WO2025140599A1 - Heterocyclic compound, and preparation method therefor and medical use thereof - Google Patents

Abstract

Provided are a heterocyclic compound, and a preparation method therefor and the medical use thereof. In particular, provided are a compound as shown in general formula (I), a preparation method therefor, a pharmaceutical composition containing the compound, and the use of the compound as a PKMYT1 inhibitor. The compound and the pharmaceutical composition containing the compound can be used for treating and/or preventing diseases related to PKMYT1 activity, such as cancers. Definitions of various groups in general formula (I) are the same as defined in the description.

Description

A heterocyclic compound and its preparation method and medical use Technical Field

The present invention belongs to the field of medical technology, and specifically relates to a heterocyclic compound, a preparation method thereof and a pharmaceutical composition containing the same, as well as use of the heterocyclic compound as a PKMYT1 inhibitor in the treatment and/or prevention of diseases associated with PKMYT1 activity.

Background Art

CCNE1 encodes cyclin E1 (CyclinE1), which forms a complex with cyclin-dependent kinase 2 (CDK2) to drive cells from G1 to S phase. In cancer, amplification of the CCNE1 gene and/or dysregulated expression of CyclinE1 often occurs in the early stages of tumor development, forcing cancer cells to enter S phase prematurely. Excessive replication, lack of origins, and insufficient nucleotide pools lead to replication fork stalling, resulting in replication stress and DNA damage. In the case of inactivation of p53 function, cells enter mitosis with damaged DNA, leading to genomic instability. Cells have evolved a complex set of mechanisms to cope with DNA damage, collectively referred to as the DNA damage response (DDR). Cell cycle checkpoint activation is an important component of the DDR, which regulates specific DNA repair mechanisms in various stages of the cell cycle, including the G1, S, G2, and mitotic checkpoints.

CCNE1 amplification is prevalent in multiple tumor types, including high-grade serous ovarian cancer, uterine cancer, and gastroesophageal cancer. In ovarian cancer, CCNE1 amplification is detected in approximately 20% of tumors, is often mutually exclusive with homologous recombination defects, and is enriched in platinum-treated recurrent tumors. Targeting CDK2, the partner protein of Cyclin E1, can effectively inhibit the proliferation of CCNE1-amplified tumor cells. Although selective CDK2 inhibitors have entered the clinical stage, due to the structural similarities between CDKs, no inhibitors with precise selectivity have entered the market. As an alternative approach, searching for potential synthetic lethal targets for Cyclin E1 may provide much-needed, new treatment options for CCNE1-amplified tumors. The latest study shows that PKMYT1 (membrane-associated tyrosine/threonine protein kinase 1) has been identified as a synthetic lethal target for CCNE1 amplification and is also an attractive target for treating some types of DDR cancers.

PKMYT1 is an evolutionarily conserved protein kinase, also known as Myt1. Its main function is to inhibit the activity of CDK1 (cdc2) by phosphorylating the Thr14 and Tyr15 sites of CDK1, regulate the G2-M cell cycle, and allow cells to enter mitosis. PKMYT1 belongs to the WEE kinase family and is structurally similar to WEE, but has been less studied. WEE1 is distributed in the nucleus and regulates the activity of CDK1 and CDK2 kinases by phosphorylating the Tyr15 residue on CDK1 and CDK2, and participates in regulating the progress of the S phase, G2/M phase, and M phase cell cycle checkpoints. PKMYT1 is mainly distributed in the cytoplasm and is located between the inner membrane of the Golgi apparatus and the endoplasmic reticulum. It is a membrane-associated inhibitory kinase that plays a role only in the G2/M phase checkpoint by selectively regulating CDK1 phosphorylation. Compared with WEE1, PKMYT1 exhibits more restricted substrate specificity because PKMYT1 only phosphorylates CDK1 but not the CDK2 complex. Targeting PKMYT1 has great clinical application prospects. Currently, only one selective inhibitor, RP-6306, has entered the clinical stage.

Summary of the invention

The present invention designs and synthesizes a series of heterocyclic compounds, and screens the PKMYT1 activity thereof. The research results show that the compounds are potent PKMYT1 inhibitors, and have a selective inhibitory effect on cells with CCNE1 gene amplification. The compounds and pharmaceutical compositions containing the compounds can be used to treat and/or prevent diseases associated with PKMYT1 activity, such as cancers carrying CCNE1 gene amplification.

Therefore, the object of the present invention is to provide a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof,

in:

X is selected from N or CR ⁷ ;

Y is selected from O, S, NR ⁷ or CR ^7a R ^7b ;

R ¹ is selected from hydrogen, halogen or -NR ^8a R ^8b ;

R ² , R ³ , R ⁴ and R ⁵ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ^7a , R ^7b and R ⁷ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ R 7a and R 7b are substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR 9 , -C(O)R 10 , -S(O) p R ¹⁰ , -C(O)NR ^9a R 9b , -S(O) p NR 9a R 9b , -NR 9a R 9b , -SR 9 , -OR 9a , -C(O)R ¹⁰ , -S(O) p R ¹⁰ , -C(O)NR 9a _R ^9b , -S(O) _p NR ^9a R ^{9b ,} -NR ^9a R ^9b , -SR ^{9 ,} -OR ^9a , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ^8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R 10 , -S(O) p R 10 , -C(O)NR 9a R ^9b , -S(O) _p NR ^{9a R 9b} , oxo, alkyl, alkoxy, ^haloalkyl , hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, ^heterocyclyl , ^aryl , heteroaryl; or, _R 8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, ^heteroaryl . ^8b together with the nitrogen atom to which it is attached forms a heterocyclic group or a heteroaryl group, wherein the heterocyclic group or the heteroaryl group is optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclic group, aryl, heteroaryl;

R ^9a , R ^9b and R ⁹ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a heterocyclyl or heteroaryl, and the heterocyclyl or heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ¹⁰ is selected from hydrogen, halogen, amino, nitro, hydroxyl, mercapto, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, mercapto, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

p is 1 or 2.

The present invention further provides a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof,

in,

X is selected from N;

Y is selected from O or NR ⁷ ;

R ¹ is selected from hydrogen, halogen or -NR ^8a R ^8b ;

R ² , R ³ , R ⁴ and R ⁵ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁷ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ^8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R 10 , -S(O) p R 10 , -C(O)NR 9a R ^9b , -S(O) _p NR ^{9a R 9b} , oxo, alkyl, alkoxy, ^haloalkyl , hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, ^heterocyclyl , ^aryl , heteroaryl; or, _R 8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, ^heteroaryl . ^8b together with the nitrogen atom to which it is attached forms a heterocyclic group or a heteroaryl group, wherein the heterocyclic group or the heteroaryl group is optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclic group, aryl, heteroaryl;

R ^9a , R ^9b and R ⁹ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a heterocyclyl or heteroaryl, and the heterocyclyl or heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ¹⁰ is selected from hydrogen, halogen, amino, nitro, hydroxyl, mercapto, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, mercapto, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

p is 1 or 2.

In one embodiment, the compound represented by the general formula (I) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, is a compound represented by the general formula (IA) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:

wherein X, Y, R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in the general formula (I).

In another specific embodiment, the compound represented by the general formula (I) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, is a compound represented by the general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:

in,

Y ₁ is selected from O or NR ^7c ;

R ^7c is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ R 7d and R 7e together with the carbon atom to which they are attached form an oxo group, a cycloalkyl group, a heterocyclyl group, an aryl group or a heteroaryl group, wherein the cycloalkyl group, the heterocyclyl group, the aryl group or the heteroaryl group is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) p R ¹⁰ , -C(O)NR ^{9a R 9b ,} -S(O) _{p NR} ^9a R ^9b , -NR ^9a R ^9b , -SR ^{9 , -OR 9} , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I).

In another specific embodiment, the compound represented by the general formula (I) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, is a compound represented by the general formula (IIA) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:

wherein Y ₁ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in the general formula (II).

In a preferred embodiment, the compound represented by the general formula (II) or the general formula (IIA) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

Selected from

in:

R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁷ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I).

In a preferred embodiment, the compound represented by the general formula (II) or the general formula (IIA) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

Selected from

in:

R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁷ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I).

In another preferred embodiment, the compound represented by the general formula (II) or the general formula (IIA) of the present invention, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, is a compound represented by the general formula (IIB), (IIC), (IID), (IIE), or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:

in,

R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ^7c is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl;

R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I).

In another preferred embodiment, the compound represented by the general formula (II), (IIA), (IIB), (IIC), (IID), (IIE) according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated product thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from hydrogen, halogen, amino, hydroxyl, mercapto, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally selected from halogen, amino, cyano, hydroxyl, mercapto, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C substituted by one or more of _3-6 membered cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, or 5-10 membered heteroaryl;

R ^7c is selected from hydrogen, halogen, amino, hydroxyl, mercapto, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally selected from deuterated, halogen, amino, cyano, hydroxyl, mercapto, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _{2-6 alkenyl, C 2-6 alkynyl} , C substituted by one or more of _3-6 membered cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, or 5-10 membered heteroaryl;

R is selected from ^C3-6 cycloalkyl, _4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl, wherein the _C3-6 cycloalkyl, 4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl is optionally substituted by one or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, oxo, _C1-6 alkyl, _C1-6 alkoxy, _C1-6 haloalkyl, _C1-6 hydroxyalkyl, _C1-6 aminoalkyl, _C1-6 haloalkoxy, _C2-6 alkenyl, _C2-6 alkynyl, C3-6 cycloalkyl, 4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl _;

R ^9a and R ^9b are each independently selected from hydrogen, C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or a 5-10 membered heteroaryl group, and the _4-6 membered heterocyclic group or the 5-10 membered heteroaryl group is optionally substituted by one _{or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, C 1-6 alkyl} , C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C 2-6 alkynyl, C _{3-6 cycloalkyl, 4-6 membered heterocyclic group, C 6-10} aryl, or 5-10 membered heteroaryl.

In another preferred embodiment, the compound represented by the general formula (II), (IIA), (IIB), (IIC), (IID), (IIE) according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated product thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally substituted with one or more groups selected from halogen, C _1-6 alkyl;

R ^7c is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally substituted with one or more groups selected from deuterated, halogen, C _1-6 alkyl;

R ⁹ is selected from C _6-10 aryl, 5-10 membered heteroaryl, wherein the C _6-10 aryl, 5-10 membered heteroaryl is optionally substituted by one or more groups selected from halogen, amino, cyano, hydroxyl, mercapto, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy;

R ^9a and R ^9b are each independently selected from hydrogen, C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are connected form a 4-6 membered heterocyclic group, and the 4-6 membered heterocyclic group is optionally substituted by one _{or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, C 1-6 alkyl, C 1-6 alkoxy, C 1-6 haloalkyl, C 1-6 hydroxyalkyl, C 1-6 aminoalkyl, C 1-6 haloalkoxy , C 2-6 alkenyl , C} 2-6 alkynyl, C 3-6 cycloalkyl, 4-6 membered heterocyclic group, C _6-10 aryl, or 5-10 membered heteroaryl.

In another preferred embodiment, the compounds represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or their tautomers, mesomers, racemates, enantiomers, diastereomers, or mixtures thereof, or deuterated substances, or pharmaceutically acceptable salts thereof, wherein: ^R6 is selected from hydrogen, halogen, cyano, _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-6 membered heteroaryl, and the _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, C3-6 cycloalkyl, _4-6 membered heterocyclyl, phenyl, 5-6 membered heteroaryl are optionally substituted by one or more groups selected from halogen and _C1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ^7c is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, _{C 3-6 cycloalkyl} , 4-6 membered heterocyclyl, phenyl, 5-6 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally selected from deuterated, halogen, C 2-6 substituted by one or more _1-6 alkyl groups;

R ⁹ is selected from phenyl, 5-6 membered heteroaryl, wherein the phenyl, 5-6 membered heteroaryl is optionally substituted by one or more groups selected from halogen, amino, cyano, hydroxyl, mercapto, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy;

R ^9a and R ^9b are each independently selected from hydrogen, C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are connected form a 4-6 membered heterocyclic group, and the 4-6 membered heterocyclic group is optionally substituted by one _{or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, C 1-6 alkyl, C 1-6 alkoxy, C 1-6 haloalkyl, C 1-6 hydroxyalkyl, C 1-6 aminoalkyl, C 1-6 haloalkoxy , C 2-6 alkenyl , C} 2-6 alkynyl, C 3-6 cycloalkyl, 4-6 membered heterocyclic group, C _6-10 aryl, or 5-10 membered heteroaryl.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the C _6-10 aryl and 5-10 membered heteroaryl are optionally substituted with halogen or C _1-6 alkyl;

R ^9a and R ^9b are each independently selected from hydrogen and C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group;

Preferably,

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the 5-10 membered heteroaryl is optionally substituted with halogen or C _1-6 alkyl;

R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group;

More preferably,

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, 5-10 membered heteroaryl, wherein the 5-10 membered heteroaryl is optionally substituted by halogen or C _1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ⁷ or R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, -CH ₂ R ⁹ , and the C _1-6 alkyl is optionally substituted with deuteration;

R ⁹ is selected from phenyl, which is optionally substituted by one or more groups selected from halogen, C _1-6 alkyl;

Preferably, R ⁷ or R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, and the C _1-6 alkyl is optionally substituted by deuterium.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R2 and ^R3 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-7 cycloalkyl, 4-7 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl, ^-NR9aR9b , ^-SR9 , ^-OR9 ; the _C1-6 alkyl, _C2-6 ^alkenyl , _C2-6 alkynyl, _C3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 membered aryl, 5-10 membered heteroaryl are optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl;

R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I);

Preferably, R ² and R ³ are each independently selected from C _1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R4 and ^R5 are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-7 cycloalkyl, 4-7 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl, ^-NR9aR9b , ^-SR9 , ^-OR9 ; the _C1-6 alkyl, _C2-6 ^alkenyl , _C2-6 alkynyl, _C3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 membered aryl, 5-10 membered heteroaryl are optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl;

R ⁹ , R ^9a , R ^9b , R ¹⁰ and p are as defined in the general formula (I);

Preferably, ^R4 and ^R5 are each independently selected from hydrogen.

In another preferred embodiment, the compound represented by the general formula (II) or (IIA) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: Selected from

R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl;

R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, -CH ₂ R ⁹ , wherein the C _1-6 alkyl is optionally substituted with deuterium;

R ⁹ is selected from phenyl, which is optionally substituted by one or more groups selected from halogen, C _1-6 alkyl;

Preferably,

R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl;

R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl and phenyl, and the C _1-6 alkyl is optionally substituted by deuterium.

