WRN helicase-inhibiting bis-heterocyclic derivative, pharmaceutical composition containing same, and use thereof

Abstract

A WRN helicase-inhibiting bis-heterocyclic derivative, a pharmaceutical composition containing same, and use thereof. Specifically, a compound represented by formula (I), a stereoisomer thereof, an optical isomer thereof, a pharmaceutically acceptable salt thereof, a crystal form thereof, an isotopic derivative thereof, a prodrug thereof, a metabolite thereof, a solvate thereof, or a hydrate thereof, a composition containing the compound, and use thereof as a WRN helicase inhibitor, which can be used to prepare a drug for treating WRN helicase-mediated diseases.

Description

A bicyclic derivative for inhibiting WRN helicase, a pharmaceutical composition containing the same, and its application Technical Field

[0001] The present invention belongs to the field of medicinal chemistry, and particularly relates to a biheterocyclic derivative for inhibiting WRN helicase, a pharmaceutical composition containing the same, and applications thereof. Background Art

[0002] Microsatellite instability (MSI) refers to a phenomenon in which DNA mismatch repair deficiency (dMMR) causes insertions, deletions, and incorporations of bases during DNA replication and recombination, leading to altered microsatellite sequences (Cell 155, 858–868 (2013)). MSI is present in many tumors, but is particularly common in colorectal, gastric, endometrial, and ovarian cancers (JCO Precis Oncol 2017; 2017: 17.00073-17.00073). MSI is significantly correlated with response to immune checkpoint blockade, a finding often exploited in clinical treatment of MSI tumors. However, 45–60% of MSI tumors remain unresponsive to immune checkpoint blockade (Science 357, 409–413 (2017)). Therefore, the search for novel treatments for MSI tumors is urgent.

[0003] Werner syndrome RecQ helicase (WRN), a member of the RecQ family of DNA helicases, unwinds non-canonical secondary DNA structures encountered during replication and recombination. Mutations in WRN can cause chromosomal instability disorders such as Werner syndrome. Studies have shown that WRN is an important synthetic lethal target in MSI tumor cells. Loss of WRN helicase activity in MSI tumor cells causes DNA double-strand breaks, promoting apoptosis and cell cycle arrest (Nature 568, 551–556, 2019). Studies have demonstrated WRN-dependent mechanisms in MSI tumor cells (Nature 586, 292–298, 2020). TA-dinucleotide repeat sequences are highly unstable and undergo extensive amplification in MSI tumor cells. When WRN is lost, these repeat sequences are susceptible to cleavage by the MUS81 nuclease, leading to extensive chromosome breaks. This research provides strong support for the development of WRN-targeted therapeutics for MSI cancers. The research group of Professor Zhang Lin / Yu Jian from the University of Pittsburgh reported (PNAS.2022;119(51):e2211775119.) the molecular mechanism by which WRN deficiency causes apoptosis in microsatellite unstable colorectal cancer cells. In microsatellite unstable colorectal cancer cells, WRN deficiency produces a synthetic lethal effect through the p53 / PUMA-mediated apoptosis signaling pathway, while p53 mutations cause the synthetic lethal effect to be significantly weakened or even eliminated. In addition, Dr. Mathew J Garnett of the Wellcome Sanger Institute reported (Cancer Discov.2021 Aug;11(8):1923-1937) that WRN gene knockout was performed on 60 drug-resistant colorectal cancer experimental models and found that more than 90% of the models were dependent on WRN. Targeting the WRN gene is expected to help develop new targeted therapies for the treatment of drug-resistant colorectal cancer. Therefore, the development of WRN inhibitors as a single therapy or in combination with targeted drugs, chemotherapy or immunotherapy for dMMR / MSI tumors has great value and prospects.

[0004] Summary of the Invention

[0005] The present invention provides a biheterocyclic derivative for inhibiting WRN helicase, a pharmaceutical composition containing the same and application thereof.

[0006] In a first aspect of the present invention, there is provided a compound represented by formula (I), its stereoisomers, its optical isomers, its pharmaceutically acceptable salts, its crystal forms, its isotopic derivatives, its prodrugs, its metabolites, its solvates or its hydrates,

[0007] in,

[0008] X is selected from CH or N; and when X is CH, said CH may be substituted by R5 or R6 (i.e., X is C-R5 or C-R6);

[0009] m, n, e and q are each independently selected from: 0, 1, 2 or 3;

[0010] R1 is selected from the following group: C 3-6 Cycloalkyl, C 4-6 Alkynyl, C 4-10 Cycloalkenyl, C6-C 10 Aryl, 5-10 membered heteroaryl, saturated or partially unsaturated 5-10 membered heterocyclic group; wherein, said R1 can be optionally replaced by 0, 1, 2 or 3 R 1a 、R 1b 、R 1c 、R 1d 、R 1e 、R 1f 、R 1g 、R i or R h replace;

[0011] R2 is selected from the following group: C 1-4 Alkyl, C 1-4 Deuterated alkyl or C 1-4 alkyl halide;

[0012] R3 and R4 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 3-6 Alkoxy, C 6-10 Alkylamino, C 1-4 R3 and R4 can form a 4-6 membered carbocyclic ring or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently optionally substituted by 0, 1, 2 or 3 R7;

[0013] R5 and R6 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, -SO2CF3, -SF5, C 3-6 Cycloalkyl, C 6-10 Alkylamino, C 1-4 Alkenyl, C 2-6 Alkynyl, C 1-4 Halogenated alkenyl, C 2-6Alkynyl; R5 and R6 can form a 4-6 membered carbocyclic or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above contains 0, 1, 2 or 3 unsaturated double bonds, each of which is independently optionally substituted by 0, 1, 2 or 3 R8;

[0014] R7 and R8 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy, or two R7 or R8 together form =O;

[0015] Cy A Select from the following groups: Among them, the Cy A Can optionally be replaced by one or more R 12 Substituent substituted; d is selected from 0, 1, 2 or 3; each R 12 Each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy, cyano substituted C 1-4 Alkyl, methoxy substituted C 1- 4 alkyl, C 1-2 Haloalkoxy substituted C 1-4 Alkyl, or two R 12 Together they form =O; B is selected from O or S;

[0016] Cy B Selected from the following group: 6-9 membered heteroaryl; R3 and R4 are each independently selected from the following group: H, D, hydroxyl, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 6-10 Alkylamino, C 1-4 a halogenated alkoxy group, or two R3 or R4 together form =O; R3 and R4 together with the carbon atoms to which they are attached can form a 4-6 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above can be independently optionally replaced by 0, 1, 2 or 3 R 42 replace;

[0017] R 42 Each independently selected from the group consisting of hydroxy, halogen, cyano, C1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy, or two R 42 Together form = O;

[0018] Wherein, the heteroaryl or heterocyclic group includes 1, 2, 3, 4 or 5 heteroatoms selected from the following group as ring atoms: N, O, S or Se; the carbocyclic ring, heterocyclic ring, heterocyclic group can be a monocyclic ring, a fused ring, a spirocyclic ring or a bridged ring structure, which can be saturated or partially unsaturated, but not aromatic;

[0019] R 1a and R 1b Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 1b and R 1c Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0020] R 1c and R 1d Each independently selected from the group consisting of H, D, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, sulfone, C 1-6 Alkylamino;

[0021] R 1e and R 1f Each independently selected from the group consisting of H, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 3-6 Halogenated cycloalkyl, Methylamino-substituted C 1-4 Alkyl, methoxy substituted C 1-4 alkyl;

[0022] Each R 1gEach independently selected from the group consisting of H, D, hydroxy, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 haloalkoxy;

[0023] R i C 3-6 cycloalkoxy;

[0024] Each R h Each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy.

[0025] In another preferred embodiment, R1 is selected from the following group:

[0026] (1)

[0027] R is selected from CH, CD, CR 1e or N;

[0028] Z and W are independently selected from CH2, CR 1e R 1f NR 1e or O, and Z and W are not CH2 or CR at the same time 1e R 1f ; Among them, R 1e and R 1f Each independently selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 deuterated alkyl;

[0029] Y and U are independently selected from CH2 or CR 1e R 1f , and Y and U are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 Deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0030] or

[0031] Z and W are independently selected from CH2 or NR 1e , and Z and W are not CH2 at the same time; where R 1e Selected from the group consisting of deuterium, C 1-4 deuterated alkyl;

[0032] Both Y and U are CR 1e R 1f ; Among them, R 1e and R 1f are independently selected from the following groups: H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0033] (2)

[0034] R is selected from CH, CD, CR 1e or N;

[0035] F is selected from NR 1e or O; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 deuterated alkyl;

[0036] K and G are independently selected from CH2, CR 1e R 1f , and K and G are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 Deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatom contained in the heterocyclic ring is selected from O, N and S.

[0037] In another preferred embodiment, R1 is

[0038] R is selected from CH, CD, CR 1e or N;

[0039] Z and W are independently selected from CH2, CR 1e R 1f or O, and at least one of Z and W is O;

[0040] R 1e 、R 1f , Y and U are defined as above.

[0041] In another preferred embodiment, R1 is selected from the following group:

[0042] (1)C 3-6 Cycloalkyl, the C 3-6 The cycloalkyl group may be optionally substituted with 0, 1, 2 or 3 R 1a or R 1b Replacement; R 1a and R 1b Each independently selected from the group consisting of H, D, oxo, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 1b and R 1c Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0043] (2)C 4-6 Alkynyl, C 4-6 Alkynyl is optionally substituted with 0, 1, 2 or 3 R 1c and R 1d Replacement; R 1c and R 1d Each independently selected from the group consisting of H, D, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, sulfone, C 1-6 Alkylamino;

[0044] (3) R is selected from CH, CD, CR 1e or N; Y, Z, W, U are independently selected from CR 1e R 1f NR 1e or oxygen atoms, R 1e and R 1f Each independently selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4Halogenated alkyl, C 3-6 Halogenated cycloalkyl, Methylamino-substituted C 1-4 Alkyl, methoxy substituted C 1-4 Alkyl; R 1e and R 1f Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0045] (4) R is selected from CH, CD, CR 1e or N; K, F, G are independently selected from CR 1e R 1f NR 1e or oxygen atoms, R 1e and R 1f Each independently selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 3-6 Halogenated cycloalkyl, Methylamino-substituted C 1-4 Alkyl, methoxy substituted C 1-4 Alkyl; R 1e and R 1f Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0046] (5)

[0047] (6) a selenium-containing aromatic heterocycle, wherein the selenium-containing aromatic heterocycle is independently optionally substituted with 0, 1, 2 or 3 R 1g Replacement; R 1g Each independently selected from the group consisting of H, D, hydroxy, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 haloalkoxy;

[0048] (7) R i Selected from C 3-6 Cycloalkoxy or halogen-substituted C 3-6 Cycloalkoxy, and Cy C Selected from the group consisting of phenyl, 5-12 membered heteroaryl, C 4-10 Cycloalkenyl, C 4-10heterocyclyl; f is selected from 0, 1, 2 or 3; R h Each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy.

[0049] (8) Cy D is selected from the group consisting of a benzo five-membered heterocycle, a benzo five-membered aromatic heterocycle, a benzo six-membered heterocycle, a benzo six-membered aromatic heterocycle, a pyrido five-membered heterocycle, a pyrido five-membered aromatic heterocycle, a pyrimido five-membered heterocycle, a pyrimido five-membered aromatic heterocycle, a pyrido six-membered heterocycle, a pyrido six-membered aromatic heterocycle, a pyrimido six-membered heterocycle, and a pyrimido six-membered aromatic heterocycle; the heteroatoms of the heterocycles and aromatic heterocycles are selected from O, N, and S; p and t are each independently selected from 0, 1, 2, 3, or 4; R m and R n Each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy; R m and R n Together with the carbon atoms to which they are attached, they can form a 3-6 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; R m Can be 0, 1, 2, 3 or 4 R m1 Replacement, R n Can be 0, 1, 2, 3 or 4 R n1 Replacement; R m1 and R n1 Each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy, C 3-6 cycloalkoxy;

[0050] R2 is selected from the following group: C 1-4 Alkyl, C 1-4 Deuterated alkyl or C 1-4 alkyl halide;

[0051] Cy A Select from the following groups:

[0052] R 12Each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy, cyano substituted C 1-4 Alkyl, methoxy substituted C 1-4 Alkyl, C 1-2 Haloalkoxy substituted C 1- 4 alkyl; B is selected from O or S; d is selected from 0, 1, 2 or 3;

[0053] in represents a covalent bond to the parent nucleus, and represents a covalent bond to the carbonyl group; Covalent bonds representing connections;

[0054] Cy B Selected from the group consisting of 6-9 membered heteroaryl;

[0055] R3 and R4 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 6-10 Alkylamino, C 1-4 R3 and R4 can form a 4-6 membered carbocyclic ring or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently optionally substituted by 0, 1, 2 or 3 R7;

[0056] R7 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 haloalkoxy;

[0057] X is selected from CH or N; and when X is CH, said CH may be substituted by R5 or R6 (i.e., X is C-R5 or C-R6);

[0058] R5 and R6 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, -SO2CF3, -SF5, C 3-6 Cycloalkyl, C 6-10 Alkylamino, C1-4 Alkenyl, C 2-6 Alkynyl, C 1-4 Halogenated alkenyl, C 2-6 Alkynyl; R5 and R6 can form a 4-6 membered carbocyclic or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above contains 0, 1, 2 or 3 unsaturated double bonds, each of which is independently optionally substituted by 0, 1, 2 or 3 R8;

[0059] R8 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy.

[0060] In another preferred embodiment, R1 is selected from the following group:

[0061] In another preferred embodiment, Cy A Select from the following groups:

[0062] Among them, R 12 and d are defined as above.

[0063] In another preferred embodiment, Cy A Select from the following groups:

[0064] in represents a covalent bond to the parent nucleus, and represents a covalent bond to the carbonyl group;

[0065] R1 is selected from the group consisting of:

[0066] (1)C 3-6 Cycloalkyl, C 3-6 Cycloalkyl is optionally substituted by 0, 1, 2 or 3 R 1a and R 1b Replacement; R 1a and R 1b Each independently selected from the group consisting of H, D, oxo, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C1-6 Alkylamino; R 1a and R 1b Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0067] (2)C 4-6 Alkynyl, C 2-6 Alkynyl is optionally substituted with 0, 1, 2 or 3 R 1c and R 1d Replacement; R 1c and R 1d Each independently selected from the group consisting of H, D, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, sulfone, C 1-6 Alkylamino;

[0068] (3) Cycloalkenyl, wherein the cycloalkenyl is a partially unsaturated monocyclic ring containing 5 or 6 ring carbon atoms, and the cycloalkenyl is unsubstituted or substituted with 1, 2, 3 or 4, preferably 1 or 2 R 33 and R 34 Replacement; R 33 and R 34 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 33 and R 34 Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 3-5 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 35 Replacement; R 35 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-4 Alkoxy, C 1- 4-haloalkoxy;

[0069] (4) Heterocyclyl, wherein the heterocyclyl is a 5- or 6-membered fully saturated or partially unsaturated group comprising ring carbon atoms and 1 or 2 ring heteroatoms independently selected from N, O and S, and wherein the heterocyclyl is unbridged or bridged, and the bridge is 1 or 2 carbon atoms, wherein the heterocyclyl is unsubstituted or substituted with 1, 2, 3 or 4 R 36 or R 37 Replacement; R 36 and R 37 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1- 4 alkoxy, C 1-4 Halogenated alkoxy, C 2-6 Halogenated alkenyl, C 1-6 Alkylamino; R 36 and R 37 Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 3-5 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 38 Replacement; R 38 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkoxy, C 1-4 haloalkoxy;

[0070] (5) phenyl, heteroaryl; wherein the heteroaryl is a 5- or 6-membered fully unsaturated monocyclic group comprising ring carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O, S and Se, preferably 1 or 2 ring heteroatoms, wherein the total number of ring S atoms does not exceed 1, and the total number of ring O atoms does not exceed 1, wherein the phenyl and heteroaryl are unsubstituted or replaced by 1, 2 or 3 ring heteroatoms independently selected from R 39 and R 40 substituted by a substituent, wherein R 39 and R 40 Independently selected from H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkoxy, C 3-6 Cycloalkoxy, C 2-6 Halogenated alkenyl, C 1-6 Alkylamino; R 39 and R 40Together with the carbon atoms to which they are attached, they can form a 3-6 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the formed 3-6 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 41 Replacement; R 41 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkoxy, C 1-4 haloalkoxy;

[0071] Cy B Selected from the following group: 6-membered heteroaryl; R3 and R4 are each independently selected from the following group: H, D, hydroxyl, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 6-10 Alkylamino, C 1-4 R3 and R4 can form a 4-6 membered carbocyclic ring or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently optionally replaced by 0, 1, 2 or 3 R 42 Replacement; R 42 Each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 haloalkoxy;

[0072] R2 is selected from the following group: C 1-4 Alkyl, C 1-4 Deuterated alkyl or C 1-4 alkyl halide;

[0073] R3 is selected from the group consisting of halogen, cyano, C 3-6 Alkoxy, C 1-4 alkyl halide;

[0074] X is selected from CH or N; and when X is CH, said CH may be substituted by R5 or R6 (i.e., X is C-R5 or C-R6);

[0075] R5 and R6 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C1-4 Halogenated alkoxy, -SO2CF3, -SF5, C 3-6 Cycloalkyl, C 6-10 Alkylamino, C 1-4 Alkenyl, C 2-6 Alkynyl, C 1-4 Halogenated alkenyl, C 2-6 Alkynyl; R5 and R6 can form a 4-6 membered carbocyclic or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above contains 0, 1, 2 or 3 unsaturated double bonds, each of which is independently optionally substituted by 0, 1, 2 or 3 R8;

[0076] R8 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy.

