. ¹ HNMR (400 MHz, DMSO) δ8.01 (d, J = 8.8 Hz, 1H), 7.73 (d, J = 8.8 Hz, 1H), 6.36 (s, 2H), 3.83 (s, 3H). LCMS (ESI) m/z = 271.0 [M + H] ⁺ .

Step 3: Synthesis of Compound 4-22

The reaction was carried out in a 100 mL three-necked flask. Compound 3-22 (2.28 g, 8.4 mmol), 3-methylphenylboronic acid (2.28 g, 16.8 mmol), copper acetate (840 mg, 4.21 mmol), triethylamine (1.9 g, 16.8 mmol), molecular sieve (0.2 g) were dissolved in 50 mL DCM, oxygen was replaced with an oxygen balloon, and stirred at room temperature for 12 h. The reaction solution was filtered, and the filtrate was concentrated to obtain a crude product, which was purified by Flash column (mobile phase: 80% DCM + 20% PE) to obtain compound 4-22 (1.14 g, yield: 35%) as a yellow solid. ¹ H NMR (400MHz, DMSO) δ8.27(s,1H),8.13(d,J＝8.8Hz,1H),7.74(d,J＝8.8Hz,1H),7.10(t,J＝7.8Hz,1H),6.96(s,1H),6.89(d,J＝8.0Hz,1H),6.76(d,J＝7.5Hz ,1H),3.85(s,3H),2.24(s,3H).LCMS(ESI)m/z＝361.0[M+H] ⁺ .

Step 4: Synthesis of compound 5-22

The reaction was carried out in a 100 mL three-necked flask. Compound 4-22 (1.14 g, 3.17 mmol) was added to THF (10 mL), the temperature was lowered to -15°C under nitrogen protection, CSI (673 mg, 4.75 mmol) was added and stirred for 1 hour. After the reaction was completed, saturated NH ₄ Cl solution was added to quench the reaction, and the mixture was extracted with EA (50 mL×3). The organic phase was washed with saturated NaCl, and the organic phase was concentrated to obtain crude 5-22 (1.48 g) as a yellow oil. LCMS (ESI) m/z=406.1[M+H] ⁺ .

Step 5: Synthesis of compound 6-22

The reaction was carried out in a 100 mL three-necked flask. The crude product 5-22 (1.48 g, 3.66 mmol) was dissolved in methanol (30 mL), sodium hydroxide (293 mg, 7.33 mmol) was added, and the mixture was stirred at 60 ° C for 2 h under nitrogen protection. Methanol was removed by rotary evaporation, water was added, and the pH was adjusted to 3-4 with dilute hydrochloric acid, and the crude product was filtered and slurried to obtain compound 6-22 (850 mg, yield: 63%) as a white solid. ¹ HNMR (400MHz, DMSO) δ12.03 (s, 1H), 8.24 (d, J = 8.8Hz, 1H), 7.75 (d, J = 8.8Hz, 1H), 7.40 (t, J = 7.4Hz, 1H), 7.32 (dd, J ₁ = 14.4Hz, J ₂ = 7.8Hz, 3H), 2.38 (s, 3H) .LCMS(ESI)m/z=373.9[M+H] ⁺ .

Step 6: Synthesis of compound 7-22

The reaction was carried out in a 100 mL single-mouth bottle. Compound 6-22 (430 mg, 1.16 mmol), CuI (0.66 g, 3.47 mmol), MFSDA (0.67 g, 1.08 mmol), triethylamine (0.35 g, 3.46 mmol) were dissolved in DMF (5 mL), and reacted at 80 ° C for 16 h under nitrogen protection. The reaction solution was cooled to room temperature and poured into water (50 mL), extracted with ethyl acetate three times (50 mL/time), and the organic phases were combined, washed with water three times (50 mL/time), washed with saturated brine once (50 mL), dried over anhydrous sodium sulfate, and purified by column chromatography (DCM: MeOH = 20: 1) to obtain 260 mg of crude compound 7-22. LCMS (ESI) m/z = 361.9 [M + H] ⁺ .

Step 7: Synthesis of compound 22

The crude compound 7-22 (220 mg, 0.588 mmol) was dissolved in acetonitrile (5 mL), and DBU (179 mg, 1.18 mmol), PyBOP (459 mg, 0.88 mmol), and 1.25 mL (2 M) of methylamine tetrahydrofuran solution were added, and the mixture was reacted at 50° C. for 1 h under microwave conditions. Purification by Prep-HPLC (H ₂ O:ACN, 0.1% TFA) gave 11.55 mg of white solid 22. ¹ H NMR (400MHz, DMSO) δ8.82–8.73(m,1H),8.44(d,J＝8.8Hz,1H),8.01(d,J＝8.8Hz,1H),7.35(t,J＝7.6Hz,1H),7.29–7.23(m,2H),7.22(d,J＝8.0Hz,1H),3. 00(d,J＝4.6Hz,3H),2.34(s,3H).LCMS(ESI)m/z＝375.1[M+H] ⁺ .