In another preferred embodiment, the compound represented by the general formula (II) or (IIA) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: Selected from

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the 5-10 membered heteroaryl is optionally substituted with halogen or C _1-6 alkyl;

R ^7c is selected from C _1-6 alkyl;

R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group;

Preferably,

^R6 is selected from hydrogen, halogen, cyano, _C1-6 alkyl, _C1-6 haloalkyl, 5-10 membered heteroaryl, wherein the 5-10 membered heteroaryl is optionally substituted by halogen or _C1-6 alkyl;

R ^7c is selected from C _1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (II) or (IIA) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: Selected from

R ⁶ is selected from C _1-6 alkyl, C _1-6 haloalkyl, 5-6 membered heteroaryl, preferably C _1-6 alkyl, C _1-6 haloalkyl.

In another preferred embodiment, the compound represented by the general formula (IIB) or (IID) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl;

R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, -CH ₂ R ⁹ , wherein the C _1-6 alkyl is optionally substituted with deuterium;

R ⁹ is selected from phenyl, which is optionally substituted by one or more groups selected from halogen, C _1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (IIB) or (IID) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl;

R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl and phenyl, and the C _1-6 alkyl is optionally substituted by deuterium.

In another preferred embodiment, the compound represented by the general formula (IIB) or (IID) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the 5-10 membered heteroaryl is optionally substituted with halogen or C _1-6 alkyl;

R ^7c is selected from C _1-6 alkyl;

R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group.

In another preferred embodiment, the compound represented by the general formula (IIB) or (IID) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

^R6 is selected from hydrogen, halogen, cyano, _C1-6 alkyl, _C1-6 haloalkyl, 5-10 membered heteroaryl, wherein the 5-10 membered heteroaryl is optionally substituted by halogen or _C1-6 alkyl;

R ^7c is selected from C _1-6 alkyl.

In another preferred embodiment, the compound represented by the general formula (IIC) or (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R6 is selected from _C1-6 alkyl, C1-6 haloalkyl, _5-6 membered heteroaryl.

In another preferred embodiment, the compound represented by the general formula (IIC) or (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R6 is selected from _C1-6 alkyl, _C1-6 haloalkyl.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

^R2 is selected from halogen and _C1-6 alkyl;

^R3 is selected from halogen, _C1-6 alkyl, _C1-6 haloalkyl, _C3-6 cycloalkyl;

R ⁴ is selected from hydrogen, halogen, cyano;

^R5 is selected from hydrogen and halogen.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R2 and ^R3 are _C1-6 alkyl; ^R4 and ^R5 are hydrogen.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: ^R2 is _C1-6 alkyl, ^R3 is _C1-6 alkyl; ^R4 is selected from hydrogen and halogen; ^R5 is selected from hydrogen and halogen.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ² is C _1-6 alkyl, R ³ is halogen; R ⁴ and R ⁵ are hydrogen.

In another preferred embodiment, the compounds represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or their tautomers, mesomers, racemates, enantiomers, diastereomers, or mixtures thereof, or deuterated substances, or pharmaceutically acceptable salts thereof, wherein: R ² is selected from halogen, R ³ is halogen and C _1-6 alkyl; R ⁴ and R ⁵ are hydrogen.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

^R2 is selected from _C1-6 alkyl;

^R3 is selected from halogen, _C1-6 alkyl, _C1-6 haloalkyl, _C3-6 cycloalkyl;

R ⁴ is selected from hydrogen, halogen, cyano;

^R5 is selected from hydrogen.

In another preferred embodiment, the compound represented by the general formula (I), (II), (IIA), (IIB), (IIC), (IID), (IIE) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:

^R2 is selected from _C1-6 alkyl;

^R3 is selected from _C1-6 alkyl;

^R4 is selected from hydrogen and halogen;

^R5 is selected from hydrogen and halogen.

Typical compounds of the present invention include, but are not limited to:

Its tautomer, meso racemate, racemate, enantiomer, diastereomer, or mixture thereof, or its pharmaceutically acceptable salt.

Another aspect of the present invention provides a method for preparing the compound represented by general formula (II) according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, which comprises the following steps:

The compound represented by the general formula D-1 or a salt thereof is reacted in a solvent, optionally in the presence of a catalyst, to obtain a compound represented by the general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or a mixture thereof, or a pharmaceutically acceptable salt thereof,

wherein Y ₁ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in the general formula (II).

Another aspect of the present invention provides a pharmaceutical composition comprising the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient.

The present invention further provides use of the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same, in the preparation of a PKMYT1 inhibitor.

The present invention further provides use of the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same, in the preparation of a medicament for preventing and/or treating a disease associated with PKMYT1 activity.

The present invention further provides the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or pharmaceutical composition containing the same, for use as a drug.

The present invention further provides the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or pharmaceutical composition containing the same, which is used as a PKMYT1 inhibitor.

The present invention further provides the compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same, which is used for preventing and/or treating diseases related to PKMYT1 activity.

The present invention further provides a method for inhibiting PKMYT1 activity, which comprises administering to a subject in need thereof an effective amount of a compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same.

The present invention further provides a method for preventing and/or treating diseases associated with PKMYT1 activity, comprising administering to a subject in need thereof a preventive or therapeutically effective amount of a compound according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing the same.

In a preferred embodiment of the present invention, the disease associated with PKMYT1 activity according to the present invention may be a solid tumor, such as ovarian cancer, breast cancer, cervical cancer, endometrial cancer, prostate cancer, colorectal cancer, esophageal cancer, liver cancer, lung cancer or thyroid cancer.

According to the conventional method in the field of the present invention, the compounds of the present invention can form pharmaceutically acceptable acid addition salts with acids. The acid includes inorganic acids and organic acids, and particularly preferably hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, naphthalene disulfonic acid, acetic acid, propionic acid, lactic acid, trifluoroacetic acid, maleic acid, citric acid, fumaric acid, oxalic acid, tartaric acid, benzoic acid, etc.

According to the conventional methods in the field of the present invention, the compounds of the present invention can form pharmaceutically acceptable basic addition salts with bases. The bases include inorganic bases and organic bases, and acceptable organic bases include diethanolamine, ethanolamine, N-methylglucamine, triethanolamine, tromethamine, etc., and acceptable inorganic bases include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, sodium hydroxide, etc.

The pharmaceutical composition containing the active ingredient may be in a form suitable for oral administration, such as tablets, troches, lozenges, aqueous or oil suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Oral compositions may be prepared according to any method known in the art for preparing pharmaceutical compositions, and such compositions may contain one or more ingredients selected from the following: sweeteners, flavoring agents, colorants and preservatives to provide pleasing and palatable pharmaceutical preparations. Tablets contain the active ingredient and non-toxic pharmaceutically acceptable excipients suitable for preparing tablets for mixing. These excipients may be inert excipients such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating agents and disintegrants such as microcrystalline cellulose, crosslinked sodium carboxymethyl cellulose, corn starch or alginic acid; binders such as starch, gelatin, polyvinyl pyrrolidone or gum arabic; and lubricants such as magnesium stearate, stearic acid or talc. These tablets may be uncoated or may be coated by known techniques which mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract, thereby providing a sustained release action over a longer period of time. For example, water soluble taste masking materials such as hydroxypropylmethylcellulose or hydroxypropylcellulose, or time extending materials such as ethylcellulose, cellulose acetate butyrate may be used.

Oral preparations may also be provided in hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent such as calcium carbonate, calcium phosphate or kaolin, or in soft gelatin capsules wherein the active ingredient is mixed with a water-soluble carrier such as polyethylene glycol or an oily vehicle such as peanut oil, liquid paraffin or olive oil.

Aqueous suspensions contain the active substance and excipients suitable for preparing aqueous suspensions for mixing. Such excipients are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone and gum arabic; dispersing agents or wetting agents, which may be naturally occurring phosphatides such as lecithin, or condensation products of alkylene oxides with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain fatty alcohols, for example heptadecaethyleneoxy cetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol, for example polyethylene oxide sorbitan monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene oxide anhydrosorbitan monooleate. The aqueous suspension may also contain one or more preservatives, for example ethylparaben or n-propylparaben, one or more coloring agents, one or more flavoring agents and one or more sweetening agents, for example sucrose, saccharin or aspartame.

Oil suspensions can be prepared by suspending the active ingredient in a vegetable oil such as peanut oil, olive oil, sesame oil or coconut oil, or a mineral oil such as liquid paraffin. The oil suspension may contain a thickener such as beeswax, hard paraffin or cetyl alcohol. The above-mentioned sweeteners and flavoring agents may be added to provide a palatable preparation. These compositions may be preserved by adding an antioxidant such as butylated hydroxyanisole or alpha-tocopherol.

Dispersible powders and granules suitable for preparing aqueous suspensions can provide the active ingredient and a dispersant or wetting agent, a suspending agent or one or more preservatives for mixing by adding water. Suitable dispersants or wetting agents and suspending agents are as described above. Other excipients such as sweeteners, flavoring agents and coloring agents can also be added. These compositions can be preserved by adding antioxidants such as ascorbic acid.

The pharmaceutical composition of the present invention can also be in the form of an oil-in-water emulsion. The oil phase can be a vegetable oil such as olive oil or peanut oil, or a mineral oil such as liquid paraffin or a mixture thereof. Suitable emulsifiers can be naturally occurring phospholipids, such as soybean lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, such as sorbitan monooleate, and condensation products of the partial esters and ethylene oxide, such as polyethylene oxide sorbitol monooleate. Emulsions can also contain sweeteners, flavoring agents, preservatives, and antioxidants. Syrups and elixirs prepared with sweeteners such as glycerol, propylene glycol, sorbitol, or sucrose can be used. Such preparations can also contain a demulcent, a preservative, a coloring agent, and an antioxidant.

The pharmaceutical composition of the present invention may be in the form of a sterile injectable aqueous solution. Acceptable vehicles and solvents that may be used are water, Ringer's solution and isotonic sodium chloride solution. The sterile injectable preparation may be a sterile injectable oil-in-water microemulsion in which the active ingredient is dissolved in an oil phase. For example, the active ingredient is dissolved in a mixture of soybean oil and lecithin. The oil solution is then added to a mixture of water and glycerol and processed to form a microemulsion. The injection or microemulsion may be injected into the patient's bloodstream by local mass injection. Alternatively, the solution and microemulsion may be preferably administered in a manner that maintains a constant circulating concentration of the compound of the present invention. To maintain this constant concentration, a continuous intravenous drug delivery device may be used.

The pharmaceutical composition of the present invention can be in the form of a sterile injection water or oil suspension for intramuscular and subcutaneous administration. The suspension can be prepared according to known techniques with the above-mentioned suitable dispersants or wetting agents and suspending agents. The sterile injection preparation can also be a sterile injection solution or suspension prepared in a non-toxic parenterally acceptable diluent or solvent, such as a solution prepared in 1,3-butanediol. In addition, sterile fixed oils can be conveniently used as solvents or suspension media. For this purpose, any blended fixed oil including synthetic mono- or diglycerides can be used. In addition, fatty acids such as oleic acid can also be used to prepare injections.

The compounds of the invention may be administered in the form of suppositories for rectal administration. These pharmaceutical compositions may be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid in the rectum and which will dissolve in the rectum to release the drug. Such materials include cocoa butter, glycerinated gelatin, hydrogenated vegetable oils, polyethylene glycols of various molecular weights and mixtures of fatty acid esters of polyethylene glycol.

It is well known to those skilled in the art that the dosage of a drug depends on a variety of factors, including but not limited to the following factors: the activity of the specific compound used, the patient's age, the patient's weight, the patient's health condition, the patient's behavior, the patient's diet, the administration time, the administration method, the excretion rate, the combination of drugs, etc. In addition, the best treatment method, such as the mode of treatment, the daily dosage of the general compound or the type of pharmaceutically acceptable salt can be verified according to traditional treatment plans.

The present invention may contain the compound represented by the general formula (I), and a pharmaceutically acceptable salt, hydrate or solvate thereof as an active ingredient, mixed with a pharmaceutically acceptable carrier or excipient to prepare a composition, and prepared into a clinically acceptable dosage form. The derivatives of the present invention may be used in combination with other active ingredients, as long as they do not produce other adverse effects, such as allergic reactions, etc. The compounds of the present invention may be used as the sole active ingredient, or in combination with other drugs for treating diseases associated with PKMYT1 activity. Combination therapy is achieved by administering the individual therapeutic components simultaneously, separately or sequentially.

Definition of terms

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

The carbon, hydrogen, oxygen, sulfur, nitrogen or halogen involved in the groups and compounds described in the present invention include their isotopes, that is, the carbon, hydrogen, oxygen, sulfur, nitrogen or halogen involved in the groups and compounds described in the present invention are optionally further replaced by one or more of their corresponding isotopes, wherein carbon isotopes include ¹² C, ¹³ C and ¹⁴ C, hydrogen isotopes include protium (H), deuterium (D, also known as heavy hydrogen), tritium (T, also known as super tritium), oxygen isotopes include ¹⁶ O, ¹⁷ O and ¹⁸ O, sulfur isotopes include ³² S, ³³ S, ³⁴ S and ³⁶ S, nitrogen isotopes include ¹⁴ N and ¹⁵ N, fluorine isotopes include ¹⁹ F, chlorine isotopes include ³⁵ Cl and ³⁷ Cl, and bromine isotopes include ⁷⁹ Br and ⁸¹ Br.

The term "alkyl" refers to a saturated aliphatic hydrocarbon group, which is a straight or branched chain group containing 1 to 20 carbon atoms, preferably an alkyl group containing 1 to 12 carbon atoms, more preferably an alkyl group containing 1 to 6 carbon atoms, an alkyl group containing 1 to 4 carbon atoms or an alkyl group containing 1 to 3 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2, 3-Dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2,2-diethylpentyl, n-decyl, 3,3-diethylhexyl, 2,2-diethylhexyl, and various branched chain isomers thereof. The alkyl group may be substituted or unsubstituted, and when substituted, the substituent may be substituted at any available point of attachment, and the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "alkylene" refers to a divalent alkyl group, wherein the alkyl group is as defined above, and has 1 to 20 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20) carbon atoms (i.e., _C1-20 alkylene). The alkylene group is preferably an alkylene group having 1 to 12 carbon atoms (i.e., _C1-12 alkylene), more preferably an alkylene group having 1 to ₆ carbon atoms (i.e., C1-6 alkylene), and further preferably an alkylene group having 1 to 4 carbon atoms (i.e., _C1-6 alkylene). Non-limiting examples of alkylene groups include, but are not limited to, methylene (—CH ₂ —), 1,1-ethylene (—CH(CH ₃ )—), 1,2-ethylene (—CH ₂ CH ₂ )—, 1,1-propylene (—CH(CH ₂ CH ₃ )—), 1,2-propylene (—CH ₂ CH(CH ₃ )—), 1,3-propylene (—CH ₂ CH ₂ CH ₂ —), 1,4-butylene (—CH ₂ CH ₂ CH ₂ CH ₂ —), and the like. The alkylene group may be substituted or unsubstituted. When substituted, it may be substituted at any available point of attachment. The substituents may be selected from one or more of alkyl, alkenyl, alkynyl, alkoxy, haloalkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio and oxo.