[0077] In another preferred embodiment, Cy A Select from the following groups:

[0078] In another preferred embodiment, Cy B Preferably from the following group:

[0079] Among them, R 44 、R 45 and R 46 Each independently selected from H, D, halogen, CN, OH, C 1-6 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-6 Alkoxy, C 1-6 Halogenated alkyl, C 1-6 Halogenated alkoxy, C 1-6 Alkylamino, C 2-4 Alkenyl, C 1-4 Halogenated alkenyl, C 2-4 Alkynyl; R 45 and R 46 Together with the carbon atoms to which they are attached, they can form a 4-6 membered carbocyclic ring or heterocyclic ring or heteroaryl group, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring or heteroaryl group formed above can be independently optionally replaced by 0, 1, 2 or 3 R 47 Replacement; R 47 Independently selected from the group consisting of H, D, halogen, hydroxy, cyano, C 1-4 Alkyl, C1-4 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated cycloalkyl, C 1-4 Halogenated alkoxy.

[0080] In another preferred embodiment, R1 is preferably selected from the following group:

[0081] In another preferred embodiment, Cy B Select from the following groups:

[0082] wherein h is selected from 0, 1, 2 or 3; R 44 、R 45 and R 46 Each independently selected from H, D, halogen, CN, OH, C 1-6 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-6 Alkoxy, C 1-6 Halogenated alkyl, C 1-6 Halogenated alkoxy, C 1-6 Alkylamino, C 2-4 Alkenyl, C 1-4 Halogenated alkenyl, C 2-4 Alkynyl; R 45 and R 46 Together with the carbon atoms to which they are attached, they can form a 4-6 membered carbocyclic ring or heterocyclic ring or heteroaryl group, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring or heteroaryl group formed above can be independently optionally replaced by 0, 1, 2 or 3 R 47 Replacement; R 47 Independently selected from the group consisting of H, D, halogen, hydroxy, cyano, C 1-4 Alkyl, C 1-4 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated cycloalkyl, C 1-4 haloalkoxy;

[0083] R1 is selected from the following group: C 3-6 Cycloalkyl, C 3-6 Cycloalkyl, C 4-6 Alkynyl, cycloalkenyl, heterocyclyl, phenyl, heteroaryl;

[0084] Among them, the C 3-6 Cycloalkyl is optionally substituted by 0, 1, 2 or 3 R 1aand R 1b Replacement; R 1a and R 1b Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 1a and R 1b Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S;

[0085] The C 4-6 Alkynyl is optionally substituted with 0, 1, 2 or 3 R 1c and R 1d Replacement; R 1c and R 1d Each independently selected from the group consisting of H, D, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, sulfone, C 1-6 Alkylamino;

[0086] The cycloalkenyl group is a partially unsaturated monocyclic ring containing 5 or 6 ring carbon atoms, and the cycloalkenyl group is unsubstituted or substituted by 1, 2, 3 or 4, preferably 1 or 2 R 33 and R 34 Replacement; R 33 and R 34 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 33 and R 34 Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 3-5 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 35 Replacement; R 35Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-4 Alkoxy, C 1-4 haloalkoxy;

[0087] The heterocyclyl is a 5- or 6-membered fully saturated or partially unsaturated group comprising ring carbon atoms and 1 or 2 ring heteroatoms independently selected from N, O and S, and wherein the heterocyclyl is unbridged or bridged and the bridge is 1 or 2 carbon atoms, wherein the heterocyclyl is unsubstituted or substituted with 1, 2, 3 or 4 R 36 or R 37 Replacement; R 36 and R 37 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 2-6 Halogenated alkenyl, C 1-6 Alkylamino; R 36 and R 37 Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 3-5 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 38 Replacement; R 38 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkoxy, C 1-4 haloalkoxy;

[0088] The heteroaryl group is a 5- or 6-membered fully unsaturated monocyclic group comprising ring carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O, S and Se, preferably 1 or 2 ring heteroatoms, wherein the total number of ring S atoms does not exceed 1 and the total number of ring O atoms does not exceed 1, wherein the phenyl and heteroaryl groups are unsubstituted or replaced by 1, 2 or 3 groups independently selected from R 39 and R 40 substituted by a substituent, wherein R 39 and R 40 Independently selected from H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkoxy, C 3-6 Cycloalkoxy, C 2-6 Halogenated alkenyl, C 1-6 Alkylamino; R 39 and R 40 Together with the carbon atoms to which they are attached, they can form a 3-6 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the formed 3-6 membered carbocyclic or heterocyclic ring can be substituted by 1, 2, 3 or 4 R 41 Replacement; R 41 Each independently selected from the group consisting of H, D, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkoxy, C 1-4 haloalkoxy;

[0089] Cy A Select from the following groups:

[0090] R 12 Each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Haloalkoxy, cyano substituted C 1-4 Alkyl, methoxy substituted C 1-4 Alkyl, C 1-2 Haloalkoxy substituted C 1- 4 alkyl; B is selected from O or S;

[0091] in represents a covalent bond to the parent nucleus, and represents a covalent bond to the carbonyl group;

[0092] R2 is selected from the following group: C 1-4 Alkyl, C 1-4 Deuterated alkyl or C 1-4 alkyl halide;

[0093] R3 is selected from the group consisting of halogen, cyano, C 3-6 Alkoxy, C 1-4 alkyl halide;

[0094] X is selected from CH or N; and when X is CH, said CH may be substituted by R5 or R6 (i.e., X is C-R5 or C-R6);

[0095] R5 and R6 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, -SO2CF3, -SF5, C 3-6 Cycloalkyl, C 6-10 Alkylamino, C 1-4 Alkenyl, C 2-6 Alkynyl, C 1-4 Halogenated alkenyl, C 2-6 Alkynyl; R5 and R6 can form a 4-6 membered carbocyclic or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above contains 0, 1, 2 or 3 unsaturated double bonds, each of which is independently optionally substituted by 0, 1, 2 or 3 R8;

[0096] R8 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkoxy.

[0097] In another preferred embodiment, R1 is preferably selected from the following group:

[0098] In another preferred embodiment, Cy A Select from the following groups:

[0099] In another preferred embodiment, Cy B Select from the following groups:

[0100] wherein h is selected from 0, 1, 2 or 3;

[0101] R 44 、R 45 and R 46 Each independently selected from H, D, halogen, CN, OH, C 1-6 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-6 Alkoxy, C 1-6 Halogenated alkyl, C 1-6 Halogenated alkoxy, C 1-6 Alkylamino, C 2-4Alkenyl, C 1-4 Halogenated alkenyl, C 2-4 Alkynyl; R 45 and R 46 Together with the carbon atoms to which they are attached, they can form a 4-6 membered carbocyclic ring or heterocyclic ring or heteroaromatic ring, wherein the heteroatoms contained in the heterocyclic ring or heteroaromatic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently optionally replaced by 0, 1, 2 or 3 R 47 replace;

[0102] R 47 Independently selected from the group consisting of H, D, halogen, hydroxy, cyano, C 1-4 Alkyl, C 1-4 Cycloalkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated alkyl, C 1-4 Halogenated cycloalkyl, C 1-4 Halogenated alkoxy.

[0103] In another preferred embodiment, Cy B Select from the following groups:

[0104] In another preferred embodiment, Select from the following groups: Select from the following groups:

[0105] Among them, R 48 、R 49 and R 50 Each independently selected from the following group: C 1-4 alkyl, C1-4 Haloalkyl, cyano, fluorine, chlorine.

[0106] In another preferred embodiment, Select from the following groups:

[0107] In some embodiments, the compound is selected from the group consisting of:

[0108] The second aspect of the present invention provides a pharmaceutical composition comprising the compound as described in the first aspect of the present invention, its stereoisomers, its optical isomers, its pharmaceutically acceptable salts, its crystal forms, its isotopic derivatives, its prodrugs, its metabolites, its solvates or its hydrates; and a pharmaceutically acceptable carrier.

[0109] In another preferred embodiment, the pharmaceutical composition further comprises other therapeutic agents.

[0110] In another preferred embodiment, the other therapeutic agents are chemotherapy drugs, immunotherapy drugs, and targeted drugs.

[0111] In another preferred embodiment, the chemotherapy drug is selected from: anastrozole, bicalutamide, bleomycin sulfate, busulfan, busulfan injection, capecitabine, N4-pentyloxycarbonyl-5-deoxy-5-fluorocytidine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, cytosine arabinoside, cytarabine liposome injection, dacarbazine, actinomycin, daunorubicin hydrochloride, daunorubicin citrate, liposome injection, dexamethasone, docetaxel, doxorubicin hydrochloride, etoposide , fludarabine phosphate, 5-fluorouracil, flutamide, tezacitabine, gemcitabine, hydroxyurea, idarubicin, ifosfamide, irinotecan, L-asparaginase, folinic acid, melphalan, 6-mercaptopurine, methotrexate, mitoxantrone, gemtuzumab ozogamicin, paclitaxel, pentostatin, tamoxifen citrate, teniposide, 6-thioguanine, thiotepa, tirapazamine, topotecan hydrochloride for injection, vinblastine, vincristine, and vinorelbine; preferably 5-fluorouracil (5-FU) and irinotecan.

[0112] In another preferred embodiment, the immunotherapy drug is selected from: PD-1 inhibitors (including small molecules and antibody drugs), PD-L1 inhibitors (including small molecules and antibody drugs), CTLA-4 inhibitors, PD-1 / CTLA-4 dual antibodies, PD-1 / TIGIT dual antibodies, PD-L1 / 4-1BB dual antibodies, PD1 / VEGF-A dual antibodies, PD-1 / LAG-3 dual antibodies;

[0113] Among them, the PD-1 inhibitor is selected from the group consisting of: nivolumab, pembrolizumab, camrelizumab, toripalimab, sintilimab, tislelizumab, penampalimab, sepalizumab, sirulimab, dostarlimab, and PDR001;

[0114] Among them, the PD-L1 inhibitor is selected from: durvalumab, atezolizumab, envolizumab, and sugemalimab;

[0115] Among them, the CTLA-4 inhibitor is selected from ipilimumab;

[0116] Among them, the PD-1 / CTLA-4 dual antibody is selected from cardunilizumab.

[0117] In another preferred embodiment, the targeted drugs include but are not limited to VEGFR inhibitors (lenvatinib), NTRK inhibitors (entrectinib), BRAFV600 inhibitors (dabrafenib), MEK inhibitors (trametinib), EGFR monoclonal antibodies (cetuximab), PARP inhibitors (olaparib, niraparib, fluzoparib and pamiparib, etc.) and ATR inhibitors.

[0118] The third aspect of the present invention provides a method for preparing a pharmaceutical composition, comprising the steps of: mixing a pharmaceutically acceptable carrier with the compound of the present invention, its stereoisomers, its optical isomers, its pharmaceutically acceptable salts, its crystal forms, its isotopic derivatives, its prodrugs, its metabolites, its solvates or its hydrates, to form the pharmaceutical composition as described in the second aspect of the present invention.

[0119] The fourth aspect of the present invention provides a compound as described in the first aspect of the present invention, its stereoisomers, its optical isomers, its pharmaceutically acceptable salts, its crystal forms, its isotopic derivatives, its prodrugs, its metabolites, its solvates or its hydrates, or the pharmaceutical composition as described in the second aspect of the present invention for the preparation of a drug for treating WRN helicase-mediated diseases.

[0120] In another preferred embodiment, the disease is tumor.

[0121] In another preferred embodiment, the disease is microsatellite unstable tumor.

[0122] In another preferred embodiment, the tumor is characterized by microsatellite instability-high (MSI-H) or a tumor with defective mismatch repair (dMMR) or "dMMR signature";

[0123] Wherein, the microsatellite unstable tumors include but are not limited to uterine body tumors, endometrial cancer, colon adenocarcinoma, gastric adenocarcinoma, rectal adenocarcinoma, adrenal cortical carcinoma, uterine carcinosarcoma, cervical squamous cell carcinoma, endocervical adenocarcinoma, esophageal cancer, breast cancer, renal clear cell carcinoma and ovarian serous cystic carcinoma;

[0124] Among them, tumors with "defective mismatch repair" (dMMR) or "dMMR characteristics" include but are not limited to lung cancer, breast cancer, kidney cancer, colorectal cancer, ovarian cancer, prostate cancer, upper aerodigestive tract cancer, gastric cancer, endometrial cancer, liver cancer, pancreatic cancer, hematopoietic and lymphoid tissue cancer, skin cancer, thyroid cancer, pleural cancer, autonomic nervous system tumors, soft tissue tumors, rhabdoid sarcoma, melanoma and other tumors.

[0125] In another preferred embodiment, the drug is administered orally, by injection, inhalation or cavity administration.

[0126] In another preferred embodiment, the dosage form of the drug is selected from the group consisting of tablets, capsules, dispersants, suspensions, granules, sprays, gels, sustained-release preparations, oral solutions, pills and nanoformulations.

[0127] In a fifth aspect, the present invention provides a method for treating a WRN helicase-mediated disease, comprising the steps of administering to a subject in need of treatment an effective amount of the compound as described in the first aspect, its stereoisomers, its optical isomers, its pharmaceutically acceptable salts, its crystal forms, its isotopic derivatives, its prodrugs, its metabolites, its solvates or its hydrates, or the pharmaceutical composition as described in the second aspect.

[0128] It should be understood that within the scope of the present invention, the above-mentioned technical features of the present invention and the technical features described in detail below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be listed here one by one. DETAILED DESCRIPTION

[0129] After extensive and in-depth research, the inventors discovered for the first time a bicyclic heterocyclic derivative with a novel structure and good WRN helicase inhibitory activity. On this basis, the inventors completed the present invention.

[0130] the term

[0131] Unless otherwise stated, the following terms used in this application (including the specification and claims) have the definitions given below.

[0132] "Alkyl" (alone or as part of another group) refers to a monovalent straight or branched chain saturated hydrocarbon group consisting solely of carbon and hydrogen atoms. "C1-C6 alkyl" refers to an alkyl group containing 1 to 6 carbon atoms, preferably a C1-C4 alkyl. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, isobutyl, sec-butyl, tert-butyl, pentyl, n-hexyl, octyl, and dodecyl. In the present invention, alkyl is also intended to include deuterated alkyl groups, examples of which include, but are not limited to, CD3, CD2CD3, and CD2CD2CD3.

[0133] "Alkoxy" (alone or as part of another group) refers to a radical of the formula -OR or -R'-OR, wherein R is an alkyl radical as defined herein and R' is an alkylene radical. Examples of alkoxy radicals include, but are not limited to, methoxy, ethoxy, isopropoxy, tert-butoxy, -CH2O-CH3, -CH2CH2-O-CH3, -CH2-O-CH2CH3, and the like.

[0134] "Halogen" (alone or as part of another group) refers to fluorine, chlorine, bromine or iodine.

[0135] "Haloalkyl" (alone or as part of another group) refers to a group in which one or more hydrogen atoms in an alkyl group as described above are replaced by the same or different halogen atoms. "Halo-C1-C6 alkyl" is preferably a halo-C1-C4 alkyl group. Examples of haloalkyl groups include, but are not limited to, -CH2Cl, -CH2CF3, -CH2CCl3, and perfluoroalkyl groups (e.g., -CF3-, -CF2CF3).

[0136] "Haloalkoxy" (alone or as part of another group) refers to a radical of the formula -OR, wherein R is a haloalkyl radical as defined herein. Examples of haloalkoxy radicals include, but are not limited to, trifluoromethoxy, difluoromethoxy, 2,2,2-trifluoroethoxy, and the like.

[0137] "Cycloalkyl" (alone or as part of another group) refers to a monovalent saturated carbocyclic group consisting of a mono- or bicyclic ring composed solely of carbon and hydrogen atoms, wherein "C3-C8 cycloalkyl" refers to a saturated carbocyclic group containing 3 to 8 carbon atoms, preferably a C3-C6 cycloalkyl. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like. Cycloalkyl groups may optionally be substituted with one or more substituents, each of which is independently hydroxy, alkyl, alkoxy, halogen, haloalkyl, amino, monoalkylamino, or dialkylamino. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

[0138] "Cycloalkyloxy" (alone or as part of another group) refers to a radical of the formula -OR, wherein R is a cycloalkyl radical as defined herein. Exemplary cycloalkyloxy radicals include cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy, and the like.