Example 12. Synthesis of Compound 13

Step 1: Synthesis of Compound 4-13

The reaction was carried out in a 100 mL three-necked flask. Compound 2-1 (2.00 g, 8.89 mmol), compound 3-13 (3.20 g, 9.78 mmol), Pd ₂ (dba) ₃ (0.81 g, 0.89 mmol), Xantphos (1.03 g, 1.78 mmol), Cs ₂ CO ₃ (4.33 g, 13.33 mmol) were added to 45 mL of 1,4-dioxane, nitrogen was protected, and the reaction was carried out at 100 ° C for 12 hours until the reaction was completed. 500 ml of water was added to the reaction solution, and 300 ml of EA was used to extract three times. The organic phase was concentrated, and the crude product was purified by silica gel flash chromatography (PE: EA = 10: 1) to obtain intermediate 4-13 (1.50 g, yield: 35.7%) as a yellow solid. LCMS (ESI) m/z = 472.0 [M + H] ⁺ .

Step 2: Synthesis of Compound 5-13

The reaction was carried out in a 100 mL three-necked flask. Intermediate 4-13 (400 mg, 0.847 mmol) was added to THF (8 ml), the temperature was lowered to -15 ° C under nitrogen protection, CSI (179 mg, 1.270 mmol) was added and stirred for 1 hour. Ammonium chloride aqueous solution (8 ml) was added to quench, water (30 ml) was added, and EA (30 ml) was used for extraction twice. The organic phase was concentrated to obtain compound 5-13 (400 mg, yield: 91.7%) as a white solid. LCMS (ESI) m/z = 515.2 [M + H] ⁺ .

Step 3: Synthesis of Compound 6-13

The reaction was carried out in a 100 mL three-necked flask. Intermediate 5-13 (400 mg, 0.776 mmol) was dissolved in methanol (8 ml), sodium hydroxide (62.1 mg, 1.552 mmol) was added and stirred at 60 ° C for 2 h under nitrogen protection. The pH was adjusted to 3-4 with 1 mol/L HCl, and the crude product was filtered to obtain the crude product, which was purified by silica gel flash chromatography (DCM: MeOH = 20: 1) to obtain compound 6-13 (260 mg, yield: 69.5%). LCMS (ESI) m/z = 483.2 [M + H] ⁺ .

Step 4: Synthesis of Compound 7-13

The reaction was carried out in a 10 mL sealed tube. The intermediate 6-13 (260 mg, 0.538 mmol) was dissolved in acetonitrile (3 ml), PyBOP (420 mg, 0.807 mmol) was added, DBU (164 mg, 1.076 mmol), 1.6 ml (2 M) of methylamine tetrahydrofuran solution was added, and the reaction was carried out at room temperature for 1 hour. After concentration under reduced pressure, Prep-TLC (DCM: MeOH = 5: 1) was purified to obtain 300 mg of white solid crude product 7-13 (containing PyBOP byproduct). LCMS (ESI) m/z = 496.1 [M + H] ⁺ .

Step 5: Synthesis of compound 13

The reaction was carried out in a 100 mL single-mouth bottle. Intermediate 7-13 (300 mg, 0.604 mmol) was dissolved in tetra-n-butylammonium fluoride TBAF (8 ml) and reacted at 60°C for 8 hours until completion. 30 ml of water was added to the reaction solution and extracted twice with 30 ml of DCM. The organic phase was concentrated and purified by Prep-TLC (DCM: MeOH = 10: 1) to obtain 50 mg of a white solid crude product, which was then purified by Prep-HPLC (0.1% TFA) to obtain 2 mg of a white solid 13 (yield: 1.0%). ¹ H NMR (400MHz, DMSO) δ13.54 (s, 1H), 8.31 (s, 1H), 8.19 (d, J = 1.6Hz, 1H), 7.88 (d, J = 8.7Hz, 1H), 7.79 (d, J = 9.0Hz, 1H), 7.57 (dd, J ₁ = 8.7Hz, J ₂ = 1.9Hz, 1H), 7.50 (dd, J ₁ = 8.9Hz, J ₂ = 2.2Hz, 1H), 6.55 (d, J = 4.5Hz, 1H), 5.74 (d, J = 2.2Hz, 1H), 2.74 (d, J = 4.4Hz, 3H). LCMS (ESI) m/z = 366.1 [M+H] ⁺ .