The term "alkenyl" refers to an alkyl group as defined above consisting of at least two carbon atoms and at least one carbon-carbon double bond, preferably an alkenyl group containing 2 to 4 carbon atoms, such as vinyl, 1-propenyl, 2-propenyl, 1-, 2- or 3-butenyl, etc. The alkenyl group may be substituted or unsubstituted, and when substituted, the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio.

The term "alkynyl" refers to an alkyl group as defined above consisting of at least two carbon atoms and at least one carbon-carbon triple bond, preferably an alkynyl group containing 2 to 4 carbon atoms or preferably an alkynyl group containing 3 to 4 carbon atoms, such as ethynyl, propynyl, butynyl, etc. Alkynyl groups may be substituted or unsubstituted, and when substituted, the substituents may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio.

The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent, the cycloalkyl ring containing 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, more preferably 3 to 6 carbon atoms or 3 to 7 carbon atoms. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, difluoro, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, etc.; polycyclic cycloalkyls include spirocyclic, fused ring and bridged ring cycloalkyls.

The term "spirocycloalkyl" refers to a polycyclic group that shares a carbon atom (called a spiral atom) between 5 to 20 monocyclic rings, which may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of spiral atoms shared between the rings, the spirocycloalkyl is divided into a single spiral cycloalkyl, a double spiral cycloalkyl or a multi-spirocycloalkyl, preferably a single spiral cycloalkyl and a double spiral cycloalkyl. More preferably, it is a 4 yuan/4 yuan, 4 yuan/5 yuan, 4 yuan/6 yuan, 5 yuan/5 yuan or 5 yuan/6 yuan single spiral cycloalkyl. Non-limiting examples of spirocycloalkyl include:

The term "condensed cycloalkyl" refers to a 5 to 20-membered, all-carbon polycyclic group in which each ring in the system shares a pair of adjacent carbon atoms with other rings in the system, wherein one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic condensed cycloalkyl, preferably a bicyclic or tricyclic, more preferably a 5-membered/5-membered or 5-membered/6-membered bicyclic alkyl. Non-limiting examples of condensed cycloalkyls include:

The term "bridged cycloalkyl" refers to a 5 to 20-membered, all-carbon polycyclic group in which any two rings share two carbon atoms that are not directly connected, which may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 members, and more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl, preferably a bicyclic, tricyclic or tetracyclic, and more preferably a bicyclic or tricyclic. Non-limiting examples of bridged cycloalkyl include:

The cycloalkyl ring may be fused to an aryl, heteroaryl or heterocyclyl ring, wherein the ring attached to the parent structure is a cycloalkyl, non-limiting examples of which include indanyl, tetrahydronaphthyl, benzocycloheptanyl, tetrahydrobenzofuranyl, tetrahydrobenzoxazolyl, tetrahydrobenzisoxazolyl, cyclopentathienyl, tetrahydrobenzothiazolyl, etc. The cycloalkyl may be optionally substituted or unsubstituted, and when substituted, the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent containing 3 to 20 ring atoms, one or more of which are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), but excluding the ring part of -OO-, -OS- or -SS-, and the remaining ring atoms are carbon. Preferably, it contains 4 to 12 ring atoms, of which 1 to 4 are heteroatoms; more preferably, it contains 4 to 7 ring atoms, of which 1 to 3 are heteroatoms, or 4 to 6 ring atoms, of which 1 to 2 are heteroatoms. Non-limiting examples of monocyclic heterocyclyls include pyrrolidinyl, imidazolidinyl, tetrahydrofuranyl, tetrahydrothienyl, dihydroimidazolyl, dihydrofuranyl, dihydropyrazolyl, dihydropyrrolyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, pyranyl, etc., preferably 1, 2, 5-oxadiazolyl, pyranyl or morpholinyl. The polycyclic heterocyclic group includes spiro, fused and bridged heterocyclic groups.

The term "spiro heterocyclic group" refers to a polycyclic heterocyclic group in which one atom (called a spiral atom) is shared between 5 to 20 monocyclic rings, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. It may contain one or more double bonds, but no ring has a completely conjugated π electron system. It is preferably 6 to 14 members, more preferably 7 to 12 members. According to the number of spiral atoms shared between rings, the spiral heterocyclic group is divided into a single spiral heterocyclic group, a double spiral heterocyclic group or a multi-spiro heterocyclic group, preferably a single spiral heterocyclic group and a double spiral heterocyclic group. More preferably, it is a 4-membered/4-membered, 4-membered/5-membered, 4-membered/6-membered, 5-membered/5-membered or 5-membered/6-membered single spiral heterocyclic group. Non-limiting examples of spiral heterocyclic groups include:

The term "fused heterocyclic group" refers to a polycyclic heterocyclic group of 5 to 20 members, each ring in the system shares a pair of atoms adjacent to other rings in the system, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron system, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. Preferably, it is 6 to 14 members, more preferably 7 to 12 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic group, preferably a bicyclic or tricyclic group, more preferably a 5-membered/5-membered or 5-membered/6-membered bicyclic fused heterocyclic group. Non-limiting examples of fused heterocyclic groups include:

The term "bridged heterocyclic group" refers to a polycyclic heterocyclic group of 5 to 14 members, wherein any two rings share two atoms that are not directly connected, which may contain one or more double bonds, but none of the rings has a completely conjugated π electron system, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. Preferably, it is 6 to 14 members, and more preferably 7 to 12 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic bridged heterocyclic group, preferably a bicyclic, tricyclic or tetracyclic group, and more preferably a bicyclic or tricyclic group. Non-limiting examples of bridged heterocyclic groups include:

The heterocyclyl ring may be fused to an aryl, heteroaryl or cycloalkyl ring, wherein the ring attached to the parent structure is the heterocyclyl.

The heterocyclyl group may be optionally substituted or unsubstituted, and when substituted, the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic (i.e., rings that share adjacent pairs of carbon atoms) group having a conjugated π electron system, preferably 6- to 10-membered, such as phenyl and naphthyl. More preferably phenyl. The aryl ring may be fused to a heteroaryl, heterocyclyl or cycloalkyl ring, wherein the ring connected to the parent structure is an aryl ring, non-limiting examples of which include:

The aryl group may be substituted or unsubstituted, and when substituted, the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

The term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 heteroatoms and 5 to 14 ring atoms, wherein the heteroatoms are selected from oxygen, sulfur and nitrogen. The heteroaryl group is preferably 5 to 10 members, containing 1 to 3 heteroatoms; more preferably 5 or 6 members, containing 1 to 2 heteroatoms; preferably, for example, imidazolyl, furanyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, pyrrolyl, tetrazolyl, pyridyl, pyrimidyl, thiadiazole, pyrazinyl, etc., preferably imidazolyl, thiazolyl, pyrazolyl or pyrimidyl, thiazolyl; more preferably pyrazolyl or thiazolyl. The heteroaryl ring can be fused to an aryl, heterocyclyl or cycloalkyl ring, wherein the ring connected to the parent structure is a heteroaryl ring, non-limiting examples of which include:

The heteroaryl group may be optionally substituted or unsubstituted, and when substituted, the substituent may be one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

The term "heteroalkyl" refers to a straight or branched chain alkyl group containing 1 to 20 carbon atoms and 1 to 3 heteroatoms selected from O, N, Si and S, wherein alkyl is as defined above, and wherein N and S may be optionally oxidized and N may be optionally quaternized.

The term "alkoxy" refers to-O-(alkyl), wherein the definition of alkyl is as described above. The non-limiting examples of alkoxy include: methoxy, ethoxy, propoxy, butoxy, cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy. Alkoxy can be optionally substituted or unsubstituted, and when substituted, substituents can be one or more of the following groups, which are independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, sulfydryl, hydroxyl, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

The term "cycloalkoxy" refers to an -O-(cycloalkyl) group, wherein cycloalkyl is as defined above.

The term "heterocycloalkoxy" refers to -O-(heterocyclyl) wherein heterocyclyl is as defined above.

The term "cycloalkylthio" refers to -S-(cycloalkyl) wherein cycloalkyl is as defined above.

The term "heterocycloalkylthio" refers to -S-(heterocyclyl) wherein heterocyclyl is as defined above.

The term "haloalkyl" refers to an alkyl group substituted with one or more halogens, wherein alkyl is as defined above.

The term "haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein alkoxy is as defined above.

The term "hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group, wherein alkyl is as defined above.

The term "hydroxy" refers to an -OH group.

The term "halogen" refers to fluorine, chlorine, bromine or iodine.

The term "amino" refers to _-NH2 .

The term "cyano" refers to -CN.

The term "nitro" refers to _-NO2 .

The term "oxo" refers to =0.

The term "carboxy" refers to -C(O)OH.

The term "thiol" refers to -SH.

The term "ester group" refers to -C(O)O(alkyl) or -C(O)O(cycloalkyl), wherein alkyl and cycloalkyl are as defined above.

The term "acyl" refers to a compound containing a -C(O)R group, wherein R is alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl.

Compounds of the present disclosure include all suitable isotopic derivatives of its compounds. The term "isotopic derivative" refers to a compound in which at least one atom is replaced by an atom having the same atomic number but different atomic masses. Examples of isotopes that can be introduced into compounds of the present disclosure include stable and radioactive isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, chlorine, bromine and iodine, such as ² H (deuterium, D), ³ H (tritium, T), ¹¹ C, ¹³ C, ¹⁴ C, ¹⁵ N, ¹⁷ O, ¹⁸ O, ³² P, ³³ P, ³³ S, ³⁴ S, ³⁵ S, ³⁶ S, ¹⁸ F, ³⁶ Cl, ⁸² Br, ¹²³ I, ¹²⁴ I, ¹²⁵ I, ¹²⁹ I and ¹³¹ I, etc., in some embodiments deuterium.

Compared with non-deuterated drugs, deuterated drugs have the advantages of reducing toxic side effects, increasing drug stability, enhancing therapeutic effects, and extending drug biological half-life. All isotopic composition changes of the compounds disclosed herein, whether radioactive or not, are included in the scope of the present disclosure. Each available hydrogen atom connected to a carbon atom can be independently replaced by a deuterium atom, wherein the replacement of deuterium can be partial or complete, and partial deuterium replacement means that at least one hydrogen is replaced by at least one deuterium.

Compounds of the present disclosure, when a position is specifically designated as "deuterium" or "D", the position is understood to have an abundance of deuterium that is at least 1000 times greater than the natural abundance of deuterium (which is 0.015%) (i.e., at least 15% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 1000 times greater than the natural abundance of deuterium (i.e., at least 15% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 2000 times greater than the natural abundance of deuterium (i.e., at least 30% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 3000 times greater than the natural abundance of deuterium (i.e., at least 45% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 3340 times greater than the natural abundance of deuterium (i.e., at least 50.1% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 3500 times greater than the natural abundance of deuterium (i.e., at least 52.5% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 4000 times greater than the natural abundance of deuterium (i.e., at least 60% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 4500 times greater than the natural abundance of deuterium (i.e., at least 67.5% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 5000 times greater than the natural abundance of deuterium (i.e., at least 75% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 5500 times greater than the natural abundance of deuterium (i.e., at least 82.5% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 6000 times greater than the natural abundance of deuterium (i.e., at least 90% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 6333.3 times greater than the natural abundance of deuterium (i.e., at least 95% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 6466.7 times greater than the natural abundance of deuterium (i.e., at least 97% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 6600 times greater than the natural abundance of deuterium (i.e., at least 99% deuterium incorporation). In some embodiments, the abundance of deuterium for each designated deuterium atom is at least 6633.3 times greater than the natural abundance of deuterium (ie, at least 99.5% deuterium incorporation).

"Optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and the description includes instances where the event or circumstance occurs or does not occur. For example, "a heterocyclic group optionally substituted with an alkyl group" means that an alkyl group may but need not be present, and the description includes instances where the heterocyclic group is substituted with an alkyl group and instances where the heterocyclic group is not substituted with an alkyl group.

"Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms in the group are replaced independently of each other by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and the skilled person can determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, amino or hydroxy groups with free hydrogens may be unstable when combined with carbon atoms with unsaturated (e.g. olefinic) bonds.

"Pharmaceutical composition" means a mixture containing one or more compounds described herein or their physiologically/pharmaceutically acceptable salts or prodrugs and other chemical components, as well as other components such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitate the absorption of the active ingredient, and thus exert biological activity.

"Pharmaceutically acceptable salt" or "pharmaceutically acceptable salt" refers to salts of the compounds of the present invention, which are safe and effective when used in mammals and have the desired biological activity.

"Carrier" refers to a carrier or diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound.

Synthesis method of the compound of the present invention

Another aspect of the present invention provides a method for preparing the compound represented by general formula (II) according to the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or pharmaceutically acceptable salt thereof, using the following technical scheme:

Solution 1

The compound represented by general formula A-1 or its salt undergoes substitution reaction with the compound represented by general formula A-2 in an organic solvent under alkaline conditions, optionally in the presence of a catalyst, to obtain the compound represented by general formula B-1 or its pharmaceutically acceptable salt.

The organic solvent is a physical mixture of one or more solvents, including methanol, ethanol, dichloromethane, N,N-dimethylformamide, toluene, acetonitrile, 1,4-dioxane, tetrahydrofuran, ethyl acetate, etc., preferably 1,4-dioxane and toluene; the base includes organic bases and inorganic bases, preferably potassium carbonate and cesium carbonate; the catalyst is a combination of a palladium catalyst and a ligand, and the palladium catalyst includes tetrakistriphenylphosphine palladium, trisdibenzylideneacetone dipalladium, [1,1'- Bis(diphenylphosphino)ferrocene] palladium dichloride, palladium acetate, etc., preferably tris dibenzylideneacetone dipalladium, ligands include 4,5-bis(diphenylphosphino)ferrocene-9,9-dimethyloxanthene, dicyclohexyl(2',4',6'-triisopropyl-3,6-dimethoxy-[1,1'-biphenyl]-2-yl)phosphine, isopropylbiphenyl 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl, etc., preferably 4,5-bis(diphenylphosphino)phosphino-9,9-dimethyloxanthene.

Solution 2

The compound represented by the general formula B-1 or its salt reacts with malondicyandiamide in an organic solvent under alkaline conditions, optionally in the presence of a catalyst, to obtain the compound represented by the general formula C-1 or its pharmaceutically acceptable salt.

The organic solvent is a physical mixture of one or more solvents, including methanol, ethanol, dichloromethane, N,N-dimethylformamide, toluene, acetonitrile, 1,4-dioxane, tetrahydrofuran, ethyl acetate, etc., preferably 1,4-dioxane and toluene; the base includes organic bases and inorganic bases, preferably potassium tert-butoxide and sodium hydride; the catalyst is a palladium catalyst and a copper catalyst, the palladium catalyst includes tetrakistriphenylphosphine palladium, trisdibenzylideneacetone dipalladium, [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, palladium acetate, etc., the copper catalyst includes cuprous iodide, copper sulfate, etc., preferably [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride.