[0139] "Heterocyclyl" or "heterocycle" (alone or as part of another group) refers to a fully saturated or partially unsaturated cyclic group (including but not limited to 3-7 membered monocyclic, 6-11 membered bicyclic, or 8-16 membered tricyclic ring systems), wherein at least one heteroatom is present in the ring having at least one carbon atom. Each heterocyclic ring containing a heteroatom carries 1, 2, 3 or 4 heteroatoms selected from nitrogen atoms, oxygen atoms or sulfur atoms, wherein the nitrogen atoms or sulfur atoms may be oxidized and the nitrogen atoms may be quaternized. Heterocycloalkane (base) refers to a fully saturated heterocycle (base). Unless otherwise specified, the heterocycle (base) may be saturated or partially unsaturated, but not aromatic. The heterocyclic group may be attached to any heteroatom or carbon atom residue of the ring or ring system molecule. Among them, "3-8 membered heterocyclyl" refers to a group having 3-8 ring members. Typical monocyclic heterocycles include, but are not limited to, azetidinyl, pyrrolidinyl, oxetanyl, pyrazolinyl, imidazolinyl, imidazolidinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolidinyl, tetrahydrofuranyl, piperidinyl, piperazinyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, hexahydroazepinyl, 4-piperidonyl, tetrahydropyranyl, morpholinyl, thiomorpholinyl, thiomorpholinyl sulfoxide, thiomorpholinyl sulfone, 1,3-dioxanyl, and tetrahydro-1,1-dioxythiophene. Polycyclic heterocyclic groups include spiro, fused, and bridged heterocyclic groups; wherein the spiro, fused, and bridged heterocyclic groups are optionally connected to other groups by single bonds, or further connected to other cycloalkyl, heterocyclic, aryl, and heteroaryl groups via any two or more atoms on the ring.

[0140] "Aryl" (alone or as part of other groups) refers to an aromatic cyclic hydrocarbon compound group with 1-5 rings, especially monocyclic and bicyclic groups. Where it contains two or more aromatic rings (bicyclic, etc.), the aromatic rings of the aryl group can be connected by a single bond (such as biphenyl) or fused (such as naphthalene, anthracene, etc.). Among them, "C6-C12 aryl" refers to an aromatic cyclic hydrocarbon compound group containing 6, 7, 8, 9, 10, 11 or 12 ring carbon atoms. Examples of aryl (especially monocyclic and bicyclic groups) include but are not limited to phenyl, biphenyl or naphthyl. Aryl can be fused to heterocyclic groups through a single bond or any two adjacent ring C atoms, for example: benzotetrahydrofuranyl, chromanyl, benzodioxanyl, wait.

[0141] "Heteroaryl," "heteroaromatic ring," or "aromatic heterocycle" (alone or as part of another group) refers to an aromatic group having heteroatoms in its backbone. "5-12 membered heterocyclyl" refers to a monocyclic, bicyclic, or tricyclic group of 5 to 12 ring atoms, containing at least one (e.g., 1, 2, or 3) ring heteroatom selected from N, O, or S, with the remaining ring atoms being C. It should be understood that the point of attachment of the heteroaryl group is located on the aromatic ring. Heteroaryl groups preferably have 5-8 ring atoms (5-8 members), and more preferably have 5-6 ring atoms (5-6 members). Examples of heteroaryl groups include, but are not limited to, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, pyrazinyl, thienyl, furanyl, pyranyl, pyridinyl, pyrrolyl, pyrazolyl, pyrimidinyl, quinolinyl, isoquinolinyl, benzofuranyl, benzothiophenyl, benzothiopyranyl, benzimidazolyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzopyranyl, indolyl, isoindolyl, triazolyl, triazinyl, quinoxalinyl, purinyl, quinazolinyl, quinolizinyl, naphthyridinyl, pteridinyl, carbazolyl, aza Base, diazepine Acridinium, etc.

[0142] "Polysubstituted" means comprising two or more substitutions.

[0143] In the present invention, the alkyl, cycloalkyl, heterocyclic, aryl, heteroaryl and other groups include substituted alkyl, cycloalkyl, heterocyclic, aryl, heteroaryl and the like unless otherwise specified, and the substituents include (but are not limited to) halogen, hydroxyl, cyano, alkyl, cycloalkyl, heterocyclic, aryl, heteroaryl and the like.

[0144] "Deuterated compound" refers to a compound in which one or more hydrogen atoms (H) are replaced by deuterium atoms (D). 1-4 Deuterated alkyl refers to a compound in which at least three hydrogen atoms (H) or more hydrogen atoms (H) are replaced by deuterium atoms (D). The substituents include (but are not limited to) deuterated methyl, deuterated ethyl, etc.

[0145] Active ingredient

[0146] As used herein, "compound of the present invention" refers to the compound represented by formula (I), and also includes its stereoisomers, optical isomers, pharmaceutically acceptable salts, crystal forms, isotopic derivatives, prodrugs, metabolites, solvates or hydrates thereof.

[0147] Unless otherwise indicated, the structural formulas described herein are intended to include all stereoisomers (e.g., cis-trans isomers, enantiomers, diastereomers, and conformational isomers): R and S configurations containing asymmetric centers, (Z) and (E) isomers of double bonds, cis-trans isomers of cycloalkanes, etc. Therefore, individual stereochemical isomers of the compounds of the present invention or mixtures of their enantiomers, diastereomers, or conformational isomers are all within the scope of the present invention.

[0148] The compounds of the present invention may contain cis- and trans-isomers, one or more chiral carbon atoms, and thus may produce stereoisomeric forms such as cis- and trans-isomers, chiral isomers, enantiomers, diastereomers, and other combinations thereof. Cis- and trans-isomerism refers to the diastereoisomerism that occurs due to restrictions on the free rotation of the compound molecule, resulting in different spatial arrangements of the various groups. These restrictions are generally caused by the presence of functional groups in the organic compound structure that cannot rotate freely, such as C=C double bonds, C=N double bonds, C=S double bonds, N=N double bonds, heterocycles, or cycloalkanes. Organic molecules containing such isomers, such as alkenes, azo compounds, and alicyclic hydrocarbons, are considered cis- and trans-isomers. The cis form refers to the presence of ligands of the same type in adjacent positions, typically designated "cis" or "cis-"; the trans form refers to the presence of ligands of the same type in diagonal positions, typically designated "trans" or "trans-". Each chiral carbon atom can be defined as either (R)- or (S)- based on stereochemistry. The present invention is intended to include all possible isomers, as well as racemates and optically pure forms thereof. The compounds of the present invention may be prepared using racemates, cis-trans isomers, chiral isomers, diastereomers, or enantiomers as starting materials or intermediates. Optically active isomers may be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, such as crystallization and chiral chromatography.

[0149] Conventional techniques for preparing / isolating individual optical isomers (i.e., cis-trans isomers and chiral isomers) include chiral synthesis from appropriate cis-trans precursors or optically pure precursors, or resolution of racemates (or racemates of salts or derivatives) using, for example, chiral high performance liquid chromatography, as described, for example, in Gerald Gübitz and Martin G. Schmid (Eds.), Chiral Separations, Methods and Protocols, Methods in Molecular Biology, Vol. 243, 2004; A.M. Stalcup, Chiral Separations, Annu. Rev. Anal. Chem. 3:341-63, 2010; Fumiss et al. (eds.), VOGEL'S ENCYCLOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5.sup.TH ED., Longman Scientific and Technical Ltd., Essex, 1991, 809-816; Heller, Acc. Chem. Res. 1990, 23, 128.

[0150] If a synthesis of a specific stereoisomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, followed by separation of the resulting stereomixture and removal of the chiral auxiliary to obtain a pure cis-trans monomer, chiral monomer, or mixed stereoisomer. If the molecule contains a cis-trans isomeric center, pure cis- or trans-forms can be obtained by purification via column chromatography (normal-phase silica gel or reverse-phase high-performance liquid chromatography). Furthermore, if the molecule contains a basic functional group, such as an amino acid, or an acidic functional group, such as a carboxyl group, diastereomeric salts can be formed with a suitable optically active acid or base, and then separated by conventional means such as fractional crystallization or chromatography to obtain the pure enantiomers.

[0151] The present invention also includes isotopically labeled compounds (i.e., isotopic derivatives) that are equivalent to the original compounds disclosed herein. However, in practice, it is common for one or more atoms to be replaced by atoms with different atomic weights or mass numbers. Examples of isotopes in the isotopic derivatives of the present invention include hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine isotopes, such as 2H, 3H, 13C, 11C, 14C, 15N, 18O, 17O, 31P, 32P, 35S, 18F, and 36Cl, respectively. Isotopic derivatives of the compounds of the present invention are all within the scope of protection of the present invention. Herein, 3H-labeled compounds and 14C-labeled compounds are useful in tissue distribution experiments of drugs and substrates. Tritium (i.e., 3H) and carbon-14 (i.e., 14C) labeled compounds are easier to prepare and detect and are the first choice among isotopes. In addition, heavier isotope substitutions, such as deuterium (2H), may be preferred in certain therapeutics due to their excellent metabolic stability, such as increasing half-life in vivo or reducing dosage. Isotopically labeled compounds can be prepared using standard methods by replacing readily available isotopically labeled reagents with non-isotopic reagents using the protocols disclosed in the examples.

[0152] As used herein, the term "pharmaceutically acceptable salt" includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0153] "Pharmaceutically acceptable acid addition salts" refer to salts formed with inorganic or organic acids that retain the biological effectiveness of the free base without the side effects. Inorganic acid salts include, but are not limited to, hydrochlorides, hydrobromides, sulfates, nitrates, and phosphates; organic acid salts include, but are not limited to, formate, acetate, 2,2-dichloroacetate, trifluoroacetate, propionate, hexanoate, octanoate, decanoate, undecylenate, glycolate, gluconate, lactate, sebacate, adipate, glutarate, malonate, oxalate, maleate, succinate, fumarate, tartrate, citrate, palmitate, stearate, oleate, cinnamate, laurate, malate, glutamate, pyroglutamate, aspartate, benzoate, methanesulfonate, benzenesulfonate, p-toluenesulfonate, alginate, ascorbate, salicylate, 4-aminosalicylate, and naphthalene disulfonate. These salts can be prepared by methods known in the art.

[0154] "Pharmaceutically acceptable base addition salts" refer to salts formed with inorganic or organic bases that retain the biological effectiveness of the free acid without other side effects. Salts derived from inorganic bases include, but are not limited to, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Preferred inorganic salts are ammonium, sodium, potassium, calcium, and magnesium salts. The salt derived from organic base includes but is not limited to following salt: primary amines, secondary amines and tertiary amines, substituted amines, including natural substituted amines, cyclic amines and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins etc. Preferred organic bases include isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline and caffeine. These salts can be prepared by methods known to the profession.

[0155] Metabolites of the compound represented by formula (I) and pharmaceutically acceptable salts thereof, as well as prodrugs that can be converted into the compound represented by formula (I) and pharmaceutically acceptable salts thereof in vivo, are also included in the scope of protection of the present invention.

[0156] As used herein, the term "solvate" refers to a complex in which the compound represented by formula (I) is coordinated with solvent molecules to form a specific ratio.

[0157] As used herein, the term "hydrate" refers to a complex formed by the compound represented by formula (I) coordinated with water molecules to form a specific ratio.

[0158] As described herein, the compounds of the present invention may be substituted with any number of substituents or functional groups to expand their scope. Generally, the term "substituted" refers to the replacement of a hydrogen radical with a substituent of a specified structure. When multiple positions in a particular structure are substituted with multiple substituents of a specified structure, the substituents may be the same or different at each position. As used herein, the term "substituted" includes all permissible organic group substitutions. Broadly speaking, permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic organic groups. Herein, for example, heteroatoms such as nitrogen may be substituted with hydrogen or any of the permissible organic groups described above to supplement their valences. Furthermore, the present invention is not intended to limit the permissible organic groups in any way. The present invention recognizes that combinations of substituents and variable groups are advantageous for the treatment of diseases by forming stable compounds. The term "stable" herein refers to compounds that are stable and maintain the structural integrity of the compound over a sufficient period of time for testing, preferably for a sufficient period of time for efficacy, and is used herein for such purposes.

[0159] Pharmaceutical compositions and methods of administration

[0160] Since the compounds of the present invention can inhibit WRN helicase and are used to treat diseases such as tumors, the compounds of the present invention and their stereoisomers, optical isomers, pharmaceutically acceptable salts, crystalline forms, isotopic derivatives, prodrugs, metabolites, solvates or hydrates thereof, as well as pharmaceutical compositions containing the compounds of the present invention as the main active ingredient can be used to prevent and / or treat (stabilize, alleviate or cure) WRN helicase-related diseases, preferably tumors; more preferably, microsatellite unstable tumors; further preferably, tumors characterized by microsatellite instability-high (MSI-H) or having "defective mismatch repair deficiency (dMMR)" or "dMMR characteristics".

[0161] Microsatellite unstable tumors are particularly present in colorectal cancer, gastric cancer and endometrial cancer, and also in adrenocortical carcinoma, uterine cancer, cervical cancer, esophageal cancer, breast cancer, kidney cancer, prostate cancer and ovarian cancer. Microsatellite unstable tumors include but are not limited to uterine body tumors, endometrial cancer, colon adenocarcinoma, gastric adenocarcinoma, rectal adenocarcinoma, adrenocortical carcinoma, uterine carcinosarcoma, cervical squamous cell carcinoma, endocervical adenocarcinoma, esophageal cancer, breast cancer, kidney renal clear cell carcinoma and ovarian serous cystic carcinoma.

[0162] Tumors with "defective mismatch repair" (dMMR) or a "dMMR signature" include cancer types associated with documented MLH1, PMS2, MSH2, MSH3, MSH6, MLH3, and PMS1 mutations or epigenetic silencing, microsatellite fragile sites, or other gene inactivation mechanisms, including but not limited to lung cancer, breast cancer, kidney cancer, colorectal cancer, ovarian cancer, prostate cancer, upper aerodigestive tract cancer, gastric cancer, endometrial cancer, liver cancer, pancreatic cancer, cancers of hematopoietic and lymphoid tissues, skin cancer, thyroid cancer, pleural cancer, autonomic nervous system tumors, soft tissue tumors, rhabdoid sarcoma, melanoma, and other tumors.

[0163] The pharmaceutical compositions of the present invention comprise a safe and effective amount of a compound of the present invention and a pharmaceutically acceptable excipient or carrier. "Safe and effective amount" means an amount of the compound sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 1-2000 mg of the compound of the present invention per dose, more preferably 10-200 mg per dose. Preferably, "one dose" is one capsule or tablet.

[0164] "Pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use and must have sufficient purity and sufficiently low toxicity. "Compatibility" here means that the components in the composition can be mixed with the compound of the present invention and with each other without significantly reducing the efficacy of the compound. Some examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tween ), wetting agents (such as sodium lauryl sulfate), colorants, flavorings, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0165] There is no particular limitation on the administration of the compound or pharmaceutical composition of the present invention. Representative administration methods include, but are not limited to, oral administration and parenteral administration (intravenous, intramuscular, or subcutaneous).

[0166] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose, and acacia; (c) humectants, for example, glycerol; (d) disintegrants, for example, agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) solubilizers, for example, paraffin; (f) absorption accelerators, for example, quaternary ammonium compounds; (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents.

[0167] Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared using coatings and shell materials, such as enteric coatings and other materials known in the art. They may contain opacifying agents, and the release of the active compound or compounds in such compositions can be delayed in a certain portion of the digestive tract. Examples of useful encapsulating components are polymeric substances and waxes. If desired, the active compound can also be microencapsulated with one or more of the above-mentioned excipients.

[0168] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compound, the liquid dosage form may contain an inert diluent conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, or mixtures thereof.

[0169] Besides such inert diluents, the composition may also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0170] Suspensions, in addition to the active compounds, may contain suspending agents such as, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances.

[0171] Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.

[0172] The compounds of the present invention can be administered alone or in combination with other pharmaceutically acceptable therapeutic agents. Drugs administered in combination with WRN inhibitors include but are not limited to chemotherapeutics, immunotherapeutics, and targeted drugs. When administered in combination, the pharmaceutical composition also includes one or more (2, 3, 4 or more) other pharmaceutically acceptable therapeutic agents. One or more (2, 3, 4 or more) of the other pharmaceutically acceptable therapeutic agents can be used simultaneously with the compounds of the present invention to treat microsatellite instability (MSI) or DNA mismatch repair defective (dMMR) tumors.

[0173] chemotherapy drugs

[0174] Chemotherapeutic drugs include but are not limited to anastrozole, bicalutamide, bleomycin sulfate, busulfan, busulfan injection, capecitabine, N4-pentyloxycarbonyl-5-deoxy-5-fluorocytidine, carboplatin, carmustine, chlorambucil, cisplatin, cladribine, cyclophosphamide, cytarabine, cytosine arabinoside, cytarabine liposome injection, dacarbazine, actinomycin, daunorubicin hydrochloride, daunorubicin citrate, liposome injection, dexamethasone, docetaxel, doxorubicin hydrochloride, etoposide, fluorophosphate Darapine, 5-fluorouracil, flutamide, tezcitabine, gemcitabine, hydroxyurea, idarubicin, ifosfamide, irinotecan, L-asparaginase, leucovorin, melphalan, 6-mercaptopurine, methotrexate, mitoxantrone, gemtuzumab ozogamicin, paclitaxel, pentostatin, tamoxifen citrate, teniposide, 6-thioguanine, thiotepa, tirapazamine, topotecan hydrochloride for injection, vinblastine, vincristine, and vinorelbine; particularly 5-fluorouracil (5-FU) and irinotecan.