Example 13. Synthesis of Compound 34

Step 1: Synthesis of intermediate 2-34

The reaction was carried out in a 50 mL single-mouth bottle. Compound 1-A (2.00 g, 6.12 mmol) was put into a single-mouth bottle, and then phosphorus oxychloride (20 ml) and two drops of catalytic amount of DMF were added. The mixture was refluxed and stirred for 12 h under nitrogen protection. The reaction was complete when the central control was detected. The phosphorus oxychloride was removed by vacuum concentration, and 25 mL of ice-water mixture was added to the reaction bottle. The mixture was extracted three times with dichloromethane (20 mL/times), and the organic phases were combined and dried over anhydrous sodium sulfate. The mixture was dried under reduced pressure without heating at room temperature to obtain 2.60 g of intermediate 2-34 as an off-white solid (yield>100%, LCMS purity 16%, most of which were isomers). LCMS (ESI) m/z=345.1[M+H] ⁺ .

Step 2: Synthesis of compound 34

The reaction was carried out in a 50 mL single-mouth bottle. Compound 2-34 (400 mg, 1.15 mmol) was dissolved in tetrahydrofuran (4 ml), 4-aminopyridine (218 mg, 2.31 mmol) and DIEA (299.5 mg, 2.3 mmol) were added, and the mixture was reacted at 60°C for 16 h. After the reaction, the concentrated reaction solution was separated and purified by Prep-TLC to obtain 9.68 mg of compound 34 as a white solid, with a yield of 2.4%. ¹ H NMR (400MHz, DMSO) δ10.80 (s, 1H), 8.52 (d, J = 5.5Hz, 2H), 8.08 (d, J = 5.1Hz, 2H), 7.85 (d, J = 8.9Hz, 1H), 7.66 (d, J = 9.1Hz, 1H), 7.57 (t, J = 7.7Hz, 1H), 7.49 ( d,J＝7.6Hz,1H),7.40(s,1H),7.37(d,J＝7.8Hz,1H),6.12(s,1H),2.43(s,3H).LCMS(ESI)m/z＝403.0[M+H] ⁺ .

Example 14. Synthesis of Compound 51

Step 1: Synthesis of Compound 2-51

The reaction was carried out in a 250 mL eggplant flask. 5-Chloro-2-fluorobenzonitrile 1-11 (5.00 g, 32.26 mmol) was dissolved in DMF (80 mL), methyl thioglycolate (3.76 g, 35.48 mmol) and potassium carbonate (8.90 g, 64.52 mmol) were added, and stirred at 80 ° C for 12 h until the reaction was complete. The reaction mixture was cooled to room temperature, and water (3 L) was added to precipitate solids, which were filtered and dried to obtain the product compound 2-51 (6.00 g, yield: 77.2%) as a yellow solid. ¹ H NMR (400MHz, DMSO) δ8.31 (d, J = 2.0 Hz, 1H), 7.89 (d, J = 8.6 Hz, 1H), 7.54 (dd, J ₁ = 8.6, J ₂ = 2.1 Hz, 1H), 7.19 (s, 2H), 3.79 (s, 3H).

Step 2: Synthesis of compound 3-51

The reaction was carried out in a 100 mL three-necked flask. Compound 2-51 (2.00 g, 8.30 mmol), 3-bromopyridine (1.42 g, 9.13 mmol), Pd ₂ (dba) ₃ (0.76 g, 0.83 mmol), Xantphos (0.96 g, 1.66 mmol), and Cs ₂ CO ₃ (5.39 g, 16.60 mmol) were dissolved in 30 mL 1,4-dioxane, and the mixture was reacted at 100 ° C for 12 h under nitrogen protection. The reaction solution was cooled to room temperature, poured into 300 ml of water, and extracted three times with 300 ml of dichloromethane (100 ml/time). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product, which was purified by silica gel flash chromatography (DCM: MeOH = 20: 1) to obtain intermediate 3-51 (2.00 g, yield: 75.8%) as a yellow solid. LCMS (ESI) m/z = 318.8 [M + H] ⁺ .

Step 3: Synthesis of intermediate 4-51

The reaction was carried out in a 100 mL three-necked flask. Intermediate 3-51 (600 mg, 1.89 mmol) was added to THF (10 ml), the nitrogen protection temperature was lowered to -15 ° C, CSI (400 mg, 2.83 mmol) was added and stirred for 1 hour until the reaction was completed. Ammonium chloride aqueous solution (10 ml) was added to quench, water (20 ml) was added, and DCM (20 ml) was extracted three times. The organic phase was concentrated to obtain the crude product of intermediate 4-51 (800 mg, yield> 100%) as a white solid. LCMS (ESI) m/z = 361.9 [M + H] ⁺ .