Solution 3

The compound represented by the general formula C-1 or its salt undergoes a hydrolysis reaction in a solvent to obtain the compound represented by the general formula D-1 or its pharmaceutically acceptable salt.

The solvent is a physical mixture of one or more solvents, including organic solvents and inorganic solvents. The organic solvents include methanol, ethanol, dichloromethane, N,N-dimethylformamide, dimethyl sulfoxide, toluene, acetonitrile, 1,4-dioxane, tetrahydrofuran, ethyl acetate, etc., and the inorganic solvents include water, concentrated sulfuric acid, etc., preferably concentrated sulfuric acid.

Solution 4

The compound represented by the general formula D-1 or its salt is reacted in a solvent, optionally in the presence of a catalyst, to obtain the compound represented by the general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or a mixture thereof, or a pharmaceutically acceptable salt thereof.

The solvent is a physical mixture of one or more solvents, including organic solvents and inorganic solvents. The organic solvents include methanol, ethanol, dichloromethane, N,N-dimethylformamide, dimethyl sulfoxide, toluene, acetonitrile, 1,4-dioxane, tetrahydrofuran, ethyl acetate, etc., and the inorganic solvents include water, etc., preferably dichloromethane; the catalyst includes hydrogen bromide, boron tribromide, etc., preferably boron tribromide.

wherein Y ₁ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in the general formula (II).

DETAILED DESCRIPTION

The present invention is further described below in conjunction with examples, but these examples are not intended to limit the scope of the present invention.

The structure of the compound is determined by nuclear magnetic resonance (NMR) and/or mass spectrometry (MS). NMR shifts are given in units of 10 ^-6 (ppm). NMR measurements are performed using a Brukerdps300 NMR spectrometer, with deuterated dimethyl sulfoxide (DMSO-d ₆ ), deuterated chloroform (CDCl ₃ ), deuterated methanol (CD ₃ OD) as the measuring solvent, and tetramethylsilane (TMS) as the internal standard.

LC-MS measurements were performed using an 1100 Series LC/MSD Trap (ESI) mass spectrometer (manufacturer: Agilent).

GC-MS determination was performed using GCMS-QP2010 SE.

The preparative liquid chromatography method used LC3000 high performance liquid chromatograph and LC6000 high performance liquid chromatograph (manufacturer: Chuangxin Tongheng). The chromatographic column was Daisogel C18 10μm 60A (20mm×250mm).

High performance liquid chromatography (HPLC) was performed using a Shimadzu LC-20AD high pressure liquid chromatograph (Agilent TC-C18 250×4.6mm 5μm column) and a Shimadzu LC-2010AHT high pressure liquid chromatograph (Phenomenex C18 250×4.6mm 5μm column).

The thin layer chromatography silica gel plate used was Qingdao Ocean Chemical GF254 silica gel plate. The silica gel plate used in thin layer chromatography (TLC) had a specification of 0.15 mm to 0.2 mm, and the specification used for thin layer chromatography separation and purification products was 0.4 mm to 0.5 mm.

Column chromatography generally uses Qingdao marine silica gel 100-200 mesh and 200-300 mesh silica gel as the carrier.

The known starting materials of the present invention can be synthesized by methods known in the art, or can be purchased from online shopping malls, Beijing Coupling, Sigma, Bailingwei, Yishiming, Shanghai Shuya, Inokai, Nanjing Yaoshi, Anaiji Chemical and other companies.

Unless otherwise specified in the examples, the reactions can be carried out under an argon atmosphere or a nitrogen atmosphere.

Argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a capacity of about 1L.

Microwave reactions were performed using a CEM Discover SP microwave reactor.

Unless otherwise specified in the examples, the solution refers to an aqueous solution.

Unless otherwise specified in the examples, the reaction temperature is room temperature, particularly 20°C to 30°C.

The reaction progress in the embodiment is monitored by thin layer chromatography (TLC), and the developing solvent systems used in the reaction are: A: dichloromethane and methanol system, B: n-hexane and ethyl acetate system, C: petroleum ether and ethyl acetate system, D: acetone, and the volume ratio of the solvent is adjusted according to the polarity of the compound.

The eluent system of column chromatography and the developing solvent system of thin layer chromatography used for purifying compounds include: A: dichloromethane and methanol system, B: petroleum ether, ethyl acetate and dichloromethane system, C: petroleum ether and ethyl acetate system. The volume ratio of the solvent is adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid can also be added for adjustment.

Unless otherwise defined, all professional and scientific terms used herein have the same meanings as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described herein can be applied to the present invention.

Example

Example 1: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (1)

Step 1: Preparation of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1,5-dimethyl-1H-pyrazol-3-amine (1a)

At room temperature, 4-bromo-1,5-dimethyl-1H-pyrazol-3-amine (1.90 g, 10.0 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (2.62 g, 10.0 mmol) were dissolved in 1,4-dioxane (38 mL), and then cesium carbonate (6.51 g, 20.0 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (694 mg, 1.20 mmol) and tris(dibenzylideneacetone)dipalladium (916 mg, 1.00 mmol) were added, and the mixture was stirred at 110° C. for 16 hours under a nitrogen atmosphere. Dilute with water (500 mL), extract with ethyl acetate (500 mL x 2), combine the organic phases, wash with saturated sodium chloride solution (1.0 L x 1), dry over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. The residue is separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 15%-25%) to obtain 1.30 g of the title compound as a yellow solid. Yield: 40.1%.

LC-MS: m/z=325 [M+H] ⁺ .

Step 2: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (1b)

Potassium carbonate (1.46 g, 10.6 mmol), L-proline (81.0 mg, 703 μmol) and cuprous iodide (70.0 mg, 352 μmol) were added to a solution of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1,5-dimethyl-1H-pyrazol-3-amine (1.14 g, 3.52 mmol) and malononitrile (279 mg, 4.22 mmol) in dimethyl sulfoxide (11 mL) at room temperature. The mixture was stirred at 90°C for 14 hours under a nitrogen atmosphere. Dilute with water (250 mL), extract with ethyl acetate (250 mL x 2), combine the organic phases, wash with saturated sodium chloride solution (500 mL x 1), dry over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. The residue is separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 35%-55%) to give 164 mg of the title compound as a yellow solid. Yield: 15.1%.

LC-MS: m/z=310 [M+H] ⁺ .

Step 3: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (1c)

5-Amino-6-(3-methoxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (164 mg, 530 μmol) was dissolved in concentrated sulfuric acid (3.0 mL) at 0°C and stirred at room temperature for 2 hours. The reaction solution was poured into ice water (100 mL), the pH was adjusted to 8-9 with aqueous ammonia, and extracted with ethyl acetate (100 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution (200 mL x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 112 mg of the title compound as a yellow solid, with a yield of 64.5%.

LC-MS: m/z=328 [M+H] ⁺ .

Step 4: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (1)

To a solution of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2,3-dimethyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (112 mg, 342 μmol) in dichloromethane (4.0 mL) was added a 1 M solution of boron tribromide in dichloromethane (1.0 mL, 1.00 mmol) at -78°C, and the mixture was stirred at room temperature for 2 hours under a nitrogen atmosphere. The reaction solution was concentrated under reduced pressure, and the residue was separated by preparative liquid chromatography (chromatographic column model: Daisogei 30 mm × 250 mm, C18, 10 um, 100 A, mobile phase: acetonitrile/water, gradient: 30%-80%) to obtain 20.0 mg of the title compound as a white solid, with a yield of 18.7%.

LC-MS: m/z=314 [M+H] ⁺ .

¹ H NMR (400MHz, CDCl ₃ ) δ 6.86 (d, J = 8.4 Hz, 1H), 6.65 (d, J = 8.4 Hz, 1H), 3.86 (s, 3H), 2.62 (s, 3H), 1.96 (s, 3H), 1.84 (s, 3H).

Example 2: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (2)

Step 1: Preparation of 4-bromo-1-methyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (2a)

At 0°C, N-bromosuccinimide (NBS) (1.29 g, 7.27 mmol) was added to a solution of 1-methyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (1.0 g, 6.06 mmol) in acetonitrile (10 mL), and the reaction solution was stirred at room temperature for 0.5 hours. The reaction solution was diluted with ethyl acetate (100 mL), concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 20%-35%) to obtain 1.34 g of the title compound as a yellow solid, with a yield of 90.7%.

LC-MS: m/z=244,246[M+H] ⁺ .

Step 2: Preparation of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (2b)

To a solution of 4-bromo-1-methyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (1.14 g, 4.67 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (1.22 g, 4.67 mmol) in 1,4-dioxane (22 mL) were added cesium carbonate (3.04 g, 9.34 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (541 mg, 934 μmol) and tetrakis(dibenzylideneacetone)dipalladium (532 mg, 467 μmol) at room temperature, and the reaction solution was stirred at 110° C. for 16 hours under a nitrogen atmosphere. The reaction solution was diluted with water (500 mL) and extracted with ethyl acetate (500 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution (1.0 L x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 15%-25%) to obtain 1.29 g of the title compound as a yellow solid. Yield: 73.0%.

LC-MS: m/z=378,380[M+H] ⁺ .

Step 3: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (2c)

To a solution of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (1.00 g, 2.64 mmol) and malononitrile (210 mg, 3.17 mmol) in dimethyl sulfoxide (10 mL) were added potassium carbonate (1.10 g, 7.93 mmol), L-proline (61.0 mg, 529 μmol) and cuprous iodide (50.0 mg, 264 μmol) at room temperature, and the reaction solution was stirred at 90 ° C for 16 hours under nitrogen atmosphere. The reaction solution was diluted with water (250 mL), extracted with ethyl acetate (250 mL x 2), the organic phases were combined, washed with saturated sodium chloride solution (500 mL x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 35%-55%) to obtain 176 mg of the title compound as a yellow solid. Yield: 18.3%.

LC-MS: m/z=364 [M+H] ⁺ .

Step 4: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (2d)

5-Amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (141 mg, 388 μmol) was dissolved in concentrated sulfuric acid (4.0 mL) at 0°C and stirred at room temperature for 2 hours. The reaction solution was added dropwise to ice water (100 mL), the pH was adjusted to 8-9 with aqueous ammonia, and extracted with ethyl acetate (100 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution (200 mL x 1), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain 76.0 mg of the title compound as a yellow solid, with a yield of 51.4%.

LC-MS: m/z=382 [M+H] ⁺ .

Step 5: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (2)

At -78°C, 1M dichloromethane solution of boron tribromide (598μL, 598μmol) was added dropwise to a dichloromethane (6.0mL) solution of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (76.0mg, 199μmol), and the reaction solution was stirred at room temperature for 2 hours under nitrogen atmosphere. The reaction solution was concentrated under reduced pressure, and the residue was separated by preparative liquid chromatography (chromatographic column model: Daisogei 30mm×250mm, C18, 10um, 100A, mobile phase: acetonitrile/water, gradient: 30%-80%) to obtain 20.0mg of the title compound as a white solid. Yield: 27.3%.

LC-MS: m/z=368 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.57(s,1H),7.05(d,J＝8.4Hz,1H),6.99(s,2H),6.90(d,J＝8.4Hz,1H),6.26(s,2H),3.87(d,J＝1.6Hz,3H),1.85(s,3H),1.76(s,3H).

Example 3: Preparation of S-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide and R-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (2-1 and 2-2)

Compound 2 was separated by SFC chiral preparative separation under the following conditions: equipment: SFC-150 mgm (waters), chiral column: CHIRALPAK IJ (20×250 mm, 5 um), temperature: 25° C., mobile phase: Hex (0.5% 2 mM NH ₃ -MeOH)/EtOH=65/35, flow rate: 20 mL/min, back pressure: 100 bar, detection wavelength: 220 nm, cycle time: 5 min, to obtain 2 isomers: compound 2-1 (RT=2.774 min) and compound 2-2 (RT=3.089 min).

Compound 2-1: LC-MS: m/z=368 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.57(s,1H),7.05(d,J＝8.4Hz,1H),6.99(s,2H),6.90(d,J＝8.4Hz,1H),6.26(s,2H),3.87(d,J＝1.6Hz,3H),1.85(s,3H),1.76(s,3H).

Compound 2-2: LC-MS: m/z=368 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.57(s,1H),7.05(d,J＝8.4Hz,1H),6.99(s,2H),6.90(d,J＝8.4Hz,1H),6.26(s,2H),3.87(d,J＝1.6Hz,3H),1.85(s,3H),1.76(s,3H).

Example 4: Preparation of 5-amino-2-ethyl-6-(3-hydroxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (3)

Step 1: Preparation of 2-(5-(trifluoromethyl)-1H-pyrazol-3-yl)isoindole-1,3-dione (3a)

At room temperature, 5-(trifluoromethyl)-1H-pyrazole-3-amine (10.0 g, 66.2 mmol) and phthalic anhydride (9.80 g, 66.2 mmol) were dissolved in 1,4-dioxane (100 mL) and reacted at 80°C for 10 hours. The reaction solution was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 100:1-3:1) to obtain 12.0 g of the title compound as a yellow solid, with a yield of 64.4%.

LC-MS: m/z=282 [M+H] ⁺ .

Step 2: Preparation of 2-(1-ethyl-5-trifluoromethyl)-1H-pyrazol-3-ylisoindole-1,3-dione (3b)

At room temperature, 2-(5-(trifluoromethyl)-1H-pyrazol-3-yl)isoindole-1,3-dione (10.0 g, 35.4 mmol) was dissolved in N,N-dimethylformamide (100 mL), potassium carbonate (14.7 g, 106 mmol) was added, and the reaction was allowed to react at room temperature overnight. 300 mL of water was added, and the mixture was extracted with ethyl acetate (150 mL x 3). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography to obtain 8.33 g of the title compound as a yellow solid in a yield of 91.6%.

LC-MS: m/z=310 [M+H] ⁺ .

Step 3: Preparation of 2-(4-bromo-1-ethyl-5-(trifluoromethyl)-1H-pyrazol-3-yl)isoindole-1,3-dione (3c)

At room temperature, 2-(1-ethyl-5-trifluoromethyl)-1H-pyrazol-3-ylisoindole-1,3-dione (8.20 g, 26.4 mmol) was dissolved in acetonitrile (80 mL), and N-bromosuccinimide (5.70 g, 31.7 mmol) was added, and the mixture was reacted at room temperature for 16 hours. 200 mL of water was added, and the mixture was extracted with ethyl acetate (150 mL x 3). The organic phase was washed with saturated NaCl solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography to obtain 3.20 g of the title compound as a yellow solid, with a yield of 40.0%.

LC-MS: m/z=388 [M+H] ⁺ .