[0175] Immunotherapy drugs

[0176] Immunotherapy drugs include but are not limited to PD-1 inhibitors (including small molecules and antibody drugs), PD-L1 inhibitors (including small molecules and antibody drugs), CTLA-4 inhibitors, PD-1 / CTLA-4 dual antibodies, PD-1 / TIGIT dual antibodies, PD-L1 / 4-1BB dual antibodies, PD1 / VEGF-A dual antibodies, PD-1 / LAG-3 dual antibodies and other immunotherapy drugs.

[0177] Among them, PD-1 inhibitors include but are not limited to nivolumab, pembrolizumab, carrelizumab, toripalimab, sintilimab, tislelizumab, penampalimab, sepalizumab, serotonin, dostarlimab and PDR001.

[0178] PD-L1 inhibitors include, but are not limited to, durvalumab, atezolizumab, envoiriumab, and sugemalimab.

[0179] CTLA-4 inhibitors include, but are not limited to, ipilimumab.

[0180] PD-1 / CTLA-4 dual antibodies include but are not limited to candidizumab.

[0181] targeted drugs

[0182] Targeted drugs include but are not limited to VEGFR inhibitors (lenvatinib), NTRK inhibitors (entrectinib), BRAFV600 inhibitors (dabrafenib), MEK inhibitors (trametinib), EGFR monoclonal antibodies (cetuximab), PARP inhibitors (olaparib, niraparib, fluzoparib and pamiparib, etc.) and ATR inhibitors.

[0183] When using a pharmaceutical composition, a safe and effective amount of the compound of the present invention is administered to a mammal (e.g., a human) in need of treatment, wherein the dosage is a pharmaceutically effective dosage. For a 60 kg human, the daily dosage is generally 1-2000 mg, preferably 20-500 mg. Of course, the specific dosage will also take into account factors such as the route of administration and the patient's health condition, all of which are within the skill of a skilled physician.

[0184] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples where specific conditions are not specified are generally carried out under conventional conditions, such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or according to the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are weight percentages and parts by weight. The compounds of the present invention are all determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). The unit of NMR shift is 10 -6 The solvents for NMR measurements are deuterated dimethyl sulfoxide, deuterated chloroform, deuterated methanol, etc., and tetramethylsilane (TMS) is the internal standard.

[0185] The abbreviations of the present invention are defined as follows:

[0186] M: molar concentration, such as 1M hydrochloric acid means 1 mol / L hydrochloric acid solution

[0187] HATU: O-(7-azabenzotriazol-1-yl)-N,N,N,N-tetramethyluronium hexafluorophosphonate

[0188] T3P: 1-propylphosphoric anhydride

[0189] DIPEA: also written as DIEA, diisopropylethylamine, also known as N,N-diisopropylethylamine

[0190] EA: Ethyl acetate

[0191] TFA: trifluoroacetic acid

[0192] DMF: N,N-dimethylformamide

[0193] THF: Tetrahydrofuran

[0194] PE: Petroleum ether

[0195] LC-MS: Liquid chromatography-mass spectrometry

[0196] DMSO: dimethyl sulfoxide

[0197] CbzCl: benzyl chloroformate

[0198] Xantphos: 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene

[0199] TLC: Thin layer chromatography

[0200] IC 50 : Half-maximal inhibitory concentration refers to the concentration at which half of the maximum inhibitory effect is achieved.

[0201] Unless otherwise indicated, compounds exemplified herein are named and numbered using ChemBioDraw Ultra 14.0.

[0202] Preparative liquid chromatography separation conditions: preparative column: Agilent 10 Prep–C18 250*21.2 mm, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 45%~55% acetonitrile, 30 mL / min.

[0203] Intermediate 8b: 2-Bromo-N-(2-methoxy-4-(trifluoromethyl)phenyl)acetamide

[0204] 2-Methoxy-4-(trifluoromethyl)aniline (8b-1) (200 mg, 1.05 mmol) and 4-dimethylaminopyridine (128 mg, 1.05 mmol) were dissolved in dry dichloromethane (2 mL), cooled to about 0°C, and bromoacetyl bromide (252 mg, 1.25 mmol) was added dropwise. After the addition, the mixture was returned to room temperature and reacted for 18 h. The reaction solution was concentrated and the crude product was flash separated (PE:EA=20:1) to give compound 8b (320 mg) as a white solid.

[0205] MS (ESI) m / z 312 [M+H] + .

[0206] Intermediate 16a: 4-Hydroxy-1,5-dimethyl-2-oxo-1,2-dihydropyridine-3-carboxylic acid

[0207] The first step 5-methyl-2H-1,3-oxazine-2,6(3H)-dione 16a-2

[0208] Compound 16a-1 (5.6 g, 50 mmol) was dissolved in THF (10 mL) at room temperature, and TMSN3 (6.0 g, 52 mmol) was added. The mixture was stirred at 70°C under N2 protection for 2 hours. After completion of the reaction, as monitored by TLC, the mixture was cooled to room temperature, and the organic phase was concentrated under reduced pressure to afford crude product 16a-2 (1.2 g).

[0209] MS (ESI) m / z 128.10 [M+H] + .

[0210] Step 2 3,5-dimethyl-2H-1,3-oxazine-2,6(3H)-dione 16a-3

[0211] Compound 16a-2 (1.0 g, 7.8 mmol) was dissolved in acetonitrile (10 mL), and KCO (1.3 g, 9.45 mmol) was added, followed by MeI (1.88 g, 13.4 mmol). The mixture was reacted at 70°C for 16 h. After completion of the reaction, as monitored by TLC, ethyl acetate (30 mL) was added and the insoluble solid was removed by filtration. The organic phase was concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 2 / 1) to obtain compound 16a-3 (320 mg).

[0212] MS (ESI) m / z 142.10 [M+H] + .

[0213] Step 3 4-hydroxy-1,5-dimethyl-2-oxo-1,2-dihydropyridine-3-carboxylic acid ethyl ester 16a-4

[0214] Dissolve diethyl malonate (312 mg, 1.95 mmol) in anhydrous THF (20 mL). Add NaH (78 mg, 60% in mineral oil) under ice-cooling and allow to react at room temperature for 20 min. Add dropwise a THF solution of compound 16a-3 (250 mg of 16a-3 dissolved in 1 mL of THF). Allow to react at room temperature for 2 hours. After completion of the reaction, as monitored by TLC, add saturated NH4Cl solution (20 mL). Extract with ethyl acetate (30 mL x 3). Combine the organic phases and concentrate under reduced pressure to obtain the crude product. The crude product is purified by silica gel column chromatography (PE / EtOAc = 2 / 1) to obtain compound 16a-4 (130 mg).

[0215] MS (ESI) m / z 212.10 [M+H] + .

[0216] Step 4: 4-Hydroxy-1,5-dimethyl-2-oxo-1,2-dihydropyridine-3-carboxylic acid 16a

[0217] Compound 16a-4 (130 mg, 0.61 mmol) was dissolved in HBr (2 mL, 33 wt.% in HOAc), and then reacted at 70° C. for 3 h. The mixture was concentrated under reduced pressure to obtain a crude product, Compound 16a (130 mg).

[0218] MS (ESI) m / z 182.20 [MH] -.

[0219] Intermediate 23a 6-(Benzyloxy)-7-methylpyrazolo[1,5-a]pyrimidine-5-carboxylic acid

[0220] Step 1: Diethyl 2-(benzyloxy)-3-hydroxyfumarate 23a-1

[0221] Ethyl 2-(benzyloxy)acetate (19a) (70 g, 360.66 mmol) and diethyl oxalate (63.25 g, 432.79 mmol) were dissolved in tetrahydrofuran (700 mL). The reaction solution was cooled to -78°C and slowly added dropwise with 2.0 M lithium diisopropylamide (216.4 ml, 432.79 mmol). The reaction was allowed to react at -78°C for 1 h. The reaction was quenched by the addition of saturated aqueous ammonium chloride (300 mL). The pH was adjusted to 6 with 85% aqueous phosphoric acid. The mixture was extracted with ethyl acetate (300 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 23a-1 (106 g) as an orange-yellow oil.

[0222] MS (ESI) m / z 295.3 [M+H] + .

[0223] Step 2: Ethyl 6-(benzyloxy)-7-hydroxypyrazolo[1,5-a]pyrimidine-5-carboxylate 23a-2

[0224] 2H-pyrazol-3-amine (3 g, 36.123 mmol) was dissolved in DMF (30 mL), and compound 23a-1 (15.94 g, 54.184 mmol) and cesium carbonate (17.65 g, 54.164 mmol) were added sequentially. The mixture was reacted at 110°C for 18 h. After cooling to room temperature, ice water (30 mL) was added, and 2.0 M hydrochloric acid solution was added dropwise to adjust the pH to 5-6. The mixture was extracted with dichloromethane (30 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA = 1:2) to obtain compound 23a-2 (1 g) as a reddish-brown oil.

[0225] MS (ESI) m / z 314.3 [M+H] + .

[0226] Step 3 6-(Benzyloxy)-7-chloropyrazolo[1,5-a]pyrimidine-5-carboxylic acid ethyl ester 23a-3

[0227] Compound 23a-2 and N,N-diethylaniline (47.69 mg, 0.3198 mmol) were dissolved in phosphorus oxychloride (1.47 g, 9.5814 mmol) and reacted at 120°C for 3 h. The mixture was concentrated under reduced pressure, and the crude product was separated by column chromatography (PE:EA = 4:1) to give compound 23a-3 (420 mg) as a light yellow solid.

[0228] MS (ESI) m / z 332.3 [M+H] + .

[0229] Step 4: 6-(Benzyloxy)-7-methylpyrazolo[1,5-a]pyrimidine-5-carboxylic acid ethyl ester 23a-4

[0230] Compound 23a-3 was dissolved in DMSO (5 mL), and tricyclohexylphosphine (71.23 mg, 0.254 mmol), tris(dibenzylideneacetone)palladium (116.30 mg, 0.127 mmol) and cesium carbonate (826.93 mg, 2.538 mmol) were added in sequence. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 130°C for 3 h. The mixture was cooled to room temperature, diluted with water (5 mL), and extracted with ethyl acetate (5 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography (PE:EA=1:1) to give compound 23a-4 (200 mg) as a light yellow solid.

[0231] MS (ESI) m / z 312.0 [M+H] + .

[0232] Step 5: 6-(Benzyloxy)-7-methylpyrazolo[1,5-a]pyrimidine-5-carboxylic acid 23a

[0233] Compound 23a-4 (200 mg, 0.643 mol) was dissolved in tetrahydrofuran (2 mL) and water (2 mL), and lithium hydroxide (46.19 mg, 1.928 mmol) was added. The mixture was allowed to react at room temperature for 2 h. The reaction solution was diluted with water (2 mL), and the pH was adjusted to 6 with 2.0 M dilute hydrochloric acid. The mixture was extracted with dichloromethane (5 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford compound 23a (160 mg) as a light yellow solid.

[0234] MS (ESI) m / z 284 [M+H] + .

[0235] Intermediate 28a 4,4,5,5-tetramethyl-2-(4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-1,3,2-dioxaborolane

[0236] First step 4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]28a-2

[0237] The synthesis of (1-(thiophen-2-yl)cyclopropyl)methanol (28a-1) was described in patent WO2022 / 206706. Compound (28a-1) (1.3 g, 8.44 mmol) was dissolved in dry acetonitrile (100 mL). Paraformaldehyde (329 mg, 10.96 mmol) and indium trichloride (187 mg, 0.84 mmol) were added sequentially. Under nitrogen, the mixture was heated to 70°C for 2 h. After cooling to room temperature, the reaction solution was concentrated under reduced pressure. The crude product was purified by column chromatography (petroleum ether) to afford compound 28a-2 (600 mg) as a colorless oil.

[0238] Step 2: 4,4,5,5-tetramethyl-2-(4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-1,3,2-dioxaborolane 28a

[0239] Under nitrogen, compound 28a-2 (600 mg, 3.61 mmol) was dissolved in dry tetrahydrofuran (10 mL), cooled to -70 ° C, and 2.5 M n-butyllithium n-hexane solution (1.52 mL, 3.8 mmol) was added dropwise. The temperature was returned to room temperature for 1 h. The reaction solution was cooled to -70 ° C, and 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (740 mg, 3.98 mmol) was added dropwise. The temperature was raised to room temperature for 2 h. The reaction was quenched by adding saturated ammonium chloride solution (30 mL). The mixture was extracted with ethyl acetate (30 mL x 3), and the organic phases were combined and concentrated. The crude product was separated and purified by column chromatography (PE / EA = 3 / 2) to give compound 28a (520 mg) as a yellow oil.

[0240] MS (ESI) m / z 293 [M+H] + .

[0241] Intermediate 29a 2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane

[0242] The first step 2-(thiophen-3-yl)ethan-1,1-d2-1-ol 29a-2

[0243] LiAlD4 (1.0 g, 25.61 mmol) was added to a single-necked flask. THF (20 mL) was added at 0°C and stirred for 10 min. Then, 20 mL of a THF solution of compound 29a-1 (4.0 g, 25.61 mmol) was slowly added and the mixture was allowed to react at 0°C for 1 h. The mixture was warmed to room temperature and stirred for 1 h. The reaction solution was cooled to 0°C, and potassium sodium tartrate was added until no gas evolution was observed. The mixture was stirred for 1 h. Petroleum ether (50 mL) was added, stirred, and filtered. The filtrate was concentrated to afford compound 29a-2 (2.5 g) as a yellow oil.

[0244] 1 H NMR (400MHz, CDCl3) δ7.29(dd,J=4.9,3.0Hz,1H),7.05(d,J=2.2Hz,1H),6.98(dd,J=4.9,0.9Hz,1H),2.89(s,2H).

[0245] Step 2 4,7-dihydro-5H-thieno[2,3-c]pyran-5,5-d2 29a-3

[0246] Compound 29a-2 (2.5 g, 19.20 mmol) was dissolved in acetonitrile, and paraformaldehyde (724 mg, 24.96 mmol) and InCl3 (425 mg, 1.92 mmol) were added. The atmosphere was replaced with N2 three times, and the reaction was carried out at 85°C for 2.5 h. The reaction solution was cooled to room temperature, and water (200 mL) was added. The mixture was extracted with a mixed solvent of PE / TBME (PE / TBME = 1 / 1, 70 mL x 2). The organic phases were combined and concentrated under reduced pressure. The crude product was purified by column chromatography (petroleum ether / DCM = 10 / 1 to 2 / 1) to obtain compound 29a-3 (1.7 g) as a yellow oil.

[0247] 1 H NMR (400MHz, CDCl3) δ7.14 (d, J = 5.0 Hz, 1H), 6.82 (d, J = 5.0 Hz, 1H), 4.82 (s, 2H), 2.74 (s, 2H).

[0248] Step 3 2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane 29a

[0249] Compound 29a-3 (1.7 g, 11.95 mmol) was dissolved in THF (20 mL), cooled to -70°C, and n-BuLi (2.5 M in THF, 5.2 mL, 13.15 mmol) was added. Stir at -70°C for 15 min, then warm to room temperature. After approximately 1 h, the reaction solution turned from yellow-green to brown-green. The reaction solution was cooled to -70°C, and 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (2.45 g, 13.15 mmol) was added. The reaction solution was slowly warmed to room temperature and allowed to react overnight. The reaction solution was cooled to -70°C, quenched by the addition of saturated aqueous NH4Cl (20 mL), and extracted with ethyl acetate (25 mL x 2). The organic phases were combined, concentrated under reduced pressure, and purified by column chromatography to yield compound 29a (2.1 g) as a yellow solid.

[0250] 1 H NMR (400MHz, CDCl3) δ7.36 (s, 1H), 4.85 (t, J = 1.4Hz, 2H), 4.16-4.07 (m, 1H), 3.94-3.84 (m, 1H), 2.75 (s, 2H), 1.33 (s, 12H).

[0251] Intermediate 30a: trimethyl((3-methyloxetan-3-yl)ethynyl)silane

[0252] The first step is 3-methyloxetane-3-carboxaldehyde 30a-2

[0253] Pyridinium chlorochromate (10.1 g, 25.61 mmol) and silica gel (100-200 mesh, 10.0 g) were dissolved in DCM (100 mL). Compound 30a-1 (4.0 g, 39.16 mmol) was added at 0°C and allowed to react overnight at room temperature. The reaction solution was concentrated under reduced pressure and purified by column chromatography to afford compound 30a-2 (2.8 g) as a colorless oil.

[0254] 1 H NMR (400MHz, CDCl3) δ9.95 (s, 1H), 4.87 (d, J = 6.4Hz, 2H), 4.50 (d, J = 6.4Hz, 2H), 1.48 (s, 3H).