Step 4: Synthesis of intermediate 5-51

The reaction was carried out in a 100 mL three-necked flask. Intermediate 4-51 (800 mg, 2.22 mmol) was dissolved in methanol (10 ml), sodium hydroxide (177 mg, 4.43 mmol) was added, and stirred at 60 ° C for 1 h under nitrogen protection. LCMS detected that the reaction was complete, and the crude product was purified by silica gel flash chromatography (DCM: MeOH = 20: 1) to obtain intermediate 5-51 (240 mg, yield: 32.9%), a white solid. ¹ HNMR (400MHz, DMSO) δ12.18 (s, 1H), 8.90 (d, J = 4.8Hz, 1H), 8.84 (d, J = 2.3Hz, 1H), 8.20 (d, J = 8.8Hz, 1H), 8.14 (d, J = 8.0Hz, 1H), 7.76 (dd, J ₁ = 8.1Hz, J ₂ = 4. 8Hz, 1H), 7.58 (dd, J ₁ = 8.8Hz, J ₂ = 2.0Hz, 1H), 5.85 (d, J = 1.8Hz, 1H).

Step 5: Synthesis of Compound 51

The reaction was carried out in a 10 mL sealed tube. Intermediate 5-51 (240 mg, 0.73 mmol) was dissolved in DMF (3 ml), PyBOP (568 mg, 1.09 mmol) and DBU (221 mg, 1.46 mmol) were added, stirred at room temperature for 1 min, 1.5 ml of 2M methylamine tetrahydrofuran solution was added, and the reaction was carried out at room temperature for 1 hour. Prep-HPLC (0.1% NH ₃ H ₂ O) was used to purify the crude product to obtain 30 mg of the crude product, which was purified by Prep-HPLC (0.1% FA) to obtain 5.15 mg of 51 as a white solid, with a yield of 2.0%. ¹ H NMR (400MHz, DMSO) δ8.92 (s, 2H), 8.52 (s, 1H), 8.16 (d, J = 8.7Hz, 1H), 8.02 (d, J = 7.9Hz, 1H), 7.80 (s, 1H), 7.58 (dd, J ₁ = 8.8, J ₂ = 1.7Hz, 1H), 5.89 (s, 1H ),3.02(s,3H).LCMS(ESI)m/z＝343.0[M+1] ⁺ .

Test Case

Test Example 1: Biochemical Test

Brief introduction to the experimental principle: MAT2A can catalyze the conversion of L-methionine and ATP into SAM, inorganic phosphate and inorganic diphosphate. By adding a colorimetric agent to the enzymatic reaction system, the content of inorganic phosphate in the sample can be quantitatively detected, thereby characterizing the enzyme activity of MAT2A.

Test Materials:

1. Tris (Life science#0497)

2. BSA (Sigma#)

3.MAT2Ahis-tag (BPS#71401-1)

4. 384-well plate (Corning #3765)

5. L-methionine (Admas#1100469)

6. ATP (Sigma#A7699)

7.MgCl2 (Sigma#M8266)

8. KCl (Sigma #7447-40-7)

9. Brij35 (Sigma#B4184)

10.EDTA (Sigma#E1644)

Test method:

1. Prepare 1xAssaybuffer.

2. Preparation of compound concentration gradient: The test compound concentration is 10 μM, 3-fold dilution, 10 concentrations, and duplicate wells. Gradient dilutions are made into 10 solutions with 100-fold final concentration in a 384-well plate. Then 250 nL is transferred to a 384-well reaction plate using Echo550 for later use. 250 nL of 100% DMSO is added to the negative control well and the positive control well, respectively.

3. Use 1xAssaybuffer to prepare an enzyme solution with a final concentration of 1.67 times.

4. Add 15 μL of 1.67 times the final concentration of enzyme solution to the compound wells and positive control wells respectively; add 15 μL of 1xAssaybuffer to the negative control wells.

5. Centrifuge at 1000 rpm for 60 seconds, shake to mix, and incubate for 15 minutes.

6. Use 1xAssaybuffer to prepare a substrate mixed solution with a final concentration of 2.5 times.

7. Add 10 μL of substrate mixed solution at 2.5 times the final concentration to start the reaction.

8. Centrifuge the 384-well plate at 1000 rpm for 60 seconds, shake to mix, and incubate for 150 minutes.

9. Add 50 μL Biomol to terminate the reaction, centrifuge at 1000 rpm for 60 seconds, and incubate for 15 minutes. Read OD620 and process the data.

Data Analysis:

Calculate the compound inhibition rate (%) and obtain the _IC50 of the test compound by fitting.