Step 4: Preparation of 4-bromo-1-ethyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (3d)

At room temperature, 2-(4-bromo-1-ethyl-5-(trifluoromethyl)-1H-pyrazol-3-yl)isoindole-1,3-dione (3.00 g, 7.73 mmol) was dissolved in ethanol (30 mL), and 80% hydrazine hydrate (10 mL) was added, and the mixture was reacted at room temperature for 16 hours. The reaction solution was filtered through celite, and the filtrate was collected and concentrated under reduced pressure to obtain 620 mg of the title compound as a solid (crude product).

LC-MS: m/z=258 [M+H] ⁺ .

Step 5: Preparation of 4-bromo-1-ethyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazol-3-amine (3e)

At room temperature, 4-bromo-1-ethyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (600 mg, 2.32 mmol) was dissolved in N,N-dimethylformamide (4 mL), and 2-iodo-4-methoxy-1,3-dimethylbenzene (609 mg, 2.32 mmol), cesium carbonate (2.27 g, 6.96 mmol), methanesulfonic acid (2-dicyclohexylphosphine)-3,6-dimethoxy-2',4',6'-triisopropyl-1,1'-biphenyl)(2'-amino-1,1'-biphenyl-2-yl)palladium(II) (210 mg, 0.230 m mol) and 2-(dicyclohexylphosphino)-3,6-dimethoxy-2'-4'-6'-tri-I-propyl-11'-biphenyl (214 mg, 0.460 mmol), replaced with nitrogen three times, stirred at 110°C under nitrogen atmosphere for 16 hours, added with 50 mL of water, extracted with ethyl acetate (30 mL x 2), the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate: petroleum ether = 15%-25%) to give 300 mg of the title compound as a yellow solid, yield: 32.8%.

LC-MS: m/z=392 [M+H] ⁺ .

Step 6: Preparation of 5-amino-2-ethyl-6-(3-methoxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (3f)

Malononitrile (45.9 mg, 0.696 mmol) was dissolved in 1,4-dioxane (4 mL) at room temperature, sodium hydride (30.6 mg, 0.766 mmol) was added, and the mixture was stirred for 30 minutes. 4-Bromo-1-ethyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazol-3-amine (136 mg, 0.348 mmol), (1R,2R)-(-)-N,N'-dimethyl-1,2-cyclohexanediamine (99.4 mg, 0.700 mmol) and CuI (6.70 mg, 0.035 mmol) were added, and the atmosphere was replaced with nitrogen three times. The mixture was reacted at 100°C under a nitrogen atmosphere for 16 hours, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 20:1-3:1) to give 150 mg of the title compound as a solid (crude product).

LC-MS: m/z=378 [M+H] ⁺ .

Step 7: Preparation of 5-amino-2-ethyl-6-(3-methoxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (3 g)

At room temperature, 5-amino-2-ethyl-6-(3-methoxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrole[2,3-c]pyrazole-4-carbonitrile (150 mg, 0.397 mmol) was added to concentrated sulfuric acid (4.0 mL), stirred for 2 hours, added to ice water (100 mL), and the pH was adjusted to 8-9 with aqueous ammonia. The mixture was extracted with ethyl acetate (50 mL x 2), and the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 80.0 mg of the title compound as a yellow solid. Yield: 51.4%.

LC-MS: m/z=396 [M+H] ⁺ .

Step 8: Preparation of 5-amino-2-ethyl-6-(3-hydroxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (3)

5-amino-2-ethyl-6-(3-hydroxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (80.0 mg, 0.202 mmol) was dissolved in dichloromethane (2 mL) at 0°C, and boron tribromide (0.516 ml, 0.516 mmol, 1 M DCM solution) was slowly added dropwise, and the mixture was reacted at room temperature for 2 hours. Methanol was added to quench the reaction, and the mixture was concentrated under reduced pressure. The residue was separated by high pressure preparative liquid chromatography (chromatographic column model: Daisogei 30 mm × 250 mm, C18, 10 um 100A, mobile phase: acetonitrile/water, gradient: 10%-50%, 0.05% formic acid) to obtain 12.0 mg of the title compound as a white solid, with a yield of 14.5%.

LC-MS: m/z=382 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d6) δ9.28(s,1H),7.57(d,J＝4.2Hz,1H),7.47–7.38(m,2H),6.86(d,J＝8.2Hz,1H),6.58(d,J＝8.2Hz,1 H), 4.45 (d, J = 19.3Hz, 1H), 3.85 (qd, J = 7.2, 4.5Hz, 2H), 2.03 (d, J = 4.5Hz, 3H), 1.87 (s, 3H), 1.23 (dd, J = 7.4, 4.1Hz, 3H).

Example 5: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-isopropyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (4)

Referring to the preparation method of Example 4, the iodoethane in step 2 was replaced by iodoisopropylane to obtain the title compound 4.

LC-MS: m/z=395.9[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ7.53(s,2H),7.39(d,J＝13.1Hz,2H),6.75(s,2H),5.60(s,1H),4.37(p,J＝6.4Hz,1H),2.18(d,J＝4.5Hz,6H),1.35(d,J＝6.5Hz,6H).

Example 6: Preparation of 5-amino-2-cyclopropyl-6-(3-hydroxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (5)

Referring to the preparation method of Example 4, the title compound 5 was prepared by replacing iodoethane in step 2 with iodocyclopropane.

LC-MS: m/z=394 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.66(s,1H),7.10(d,J＝8.3Hz,1H),6.95(d,J＝8.3Hz,1H),6.54(s,2H),6.20(s,2H),2. 97(tt,J＝7.3,3.6Hz,1H),1.91(s,3H),1.83(s,3H),0.84–0.74(m,2H),0.49–0.40(m,2H).

Example 7: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-phenyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (6)

Step 1: Preparation of 1-phenyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (6a)

At room temperature, (E)-4-amino-4-ethoxy-1,1,1-trifluorobutyl-3-ene-2-one (3.50 g, 19.1 mmol) and triethylamine (3.86 g, 38.2 mmol) were added to a solution of phenylhydrazine (3.33 g, 23.0 mmol) in ethanol (20 mL), and the mixture was stirred at 95°C for 16 hours. 50 mL of water was added to the reaction solution, and the mixture was extracted with dichloromethane (50 mL x 2). The organic phase was washed with a saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether: ethyl acetate = 19:1) to obtain 1.70 g of the title compound as a yellow solid, with a yield of 39.2%.

LC-MS: m/z=227.9[M+H] ⁺ .

Step 2: Preparation of 1-phenyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazol-3-amine (6b)

To a solution of 1-phenyl-5-(trifluoromethyl)-1H-pyrazol-3-amine (1.70 g, 7.49 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (2.36 g, 8.99 mmol) in 1,4-dioxane (30 mL) was added cesium carbonate (6.10 g, 18.7 mmol), 2-(dicyclohexylphosphino)-3,6-dimethoxy-2'-4'-6'-tri-1-propyl-11'-biphenyl (1.21 g, 2 .25mmol) and tris(dibenzylideneacetone)dipalladium (1.03g, 1.12mmol), stirred at 100℃ under nitrogen atmosphere for 16 hours, added 100mL of water, extracted with dichloromethane (50mL x 2), the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 19:1) to give 2.60g of the title compound as a white solid, yield: 96.1%.

LC-MS: m/z=361.9[M+H] ⁺ .

Step 3: Preparation of 4-bromo-1-phenyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazol-3-amine (6c)

At 0°C, 1-phenyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazole-3-amine (2.60 g, 7.20 mmol) was dissolved in 20 mL of N,N-dimethylformamide, and N-bromosuccinimide (1.54 g, 8.64 mmol) was added in batches. The mixture was stirred at room temperature for 1 hour, and 100 mL of water was added. The mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 3.10 g of the title compound as a brown oil. Yield: 98.1%.

LC-MS: m/z=439.9[M+H] ⁺ .

Step 4: Preparation of 5-amino-2-phenyl-6-(3-methoxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (6d)

Malononitrile (933 mg, 14.1 mmol) was dissolved in ethylene glycol dimethyl ether (6 mL) at room temperature, sodium hydride (1.13 g, 28.2 mmol) was added, and the mixture was stirred for 30 minutes. 4-Bromo-1-phenyl-N-(3-methoxy-2,6-dimethylphenyl)-5-(trifluoromethyl)-1H-pyrazole-3-amine (3.10 g, 7.06 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (775 mg, 1.06 mmol) were added, and the mixture was replaced with nitrogen three times. The mixture was stirred at 100°C under nitrogen atmosphere for 16 hours, 30 mL of water was added, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether:ethyl acetate = 6:1) to give 1.90 g of the title compound as a yellow solid. Yield: 63.3%.

LC-MS: m/z=425.9[M+H] ⁺ .

Step 5: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-phenyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (6e).

At room temperature, 5-amino-2-phenyl-6-(3-methoxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (1.00 g, 2.35 mmol) was added to 4 mL of concentrated sulfuric acid and stirred for 2 hours. 100 mL of ice water was added and the pH was adjusted to 8-9 with aqueous ammonia. The mixture was extracted with ethyl acetate (30 mL x 2). The organic layer was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 6:5) to give 300 mg of the title compound as a yellow oil. Yield: 28.8%.

LC-MS: m/z=444 [M+H] ⁺ .

Step 6: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-phenyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (6)

At 0°C, 1M boron tribromide dichloromethane solution (4.50 mL, 4.50 mmol) was slowly added to a dichloromethane (5 mL) solution of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-phenyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (200 mg, 0.451 mmol), and the mixture was stirred at room temperature for 16 hours. Methanol was added to quench the reaction, and the mixture was concentrated under reduced pressure. The residue was separated and purified by preparative liquid chromatography (chromatographic column model: Daisogei 30 mm×250 mm, C18, 10 um, 100A, mobile phase: acetonitrile/water, gradient: 30%-80%) to obtain 24.0 mg of the title compound as a white solid. Yield: 12.4%.

LC-MS: m/z=429.90[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d6) δ8.98 (s, 1H), 7.76–7.60 (m, 3H), 7.41 (d, J = 8.3Hz, 2H), 7.37–7.25 (m, 3H), 6.60 (t, J = 10.7Hz, 1H), 6.34 (d, J = 8.1Hz, 1H), 4.82 (d, J = 13.1Hz, 1H), 2.00–1.77 (m, 6H).

Example 8: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-3-methyl-6H-pyrrolo[2,3-c]isoxazole-4-carboxamide (7)

Step 1: Preparation of 4-bromo-5-methylisoxazol-3-amine (7a)

At room temperature, 5-methylisoxazol-3-amine (0.784 g, 8.00 mmol) was dissolved in 20 mL of N,N-dimethylformamide. N-bromosuccinimide (1.44 g, 8.10 mmol) was added in batches. The mixture was stirred at room temperature for 1 hour. 100 mL of water was added and the mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 1.25 g of the title compound as a brown oil. Yield: 88.2%.

LC-MS: m/z=177,179[M+H] ⁺ .

Step 2: Preparation of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-5-methylisoxazol-3-amine (7b)

To a solution of 4-bromo-5-methylisoxazol-3-amine (1.25 g, 7.06 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (1.85 g, 7.06 mmol) in 1,4-dioxane (12 mL) was added cesium carbonate (4.60 g, 14.1 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (817 mg, 1.41 mmol) and tris(dibenzylideneacetone) dihydrate at room temperature. Palladium (804 mg, 706 μmol) was replaced with nitrogen three times, stirred at 110 °C under nitrogen atmosphere for 16 hours, diluted with 100 mL of water, extracted with ethyl acetate (50 mL x 2), and the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 15%-25%) to obtain 450 mg of the title compound as a yellow solid. Yield: 20.5%.

LC-MS: m/z=311,313[M+H] ⁺ .

Step 3: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-3-methyl-6-pyrrolo[2,3-c]isoxazole-4-carbonitrile (7c)

To a solution of malononitrile (191 mg, 2.89 mmol) in ethylene glycol dimethyl ether (15 mL) was added sodium hydride (60%, 116 mg, 2.89 mmol) at room temperature, and the mixture was stirred for 30 minutes. 4-Bromo-N-(3-methoxy-2,6-dimethylphenyl)-5-methylisoxazol-3-amine (450 mg, 1.45 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (106 mg, 145 μmol) were added. The atmosphere was replaced with nitrogen three times, and the mixture was stirred at 110° C. under a nitrogen atmosphere for 16 hours. The mixture was concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether:ethyl acetate=55%-65%) to give 420 mg of the title compound as a yellow solid. Yield: 98.0%.

LC-MS: m/z=297 [M+H] ⁺ .

Step 4: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-3-methyl-6-pyrrolo[2,3-c]isoxazole-4-carboxamide (7d)

At room temperature, 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-3-methyl-6-pyrrolo[2,3-c]isoxazole-4-carbonitrile (420 mg, 1.42 mmol) was added to 2 mL of sulfuric acid and stirred for 2 hours. 100 mL of ice water was added and the pH was adjusted to 8-9 with aqueous ammonia. The mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 97 mg of the title compound as a yellow solid. Yield: 21.8%.

LC-MS: m/z=315 [M+H] ⁺ .

Step 5: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-3-methyl-6-pyrrolo[2,3-c]isoxazole-4-carboxamide (7)

Boron tribromide (926 μL, 926 μmol, 1M DCM solution) was added dropwise to a solution of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-3-methyl-6-pyrrolo[2,3-c]isoxazole-4-carboxamide (97.0 mg, 308 μmol) in dichloromethane (6 mL) at 0°C, stirred at room temperature for 2 hours, concentrated under reduced pressure, and the residue was separated by preparative liquid chromatography (chromatographic column model: Daisogei 30 mm×250 mm, C18, 10 um, 100A, mobile phase: acetonitrile/water, gradient: 30%-80%) to obtain 10.0 mg of the title compound as a white solid. Yield: 10.8%.

LC-MS: m/z=301.85[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.57(s,1H),8.03(d,J＝26.0Hz,2H),7.03(d,J＝8.4Hz,1H),6.85(d,J＝8.4Hz,1H),5.34( qd,J＝6.8,2.4Hz,1H),1.87(d,J＝3.2Hz,3H),1.79(d,J＝3.6Hz,3H),1.51(d,J＝6.8Hz,3H).

Example 9: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-3-(trifluoromethyl)-6H-pyrrolo[2,3-c]isoxazole-4-carboxamide (8)

Referring to the preparation method of Example 8, 5-methylisoxazol-3-amine in step 1 was replaced with 5-(trifluoromethyl)isoxazol-3-amine to obtain the title compound 8.

LC-MS: m/z=355.1[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.57(s,1H),8.05(d,J＝26.0Hz,2H),7.04(d,J＝8.3Hz,1H),6.87(d,J＝8.3Hz ,1H),5.36(qd,J＝6.7,2.4Hz,1H),1.89(d,J＝3.2Hz,3H),1.76(d,J＝3.6Hz,3H).