[0255] Step 2 3-(2,2-dibromovinyl)-3-methyloxetane 30a-3

[0256] Triphenylphosphine (10.48 g, 39.95 mmol) was dissolved in DCM (50.0 mL) and cooled to 0°C. CBr (6.62 g, 19.98 mmol) was added and allowed to react at 0°C for 0.5 h. Compound 30a-2 (1.0 g, 9.99 mmol) was then added and the mixture was allowed to warm to room temperature for 1 h. After completion of the reaction, petroleum ether (150 mL) was added and stirred for 0.5 h. A large amount of solid precipitated, which was filtered. The filtrate was concentrated under reduced pressure, and the crude product was purified by column chromatography to afford compound 30a-3 (1.7 g) as a light yellow oil.

[0257] 1 H NMR (400MHz, CDCl3) δ6.69 (s, 1H), 4.81 (d, J = 5.8Hz, 2H), 4.38 (d, J = 6.0Hz, 2H), 1.60 (s, 3H).

[0258] Step 3: Trimethyl((3-methyloxetan-3-yl)ethynyl)silane 30a

[0259] Compound 30a-3 (1.0 g, 3.91 mmol) was dissolved in anhydrous THF (15 mL). The reaction solution was cooled to -70°C, and n-BuLi (2.5 M in THF, 3.6 mL, 8.99 mmol) was added. The mixture was allowed to react at -70°C for 1 h. TMSCl (1.49 g, 13.68 mmol) was then added at -70°C. The mixture was allowed to react at -70°C for 0.5 h, and then the temperature was raised to room temperature for 0.5 h. The reaction solution was concentrated under reduced pressure, and petroleum ether (15 mL) was added. The mixture was filtered through silica gel, and the filtrate was concentrated to afford compound 30a (590 mg) as a light yellow oil.

[0260] 1 H NMR (400MHz, CDCl3) δ4.82 (d, J = 5.3Hz, 2H), 4.37 (d, J = 5.4Hz, 2H), 1.61 (s, 3H), 0.18-0.15 (s, 9H).

[0261] Intermediate 37a 4,4,5,5-tetramethyl-2-(5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-1,3,2-dioxaborolane

[0262] The first step 5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]37a-2

[0263] The synthesis of compound 37a-1 is based on patent PCT / CN / 2022 / 083485. Compound 37a-1 (1.9 g, 12.33 mmol), paraformaldehyde (0.4 g, 13.33 mmol), and indium trichloride (0.13 g, 0.58 mmol) were dissolved in acetonitrile (20 mL) and purged with nitrogen. The mixture was reacted at 45°C for 0.5 hour, then heated to 65°C for 0.5 hour, and then heated to 85°C for 2 hours. The mixture was cooled to room temperature and diluted with water (20 mL). The mixture was extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 37a-2 (0.8 g) as an oil.

[0264] 1 H NMR (400MHz, CDCl3) δ7.11 (dd, J=5.1, 0.9Hz, 1H), 6.48 (d, J=5.1Hz, 1H), 4.92 (s, 2H), 3.70 (s, 2H), 0.99–0.94 (m, 2H), 0.93–0.87 (m, 2H).

[0265] Step 2: 4,4,5,5-Tetramethyl-2-(5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-1,3,2-dioxaborolane 37a

[0266] Compound 37a-2 (0.8 g, 4.81 mmol) was dissolved in THF (10 mL), the reaction solution was cooled to -78 ° C, and 2.5 M n-butyllithium n-hexane solution (1.9 mL, 5.0 mmol) was added dropwise. The reaction was allowed to react for 10 minutes, and the temperature was slowly raised to room temperature for 0.5 hour. The temperature was again lowered to -78 ° C, and isopropyl alcohol pinacol borate (0.98 g, 5.26 mmol) was added dropwise. The reaction was allowed to react for 10 minutes. Water (20 mL) was added at low temperature to quench the reaction. The reaction was extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 37a (0.5 g) as a white solid.

[0267] MS (ESI) m / z 293.1 [M+H] +

[0268] Intermediate 41a tert-Butyl 2-(methoxymethyl)piperazine-1-carboxylate

[0269] The first step is 1-tert-butyl-2-hydroxymethylpiperazine-1,4-dicarboxylic acid-4-benzyl ester 41a-2

[0270] Compound 41a-1 (5.0 g, 23.12 mmol) was dissolved in THF (100 mL) and water (20 mL). NaHCO₃ (7.77 g, 92.47 mmol) and CbzCl (5.9 g, 34.68 mmol) were added at 0°C. The mixture was allowed to warm to room temperature and allowed to react for 12 h. The mixture was concentrated under reduced pressure, and EtOAc (150 mL) and water (50 mL) were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography to afford compound 41a-2 (9.7 g) as a yellow oil.

[0271] MS (ESI) m / z 251.1 [M-Boc+H] +

[0272] Step 2 1-tert-Butyl-2-(methoxymethyl)piperazine-1,4-dicarboxylic acid 4-benzyl ester 41a-3

[0273] Compound 41a-2 (9.2 g, 26.25 mmol) was dissolved in THF (100 mL), cooled to -30°C, and NaH (2.1 g, 52.51 mmol, 60% wt) was added. The mixture was stirred for 10 min, and CH₃I (11.18 g, 5.0 mL, 78.76 mmol) was added. The mixture was allowed to warm to room temperature and react overnight. The reaction mixture was cooled to 0°C, and saturated aqueous NH₄Cl (20 mL) was added until no significant gas evolution occurred. Water (300 mL) was added, and the mixture was extracted with EtOAc (100 mL x 2). The organic phases were combined and concentrated under reduced pressure. The crude product was purified by column chromatography to afford compound 41a-3 (5.5 g) as a colorless oil.

[0274] MS (ESI) m / z 387.1 [M+Na] +

[0275] Step 3 tert-Butyl 2-(methoxymethyl)piperazine-1-carboxylate 41a

[0276] Compound 41a-3 (5.5 g, 15.09 mmol) was dissolved in THF (100 mL) and 10% Pd / C (1.0 g, 844.37 μmol) was added. The mixture was deoxygenated three times by displacement. The reaction mixture was allowed to react overnight at room temperature. The reaction mixture was filtered through Celite and washed with MeOH (200 mL). The filtrate was concentrated under reduced pressure to afford compound 41a (3.5 g) as a yellow-brown oil.

[0277] MS (ESI) m / z 231.1 [M+H] +

[0278] Intermediate 42a 4-Hydroxy-1,5,6-trimethyl-2-oxo-1,2-dihydropyridine-3-carboxylic acid

[0279] The synthesis of compound 42a-1 was based on patent WO2007007903. Compound 42a-1 (210 mg, 0.93 mmol) was dissolved in HBr (2 mL, 33 wt.% in HOAc) and reacted at 65°C for 3 h. The reaction solution was concentrated under reduced pressure to afford compound 42a (190 mg), which was used directly in the next step.

[0280] MS (ESI) m / z 198.2 [M+H] + .

[0281] Intermediate 43a 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-bromo-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide

[0282] The first step 2-(2-bromo-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 43a-1

[0283] Compound 1g (2.26g, 3.4mmol) was dissolved in ethyl acetate (20mL), and hydrogen chloride-ethyl acetate (20mL, 80mmol, 4M in EtOAc) was added. The mixture was allowed to react at room temperature for 6 hours. After completion of the reaction, the reaction mixture was concentrated under reduced pressure to obtain crude product 43a-1 (2.2g). The crude product was used directly in the next step.

[0284] MS (ESI) m / z 562.1 [M+H] +

[0285] Step 2: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-bromo-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 43a

[0286] Crude product 43a-1 (2.2 g, 3.4 mmol) was dissolved in 1,4-dioxane (30 mL), compound 1e (0.91 g, 3.75 mmol), triethylamine (1.02 g, 10.2 mmol), and 1-propylphosphonic acid cyclic anhydride (1.6 g, 5.1 mmol, 50% in EtOAc) and allowed to react at room temperature for 3 hours. After completion of the reaction, saturated sodium bicarbonate solution was added, and the mixture was extracted with ethyl acetate. The organic phase was concentrated to obtain the crude product, which was then purified by slurrying with ethyl acetate to afford compound 43a (1.8 g).

[0287] MS (ESI) m / z 788.1 [M+H] +

[0288] Intermediate 43b 6-methyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine

[0289] The first step is 6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 43b-2

[0290] 4,5,6,7-Tetrahydrothiophene[2,3-c]pyridine hydrochloride (43b-1) (1.37 g, 10 mmol) was dissolved in tetrahydrofuran (40 mL), and potassium carbonate (2.07 g, 15 mmol) and methyl p-toluenesulfonate (1.48 g, 8 mmol) were added. The mixture was reacted at room temperature for 16 hours. After completion of the reaction, water (30 mL) was added, and the mixture was extracted with ethyl acetate (30 mL). The organic phase was concentrated under reduced pressure to obtain the crude product. The crude product was separated and purified by column chromatography (PE / EA = 1:3) to obtain compound 43b-2 (0.25 g).

[0291] MS (ESI) m / z 154.0 [M+H] + .

[0292] Step 2: 6-Methyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 43b

[0293] Compound 43b-2 (250 mg, 1.63 mmol) was dissolved in tetrahydrofuran (10 mL). n-Butyl lithium (0.72 mL, 1.8 mmol, 2.5 M in THF) was added dropwise at -78°C. After reacting at -78°C for 1 hour, isopropyl pinacol borate (334 mg, 1.8 mmol) was added and the reaction continued at -78°C. TLC monitored the reaction completion. Saturated ammonium chloride solution (30 mL) was added, and the mixture was extracted with ethyl acetate. The organic phase was concentrated to obtain the crude product, which was then separated and purified by column chromatography (DCM / MeOH = 10:1) to afford compound 43b (220 mg).

[0294] MS (ESI) m / z 280.1 [M+H] + .

[0295] Intermediate 44a 5-methyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine

[0296] The first step is 5-methyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 44a-2

[0297] 4,5,6,7-Tetrahydrothieno[3,2-c]pyridine hydrochloride (44a-1) (500 mg, 3.60 mmol) was dissolved in dry methanol (10 mL). Acetic acid (2 mL) and 37% formaldehyde (3.17 g, 36.00 mmol) were added and stirred at room temperature for 1 h. Sodium triacetoxyborohydride (3.05 g, 7.2 mmol) was then added portionwise. The reaction was stirred at room temperature for 48 h. The reaction was complete by TLC. The reaction solution was concentrated under reduced pressure and the crude product was separated by column chromatography (DCM / MeOH = 20:1) to afford compound 44a-2 (350 mg) as a light yellow oil.

[0298] MS (ESI) m / z 154.0 [M+H] + .

[0299] Step 2: 5-Methyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 44a

[0300] Compound 44a-2 (350 mg, 2.29 mmol) was dissolved in dry tetrahydrofuran (10 mL) and cooled to -70°C. 2.5 M n-butyllithium (0.92 ml, 2.29 mmol) was slowly added dropwise. The reaction was maintained at -70°C for 30 minutes. Isopropyl pinacol borate (469.07 mg, 2.52 mmol) was then added dropwise. After addition, the temperature was naturally raised to room temperature and the reaction was allowed to proceed for 2 hours. After completion of the reaction by TLC, saturated aqueous ammonium chloride was added for quenching (5 mL). The mixture was extracted with ethyl acetate (10 mL x 3). The organic phase was dried over anhydrous sodium sulfate, and the crude product was separated by column chromatography (DCM / MeOH = 10:1) to afford compound 44a (180 mg) as a pale yellow solid.

[0301] MS (ESI) m / z 280.1 [M+H] + .

[0302] Example 1 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0303] Step 1: 4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 1c

[0304] The synthesis of intermediate 1a is described in patent WO2022249060A1, and the synthesis of intermediate 1b is described in patent WO2022268230A1. Compound 1b (115 mg, 0.68 mmol) and 1a (133 mg, 0.23 mmol) were dissolved in dry dimethyl sulfoxide (2 mL). Potassium acetate (124 mg, 1.36 mmol) was added, and the temperature was raised to 120°C for 18 h. Compound 1b (115 mg, 0.68 mmol) and potassium acetate (124 mg, 1.36 mmol) were then added, and the reaction continued at 120°C for 18 h. The mixture was cooled to room temperature, diluted with water (20 mL), and extracted with ethyl acetate (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and the crude product separated on a silica gel plate (PE:EA = 2:1) to afford compound 1c (57 mg) as a white solid.

[0305] MS (ESI) m / z 715.2 [M+H] + .

[0306] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 1d

[0307] Dissolve compound 1c (57 mg, 0.08 mmol) in dry dichloromethane (1 mL), add trifluoroacetic acid (1 mL), and stir at room temperature for 1 h. Concentrate the solution and use the crude product directly in the next reaction.

[0308] MS (ESI) m / z 615 [M+H] + .

[0309] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 1f

[0310] Compound 1d was dissolved in DMF (2 mL), and intermediate 1e (synthesis reference patent WO2022249060A1) (25.3 mg, 0.1 mmol) and DIEA (103 mg, 0.8 mmol) were added sequentially. The reaction solution was cooled to approximately 0°C, and a 50% ethyl acetate solution of T3P (76 mg, 0.12 mmol) was added dropwise. After addition, the mixture was returned to room temperature and stirred for 3 h. Water (20 mL) was added to quench the reaction, and the mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, concentrated, and the crude product was separated and purified on a silica gel plate (PE:EA = 1:3) to afford compound 1f (20 mg) as a white solid.

[0311] MS (ESI) m / z 841.2 [M+H] + .

[0312] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 1

[0313] Trifluoroacetic acid (2 mL) was added to compound 1f (20 mg, 0.042 mmol), and the temperature was raised to 50°C for 18 h. The reaction solution was concentrated, and the crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) to give compound 1 (3.0 mg) as a white solid.

[0314] MS (ESI) m / z 751.2 [M+H] + .

[0315] 1 H NMR(400MHz,DMSO-d6)δ10.34(br s,1H),8.53(s,1H),8.03(d,J=8.6Hz,1H),7.96(s,1H),7.72(d,J=8.7Hz,1H),5.21(s,2H),4.51(d,J=11.3Hz,1H),3.50(m, 7H),3.23(m,2H),2.94(m,3H),2.77(d,J=9.8Hz,1H),2.60(d,J=9.9Hz,1H),2.43(s,3H),2.17(m,4H),1.16(t,J=7.4Hz,3H).

[0316] Example 2 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0317] Step 1: 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 2b

[0318] Compound 1a (0.10 g, 0.151 mmol) was dissolved in dioxane (5 mL) and water (1 mL). Compound 2a (synthesis reference patent WO2019023468 A1) (0.06 g, 0.20 mmol), potassium phosphate (0.11 g, 0.500 mmol), and Pd(dppf)Cl2 (11.0 mg, 0.015 mmol) were added under N2 protection and reacted at 85°C for 1 h. After completion of the reaction, as monitored by TLC, the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined and concentrated under reduced pressure to obtain the crude product, compound 2b (67 mg).

[0319] MS (ESI) m / z 720.10 [M+H] + .

[0320] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-7-oxo-6-(piperazin-1-yl)-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 2c

[0321] Compound 2b (67.0 mg, 0.093 mmol) was dissolved in dichloromethane (3 mL), trifluoroacetic acid (3 mL) was added, and the mixture was allowed to react at room temperature for 1 h. After the reaction was completed as monitored by TLC, the reaction solution was concentrated under reduced pressure to give the crude product Compound 2c (67.0 mg).

[0322] MS (ESI) m / z 620.10 [M+H] + .

[0323] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 2d

[0324] Compound 2c (0.067 g, 0.153 mmol) was dissolved in THF (10 mL), and compound 1e (49.0 mg, 0.200 mmol), triethylamine (170 mg, 1.53 mmol), and T3P (204 mg, 0.3 mmol, 50% in EA) were added. The reaction mixture was allowed to react at room temperature for 1 h. After completion of the reaction, as monitored by TLC, the mixture was extracted with ethyl acetate (50 mL × 3). The organic phases were combined and concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 2d (0.050 g).

[0325] MS (ESI) m / z 846.10 [M+H] + .

[0326] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 2

[0327] Compound 2d (0.050 g, 0.061 mmol) was dissolved in trifluoroacetic acid (3 mL) and reacted at 50°C for 3 h. After completion of the reaction, monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain the crude product. The crude product was purified by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) and lyophilized to obtain compound 2 (17.0 mg).

[0328] MS (ESI) m / z 756.20 [M+H] + .

[0329] 1 H NMR (400MHz, CDCl3) δ11.79(s,1H),9.10(s,1H),8.58(s,1H),8.48(d,J=8.8Hz,1H),7 .63(d,J=2.0Hz,1H),7.53(d,J=8.8Hz,1H),6.94(s,1H),5.13(d,J=15.6Hz,1H),5.03 (d,J=15.6Hz,1H),3.94-3.75(m,5H),3.49-3.41(m,1H),3.23-3.06(m,5H),2.87-2.6 4(m,6H),2.56(s,3H),2.45-2.28(m,3H),2.04(q,J=7.5Hz,2H),1.33(t,J=7.5Hz,3H).