Compound inhibition rate (%) = (OD620_max-OD620_sample)/(OD620_max-OD620_min)X100

Among them: OD620_sample is the absorbance value of the sample well; OD620_min is the absorbance value of the positive control well, indicating the reading of the well without enzyme activity; OD620_max is the absorbance value of the negative control well, indicating the reading of the well without compound inhibition

Test results:

Under the conditions of this test, the inhibitory effect of the test compound on MAT2A enzyme activity can be expressed by the IC ₅₀ value of the inhibition of the level of phosphate production during the enzymatic reaction. The MAT2A inhibitory activity of the test compound is shown in Table 1. Among them, 0nM＜A＜100nM, 100nM≤B＜500nM, C≥500nM.

Table 1

Compound MAT2A inhibitory activity IC ₅₀ (nM) Compound MAT2A inhibitory activity IC ₅₀ (nM) 1 A 31 B 2 B 32 C 3 A 33 B 4 A 34 A 7 C 35 A 8 C 36 A 9 C 37 A 10 C 38 A 11 A 39 A 12 A 40 C 13 C 41 B 14 A 42 A 15 A 43 A 16 A 44 A 17 A 45 A 18 A 46 A 19 B 47 A 20 B 48 A twenty one A 49 A twenty two A 50 B twenty three A 52 A twenty four A 53 A 25 A 57 A 26 A 64 A 27 C 71 A 28 B 87 A 29 A 30 B

Test Example 2: Detection of intracellular SAM levels

Brief introduction to the experimental principle: After the MAT2A inhibitor to be tested is incubated with cells for a period of time, the cells are lysed with a lysis buffer to quench the MAT2A enzyme activity. The SAM content in the cell lysate is detected by LC-MS/MS to characterize the MAT2A enzyme activity in the cells.

Test Materials:

1. HCT116 MTAP ^-/- cells (Horizon#HD R02-033)

2. RPMI-1640 (BI#C3010-0500)

3. Fetal bovine serum (EXCELL#FND500)

4.Penicillin-Streptomycin (Gibco#15140-122)

5. 0.25% Typsin-EDTA (Gibco#25200-072)

6.DMSO (Sigma#D2650)

7. Methanol (Sigma#34860)

8. Acetic acid (GREAGENT#G73562B)

9. 96-well plate (Corning#3599)

Test method:

1. HCT116 MTAP ^-/- cells were cultured in RPMI-1640 medium containing 10% fetal bovine serum and 1% penicillin-streptomycin at 37°C and 5% CO ₂ in a cell culture incubator. Cells in the logarithmic growth phase were used for subsequent experiments.

2. Adjust the cell concentration to 20,000 cells per well, inoculate a 96-well plate, and culture overnight in a cell culture incubator at 37°C and 5% _CO2 .

3. Dissolve the compound in DMSO, dilute with culture medium to an appropriate concentration, and add to the cell plate. Continue culturing at 37°C and 5% CO ₂ for 6 hours.

4. Aspirate the supernatant, wash once with PBS, and then add lysis buffer to lyse the cells.

5. After the lysate is processed, the SAM concentration is determined by LC-MS/MS analysis.

Data Analysis:

Calculate the compound inhibition rate (%), and obtain the _IC50 of the test compound by fitting.

Compound inhibition rate (%) = (SAM_max-SAM_sample)/(SAM_max-SAM_min)X100

Where: SAM_sample is the SAM concentration of the sample well; SAM_min is the SAM concentration of the positive control well, indicating the reading without cells; SAM_max is the absorbance value of the negative control well, indicating the SAM concentration of the well without compound inhibition.

Test results:

Under the conditions of this experiment, the IC ₅₀ value of the test compound on the inhibition of SAM production in HCT116 MTAP ^-/- cells is used to characterize the inhibitory ability of the test compound on the MAT2A enzyme activity in the cells. The inhibitory activity of the test compound on the intracellular SAM production is shown in Table 2.

Table 2

Compound SAM production inhibitory activity IC ₅₀ (nM) 1 12.9 14 3.6 16 28.7 18 8.5 twenty three 9.6 29 29.3 39 25.2

Test Example 3: Human pancreatic cancer KP-4 cell activity inhibition test

Brief introduction to the experimental principle: After the MAT2A inhibitor to be tested is incubated with cancer cells for a period of time, a cell activity measurement method based on ATP content is used to characterize the effect of the test compound on cell activity.

Test Materials:

1. KP-4 cells (JCRB#JCRB0182)

2.IMDM (Gibco#12440061)

3. Fetal bovine serum (EXCELL#FND500)

4.Penicillin-Streptomycin (Gibco#15140-122)

5. 0.25% Typsin-EDTA (Gibco#25200-072)

6.DMSO (Sigma#D2650)

7. 96-well plate (Corning #3610)

8.CellTiter-Glo(Promega#G7571)

Test method:

1. KP-4 cells were cultured in IMDM containing 10% fetal bovine serum and 1% penicillin-streptomycin at 37°C and 5% _CO2 in a cell culture incubator. Cells in the logarithmic growth phase can be used for subsequent experiments.