Example 10: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-(methyl-d3)-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (9)

Step 1: Preparation of 1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (9a)

Triethylamine (17.9 mL, 128 mmol) was added to a solution of (E)-4-amino-4-ethoxy-1,1,1-trifluorobut-3-en-2-one (11.8 g, 64.2 mmol) and (methyl-d ₃ )hydrazine hydrochloride (6.04 g, 70.6 mmol) in ethanol (220 mL) at room temperature, and the mixture was stirred at 85° C. for 16 hours. The mixture was concentrated under reduced pressure, 300 mL of water was added, and the mixture was extracted with ethyl acetate (150 mL x 3). The organic phase was washed with a saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate: petroleum ether = 15%-45%) to obtain 2.11 g of the title compound as a yellow oil, with a yield of 19.5%.

LC-MS: m/z=169 [M+H] ⁺ .

Step 2: Preparation of 4-bromo-1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (9b)

To a solution of 1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (2.11 g, 12.6 mmol) in acetonitrile (40 mL) was added N-bromosuccinimide (2.68 g, 15.1 mmol) at room temperature. The mixture was stirred for 2 hours and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 20%-35%) to give 1.39 g of the title compound as a yellow solid. Yield: 44.8%.

LC-MS: m/z=247,249[M+H] ⁺ .

Step 3: Preparation of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (9c)

To a solution of 4-bromo-1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (1.39 g, 5.63 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (1.47 g, 5.63 mmol) in 1,4-dioxane (28 mL) were added cesium carbonate (3.67 g, 11.3 mmol), 4,5-bisdiphenylphosphino-9,9-dimethylxanthene (651 mg, 1.13 mmol) and tris(dibenzylideneacetone)dipalladium (640 mg, 563 μmol) at room temperature, the atmosphere was replaced with nitrogen three times and the mixture was stirred at 100° C. under nitrogen atmosphere for 16 hours. 100 mL of water was added, and the mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 15%-25%) to obtain 1.70 g of the title compound as a yellow solid. Yield: 79.3%.

LC-MS: m/z=381,383[M+H] ⁺ .

Step 4: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-(methyl-d ₃ )-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (9d)

Potassium carbonate (1.00 g, 7.24 mmol), L-proline (278 mg, 2.41 mmol) and cuprous iodide (230 mg, 1.21 mmol) were added to a solution of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-(methyl-d ₃ )-5-(trifluoromethyl)-1H-pyrazol-3-amine (920 mg, 2.41 mmol) and malononitrile (239 mg, 3.62 mmol) in dimethyl sulfoxide (14 mL) at room temperature. The atmosphere was replaced with nitrogen three times and stirred at 100° C. under nitrogen atmosphere for 16 hours. 100 mL of water was added and the mixture was extracted with ethyl acetate (50 mL x 100 mL). 2), the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 35%-55%) to obtain 124 mg of the title compound as a yellow solid, yield: 14.0%.

LC-MS: m/z=367 [M+H] ⁺ .

Step 5: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-(methyl-d ₃ )-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (9e)

At room temperature, 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-(methyl- _d3 )-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (124 mg, 328 μmol) was added to 4 mL of concentrated sulfuric acid and stirred for 2 hours. 100 mL of ice water was added and the pH was adjusted to 8-9 with aqueous ammonia. The mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 106 mg of the title compound as a yellow solid. Yield: 81.5%.

LC-MS: m/z=385 [M+H] ⁺ .

Step 6: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-(methyl-d ₃ )-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (9)

Boron tribromide (968 μL, 968 μmol, 1 M DCM solution) was added dropwise to a solution of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-(methyl-d ₃ )-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (93.0 mg, 242 μmol) in dichloromethane (4 mL) at 0°C, and the mixture was stirred at room temperature for 3 hours, concentrated under reduced pressure, and the residue was separated and purified by preparative liquid chromatography (chromatographic column model: Daisogei 30 mm×250 mm, C18, 10 um, 100A, mobile phase: acetonitrile/water, gradient: 30%-80%) to obtain 27.0 mg of the title compound as a white solid. Yield: 30.1%.

LC-MS: m/z=370.85[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ 9.55 (s, 1H), 7.05 (d, J = 8.4Hz, 1H), 6.99 (s, 2H), 6.90 (d, J = 8.4Hz, 1H), 6.26 (s, 2H), 1.85 (s, 3H), 1.76 (s, 3H).

Example 11: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (10)

Referring to the preparation method of Example 1, 4-bromo-1,5-dimethyl-1H-pyrazol-3-amine in step 1 was replaced with 4-bromo-1-methyl-1H-pyrazol-3-amine to prepare the title compound 10.

LC-MS: m/z=299 [M+H] ⁺ .

¹ H NMR (400MHz, CDCl ₃ ) δ7.14 (s, 1H), 6.98 (d, J = 8.8 Hz, 1H), 6.76 (d, J = 8.3 Hz, 1H), 3.93 (s, 3H), 1.99 (s, 3H), 1.92 (s, 3H).

Example 12: Preparation of 5-amino-3-chloro-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (11)

Referring to the preparation method of Example 2, 1-methyl-5-(trifluoromethyl)-1H-pyrazole-3-amine in step 1 was replaced with 5-chloro-1-methyl-1H-pyrazole-3-amine to obtain the title compound 11.

LC-MS: m/z=335 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ7.48 (d, J = 21.4 Hz, 2H), 7.34 (s, 1H), 7.27–6.87 (m, 1H), 6.72 (s, 2H), 5.35 (s, 1H), 3.53 (s, 3H), 2.18 (d, J = 3.6 Hz, 6H).

Example 13: Preparation of 5-amino-3-cyano-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (12)

Step 1: Preparation of 5-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-1H-pyrazol-3-amine (12a)

To a solution of 5-bromo-1-methyl-1H-pyrazol-3-amine (1.39 g, 5.63 mmol) and 2-iodo-4-methoxy-1,3-dimethylbenzene (1.47 g, 5.63 mmol) in 1,4-dioxane (28 mL) was added cesium carbonate (3.67 g, 11.3 mmol), 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (651 mg, 1.13 mmol) and tris(dibenzylideneacetone) at room temperature. Dipalladium (640 mg, 563 μmol) was replaced with nitrogen three times, stirred at 100°C under nitrogen atmosphere for 16 hours, 100 mL of water was added, extracted with ethyl acetate (50 mL x 2), the organic phase was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: ethyl acetate/petroleum ether = 15%-25%) to give 1.70 g of the title compound as a yellow solid. Yield: 79.3%.

LC-MS: m/z=310 [M+H] ⁺ .

Step 2: Preparation of 3-((3-methoxy-2,6-dimethylphenyl)amino)-1-methyl-1H-pyrazole-5-carbonitrile (12b)

5-Bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-1H-pyrazole-3-amine (1.00 g, 3.2 mmol) and cuprous cyanide (3.05 g, 32.3 mmol) were added to N,N-dimethylformamide (10 mL) at room temperature and stirred at 150°C for 8 hours. 100 mL of water was added and the mixture was extracted with ethyl acetate (50 mL x 3). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 100:0-20:1) to give 1.20 g of the title compound as a yellow solid.

LC-MS: m/z 257 [M+H] ⁺ .

Step 3: Preparation of 4-bromo-3-((3-methoxy-2,6-dimethylphenyl)amino)-1-methyl-1H-pyrazole-5-carbonitrile (12c)

3-((3-Methoxy-2,6-dimethylphenyl)amino)-1-methyl-1H-pyrazole-5-carbonitrile (1.15 g, 4.5 mmol) was dissolved in acetonitrile (20 ml) at room temperature, and then N-bromosuccinimide (0.63 g, 4.1 mmol) was slowly added. After reacting at room temperature for 2 hours, the mixture was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 100:0-5:1) to obtain 0.46 g of the title compound as a yellow solid. The yield was 30.7%.

LC-MS: m/z=336 [M+H] ⁺ .

Step 4: Preparation of 5-amino-3-cyano-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxylic acid methyl ester (12d)

At room temperature, 4-bromo-3-((3-methoxy-2,6-dimethylphenyl)amino)-1-methyl-1H-pyrazole-5-carbonitrile (1.06 g, 3.2 mmol), methyl cyanoacetate (0.630 g, 6.30 mmol), L-proline (73.0 mg, 0.63 mmol), cuprous iodide (60.0 mg, 0.32 mmol) and potassium carbonate (1.32 g, 9.5 mmol) were added to dimethyl sulfoxide (20 mL). The mixture was reacted at 90 °C under nitrogen atmosphere for 16 hours. 100 mL of water was added and the mixture was extracted with dichloromethane (50 mL x 3). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 100:1-5:2) to give 140 mg of the title compound as a yellow solid in a yield of 12.6%.

LC-MS: m/z=354 [M+H] ⁺ .

Step 5: Preparation of 5-amino-3-cyano-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (12e)

At room temperature, 5-amino-3-cyano-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxylic acid methyl ester (140 mg, 0.400 mmol) and 7 M ammonia methanol solution (5 mL) were added to a sealed tube, stirred at 50° C. for 48 hours, concentrated under reduced pressure, and the residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate=100:1-5:3) to give the title compound as a yellow solid (90.0 mg, yield 67.2%).

LC-MS: m/z=339 [M+H] ⁺ .

Step 6: Preparation of 5-amino-3-cyano-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (12)

5-Amino-3-cyano-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (90.0 mg, 0.025 mmol) was dissolved in dichloromethane (5 ml) at 0°C, and 1 M boron tribromide solution in dichloromethane (1 ml, 0.1 mmol) was added dropwise. The mixture was stirred at room temperature for 2 hours, and methanol was added to quench the reaction. The mixture was concentrated under reduced pressure and the residue was separated by high pressure preparative liquid separation (chromatographic column model: Daisogei 30 mm × 250 mm, C18, 10 um 100A, mobile phase: acetonitrile/water, gradient: 10%-50%, 0.05% formic acid) to give 7 mg of the title compound as a yellow solid.

LC-MS: m/z=325 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ11.07(s,1H),9.01(s,1H),7.25(s,2H),6.79(d,J＝8.1Hz,1H),6.53(d,J＝8.4Hz,1H),6.02(s,1H),3.86(s,3H),1.98(d,J＝31.6Hz,6H).

Example 14: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (13)

Step 1: Preparation of N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(thiazol-2-yl)-1H-pyrazol-3-amine (13a)

5-Bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-1H-pyrazol-3-amine (2.00 g, 6.50 mmol) was dissolved in 1,4-dioxane (100 mL) at room temperature, and tributylthiazole-5-tin (3.54 g, 13.0 mmol) and tetrakistriphenylphosphine palladium (0.746 g, 0.650 mmol) were added. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 100° C. under a nitrogen atmosphere for 16 hours. 5 g of potassium carbonate was added, and the mixture was concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate=100:0-5:1) to obtain 1.63 g of the title compound as a yellow solid. The yield was 80.7%.

LC-MS: m/z=315 [M+H] ⁺ .

Step 2: Preparation of 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(thiazol-2-yl)-1H-pyrazol-3-amine (13b)

N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(thiazol-2-yl)-1H-pyrazol-3-amine (2.00 g, 6.40 mmol) and N-bromosuccinimide (1.13 g, 6.40 mmol) were added to dichloromethane (30 mL) at room temperature and stirred for 16 hours. The mixture was concentrated under reduced pressure and the residue was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate=100:0-5:1) to give the title compound as a yellow solid (1.50 g, yield 60.0%).

LC-MS: m/z=394 [M+H] ⁺ .

Step 3: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (13c)

At room temperature, 4-bromo-N-(3-methoxy-2,6-dimethylphenyl)-1-methyl-5-(thiazol-2-yl)-1H-pyrazol-3-amine (1.40 g, 3.60 mmol), malononitrile (0.472 g, 7.20 mmol), L-proline (0.082 g, 0.720 mmol), cuprous iodide (0.068 g, 0.360 mmol) and potassium carbonate (1.48 g, 10.8 mmol) were added. l) was dissolved in dimethyl sulfoxide (20 mL), replaced with nitrogen three times, stirred at 90°C under nitrogen atmosphere for 16 hours, added with 100 mL of water, extracted with dichloromethane (50 mL x 3), the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (mobile phase: petroleum ether/ethyl acetate = 100:1-5:2) to obtain 900 mg of the title compound as a yellow solid. The yield was 67.2%.

LC-MS: m/z=379 [M+H] ⁺ .

Step 4: Preparation of 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (13d)

At room temperature, 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carbonitrile (0.9 g, 2.4 mmol) was dissolved in 3 ml of concentrated sulfuric acid, stirred at room temperature for 4 hours, poured into 100 mL of ice water, adjusted to pH = 7 with saturated sodium bicarbonate solution, extracted with dichloromethane (30 mL x 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 600 mg of the title compound as a yellow solid, with a yield of 63.8%.

LC-MS: m/z=396 [M+H] ⁺ .

Step 5: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (13)

At 0°C, 5-amino-6-(3-methoxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (400 mg, 1.04 mmol) was dissolved in dichloromethane (20 ml), and a 1M dichloromethane solution of boron tribromide (6 ml, 6.00 mmol) was added dropwise. The mixture was stirred at room temperature for 2 hours, and methanol was added to quench the reaction. The mixture was concentrated under reduced pressure, and the residue was separated by high pressure preparative liquid chromatography (chromatographic column model: Daisogei 30 mm × 250 mm, C18, 10 um 100A, mobile phase: acetonitrile/water, gradient: 10%-50%, 0.05% formic acid) to obtain 101 mg of the title compound as a yellow solid.

LC-MS: m/z=383 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.53(s,1H),8.07(d,J＝3.6Hz,1H),7.95(s,2H),7.54(d,J＝3.6Hz,1H),7.05(d,J＝8 .3Hz,1H),6.90(d,J＝8.3Hz,1H),5.05(s,2H),4.04(s,3H),1.89(s,3H),1.80(s,3H).

Example 15: Preparation of S-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide and R-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (13-1 and 13-2)

Compound 13 was separated by SFC chiral preparative separation under the following conditions: equipment: SFC-150mgm (waters), chiral column: CHIRALPAK IC (20×250mm, 5um), temperature: 25°C, mobile phase: MTBE (0.5% 2mM NH3-MeOH)/MeOH=60/40, flow rate: 20mL/min, back pressure: 100bar, detection wavelength: 220nm, cycle time: 5min, and two isomers were obtained: compound 13-1 (RT=2.63min) and compound 13-2 (RT=3.13min).

Compound 13-1: LC-MS: m/z=383 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.53(s,1H),8.07(d,J＝3.6Hz,1H),7.95(s,2H),7.54(d,J＝3.6Hz,1H),7.05(d,J＝8 .3Hz,1H),6.90(d,J＝8.3Hz,1H),5.05(s,2H),4.04(s,3H),1.89(s,3H),1.80(s,3H).

Compound 13-2: LC-MS: m/z=383 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.53(s,1H),8.07(d,J＝3.6Hz,1H),7.95(s,2H),7.54(d,J＝3.6Hz,1H),7.05(d,J＝8 .3Hz,1H),6.90(d,J＝8.3Hz,1H),5.05(s,2H),4.04(s,3H),1.89(s,3H),1.80(s,3H).