[0330] Example 3 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-cyclopropyl-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0331] Step 1: 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-cyclopropyl-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 3a

[0332] Compound 1a (0.100 g, 0.151 mmol) was dissolved in DMF (5 mL), and cyclopropyltributyltin (66.2 mg, 0.20 mmol) and tetrakistriphenylphosphine palladium (8.90 mg, 0.0151 mmol) were added. The mixture was reacted at 135°C for 8 h under N₂ protection. After completion of the reaction, the reaction solution was concentrated under reduced pressure and monitored by TLC to obtain the crude product. The crude product was then purified by column chromatography (PE / EA = 1 / 1) to afford compound 3a (80.0 mg).

[0333] MS (ESI) m / z 624.10 [M+H] + .

[0334] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-cyclopropyl-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 3b

[0335] Compound 3a (80.0 mg, 0.128 mmol) was dissolved in dichloromethane (3 mL), and trifluoroacetic acid (3 mL) was added. The mixture was allowed to react at room temperature for 1 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to give the crude product, compound 3b (80.0 mg).

[0336] MS (ESI) m / z 524.10 [M+H] + .

[0337] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-cyclopropyl-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 3c

[0338] Compound 3b (0.080 g, 0.153 mmol) was dissolved in THF (10 mL), and compound 1e (49.0 mg, 0.200 mmol), triethylamine (170 mg, 1.53 mmol), and T3P (204 mg, 0.3 mmol, 50% in EA) were added. The reaction mixture was allowed to react at room temperature for 1 h. After completion of the reaction, as monitored by TLC, the mixture was extracted with ethyl acetate (50 mL x 3). The combined organic phases were concentrated under reduced pressure to afford the crude product, which was then purified by column chromatography (PE / EtOAc = 1 / 1) to afford compound 3c (0.056 g).

[0339] MS (ESI) m / z 750.10 [M+H] + .

[0340] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-cyclopropyl-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 3

[0341] Compound 3c (0.056 g, 0.075 mmol) was dissolved in trifluoroacetic acid (3 mL) and reacted at 50°C for 3 h. After completion of the reaction, monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain the crude product. The crude product was purified by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) and lyophilized to obtain compound 3 (17.0 mg).

[0342] MS (ESI) m / z 660.20 [M+H] + .

[0343] 1 H NMR (400 MHz, CDCl3) δ 11.79 (br s,1H),8.96(s,1H),8.58(s,1H),8.47(d,J=8.6Hz,1H),7.66(d,J=1.9Hz,1H) ,7.54(d,J=8.8Hz,1H),5.64(d,J=12.8Hz,1H),5.06(s,2H),4.78(d,J=12.8Hz ,1H),3.83-3.78(m,2H),3.52-3.46(m,1H),3.22-2.98(m,3H),2.86-2.70(m, 2H), 2.56 (s, 3H), 2.15-2.08 (m, 1H), 1.31 (t, J = 7.5Hz, 3H), 1.13-1.05 (m, 4H).

[0344] Example 4 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0345] Step 1: (S)-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,4a,5,6,7,9-hexahydrothieno[2,3-f]indolizine 4c

[0346] The synthesis of compound 4b was based on Heterocycles 50 (1999) 1, 445-452. Compound 4b (323 mg, 1.8 mmol) was dissolved in tetrahydrofuran (5 mL). n-Butyl lithium (0.86 mL, 2.15 mmol, 2.5 M in THF) was added dropwise at -78°C. After reacting at -78°C for 1 hour, isopropyl pinacol borate (400 mg, 2.5 mmol) was added and the reaction continued at -78°C. After completion of the reaction, saturated ammonium chloride solution was added, and the mixture was extracted with ethyl acetate. The organic phase was concentrated to obtain the crude product, which was then purified by column chromatography to yield compound 4c (430 mg).

[0347] MS (ESI) m / z 306.2 [M+H] + .

[0348] Step 2 (S)-2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(4,4a,5,6,7,9-hexahydrothieno[2,3-f]indolizin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide 4d

[0349] Compound 4b (200 mg, 0.65 mmol) was dissolved in 1,4-dioxane (5 mL) and water (1 mL), and intermediate 43a (430 mg, 0.54 mmol), potassium phosphate (343 mg, 1.54 mmol), and Pd(dppf)Cl2 (44 mg, 0.05 mmol) were added. The atmosphere was purged with nitrogen three times, and the mixture was reacted at 100°C for 3 hours. After cooling, the reaction solution was diluted with ethyl acetate and washed three times with saturated brine. The organic phase was concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography to afford compound 4d (160 mg).

[0350] MS (ESI) m / z 887.4 [M+H] + .

[0351] Step 3 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 4

[0352] Compound 4d (160 mg, 0.18 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 3 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain a crude product. The crude product was purified by preparative HPLC to afford compound 4 (41.2 mg).

[0353] MS (ESI) m / z 797.3 [M+H] + .

[0354] 1 H NMR (400MHz, DMSO-d6) δ10.42(s,1H),8.45(s,1H),8.32(s,2H)8.05(d,J=8.6Hz,1H),7.96(s,1H),7. 73-7.69(m,1H),7.46(s,1H),5.34(s,2H),4.53(d,J=11.76Hz,1H),4.13(d,J=15.4Hz,1H),3.49(s,1H ),3.32(s,1H),3.18–3.11(m,2H),3.05-2.95(m,3H),2.79-2.90(m,2H),2.60-2.71(m,1H),2.29-2.45 (m,5H),2.26–2.17(m,1H),2.05–1.97(m,1H),1.73-1.85(m,2H)1.49–1.39(m,1H),1.15-1.23(m,3H).

[0355] Example 5 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(5-cyclopropyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0356] Step 1: 5-cyclopropyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 5d

[0357] Compound 44a-1 (500 mg, 3.59 mmol) was dissolved in methanol (5 mL), and (1-ethoxycyclopropyloxy)trimethylsilane (2.96 g, 21.54 mmol), acetic acid (1.7 g, 35.9 mmol), and sodium cyanoborohydride (802 mg, 16.15 mmol) were added. The mixture was reacted at 70°C overnight. The reaction solution was cooled to room temperature, and saturated sodium bicarbonate solution was added. The mixture was extracted with ethyl acetate, and the organic phases were combined and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EA = 4 / 1) to obtain 5d (110 mg) as a yellow oil.

[0358] MS (ESI) m / z 180 [M+H] +

[0359] Step 2 5-Cyclopropyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 5e

[0360] Compound 5d (110 mg, 0.36 mmol) was dissolved in tetrahydrofuran (5 mL), and n-butyllithium (0.28 mL, 2.5 M, 0.39 mmol) was added at -78°C. The mixture was reacted under nitrogen for 0.5 h. 2-Isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (125 mg, 0.39 mmol) was added dropwise to the reaction mixture, and the reaction was allowed to proceed at room temperature for 2 h. After completion of the reaction, saturated ammonium chloride solution was added to the reaction mixture at 0°C. The mixture was extracted with ethyl acetate, and the organic phases were combined and concentrated under reduced pressure to afford the crude product, Compound 5e (170 mg), as a yellow oil.

[0361] MS (ESI) m / z 306 [M+H] +

[0362] Step 3 2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(5-cyclopropyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide 5f

[0363] Compound 5e (170 mg, 0.55 mmol) was dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Compound 43a (249 mg, 0.44 mmol), sodium carbonate (104 mg, 1.65 mmol), and 1,1'-bis(diphenylphosphino)ferrocene (27 mg, 0.055 mmol) were added. The mixture was reacted at 100°C for 4 h under nitrogen. After completion of the reaction, water was added to the reaction mixture, and the mixture was extracted with ethyl acetate. The organic phases were combined and concentrated under reduced pressure to obtain the crude product. The crude product was separated by silica gel column chromatography to obtain compound 5f (130 mg) as a yellow oil.

[0364] MS (ESI) m / z 888 [M+H] +

[0365] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(5-cyclopropyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 5f

[0366] Compound 5f (130 mg, 0.15 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain a crude product, which was purified by preparative HPLC to obtain compound 5 (41.2 mg).

[0367] MS (ESI) m / z 797 [M+H] +

[0368] 1 H NMR(400MHz,DMSO-d6)δ10.40(s,1H),10.25(s,1H),8.58(s,1H),8.06(d,J=8.6Hz,1H),7 .97(d,J=2.0Hz,1H),7.72(dd,J=8.7,2.0Hz,1H),7.53(s,1H),5.34(s,2H),4.53(d,J=12. 5Hz,1H),3.54-3.46(m,5H),3.34(s,2H),3.29-3.22(m,2H),3.03-2.97(m,4H),2.84-2.8 0(m,2H),2.68-2.62(m,1H),2.44(s,3H),1.19(t,J=7.6Hz,3H),0.85(m,2H),0.52(m,2H).

[0369] Example 6 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0370] Step 1: Methyl 2-(methylamino)cyclopent-1-ene-1-carboxylate 6b

[0371] Compound 6a (0.71 g, 5 mmol) was dissolved in THF (10 mL), and MeNH2-THF (12.5 mL, 25 mmol, 2 M in THF) was added. The mixture was stirred under N2 protection for 2 hours. After completion of the reaction, as monitored by TLC, ethyl acetate (20 mL) was added, and the organic phase was washed with water (10 mL) and brine (10 mL). The organic phase was concentrated under reduced pressure to yield crude product 6b (560 mg).

[0372] MS (ESI) m / z 156.10 [M+H] + .

[0373] Step 2: Methyl 2-(3-ethoxy-N-methyl-3-oxopropionylamino)cyclopent-1-ene-1-carboxylate 6c

[0374] Compound 6b (560 mg, 3.6 mmol) was dissolved in toluene (10 mL), and NaOAc (590 mg, 7.2 mmol) was added, followed by ethyl 3-chloro-3-oxopropanoate (538 mg, 3.6 mmol). The reaction was allowed to proceed at room temperature for 6 h. After completion of the reaction, as monitored by TLC, ethyl acetate (30 mL) was added and the mixture was washed with water (10 mL) and then brine (10 mL). The organic phase was concentrated under reduced pressure to yield the crude product, Compound 6c (460 mg).

[0375] MS (ESI) m / z 270.10 [M+H] + .

[0376] Step 3: 4-Hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridine-3-carboxylic acid methyl ester 6d

[0377] Compound 6c (460 mg, 0.153 mmol) was dissolved in anhydrous methanol (30 mL), and MeONa solution (0.5 mL, 30 wt% in MeOH) was added dropwise. The mixture was allowed to react at room temperature for 1 h. After completion of the reaction, the reaction solution was evaporated to approximately 10 mL, and 30 mL of water was added. The pH was adjusted to approximately 7 with dilute HCl in an ice bath. The mixture was extracted with dichloromethane (30 mL x 3). The organic phases were combined and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 2 / 1) to afford compound 6d (390 mg).

[0378] MS (ESI) m / z 224.10 [M+H] + .

[0379] Step 4: 4-Hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridine-3-carboxylic acid 6e

[0380] Compound 6d (38 mg, 0.17 mmol) was dissolved in anhydrous THF (1 mL). Potassium tert-butoxide (1 mL, 1 M in THF) was added at room temperature, and the mixture was reacted at 80°C for 3 h. The pH was adjusted to approximately 7 with dilute HCl in an ice bath. The mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined and concentrated under reduced pressure to afford crude product 6e (24 mg).

[0381] MS (ESI) m / z 210.10 [M+H] + .

[0382] Step 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 6

[0383] Compound 6e (24 mg, 0.11 mmol) was dissolved in DMF (2 mL), and EDCI (22 mg, 0.11 mmol), HOBt (15 mg, 0.11 mmol), and DIEA (36 mg, 0.28 mmol) were added at room temperature. Compound 6f (synthesis reference patent WO2022249060A1) was then added, and the mixture was allowed to react at room temperature for 16 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain the crude product, which was then separated and purified by silica gel column chromatography (DCM / MeOH = 10:1) to obtain compound 6 (1.4 mg).

[0384] MS (ESI) m / z 757.20 [M+H]+ .

[0385] 1 H NMR (400MHz, CDCl3) δ9.00(s,1H),8.49(d,J=8.8Hz,1H),7.64(s,1H),7.54(d,J=8.8Hz,1H),7.36(s,1H),6.97(s,1H),5.09(s,2H),4.37(m,2 H),3.92(m,2H),3.74-3.65(m,4H),3.43(s,3H),3.16(m,2H),2.95-2. 84(m,4H),2.73(m,4H),2.16(m,2H),2.03(m,2H),1.32(t,J=7.6Hz,3H)

[0386] Example 7 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(4,6-dimethyl-3-oxo-3,4-dihydropyrazine-2-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0387] Step 1: 3-Chloro-1,5-methylpyrazin-2(1H)-one 7b

[0388] Compound 7a (100 mg, 0.69 mmol) and potassium carbonate (191 mg, 1.38 mmol) were dissolved in DMF (5 mL). To the reaction solution was added iodomethane (118 mg, 0.83 mmol) and stirred at room temperature for 4 h. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The mixture was extracted three times with ethyl acetate / H2O. The organic layers were combined, washed with saturated brine, and dried over anhydrous sodium sulfate to obtain crude compound 7b (100 mg) as a yellow oil, which was used directly in the next step.

[0389] MS (ESI) m / z 159.2 [M+H] + .

[0390] Step 2: 4,6-Dimethyl-3-oxo-3,4-dihydropyrazine-2-carboxylic acid phenyl ester 7c

[0391] Compound 7b (100 mg, 0.63 mmol), phenyl formate (231 mg, 1.89 mmol), Xantphos (36 mg, 0.063 mmol), palladium acetate (7 mg, 0.031 mmol), and TEA (159 mg, 1.57 mmol) were dissolved in anhydrous acetonitrile (2 mL). The atmosphere was replaced with nitrogen three times and the mixture was refluxed at 80°C for 4 h. The reaction solution was cooled to room temperature and filtered through celite. The solvent was removed by concentration under reduced pressure and the mixture was extracted three times with DCM / H2O. The organic layers were combined, washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography (PE / EA = 1 / 1) to afford compound 7c (40 mg) as an off-white solid.

[0392] MS (ESI) m / z 245.1 [M+H] + .

[0393] 1 H NMR (400MHz, CDCl3) δ7.42(m,3H),7.30-7.22(m,3H),3.61(s,3H),2.39(s,3H).

[0394] Step 3 4,6-dimethyl-3-oxo-3,4-dihydropyrazine-2-carboxylic acid 7d

[0395] Compound 7c (25 mg, 0.10 mmol) was dissolved in THF / MeOH / H₂O (1 / 1 / 1). Lithium hydroxide (7 mg, 0.30 mmol) was added and allowed to react at room temperature for 1 h. After completion of the reaction, the pH was adjusted to 6 with 1N hydrochloric acid (1.0 M). The solvent was then concentrated under reduced pressure to afford crude compound 7d (20 mg) as a yellow solid.

[0396] MS (ESI) m / z 169.1 [M+H] + .

[0397] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-6-(4-(4,6-dimethyl-3-oxo-3,4-dihydropyrazine-2-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 7

[0398] Compound 7d (20 mg, 0.12 mmol) and compound 6f (68 mg, 0.12 mmol) were dissolved in DMF (2 mL). TEA (48 mg, 0.48 mmol) was added, and a T3P / EA solution (57 mg, 0.18 mmol) was added to the mixture. The mixture was allowed to react at room temperature for 2 h. After completion of the reaction, the mixture was extracted three times with EA / H2O. The organic layers were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) to afford compound 7 (3 mg) as a white solid.

[0399] MS (ESI) m / z 716.2 [M+H] + .

[0400] 1 H NMR (400MHz, CDCl3) δ8.99(s,1H),8.48(d,J=8.7Hz,1H),7.64(s,1H),7.53(d,J=8.5Hz,1H ),7.00(s,1H),6.96(s,1H),5.38-5.32(m,1H),5.11(d,J=15.2Hz,1H),5.06(d,J=15.2Hz,1 H),4.74-4.67(m,1H),4.36(m,2H),3.92(m,2H),3.76(m,2H),3.53(s,3H),3.48(m,1H),3. 18-3.12(m,3H),2.78(m,1H),2.72(m,2H),2.63(m,1H),2.30(s,3H),1.30(t,J=7.6Hz,3H).

[0401] Example 8 2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methoxy-4-(trifluoromethyl)phenyl)acetamide

[0402] Step 1: 4-(5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 8a

[0403] Compound 5c (200 mg, 0.47 mmol) and compound 2a (185 mg, 0.70 mmol) were dissolved in 1,4-dioxane (5 mL) and water (1.5 mL), and Pd(dppf)Cl2 (19 mg, 0.023 mmol) and sodium carbonate (99.2 mg, 0.94 mmol) were added. The atmosphere was replaced with nitrogen three times, and the temperature was raised to 100°C for 16 h. The mixture was cooled to room temperature, diluted with water (20 mL), and extracted with dichloromethane (30 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated. The crude product was flash separated (DCM / MeOH = 30 / 1) to give compound 8a (120 mg) as a brown solid.