2. Adjust the cell concentration to 500 cells per well, inoculate a 96-well plate, and culture overnight in a cell culture incubator at 37°C and 5% _CO2 .

3. Dissolve the compound in DMSO, dilute with culture medium to an appropriate concentration, and add to the cell plate. Continue culturing at 37°C, 5% CO ₂ for 5 days.

4. Add CellTiter-Glo reagent and use an ELISA reader to detect cell activity.

Data Analysis:

Calculate the compound inhibition rate (%) and obtain the _IC50 of the test compound by fitting.

Compound inhibition rate (%) = (Signal_max-Signal_sample)/(Signal_max-Signal_min)X100

Among them: Signal_sample is the reading of the sample well, indicating the cell activity of the compound inhibiting well; Signal_min is the reading of the positive control well, indicating the background activity without cells; Signal_max is the reading of the negative control well, indicating the cell activity of the compound not inhibiting well.

Test results:

Under the conditions of this experiment, the IC ₅₀ values of the test compounds for inhibiting the activity of KP-4 cells are shown in Table 3.

Table 3

Compound KP-4 cell antiproliferative activity IC ₅₀ (nM) 1 131.2 11 30.0 12 60.7 14 159.2 15 209.1 17 20.4 twenty three 126.9 29 91.6 39 25.2 45 15.5 47 11.3

Claims (14)