Example 16: Preparation of 5-amino-3-(5-fluoropyridin-3-yl)-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (14)

Referring to the preparation method of Example 14, the title compound 14 was prepared by replacing tributylthiazole-5-tin in step 2 with (5-fluoropyridin-3-yl)boric acid.

LC-MS: m/z=394 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.56(s,1H),8.77–8.58(m,2H),8.03(dt,J＝9.6,2.3Hz,1H),7.05(d,J＝8.3Hz,1H),6 .90(d,J=8.3Hz,1H),6.64(s,2H),5.48(s,2H),3.66(s,3H),1.91(s,3H),1.81(s,3H).

Example 17: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(5-methylthiazol-2-yl)-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (15)

Referring to the preparation method of Example 14, the title compound 15 was prepared by replacing tributylthiazole-5-tin in step 2 with 5-methyl-2-(tri-n-butyltin)thiazole.

LC-MS: m/z=397 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.53(s,1H),7.55(d,J＝8.3Hz,1H),7.05(s,1H),6.90(d,J＝8.3Hz,1H),5.05(s,2H),4.04(s,3H),2.31(s,3H),1.92(s,3H),1.80(s,3H).

Example 18: Preparation of 5-amino-6-(2-chloro-3-hydroxy-6-methylphenyl)-2-methyl-3-(thiazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (16)

Referring to the preparation method of Example 14, the title compound 16 was prepared by replacing 2-iodo-4-methoxy-1,3-dimethylbenzene in step 1 with 2-chloro-3-iodo-1-methoxy-4-methylbenzene.

LC-MS: m/z=404 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ7.98–7.94 (m, 1H), 7.21 (d, J = 8.4 Hz, 1H), 7.15 (s, 2H), 7.08 (d, J = 8.4 Hz, 1H), 4.04 (s, 3H), 1.96 (s, 3H).

Example 19: Preparation of 5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(oxazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (17)

Referring to the preparation method of Example 14, the title compound 17 was prepared by replacing tributylthiazole-5-tin in step 2 with 2-(tri-n-butylstannyl)oxazole.

LC-MS: m/z=367 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.40(d,J＝104.6Hz,2H),8.31(d,J＝0.9Hz,1H),7.46(d,J＝0.8Hz,1H),7.05(d,J＝7 .2Hz,3H),6.90(d,J=8.3Hz,1H),6.61(s,1H),4.04(s,3H),1.87(s,3H),1.78(s,3H).

Example 20: Preparation of S-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(oxazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide and R-5-amino-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(oxazol-2-yl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (17-1 and 17-2)

Compound 17 was subjected to SFC chiral preparative separation under the following conditions: equipment: Waters 150 preparative SFC (SFC-26), chiral column: CHIRALPAK IM (30×250 mm, 10 um), temperature: 38° C., mobile phase A: Supercritical CO ₂ , mobile phase B: methanol, A/B=60/40, flow rate: 120 mL/min, back pressure: 100 bar, detection wavelength: 220 nm, cycle time: 7.50 min, to obtain 2 isomers: compound 17-1 (RT=2.08 min) and compound 17-2 (RT=2.77 min).

Compound 17-1: LC-MS: m/z=367 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.40(d,J＝104.6Hz,2H),8.31(d,J＝0.9Hz,1H),7.46(d,J＝0.8Hz,1H),7.05(d,J＝7 .2Hz,3H),6.90(d,J=8.3Hz,1H),6.61(s,1H),4.04(s,3H),1.87(s,3H),1.78(s,3H).

Compound 17-2: LC-MS: m/z=367 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.40(d,J＝104.6Hz,2H),8.31(d,J＝0.9Hz,1H),7.46(d,J＝0.8Hz,1H),7.05(d,J＝7 .2Hz,3H),6.90(d,J=8.3Hz,1H),6.61(s,1H),4.04(s,3H),1.87(s,3H),1.78(s,3H).

Example 21: Preparation of 5-amino-6-(3-fluoro-5-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (18)

Referring to the preparation method of Example 2, the title compound 18 was prepared by replacing 2-iodo-4-methoxy-1,3-dimethylbenzene in step 2 with 1-fluoro-3-iodo-5-methoxy-2,4-dimethylbenzene.

LC-MS: m/z=386 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ 10.08 (s, 1H), 7.21–7.03 (m, 2H), 6.79 (d, J = 11.1Hz, 1H), 6.30 (s, 2H), 3.88 (d, J = 1.4Hz, 3H), 1.83–1.54 (m, 6H).

Example 22: Preparation of 5-amino-6-(4-fluoro-3-hydroxy-2,6-dimethylphenyl)-2-methyl-3-(trifluoromethyl)-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (19)

Referring to the preparation method of Example 2, the title compound 19 was prepared by replacing 2-iodo-4-methoxy-1,3-dimethylbenzene in step 2 with 1-fluoro-4-iodo-2-methoxy-3,5-dimethylbenzene.

LC-MS: m/z=385.95[M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ ) δ 9.64 (s, 1H), 7.09 (t, J = 12.4 Hz, 3H), 6.27 (s, 2H), 3.87 (d, J = 1.6 Hz, 3H), 1.86 (s, 3H), 1.81 (s, 3H).

Example 23: Preparation of 5-amino-3-(benzofuran-2-yl)-6-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-2,6-dihydropyrrolo[2,3-c]pyrazole-4-carboxamide (20)

Referring to the preparation method of Example 14, the title compound 20 was prepared by replacing tributylthiazole-5-tin in step 2 with benzofuran-2-boronic acid.

LC-MS: m/z=416 [M+H] ⁺ .

¹ H NMR (400MHz, DMSO-d ₆ )δ9.54(s,1H),7.75(dd,J＝7.7,1.3Hz,1H),7.67–7.60(m,1H),7.45–7.28(m,3H),7.06( d,J＝8.3Hz,1H),6.94–6.78(m,3H),6.35(s,2H),3.86(s,3H),1.90(s,3H),1.81(s,3H).

Biological tests

Experimental Example 1: PKMYT1 Enzyme Experiment

Experimental materials: compounds of the present invention, PKMYT1 (Carna, 05-176), ATP (Promega, V9102), DTT (Sigma, 646563), ADP-GloTM (Promega, V9102, containing DTT, HEPES, MgCl ₂ , BRIJ-35, EGTA), Inactive CDK1 (Signalchem, C22-14G).

1.1. Reagent preparation

1× Kinase Reaction Buffer:

To prepare 1 mL of kinase reaction buffer, use 1M DTT, 1M HEPES, 1M MgCl ₂ , 10% BRIJ-35(%) and 1M EGTA stock solutions, dilute 500X, 20X, 100X, 1000X and 1000X to a final concentration of 2mM DTT, 50mM HEPES, 10mM MgCl ₂ , 0.01% BRIJ-35(%) and 1mM EGTA. Then add 2uL DTT, 50uL HEPES, 10uL MgCl ₂ , 1uL BRIJ-35(%) and 1uL EGTA buffer, and finally add 936uL water and mix well.

Preparation of working solution of test compound: Add test compound to DMSO and shake to dissolve, and prepare a stock solution with a concentration of 10 mM for later use.

1.2 Experimental Methods

Prepare the buffer solution by the above method, and transfer 50nL of the prepared test compound working solution to each well of the reaction plate using Echo 655. Seal the reaction plate with a sealing film and centrifuge at 1000g for 1 minute. Prepare 2× kinase reaction buffer (5ng/ul) with 1× kinase reaction buffer. Add 2.5μl 2× kinase reaction buffer to each well of the reaction plate. Seal the plate with a sealing film, centrifuge at 1000g for 30 seconds, and let it stand at room temperature for 10 minutes. Prepare 2× kinase substrate Unactive CDK1 (0.02ng/ul) and ATP (400uM) mixed solution with 1× kinase reaction buffer. Add 2.5μL 2× kinase substrate and ATP mixed solution to the reaction plate, centrifuge at 1000g for 30 seconds, and start the reaction. Seal the plate with a sealing film and react the kinase at room temperature for 120 minutes. Add 4μL ADP-Glo reagent. React at room temperature for 60 minutes. Add 8μL kinase detection reagent and react at room temperature for 60 minutes. Luminescence signal values were read using Envision (2104) multi-function plate reader.

1.3 Data Analysis

The % inhibition rate was calculated as follows:
Suppression%=100-(Signal cmpd-Signal Ave_PC)/(Signal Ave_VC-Signal
Ave_PC)×100

Signal cmpd: chemiluminescence value of each concentration of compound

SignalAve_PC: Average value of system chemiluminescence value

SignalAve_VC: Average value of negative control chemiluminescence value

Calculate IC ₅₀ and draw compound effect dose curve:

_IC50 was calculated by fitting the percent inhibition values and logarithms of compound concentration to non-linear regression (dose response - variable slope) using Graphpad 6.0.

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)×HillSlope))

X: Logarithmic value of inhibitor concentration; Y: Percentage of inhibition rate

Table 1 provides the in vitro enzymatic activities (IC ₅₀ ) of the compounds of the present invention against PKMYT1.

In Table 1, the PKMYT1 in vitro enzymatic activity values of the compounds are as follows: A refers to IC ₅₀ <10 nM; B refers to 10 nM<IC ₅₀ <100 nM; C refers to 100 nM<IC ₅₀ <1000 nM; and D refers to IC ₅₀ >1000 nM.

Table 1. Enzymatic activity of the compounds of the present invention against PKMYT1

Conclusion: The compounds of the present invention have good inhibitory effects on the enzymatic activity of PKMYT1.

Experimental Example 2: OVCAR3 cell anti-proliferation experiment

Experimental materials: compounds of the present invention, RPMI1640 culture medium (Invitrogen, A10491-01), fetal bovine serum (Gibco, 10099141), human insulin (aladdin, I302196), penicillin/streptomycin antibiotics (Invitrogen, 15140122), CelltiterGlo assay kit (CTG) (Promega, G7573), OVCAR3 cell line (ATCC, HTB-161TM).

2.1 Experimental methods:

On day 1, add the cell suspension to a 384-well plate, 45 μL per well, 250 live cells/well. Add 2uL of the test compound working solution to 198uL of culture medium for a 100-fold dilution, then use a gun to dilute 3 times in the culture medium, 9+0 concentrations, and start the dilution from 100μM. Take 5uL of the compound diluted in the middle of the culture medium and add it to 45uL of cells in a 384-well plate according to the corresponding position, and set up a double-well experiment. The cell plate was placed in a 37, 5% carbon dioxide incubator for 7 days.

Then, Promega CellTiter-Glo assay was performed. The cell plate was removed and equilibrated at room temperature for 30 minutes. 20 μL CTG was added to each well and mixed by vortexing. The cells were incubated at room temperature for 10 minutes and luminescence was read using Biotek (Cytation 3) multi-label analyzer.

2.2 Data Analysis

The inhibition rate was calculated as follows:
Suppression%=100-(Signal cmpd-Signal Ave_PC)/(Signal Ave_VC-Signal
Ave_PC)×100

Signal cmpd: chemiluminescence value of each concentration of compound

SignalAve_PC: average value of system chemiluminescence value

SignalAve_VC: Average value of negative control chemiluminescence value

Calculate IC ₅₀ and draw compound effect dose curve:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*HillSlope))

X: logarithmic value of compound concentration; Y: percentage of inhibition rate.

Table 2 provides the inhibitory activity of the compounds of the present invention on the proliferation of OVCAR3 cells.

In Table 2, the inhibitory activity values of the compounds on OVCAR3 cell proliferation are as follows: A refers to IC ₅₀ <100 nM; B refers to 100 nM<IC ₅₀ <1000 nM; C refers to 1000 nM<IC ₅₀ <10000 nM; and D refers to IC ₅₀ >10000 nM.

Table 2. Inhibitory activity of the compounds of the present invention on OVCAR3 cell proliferation

Conclusion: The compounds of the present invention have good antiproliferative activity against OVCAR3 cells.

Test Example 3: WEE1 Enzyme Experiment

Experimental materials: compounds of the present invention, WEE1 (BPS, 40412), ATP (Promega, V9103), HEPES (Beyotime, C0217), MgCl ₂ (Sigma, M1028), MnCl ₂ (Sigma, M1787), PEG20000 (Sigma, 95172-250G-F), Na ₃ VO ₄ (NEB, P0758S), DTT (Sigma, 646563), ADP-Glo™ (Promega, V9103).

3.1 Reagent preparation:

1× Kinase Reaction Buffer:

Prepare 1 mL of kinase reaction buffer using 1M DTT, 1M HEPES, 1M MgCl ₂ , 1MMnCl ₂ , 1mg/mL PEG20000 and 100mM Na ₃ VO ₄ stock solutions, dilute to 833.33X, 14.29X, 333.33X, 333.33X, 20X and 33333.33X to a final concentration of 1.2mM DTT, 70mM HEPES, 3mM MgCl ₂ , 3mM MnCl 2, 0.05mg/mL PEG20000 and 0.003mM Na ₃ VO ₄ , then add 1.2ul DTT, 70ul HEPES, 3ul MgCl ₂ , 3ul MnCl ₂ , 50ul PEG20000 and 0.03ul Na ₃ VO 4 respectively. ₄ buffer, and finally add 869.8ul of water and mix well.

3.2 Experimental methods:

Prepare the buffer by the above method, and transfer 150nL of DMSO-diluted compound solution to each well of the reaction plate using Echo 655. Seal the reaction plate with a sealing film and centrifuge at 1000g for 1 minute. Prepare 2× kinase reaction buffer (12nM) with 1× kinase reaction buffer. Add 7.5μl 2× kinase reaction buffer to each well of the reaction plate. Seal the plate with a sealing film, centrifuge at 1000g for 1 minute, and let it stand at room temperature for 15 minutes. Prepare 2× kinase substrate ATP (20uM) with 1× kinase reaction buffer. Add 7.5μL 2× kinase substrate reaction buffer to the reaction plate, centrifuge at 1000g for 1 minute, and start the reaction. Seal the plate with a sealing film and react the kinase at 30℃ for 60 minutes. Add 15μL ADP-Glo reagent. React at room temperature for 60 minutes. Add 30μL kinase detection reagent and react at room temperature for 60 minutes. Read the luminescence signal value using Envision (2104) multi-function plate reader.

3.3 Data Analysis:

The % inhibition rate was calculated as follows:
Suppression%=100-(Signal cmpd-Signal Ave_PC)/(Signal Ave_VC-Signal
Ave_PC)×100

Signal cmpd: chemiluminescence value of each concentration of compound

SignalAve_PC: Average value of system chemiluminescence value

SignalAve_VC: Average value of negative control chemiluminescence value

Calculate IC ₅₀ and draw compound effect dose curve:

_IC50 was calculated by fitting the percent inhibition values and logarithms of compound concentration to non-linear regression (dose response - variable slope) using Graphpad 6.0.