[0404] Step 2: tert-Butyl 4-(5-ethyl-4-(2-(2-methoxy-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 8c

[0405] Compound 8a (120 mg, 0.25 mmol) and compound 8b (85 mg, 0.27 mmol) were dissolved in dry acetonitrile (2 mL), and N,N-diisopropylethylamine (96 mg, 0.74 mmol) was added. The mixture was reacted at room temperature for 18 h. The reaction solution was concentrated and separated on a silica gel plate (PE:EA=2:1) ​​to give compound 8c (40 mg) as a white solid.

[0406] MS (ESI) m / z 716.4 [M+H] + .

[0407] Step 3: 2-(5-ethyl-7-oxo-6-(piperazin-1-yl)-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methoxy-4-(trifluoromethyl)phenyl)acetamide 8d

[0408] Compound 8c (40 mg, 0.056 mmol) was dissolved in dry dichloromethane (1 mL). Trifluoroacetic acid (1 mL) was added and the mixture was stirred at room temperature for 1 h. After concentration, saturated aqueous sodium bicarbonate (10 mL) was added to the residue, and the mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product 8d was directly used in the next reaction.

[0409] MS (ESI) m / z 616 [M+H] + .

[0410] Step 4: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methoxy-4-(trifluoromethyl)phenyl)acetamide 8e

[0411] Compound 8d was dissolved in DMF (2 mL), and compound 1e (17.7 mg, 0.073 mmol) and N,N-diisopropylethylamine (72 mg, 0.56 mmol) were added sequentially. The reaction solution was cooled to approximately 0°C, and a 50% ethyl acetate solution of T3P (53.4 mg, 0.084 mmol) was added dropwise. After addition, the mixture was returned to room temperature and stirred for 3 h. Water (20 mL) was added to quench the mixture, and the mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was separated and purified on a silica gel plate (PE:EA = 1:3) to afford compound 8e (19 mg) as a white solid.

[0412] MS (ESI) m / z 842.5 [M+H] + .

[0413] Step 5: 2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methoxy-4-(trifluoromethyl)phenyl)acetamide 8

[0414] Trifluoroacetic acid (1 mL) was added to compound 8e (19 mg, 0.023 mmol), and the temperature was raised to 50°C for 18 h. The reaction solution was concentrated, and the crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) to give compound 8 (10 mg) as a white solid.

[0415] MS (ESI) m / z 752.4 [M+H] + .

[0416] 1H NMR (400MHz, DMSO-d6) δ10.11(s,1H),8.55(s,1H),8.20(d,J=8.4Hz,1H),7.36(s,1H),7.27( d,J=8.4Hz,1H),6.71(s,1H),5.30(s,2H),4.52(d,J=12.1Hz,1H),3.98(s,3H),3.73(t,J=6.8 Hz,2H),3.55-3.38(m,4H),3.00-2.88(m,3H),2.81(m,1H),2.66-2.54(m,2H),2.44(s,3H),2. 27(m,2H),2.17(t,J=7.2Hz,1H),1.90-1.85(m,2H),1.76-1.60(m,4H),1.16(t,J=7.4Hz,3H).

[0417] Example 9 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0418] Step 1: (S)-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizine 9b

[0419] The synthesis of compound 9a is based on Heterocycles 50 (1999) 1, 445-452. Compound 9a (330 mg, 1.8 mmol) was dissolved in tetrahydrofuran (10 mL) and n-butyllithium (0.88 mL, 2.21 mmol, 2.5 M in THF) was added dropwise at -78°C. After reacting at -78°C for 1 hour, isopropyl pinacol borate (411 mg, 2.21 mmol) was added and the reaction continued at -78°C. TLC monitored the reaction completion, and saturated ammonium chloride solution (30 mL) was added, followed by extraction with ethyl acetate. The organic phase was concentrated to obtain the crude product, which was then separated by column chromatography (DCM / MeOH = 10:1) to afford compound 9b (500 mg).

[0420] MS (ESI) m / z 306.2 [M+H] + .

[0421] Step 2 (S)-2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide 9c

[0422] Compound 9b (200 mg, 0.65 mmol) was dissolved in 1,4-dioxane (7 mL) and water (1 mL), and intermediate 43a (469 mg, 0.59 mmol), potassium phosphate (375 mg, 1.71 mmol), and Pd(dppf)Cl2 (48 mg, 0.05 mmol) were added. The atmosphere was purged with nitrogen three times, and the reaction was continued at 100°C for 3 hours. After cooling, the reaction solution was diluted with ethyl acetate (40 mL) and washed three times with saturated brine (15 mL). The organic phase was concentrated under reduced pressure to obtain the crude product. The crude product was separated by column chromatography (DCM / MeOH = 10:1) to obtain compound 9c (240 mg).

[0423] MS (ESI) m / z 887.5 [M+H] + .

[0424] Step 3 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(4,6,7,8,8a,9-hexahydrothieno[3,2-f]indolizin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 9

[0425] Compound 9c (240 mg, 0.27 mmol) was dissolved in trifluoroacetic acid (4 mL) and reacted at 50°C for 3 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain a crude product, which was then purified by preparative HPLC to obtain compound 9 (45.9 mg).

[0426] MS (ESI) m / z 797.3 [M+H] + .

[0427] 1H NMR (400MHz, CDCl3) δ9.11 (s, 1H), 8.60 (s, 1H), 8.48 (d, J = 8.6Hz, 1H), 7.61 (s, 1H), 7.5 7-7.50(m,2H),5.71–5.65(m,1H),5.15(s,2H),4.82-4.79(m,1H),4.23(d,J=13.4Hz,1 H),3.85–3.79(m,2H),3.50-3.40(m,2H),3.21-3.08(m,4H),2.67-2.93(m,4H),2.58(s ,3H),2.54-2.40(m,1H),2.21–2.16(m,1H),2.06–1.90-2.11(m,4H)1.33-1.36(m,3H).

[0428] Example 10 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyanocyclohex-1-en-1-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0429] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4-cyanocyclohex-1-en-1-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 10b

[0430] The synthesis of intermediate 10a was based on patent WO2015144001 A1. Compounds 1a (200 mg, 0.30 mmol) and 10a (105 mg, 0.45 mmol) were dissolved in 1,4-dioxane (4 mL) and water (0.4 mL). 1,1-Bis(diphenylphosphino)ferrocenepalladium dichloride dichloromethane complex (24.5 mg, 0.03 mmol) and sodium carbonate (95.4 mg, 0.90 mmol) were added. The atmosphere was purged with nitrogen three times, and the temperature was raised to 100°C for 16 h. The mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (10 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated on a silica gel column (PE:EA = 1:4) to afford compound 10b (47 mg) as a tan solid.

[0431] MS (ESI) m / z 689 [M+H] + .

[0432] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyanocyclohex-1-en-1-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 10c

[0433] Compound 10b (47 mg, 0.068 mmol) was dissolved in dry dichloromethane (1 mL), and trifluoroacetic acid (1 mL) was added. The mixture was stirred at room temperature for 1 h. The mixture was concentrated under reduced pressure, and the residue was dissolved in dichloromethane (5 mL). The reaction was concentrated under reduced pressure at least three times. The crude product was directly used in the next reaction to obtain compound 10c (54 mg) as a yellow-brown solid.

[0434] MS (ESI) m / z 589 [M+H] + .

[0435] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4-cyanocyclohex-1-en-1-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 10d

[0436] Compound 10c (54 mg, crude) was dissolved in DMF (2 mL), followed by the addition of compound 1e (26.85 mg, 0.110 mmol) and triethylamine (46.55 mg, 0.46 mmol). The reaction mixture was cooled to approximately 0°C, and a 50% ethyl acetate solution of 1-propylphosphoric acid cyclic anhydride (43.9 mg, 0.138 mmol) was added dropwise. After addition, the mixture was allowed to return to room temperature and allowed to react for 2 h. Water (10 mL) was added to quench the mixture, followed by extraction with dichloromethane (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified on a silica gel plate (PE / EA = 1 / 2) to afford compound 10d (20 mg) as a pale yellow solid.

[0437] MS (ESI) m / z 815 [M+H] + .

[0438] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyanocyclohex-1-en-1-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 10

[0439] Compound 10d (20 mg, 0.025 mmol) was dissolved in trifluoroacetic acid (1 mL), heated to 50°C for 3 h, and the reaction solution was concentrated under reduced pressure and purified on a silica gel plate (DCM / MeOH = 30 / 1) to give compound 10 (12.2 mg) as a light yellow solid.

[0440] MS (ESI) m / z 725 [M+H] + .

[0441] 1 H NMR (400MHz, DMSO-d6) δ10.35(s,1H),10.23(s,1H),8.57(s,1H),8.05(d,J=8.0Hz, 1H),7.96(s,1H),7.71(d,J=8.0Hz,1H),6.79(s,1H),5.31(s,2H),5.08-4.98(m,1H ),4.52(d,J=12.8Hz,1H),3.54-3.43(m,4H),3.16(m,1H),2.99(m,2H),2.81(d,J=1 1.2Hz,1H),2.67-2.54(m,3H),2.44(s,3H),2.02-1.93(m,4H),1.26(t,J=7.5Hz,3H)

[0442] Example 11 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-(difluoromethylene)piperidin-1-yl)-5-ethyl-6-(4-(4-hydroxy-1-methyl)-2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0443] Compound 6e (38 mg, 0.18 mmol) was dissolved in DMF (2 mL), and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (34.8 mg, 0.18 mmol), 1-hydroxybenzotriazole (24.5 mg, 0.18 mmol) and N,N-diisopropylethylamine (58.6 mg, 0.45 mmol) were added sequentially. The mixture was stirred at room temperature for 10 minutes, and compound 1d (55.9 mg, 0.09 mmol) was added. The reaction was carried out at room temperature for 48 hours, and water (20 mL) was added to quench the mixture. The mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) to give compound 11 (10.9 mg) as a light yellow solid.

[0444] MS (ESI) m / z 806.4 [M+H] + .

[0445] 1 H NMR (400MHz, DMSO-d6) δ10.33(s,1H),8.03(d,J=8.6Hz,1H),7.96(s,1H),7.72(d,J=8.3Hz,1H),5.21(s,2H) ,3.59-3.39(m,8H),3.30(s,3H),2.92(m,6H),2.69(m,4H),2.18(m,4H),2.00(m,2H),1.16(t,J=7.4Hz,3H).

[0446] Example 12 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(1,1-difluorospiro[2.5]oct-5-en-6-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0447] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(1,1-difluorospiro[2.5]oct-5-en-6-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 12b

[0448] The synthesis of compound 12a is based on patent WO2021141041A1. Compound 1a (150 mg, 0.23 mmol) was dissolved in dioxane (4 mL) and water (0.5 mL). Compound 12a (124 mg, 0.46 mmol), sodium carbonate (48 mg, 0.45 mmol), and Pd(dppf)Cl2 (17 mg, 0.023 mmol) were added. The mixture was protected by nitrogen and reacted at 100°C for 1 h. After completion of the reaction, the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined and concentrated under reduced pressure to yield the crude product, compound 12b (80 mg).

[0449] MS (ESI) m / z 671.1 [M–C4H7] +

[0450] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(1,1-difluorospiro[2.5]oct-5-en-6-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 12c

[0451] Compound 12b (80 mg, 0.11 mmol) was dissolved in dichloromethane (3 mL), and trifluoroacetic acid (3 mL) was added. The mixture was allowed to react at room temperature for 1 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to give the crude product, Compound 12c (80 mg).

[0452] MS (ESI) m / z 625.10 [M+H] + .

[0453] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(1,1-difluorospiro[2.5]oct-5-en-6-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 12d

[0454] Compound 12c (80 mg, 0.128 mmol) was dissolved in THF (10 mL), and compound 1e (38 mg, 0.154 mmol), triethylamine (130 mg, 1.28 mmol), and T3P (61 mg, 0.192 mmol, 50% in EA) were added. The reaction mixture was allowed to react at room temperature for 1 h. After completion of the reaction, as monitored by TLC, the mixture was extracted with ethyl acetate (20 mL x 3). The combined organic phases were concentrated under reduced pressure to afford the crude product, which was then purified by column chromatography (PE / EtOAc = 1 / 1) to afford compound 12d (70 mg).

[0455] MS (ESI) m / z 852.20 [M+H] + .

[0456] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(1,1-difluorospiro[2.5]oct-5-en-6-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 12

[0457] Compound 12d (50 mg, 0.059 mmol) was dissolved in trifluoroacetic acid (3 mL) and reacted at 50°C for 3 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain the crude product, which was then purified by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) and lyophilized to afford compound 12 as a white solid (35 mg).

[0458] MS (ESI) m / z 762.20 [M+H] + .

[0459] 1 H NMR (400MHz, CDCl3) δ11.78(s,1H),9.07(s,1H),8.58(s,1H),8.49(d,J=8.7Hz,1H),7 .64(s,1H),7.53(d,J=8.7Hz,1H),7.01(s,1H),5.65(m,1H),5.10(s,2H),4.78(m,1H) ,3.80(m,2H),3.49(m,1H),3.25-3.00(m,3H),2.90-2.65(m,4H),2.56(s,3H),2.54-2 .44(m,1H),2.30-2.18(m,1H),1.82(m,2H),1.32(t,J=7.6Hz,3H),1.19-1.08(m,1H), 0.90-0.80(m,1H).

[0460] Example 13 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(6-cyclopropyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0461] Step 1: 6-cyclopropyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 13a

[0462] Compound 43b-1 (0.5 g, 2.85 mmol) was dissolved in ethanol (5 mL), and acetic acid (1.7 g, 28.5 mmol) and 1-ethoxy-1-trimethylsilyloxycyclopropane (3.0 g, 17.1 mmol) were added. Sodium cyanoborocyanide (0.89 g, 12.8 mmol) was then added portionwise. The mixture was reacted at 60°C for 16 hours. After completion of the reaction, water (20 mL) was added, the mixture was extracted with ethyl acetate, and the organic phase was concentrated under reduced pressure to obtain the crude product. The crude product was isolated and purified by column chromatography (PE / EA = 5:1) to obtain compound 13a (0.47 g).

[0463] MS (ESI) m / z 180.0 [M+H] + .

[0464] Step 2: 6-Cyclopropyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 13b

[0465] Compound 13a (470 mg, 2.62 mmol) was dissolved in tetrahydrofuran (15 mL), and n-butyllithium (1.26 mL, 3.15 mmol, 2.5 M in THF) was added dropwise at -78°C. After reacting at -78°C for 1 hour, isopropyl pinacol borate (585 mg, 3.15 mmol) was added and the reaction continued at -78°C. After completion of the reaction, saturated aqueous ammonium chloride (30 mL) was added, and the mixture was extracted with ethyl acetate. The organic phase was concentrated to obtain the crude product, which was then separated by column chromatography to afford compound 13b (620 mg).

[0466] MS (ESI) m / z 306.2 [M+H] + .

[0467] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6-cyclopropyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 13c

[0468] Compound 13b (200 mg, 0.65 mmol) was dissolved in 1,4-dioxane (6 mL) and water (1 mL). Intermediate 43a (426 mg, 0.54 mmol), potassium phosphate (343 mg, 1.62 mmol), and Pd(dppf)Cl2 (48 mg, 0.06 mmol) were added. The atmosphere was purged with nitrogen three times and the reaction was carried out at 100°C for 3 hours. After cooling, the reaction solution was diluted with ethyl acetate (30 mL) and washed three times with saturated brine (15 mL). The organic phase was concentrated under reduced pressure to obtain the crude product. The crude product was separated by column chromatography to obtain compound 13c (400 mg).

[0469] MS (ESI) m / z 887.3 [M+H] + .

[0470] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(6-cyclopropyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide KTH051173

[0471] Compound 13c (400 mg, 0.45 mmol) was dissolved in TFA (5 mL) and reacted at 50°C for 3 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain a crude product. The crude product was purified by preparative HPLC to afford compound 13 (32.1 mg).

[0472] MS (ESI) m / z 797.2 [M+H] + .

[0473] 1 H NMR (400MHz, DMSO-d6) δ10.38(s,1H),10.22(br.1H),8.58(s,1H),8.05(d,J=8.7Hz,1H),7. 97(s,1H),7.72(d,J=7.3Hz,1H),7.45(s,1H),5.33(s,2H),4.53(d,J=11.3Hz,1H),3.79(s, 2H),3.60–3.42(m,3H),3.29-3.19(m,2H),3.05-2.93(m,3H),2.93-2.77(m,3H),2.66(m,3H ),2.44(s,3H),1.19-1.14(m,3H),0.50-0.48(d,J=6.4Hz,2H),0.43-0.36(d,J=6.4Hz,2H).

[0474] Example 14 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(3-(methylsulfonyl)azetidin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0475] Step 1: 4-(4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-2-(3-(methylsulfonyl)azetidin-1-yl)-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 14a

[0476] 1a (200 mg, 0.302 mmol), AcOK (178 mg, 1.81 mmol), and 3-methylsulfonyl-azetidine hydrochloride (156 mg, 0.906 mmol) were dissolved in DMSO (4 mL) and reacted at 120°C for 16 h. After the reaction was complete, H2O (30 mL x 3) was added and extracted with ethyl acetate (30 mL). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product, which was purified by silica gel column chromatography to afford compound 14a (100 mg) as a yellow solid.