1. A compound as represented by formula (I), its stereoisomer or a pharmaceutically acceptable salt thereof,

in, _X1 is ^CR3 or N, _X2 is ^CR4 or N, _X3 is ^CR5 or N, _X4 is ^CR6 or N; and at most two of _X1 , _X2 , _X3 , and _X4 are N at the same time; R ³ , R ⁴ , R ⁵ , and R ⁶ are each independently selected from H, D, halogen, C ₁ -C ₆ alkyl, C ₂ -C ₆ alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₁ -C ₆ alkylsulfonyl, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, mercapto, C ₁ -C ₆ alkylamino, amino, nitro, carboxyl, and NHCOR ^a , wherein the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen, and R ^a is selected from C ₁ -C ₁₀ alkyl and C ₃ -C ₁₀ cycloalkyl; W is selected from O, NR ^b , S, CHR ^b ; R ^b is selected from H, D, C ₁ -C ₄ alkyl, C ₃ -C ₆ cycloalkyl, and the alkyl and cycloalkyl are unsubstituted or substituted with one or more substituents selected from D, halogen, -OH; R ¹ is selected from unsubstituted or substituted C ₃ -C ₁₀ cycloalkyl, unsubstituted or substituted C ₆ -C ₁₀ aryl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, wherein the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C 1 -C ₄ alkoxy, C 1 -C 4 haloalkyl, COOH, CONHR ^c , NHCOR ^c , NHSO ₂ R ^c , R ^c is selected from H, C ₁ -C 4 alkyl, C 3 -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl, wherein the C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C _{4 alkoxy, C 1 -C 4 haloalkyl ,} COOH, CONHR c , NHCOR c , _{NHSO 2} R ^c -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl is unsubstituted or substituted by one or more selected from halogen, hydroxyl, cyano; R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ; R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, D, unsubstituted or substituted C ₁ -C ₆ alkyl, unsubstituted or substituted C ₁ -C ₆ alkoxy, unsubstituted or substituted C ₂ -C ₆ alkenyl, unsubstituted or substituted C ₂ -C ₆ alkynyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted C ₅ -C ₁₀ aryl, unsubstituted or substituted 3-7 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₆ alkyl), -NHSO ₂ -(C ₁ -C ₆ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution by one or more substituents selected from the following: D, halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C ₃ alkyl which is unsubstituted or substituted by one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted by one or more substituents selected from Group B, or 5-10 membered heteroaryl which is unsubstituted or substituted by one or more substituents selected from Group B; R ^d is H, COOH, Boc, C ₁ -C ₃ alkyl which is unsubstituted or substituted by one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted by one or more substituents selected from Group B, or 5-10 membered heteroaryl which is unsubstituted or substituted by one or more substituents selected from Group B; the substituents in Group B include: C ₁ -C ₆ alkyl, halogen, OH, and amino; or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 4-6 membered heterocycloalkyl group which is unsubstituted or substituted with one or more substituents selected from the following: OH, halogen, C ₁ -C ₁₀ alkyl, C ₃ -C ₁₀ cycloalkyl; The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively. 2. The compound of formula (I) according to claim 1, its stereoisomer or a pharmaceutically acceptable salt thereof, in, R ³ , R ⁴ , R ⁵ , R ⁶ are each independently selected from H, halogen, C ₁ -C ₆ alkyl, C 2 -C 6 alkenyl, C ₂ -C ₆ alkynyl, C ₁ -C ₆ alkoxy, C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, amino, and the alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen; further preferably, R ₃ , R ⁵ , R ₆ are each independently selected from H and halogen, and R ⁴ is selected from H, halogen, C ₁ -C 6 alkyl, C 2 -C 6 alkenyl, C ₂ -C 6 alkynyl, C 1 -C ₆ alkoxy, C 3 -C 7 cycloalkyl, cyano, hydroxyl, amino, and the alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from ^D and halogen; further preferably, R 3 , R 5 , R ^{6 are each independently selected from H and halogen, and R 4} is selected from H, halogen, C _{1 -C 6 alkyl, C 2 -C 6 alkenyl ,} C 2 -C ₆ alkynyl, C 1 -C 6 alkoxy, ^C ₃ -C ₇ cycloalkyl, cyano, hydroxyl, amino, and the alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen; further preferably, ^{5. R6} is H, ^R4 is selected from H, halogen, _C1 - _C6 alkyl, _C2 - _C6 alkenyl, _C2 - _C6 alkynyl, _C1 - _C6 alkoxy, _C3 - _C7 cycloalkyl, cyano, hydroxyl, amino, and the alkyl, alkenyl, alkynyl, alkoxy, and cycloalkyl are unsubstituted or optionally substituted with one or more substituents selected from D and halogen; The ^Rb is selected from H, D, _C1 - _C3 alkyl, _C3 - _C6 cycloalkyl, wherein the alkyl and cycloalkyl are unsubstituted or substituted by one or more substituents selected from halogen and -OH; more preferably, the ^Rb is selected from H, D, methyl, ethyl, n-propyl, isopropyl, cyclopropyl and cyclobutyl, wherein the methyl, ethyl, n-propyl, isopropyl, cyclopropyl and cyclobutyl are unsubstituted or substituted by one or more substituents selected from D, halogen and -OH; R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, wherein the substituents in Group A include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl, COOH, CONHR ^c , NHCOR ^c , NHSO ₂ R ^c , R ^c is selected from H, C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C 1 -C 10 alkoxy or C ₆ -C ₁₀ aryl, wherein the C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl in R ^c ₆ - _C10 aryl is unsubstituted or substituted by one or more selected from halogen, hydroxyl, cyano; Preferably, R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, wherein the substitution in R ¹ refers to substitution by one or more substituents selected from Group A, and Group A substituents include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl; R ² is selected from NR ⁷ R ⁸ , -OR ⁹ , or -SR ⁹ ; R ⁷ , R ⁸ , and R ⁹ are each independently selected from H, unsubstituted or substituted C ₁ -C ₃ alkyl, unsubstituted or substituted C ₁ -C ₃ alkoxy, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted phenyl, unsubstituted or substituted 3-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl, -NHCO-(C ₁ -C ₃ alkyl), and -NHSO ₂ -(C ₁ -C ₃ alkyl); wherein the substitution in R ⁷ , R ⁸ , and R ⁹ refers to substitution with one or more substituents selected from the following: halogen, -NHR ^d , -N(R ^d ) ₂ , -OR ^d , -SR ^d , C ₁ -C 3 alkyl unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C ₇ alkyl unsubstituted or substituted with one or more substituents selected from Group B, R ^d is H, COOH, Boc, C 1 -C 3 alkyl which is unsubstituted or substituted with one or more substituents selected from Group B, C ₃ -C ₇ cycloalkyl which is unsubstituted or substituted with one or more substituents selected from Group B, or 5-10 _membered heteroaryl which is unsubstituted or substituted with one _{or more substituents selected from Group B; the substituents in Group B include: C 1 -C 3 alkyl , halogen} , OH, and amino; or R ⁷ , R ⁸ together with the nitrogen atom to which they are attached form a 5-6 membered heterocycloalkyl group which is unsubstituted or substituted by one or more substituents selected from the following: OH, halogen, C ₁ -C ₃ alkyl, C ₃ -C ₆ cycloalkyl; The heteroaryl and heterocycloalkyl groups contain one or more heteroatoms selected from N, O, and S, respectively. 3. The compound of formula (I) according to claim 2, its stereoisomer or a pharmaceutically acceptable salt thereof, Wherein, _X1 is CH or N, _X2 is ^CR4 or N, _X3 is CH or N, and _X4 is CH or N; and at most two of _X1 , _X2 , _X3 , and _X4 are N at the same time; and ^R4 is selected from F, Cl, Br, _CH3 , CD3, _CF3 , _CF2H , _CF2D , _OH , SH, _NH2 , CN, _OCH3 , _CH3CH2 , _-CH2CH2CH3 , cyclopropyl, _{-CH2 ( CH3 ) CH3 ,} and _NO2 . 4. The compound of formula (I) according to claim 2, its stereoisomer or a pharmaceutically acceptable salt thereof, Wherein, W is selected from O, NH, NCH ₃ , NC ₂ H ₄ OH, NCH(CH ₃ ) ₂ , NC ₂ H ₅ , S, and CH ₂ . 5. The compound of formula (I) according to any one of claims 1 to 4, its stereoisomer or a pharmaceutically acceptable salt thereof, Wherein, R ¹ is selected from unsubstituted or substituted phenyl, unsubstituted or substituted C ₃ -C ₇ cycloalkyl, unsubstituted or substituted 5-6 membered heterocycloalkyl, unsubstituted or substituted 5-10 membered heteroaryl; the 5-10 membered heteroaryl is selected from the following groups:

Wherein, the substitution in R ¹ refers to substitution by one or more substituents selected from Group A; Group A substituents include: halogen, CN, OH, oxo, SH, NH ₂ , NO ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl (such as CF ₃ , CF ₂ H), COOH, CONHR ^c , NHCOR ^c , NHSO ₂ R ^c , R ^c is selected from H, C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl, wherein the C ₁ -C ₄ alkyl, C ₃ -C ₁₀ cycloalkyl, C ₁ -C ₁₀ alkoxy, C ₆ -C ₁₀ aryl in R ^c is unsubstituted or substituted by one or more substituents selected from halogen, hydroxyl, and cyano. 6. The compound of formula (I) according to any one of claims 1 to 4, its stereoisomer or a pharmaceutically acceptable salt thereof, Wherein, the R ¹ is selected from

wherein R ¹⁰ is selected from halogen, CN, OH, SH, NH ₂ , C ₁ -C ₄ alkyl, C ₃ -C ₇ cycloalkyl, C ₁ -C ₄ alkoxy, C ₁ -C ₄ haloalkyl, COOH, CONH ₂ , CONHR ^c , NHCOR ^c , and R ^c is selected from C ₁ -C ₄ alkyl, C ₃ -C ₆ cycloalkyl, C ₁ -C ₄ alkoxy or phenyl which is unsubstituted or substituted with one or more groups selected from halogen, hydroxyl, cyano. 7. The compound of formula (I) according to any one of claims 1 to 4, its stereoisomer or a pharmaceutically acceptable salt thereof, wherein R ¹ is selected from

8. The compound of formula (I) according to any one of claims 1 to 4, its stereoisomer or a pharmaceutically acceptable salt thereof, wherein R ² is selected from NH ₂ ,

9. The compound according to any one of claims 1 to 8, its stereoisomer or a pharmaceutically acceptable salt thereof, wherein the compound represented by formula (I) is represented by the following formula I-A,

Wherein, W, R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as described in the corresponding claims respectively. 10. The compound according to claim 1, its stereoisomer or a pharmaceutically acceptable salt thereof, wherein the compound represented by formula (I) is selected from the following structures:

11. A composition comprising at least one compound of formula (I) according to any one of claims 1 to 10, a stereoisomer thereof or a pharmaceutically acceptable salt thereof. 12. A pharmaceutical preparation comprising a therapeutically effective amount of a compound of formula (I) according to any one of claims 1 to 10, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients. 13. A pharmaceutical combination comprising at least one therapeutically effective amount of a compound of formula (I) according to any one of claims 1 to 10, a stereoisomer thereof or a pharmaceutically acceptable salt thereof, and one or more other therapeutic agents. 14. Use of the compound represented by formula (I) according to any one of claims 1 to 10, its stereoisomers or pharmaceutically acceptable salts thereof in the preparation of a drug for treating and/or preventing tumors, Preferably, the tumor includes: MTAP-deficient tumors; MTAP-lowly expressed tumors; MAT2A-abnormally expressed tumors; other MAT2A-dependent tumors. Preferably, the tumor includes: breast cancer, lung cancer, glioblastoma, brain cancer and spinal cancer, head and neck cancer, skin cancer, reproductive system cancer, gastrointestinal system cancer, esophageal cancer, nasopharyngeal cancer, pancreatic cancer, rectal cancer, hepatocellular carcinoma, bile duct cancer, gallbladder cancer, colon cancer, multiple myeloma, kidney and bladder cancer, bone cancer, malignant mesothelioma, sarcoma, lymphoma, adenocarcinoma, thyroid cancer, heart tumors, germ cell tumors, malignant neuroendocrine tumors, malignant rhabdoid tumors, soft tissue sarcomas, midline tract cancer and unknown primary cancer.