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*HillSlope))

X: Logarithmic value of inhibitor concentration; Y: Percentage of inhibition rate

Table 3 provides the in vitro enzymatic activities (IC ₅₀ ) of the compounds of the present invention against WEE1.

In Table 3, the WEE1 in vitro enzymatic activity values of the compounds are as follows: A refers to IC ₅₀ <10 nM; B refers to 10 nM<IC ₅₀ <100 nM; C refers to 100 nM<IC ₅₀ <1000 nM; and D refers to IC ₅₀ >1000 nM.

Table 3 Enzymatic activity of the compounds of the present invention against WEE1

Conclusion: The compounds of the present invention have weaker inhibitory effects on WEE1 kinases of the same family and have good enzyme selectivity.

Experimental Example 4: HCC1569 cell proliferation inhibition experiment

Experimental materials: compounds of the present invention, RPMI1640 culture medium (ATCC, 30-2001), fetal bovine serum (Gibco, 10099141), penicillin/streptomycin antibiotics (Invitrogen, 15140122), Celltiter Glo assay kit (CTG) (Promega, G7572), HCC1569 cell line (Kebai, CBP60372).

4.1 Experimental methods:

On the first day, the cell suspension was added to a 384-well plate using a Multidrop dispenser, 50 μL per well, 500 live cells/well. The test compound dissolved in DMSO was added to 50 μL of cells in a 384-well plate using an ultra-micropipette, starting from 10000 nM, 3-fold serial dilution, 9+0 concentrations, and a double-well experiment was set up. The cell plate was placed in a 37°C, 5% carbon dioxide incubator for 7 days.

Then, Promega CellTiter-Glo assay was performed. The cell plate was removed and equilibrated at room temperature for 30 minutes. 20 μL CTG was added to each well and mixed by vortexing. The cells were incubated at room temperature for 10 minutes and luminescence was read using Biotek (Cytation 3) multi-label analyzer.

4.2 Data Analysis:

The inhibition rate was calculated as follows:
Suppression%=100-(Signal cmpd-Signal Ave_PC)/(Signal Ave_VC-Signal
Ave_PC)×100

Signal cmpd: chemiluminescence value of each concentration of compound

SignalAve_PC: average value of system chemiluminescence value

SignalAve_VC: Average value of negative control chemiluminescence value

Calculate IC ₅₀ and draw compound effect dose curve:

Y＝Bottom+(Top-Bottom)/(1+10^((LogIC ₅₀ -X)*HillSlope))

X: logarithmic value of compound concentration; Y: percentage of inhibition rate.

Table 4 provides the inhibitory activity of the compounds of the present invention on HCC1569 cell proliferation. In Table 5, the inhibitory activity values of the compounds on HCC1569 cell proliferation: A means IC ₅₀ <100nM; B means 100nM<IC ₅₀ <1000nM; C means 1000nM<IC ₅₀ <10000nM; D means IC ₅₀ >10000nM.

Table 4. Inhibitory activity of the compounds of the present invention on HCC1569 cell proliferation

Conclusion: The compounds of the present invention have good antiproliferative activity against HCC1569 cells.

Claims (20)

A compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof,
in: X is selected from N or CR ⁷ ; Y is selected from O, S, NR ⁷ or CR ^7a R ^7b ; R ¹ is selected from hydrogen, halogen or -NR ^8a R ^8b ; R ² , R ³ , R ⁴ and R ⁵ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ^7a , R ^7b and R ⁷ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ R 7a and R 7b are substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR 9 , -C(O)R 10 , -S(O) p R ¹⁰ , -C(O)NR ^9a R 9b , -S(O) p NR 9a R 9b , -NR 9a R 9b , -SR 9 , -OR 9a , -C(O)R ¹⁰ , -S(O) p R ¹⁰ , -C(O)NR 9a _R ^9b , -S(O) _p NR ^9a R ^{9b ,} -NR ^9a R ^9b , -SR ^{9 ,} -OR ^9a , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ^8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R 10 , -S(O) p R 10 , -C(O)NR 9a R ^9b , -S(O) _p NR ^{9a R 9b} , oxo, alkyl, alkoxy, ^haloalkyl , hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, ^heterocyclyl , ^aryl , heteroaryl; or, _R 8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, ^heteroaryl . ^8b together with the nitrogen atom to which it is attached forms a heterocyclic group or a heteroaryl group, wherein the heterocyclic group or the heteroaryl group is optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclic group, aryl, heteroaryl; R ^9a , R ^9b and R ⁹ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a heterocyclyl or heteroaryl, and the heterocyclyl or heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ¹⁰ is selected from hydrogen, halogen, amino, nitro, hydroxyl, mercapto, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, mercapto, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; p is 1 or 2. The compound represented by the general formula (I) according to claim 1, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, in, X is selected from N; Y is selected from O or NR ⁷ ; R ¹ is selected from hydrogen, halogen or -NR ^8a R ^8b ; R ² , R ³ , R ⁴ and R ⁵ are each independently selected from hydrogen, halogen, amino, nitro, hydroxyl, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ⁷ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ^8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R 10 , -S(O) p R 10 , -C(O)NR 9a R ^9b , -S(O) _p NR ^{9a R 9b} , oxo, alkyl, alkoxy, ^haloalkyl , hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, ^heterocyclyl , ^aryl , heteroaryl; or, _R 8a and R ^8b are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, ^heteroaryl . ^8b together with the nitrogen atom to which it is attached forms a heterocyclic group or a heteroaryl group, wherein the heterocyclic group or the heteroaryl group is optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclic group, aryl, heteroaryl; R ^9a , R ^9b and R ⁹ are each independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a heterocyclyl or heteroaryl, and the heterocyclyl or heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, sulfhydryl, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ¹⁰ is selected from hydrogen, halogen, amino, nitro, hydroxyl, mercapto, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl are optionally substituted with one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, mercapto, carboxyl, ester, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; p is 1 or 2. The compound represented by the general formula (I) according to claim 1 or 2, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, which is a compound represented by the general formula (IA) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:
wherein X, Y, R ¹ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in claim 1 or 2. The compound represented by the general formula (I) according to any one of claims 1 to 3, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, which is a compound represented by the general formula (II), or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:
in, Y ₁ is selected from O or NR ^7c ; R ^7c is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ R 7d and R 7e together with the carbon atom to which they are attached form an oxo group, a cycloalkyl group, a heterocyclyl group, an aryl group or a heteroaryl group, wherein the cycloalkyl group, the heterocyclyl group, the aryl group or the heteroaryl group is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) p R ¹⁰ , -C(O)NR ^{9a R 9b ,} -S(O) _{p NR} ^9a R ^9b , -NR ^9a R ^9b , -SR ^{9 , -OR 9} , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁹ , R ^9a , R ^9b , R ¹⁰ , and p are as defined in claim 1. The compound represented by the general formula (I) according to claim 4, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt, which is a compound represented by the general formula (IIA) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated product, or its pharmaceutically acceptable salt:
wherein Y ₁ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in claim 3. The compound represented by the general formula (I) according to claim 4 or 5, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein:
Selected from in: R ⁶ is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ^7c is selected from hydrogen, halogen, amino, nitro, hydroxyl, thiol, cyano, alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , wherein the alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl is optionally selected from deuterated, halogen, amino, nitro, cyano, hydroxyl, thiol, -COOR ⁹ , -C(O)R ¹⁰ , -S(O) _p R ¹⁰ , -C(O)NR ^9a R ^9b , -S(O) _p NR ^9a R ^9b , -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ , oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, aminoalkyl, haloalkoxy, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl; R ⁹ , R ^9a , R ^9b , R ¹⁰ , and p are as defined in claim 1. The compound represented by the general formula (I) according to any one of claims 4 to 6, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: R ⁶ is selected from hydrogen, halogen, amino, hydroxyl, mercapto, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally selected from halogen, amino, cyano, hydroxyl, mercapto, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C substituted by one or more of _3-6 membered cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, or 5-10 membered heteroaryl; R ^7c is selected from hydrogen, halogen, amino, hydroxyl, mercapto, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -CH ₂ R ⁹ , -NR ^9a R ^9b , wherein the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl is optionally selected from deuterated, halogen, amino, cyano, hydroxyl, mercapto, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _{2-6 alkenyl, C 2-6 alkynyl} , C substituted by one or more of _3-6 membered cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, or 5-10 membered heteroaryl; R is selected from ^C3-6 cycloalkyl, _4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl, wherein the _C3-6 cycloalkyl, 4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl is optionally substituted by one or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, oxo, _C1-6 alkyl, _C1-6 alkoxy, _C1-6 haloalkyl, _C1-6 hydroxyalkyl, _C1-6 aminoalkyl, _C1-6 haloalkoxy, _C2-6 alkenyl, _C2-6 alkynyl, C3-6 cycloalkyl, 4-6 membered heterocyclyl, _C6-10 aryl, 5-10 membered heteroaryl _; R ^9a and R ^9b are each independently selected from hydrogen, C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group or a 5-10 membered heteroaryl group, and the _4-6 membered heterocyclic group or the 5-10 membered heteroaryl group is optionally substituted by one _{or more groups selected from halogen, amino, cyano, hydroxyl, thiol, carboxyl, C 1-6 alkyl} , C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C 2-6 alkynyl, C _{3-6 cycloalkyl, 4-6 membered heterocyclic group, C 6-10} aryl, or 5-10 membered heteroaryl. The compound represented by the general formula (I) according to any one of claims 1 to 7, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the C _6-10 aryl and 5-10 membered heteroaryl are optionally substituted with halogen or C _1-6 alkyl; R ^9a and R ^9b are each independently selected from hydrogen and C _1-6 alkyl; or, R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group; Preferably, R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the 5-10 membered heteroaryl is optionally substituted with halogen or C _1-6 alkyl; R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group; More preferably, R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, 5-10 membered heteroaryl, wherein the 5-10 membered heteroaryl is optionally substituted by halogen or C _1-6 alkyl. The compound represented by the general formula (I) according to any one of claims 1 to 8, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ⁷ or R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, -CH ₂ R ⁹ , and the C _1-6 alkyl is optionally substituted with deuterium; R ⁹ is selected from phenyl, which is optionally substituted by one or more groups selected from halogen, C _1-6 alkyl; Preferably, R ⁷ or R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, and the C _1-6 alkyl is optionally substituted by deuterium. The compound represented by the general formula (I) according to any one of claims 1 to 9, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ² and R ³ are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 membered aryl, 5-10 membered heteroaryl are optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl; R ⁹ , R ^9a , R ^9b , R ¹⁰ , and p are as defined in claim 1; Preferably, R ² and R ³ are each independently selected from C _1-6 alkyl. The compound represented by the general formula (I) according to any one of claims 1 to 10, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein: R ⁴ and R ⁵ are each independently selected from hydrogen, halogen, amino, hydroxyl, cyano, C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl, -NR ^9a R ^9b , -SR ⁹ , -OR ⁹ ; the C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 membered aryl, 5-10 membered heteroaryl are optionally substituted by one or more groups selected from halogen, amino, nitro, cyano, hydroxyl, thiol, carboxyl, ester, oxo, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 haloalkyl, C _1-6 hydroxyalkyl, C _1-6 aminoalkyl, C _1-6 haloalkoxy, C _2-6 alkenyl, C _2-6 alkynyl, C _3-7 cycloalkyl, 4-7 membered heterocyclyl, C _6-10 aryl, 5-10 membered heteroaryl; R ⁹ , R ^9a , R ^9b , R ¹⁰ , and p are as defined in claim 1; Preferably, ^R4 and ^R5 are each independently selected from hydrogen. The compound represented by the general formula (I) according to any one of claims 4 to 11, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: Selected from R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl; R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl, phenyl, -CH ₂ R ⁹ , wherein the C _1-6 alkyl is optionally substituted with deuterium; R ⁹ is selected from phenyl, which is optionally substituted by one or more groups selected from halogen, C _1-6 alkyl; Preferably, R ⁶ is selected from C _1-6 alkyl or C _1-6 haloalkyl; R ^7c is selected from C _1-6 alkyl, C _3-6 cycloalkyl and phenyl, and the C _1-6 alkyl is optionally substituted by deuterium. The compound represented by the general formula (I) according to any one of claims 4 to 11, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: Selected from R ⁶ is selected from hydrogen, halogen, cyano, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 4-6 membered heterocyclyl, phenyl, 5-10 membered heteroaryl, -NR ^9a R ^9b , wherein the 5-10 membered heteroaryl is optionally substituted with halogen or C _1-6 alkyl; R ^7c is selected from C _1-6 alkyl; R ^9a and R ^9b together with the nitrogen atom to which they are attached form a 4-6 membered heterocyclic group; Preferably, ^R6 is selected from hydrogen, halogen, cyano, _C1-6 alkyl, _C1-6 haloalkyl, 5-10 membered heteroaryl, wherein the 5-10 membered heteroaryl is optionally substituted by halogen or _C1-6 alkyl; R ^7c is selected from C _1-6 alkyl. The compound represented by the general formula (I) according to any one of claims 4 to 11, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: Selected from R ⁶ is selected from C _1-6 alkyl, C _1-6 haloalkyl, 5-6 membered heteroaryl, preferably C _1-6 alkyl, C _1-6 haloalkyl. The compound represented by the general formula (I) according to any one of claims 1 to 14, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or its deuterated substance, or its pharmaceutically acceptable salt, wherein: ^R2 is selected from halogen and _C1-6 alkyl; R ³ is selected from halogen, C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, preferably C _1-6 alkyl; R ⁴ is selected from hydrogen, halogen, cyano, preferably hydrogen and halogen; ^R5 is selected from hydrogen and halogen. The compound represented by the general formula (I) according to any one of claims 1 to 15, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, wherein the compound is selected from:

A method for preparing a compound represented by general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, comprising the following steps:
The compound represented by the general formula D-1 or a salt thereof is reacted in a solvent, optionally in the presence of a catalyst, to obtain a compound represented by the general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or a mixture thereof, or a pharmaceutically acceptable salt thereof, wherein Y ₁ , R ² , R ³ , R ⁴ , R ⁵ and R ⁶ are as defined in claim 4. A pharmaceutical composition comprising a compound represented by the general formula (I) according to any one of claims 1 to 16, or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or a deuterated substance thereof, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient. Use of a compound represented by general formula (I) according to any one of claims 1 to 16 or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 18 in the preparation of a membrane-associated tyrosine/threonine protein kinase 1 (PKMYT1) inhibitor. Use of a compound represented by general formula (I) according to any one of claims 1 to 16 or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or deuterated substance thereof, or pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 18 in the preparation of a medicament for preventing and/or treating a disease associated with the activity of membrane-associated tyrosine/threonine protein kinase 1 (PKMYT1), wherein the disease is preferably a tumor disease, such as ovarian cancer, breast cancer, cervical cancer, endometrial cancer, prostate cancer, colorectal cancer, esophageal cancer, liver cancer, lung cancer or thyroid cancer.