[0477] MS (ESI) m / z 717.3 [M+H] + .

[0478] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(3-(methylsulfonyl)azetidin-1-yl)-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 14b

[0479] Compound 14a (100 mg, 0.139 mmol) was dissolved in TFA (2 mL) and reacted at room temperature for 1 h. The mixture was concentrated under reduced pressure, saturated sodium carbonate solution was added, and the mixture was extracted with dichloromethane (30 mL x 3). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product, which was purified by silica gel column chromatography to obtain compound 14b (41 mg).

[0480] MS (ESI) m / z 617.2 [M+H] + .

[0481] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(3-(methylsulfonyl)azetidin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 14c

[0482] Compound 14b (41 mg, 0.0655 mmol), compound 1e (21 mg, 0.0865 mmol), and DIPEA (43 mg, 0.333 mmol) were dissolved in DMF (2 mL). T3P (50% wt, 64 mg, 0.0998 mmol) was added and allowed to react at room temperature for 16 h. After completion of the reaction, H2O was added and the mixture was extracted with dichloromethane. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product, which was then purified by silica gel column chromatography to obtain compound 14c (45 mg).

[0483] MS (ESI) m / z 843.4 [M+H] + .

[0484] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(3-(methylsulfonyl)azetidin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 14

[0485] Compound 14c (45 mg, 0.0534 mmol) was dissolved in TFA (2 mL) and reacted at 50°C for 3 h. After concentration under reduced pressure, the crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 20%-30% acetonitrile, 50 mL / min) and purified by reverse phase chromatography to obtain compound 14 (3 mg) as a white solid.

[0486] MS (ESI) m / z 753.3 [M+H] + .

[0487] 1H NMR(400MHz,DMSO-d6)δ10.79–9.80(m,1H),8.50(s,1H),7.99(d,J=8.5Hz,1H),7 .89(s,1H),7.64(d,J=7.2Hz,1H),5.16(s,2H),4.52–4.33(m,2H),4.21(t,J=8.7 Hz,2H),4.10(dd,J=9.2,5.6Hz,2H),3.42(d,J=11.0Hz,3H),2.96(s,3H),2.92–2.50(m,6H),2.37(s,3H),1.09(t,J=7.4Hz,3H).

[0488] Example 15 2-(2-(3-oxabicyclo[3.1.0]hexan-6-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide

[0489] Step 1: 4-(2-(3-oxabicyclo[3.1.0]hexan-6-yl)-4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 15b

[0490] The synthesis of intermediate 15a is described in patent WO2020072695A1. Compound 1a (530 mg, 0.8 mmol), compound 15a (264 mg, 1.2 mmol), Pd(dppf)Cl2 (59 mg, 0.08 mmol), and potassium phosphate (424 mg, 2.0 mmol) were added to a sealed tube. The atmosphere was replaced with argon three times, and 1,4-dioxane (10 mL) and H2O (2 mL) were added. The tube was sealed and heated to 80°C with stirring for 16 h. Ethyl acetate (100 mL) and saturated brine (10 mL) were added, and the mixture was extracted. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 2) to obtain compound 15b (31 mg) as a pale yellow solid.

[0491] MS (ESI) m / z 666 [M+H] + .

[0492] Step 2: 2-(2-(3-oxabicyclo[3.1.0]hexan-6-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 15c

[0493] Compound 15b (31 mg, 0.047 mmol) was dissolved in DCM (2 mL) and TFA (2 mL) was added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, the reaction solution was concentrated, ethyl acetate (30 mL) was added, and saturated sodium bicarbonate solution was adjusted to pH 8 for extraction. The organic phase was washed with saturated brine and concentrated under reduced pressure to afford crude compound 15c (33 mg), which was used directly in the next step without purification.

[0494] MS (ESI) m / z 566 [M+H] + .

[0495] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(3-oxabicyclo[3.1.0]hexan-6-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 15d

[0496] Compound 15c (33 mg, 0.047 mmol) was dissolved in THF (2 mL), and compound 1e (14 mg, 0.056 mmol), DIPEA (30 mg, 0.23 mmol), and T3P (44 mg, 0.07 mmol, 50% in EA) were added. The reaction mixture was allowed to react at room temperature for 3 h. After completion of the reaction, as monitored by TLC, saturated sodium bicarbonate solution was added to quench the reaction. The mixture was extracted with ethyl acetate (50 mL), and the organic phase was washed three times with saturated brine and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (EtOAc) to afford compound 15d (21 mg) as a pale yellow solid.

[0497] MS (ESI) m / z 792 [M+H] + .

[0498] Step 4: 2-(2-(3-oxabicyclo[3.1.0]hexane-6-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 15

[0499] Compound 15d (21 mg, 0.026 mmol) was dissolved in trifluoroacetic acid (2 mL), and the reaction mixture was reacted at 50°C for 3 h. After completion of the reaction as monitored by TLC, the reaction mixture was concentrated under reduced pressure, ethyl acetate (30 mL) was added, and saturated sodium bicarbonate was adjusted to alkalinity. Extraction was performed, and the organic phase was washed once with saturated brine and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography (MeOH / EtOAc = 1 / 10) to obtain compound 15 (11 mg).

[0500] MS (ESI) m / z 702 [M+H] + .

[0501] Example 16 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-1,5-dimethyl-2-oxo-1,2-dihydropyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0502] Compound 16a (64 mg, 0.35 mmol) was dissolved in DMF (3 mL). EDCI-HCl (86 mg, 0.45 mmol), HOBt (60 mg, 0.45 mmol), and DIEA (117 mg, 0.9 mmol) were added at room temperature. Compound 6f was then added, and the mixture was allowed to react at 50°C for 5 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain the crude product, which was then purified by silica gel column chromatography (DCM / MeOH = 10:1) to afford compound 16 (31.2 mg).

[0503] MS (ESI) m / z 731.40 [M+H] + .

[0504] 1 H NMR (400MHz, CDCl3) δ9.03 (s, 1H), 8.48 (d, J = 8.6Hz, 1H), 7.63 (s, 1H), 7.54 (d,J=8.6Hz,1H),7.14(s,1H),6.95(s,1H),5.12(s,2H),4.36(m,2H),4.18 (m,1H),3.91(t,J=5.6Hz,2H),3.66(m,2H),3.46(s,3H),3.34(m,2H),3.20 -3.10(m,2H),2.71(m,4H),2.04(s,3H),2.00(m,1H),1.30(t,J=7.6Hz,3H)

[0505] Example 17 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(6-ethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0506] Step 1: 6-ethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 17a

[0507] Dissolve 4,5,6,7-tetrahydrothieno[2,3-c]pyridine hydrochloride (43b-1) (2 g, 11.38 mmol) in dry tetrahydrofuran (40 mL). Add potassium carbonate (4.7 g, 34.15 mmol). Cool to 0°C, then add ethyl 4-methylbenzenesulfonate (2.3 g, 11.38 mmol) dropwise. Return to room temperature and react for 18 hours. After completion, concentrate under reduced pressure. The crude product is separated by column chromatography (PE:EA = 2:1) to afford compound 17a (460 mg) as a light yellow oil.

[0508] MS (ESI) m / z 168.0 [M+H] + .

[0509] Step 2: 2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-6-ethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 17b

[0510] Compound 17a (200 mg, 1.20 mmol) was dissolved in dry tetrahydrofuran (2 mL), cooled to -70°C, and 2.5 M n-butyllithium (0.7 ml, 1.80 mmol) was slowly added dropwise. Stirring was continued at -70°C for 30 min. Isopropyl pinacol borate (334.1 mg, 1.80 mmol) was added dropwise at -70°C. After addition, the mixture was allowed to warm to room temperature and allowed to react for 2 h. After completion, the reaction was quenched with aqueous solution and extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to yield the crude product, compound 17b (230 mg), as a yellow-brown oil.

[0511] MS (ESI) m / z 294.2 [M+H] + .

[0512] Step 3: 2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(6-ethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide 17c

[0513] Compound 17b (214 mg, 0.27 mmol) and 43a (100 mg, 0.34 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL), and 1,1-bis(diphenylphosphino)ferrocenepalladium dichloride dichloromethane complex (27.8 mg, 0.04 mmol) and sodium carbonate (108.4 mg, 1.02 mmol) were added. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 100 ° C for 4 h. The mixture was cooled to room temperature, diluted with water (5 mL), and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 17c (130 mg) as a reddish-brown solid.

[0514] MS (ESI) m / z 875.3 [M+H] + .

[0515] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(6-ethyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 17

[0516] Compound 17c (130 mg, 0.15 mmol) was dissolved in trifluoroacetic acid (2 mL), heated to 50°C for 2 h, and the reaction solution was concentrated under reduced pressure. The crude product was separated by preparative HPLC to obtain compound 17 (19.7 mg) as a light yellow solid.

[0517] MS (ESI) m / z 785.3 [M+H] + .

[0518] 1H NMR (400MHz, DMSO-d6) δ10.40(s,1H),10.24(s,1H),8.59(s,1H),8.06(d,J=8.0 Hz,1H),7.98(d,J=8.0Hz,1H),7.73(dd,J=8.0Hz,1H),7.61(s,1H),5.35(s,2H) ,4.53(d,J=8Hz,1H),3.51(d,J=8Hz,3H),3.35-3.42(m,4H),3.32~3.30(m,4H), 3.01(s,2H),2.84(d,J=8Hz,1H),2.65(s,1H),2.45(s,3H),1.20(t,J=8Hz,3H).

[0519] Example 18 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyclopropyloxyphenyl)-5-ethyl-6-(4-(4-hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopentadienyl[b]pyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0520] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4-cyclopropyloxyphenyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 18a

[0521] Compound 1a (100 mg, 0.15 mmol) was dissolved in 1,4-dioxane (4 mL), and 2-(4-cyclopropyloxyphenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (60 mg, 0.23 mmol), Pd(PPh3)4 (18 mg, 0.015 mmol), potassium phosphate (70 mg, 0.30 mmol), and water (1 mL) were added. Under nitrogen protection, the reaction was continued at 85°C for 2 h. After completion of the reaction, as monitored by TLC, water (20 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined and concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 18a (70 mg).

[0522] MS (ESI) m / z 660.1 [M–C4H7] + .

[0523] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyclopropyloxyphenyl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 18b

[0524] Compound 18a (70 mg, 0.098 mmol) was dissolved in dichloromethane (5 mL), and trifluoroacetic acid (1 mL) was added. The mixture was allowed to react at room temperature for 1 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to give compound 18b (60 mg).

[0525] MS (ESI) m / z 616.20 [M+H] + .

[0526] Step 3 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4-cyclopropyloxyphenyl)-5-ethyl-6-(4-(4-hydroxy-1-methyl-2-oxo-2,5,6,7-tetrahydro-1H-cyclopentadienyl[b]pyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 18

[0527] Compound 18b (60 mg, 0.097 mmol) was dissolved in DMF (3 mL), and compound 6e (24 mg, 0.117 mmol), DIPEA (75 mg, 0.58 mmol), EDCI-HCl (44 mg, 0.23 mmol), and HOBT (31 mg, 0.23 mmol) were added. The reaction mixture was reacted at 60°C for 8 h. After completion of the reaction, as monitored by TLC, extraction with ethyl acetate (20 mL x 3) was performed. The organic phases were combined and concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (DCM / MeOH = 10 / 1) to afford compound 18 (5 mg).

[0528] MS (ESI) m / z 807.20 [M+H] + .

[0529] 1H NMR (400MHz, CDCl3) δ9.19 (s, 1H), 8.49 (d, J = 8.6Hz, 1H), 8.20 (d, J = 8.5Hz, 2H) ,7.57(s,1H),7.51(d,J=8.0Hz,1H),7.11(d,J=8.6Hz,2H),5.35(m,1H),5.15(s ,2H),3.80(m,1H),3.66(m,4H),3.43(s,3H),3.35(m,2H),2.93(m,2H),2.85(m, 2H),2.73(m,2H),2.35(m,1H),2.15(m,2H),1.31(t,J=7.5Hz,3H),0.84(m,4H).

[0530] Example 19 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(6-hydroxypyrazolo)[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0531] Step 1: Sodium (Z)-2-(benzyloxy)-3-ethoxy-3-oxoprop-1-en-1-olate 19b

[0532] Ethyl (benzyloxy)acetate (19a) (5 g, 25.7 mmol) was dissolved in tetrahydrofuran (30 mL). The reaction mixture was cooled to 0°C and sodium hydride (1.13 g, 60% in mineral oil) was added. The mixture was allowed to return to room temperature and react for 1 hour. The reaction mixture was cooled to 0°C again and ethyl formate (2.14 g, 28.8 mmol) was added dropwise. After the addition was complete, the mixture was allowed to return to room temperature and react for 3 hours. The reaction mixture was concentrated under reduced pressure and the crude product was directly used in the next reaction. This resulted in compound 19b (6.3 g) as a gray solid.

[0533] MS (ESI) m / z 245 [M+H] + .

[0534] Step 2: Ethyl (Z)-2-(Benzyloxy)-3-ethoxyacrylate 19c

[0535] Compound 19b (6.3 g, 25.8 mmol) was dissolved in DMF (30 mL), and iodoethane (4 g, 25.8 mmol) was added. After complete addition, the temperature was raised to 35°C and stirred for 1 h. Water (30 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified on a silica gel plate (PE / EA = 2 / 1) to afford compound 19c (6 g) as a light yellow oil.

[0536] MS (ESI) m / z 251 [M+H] + .

[0537] Step 3 6-(Benzyloxy)pyrazolo[1,5-a]pyrimidin-5-ol 19d

[0538] Dissolve 2H-pyrazol-3-amine (1 g, 12.04 mmol) in DMF (30 mL), add compound 19c (4.52 g, 18.06 mmol) and cesium carbonate (5.88 g, 18.06 mmol), and heat to 110°C for 18 hours. After cooling to room temperature, add ice water (100 mL), and adjust the pH to approximately 5-6 with 2.0 M hydrochloric acid dropwise. Filter the mixture under reduced pressure, wash the filter cake with water (10 mL x 3), and dry to yield compound 19d (670 mg) as a yellow-brown solid.

[0539] MS (ESI) m / z 242 [M+H] + .

[0540] Step 4: 6-(Benzyloxy)-5-chloropyrazolo[1,5-a]pyrimidine 19e

[0541] Compound 19d (670 mg, 2.78 mmol) was dissolved in dry acetonitrile (7 mL), and phosphorus oxychloride (1.29 g, 8.34 mmol) was added. After complete addition, the temperature was raised to 35°C and stirred for 5 h. The mixture was concentrated under reduced pressure, and the crude product was purified on silica gel (PE / EA = 4 / 1) to afford compound 19e (200 mg) as a pale yellow solid.

[0542] MS (ESI) m / z 260 [M+H] + .

[0543] Step 5: 6-(Benzyloxy)pyrazolo[1,5-a]pyrimidine-5-carboxylic acid phenyl ester 19f

[0544] Compound 19e (200 mg, 0.78 mmol) was dissolved in acetonitrile (2 mL), and tri-tert-butylphosphine tetrafluoroborate (22.68 mg, 0.078 mmol), palladium acetate (8.76 mg, 0.039 mmol) and triethylamine (394.64 mg, 3.9 mmol) were added sequentially. The atmosphere was replaced with nitrogen three times, and the temperature was raised to 80°C for 16 h. The mixture was cooled to room temperature, diluted with water (4 mL), and extracted with ethyl acetate (5 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by silica gel column chromatography (PE / EA = 1 / 1) to give compound 19g (140 mg) as a white solid.

[0545] MS (ESI) m / z 345 [M+H] + .

[0546] Step 6: 6-(Benzyloxy)pyrazolo[1,5-a]pyrimidine-5-carboxylic acid 19h

[0547] Compound 19g (140 mg, 0.406 mol) was dissolved in tetrahydrofuran (1 mL) and water (1 mL). Lithium hydroxide (29.6 mg, 1.218 mmol) was added and stirred at room temperature for 2 h. The reaction solution was concentrated under reduced pressure, and the crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 5%-20% acetonitrile, 30 mL / min) to afford compound 19h (70 mg) as a white solid.

[0548] MS (ESI) m / z 270 [M+H] + .

[0549] Step 7: 2-(6-(4-(6-(benzyloxy)pyrazolo[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 19i

[0550] Compound 1e (50 mg, 0.088 mmol) was dissolved in DMF (2 mL), and compound 19h (28.61 mg, 0.106 mmol) and N,N-diisopropylethylamine (44.5 mg, 0.44 mmol) were added. The reaction solution was cooled to approximately 0°C, and a 50% ethyl acetate solution of 1-propylphosphonic acid cyclic anhydride (84.6 mg, 0.133 mmol) was added dropwise. After addition, the mixture was allowed to return to room temperature and react for 2 h. Water (5 mL) was added to quench the reaction, and the mixture was extracted with dichloromethane (5 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (PE:EA = 1:2) to afford compound 19i (30 mg) as a white solid.

[0551] MS (ESI) m / z 817 [M+H] + .

[0552] Step 8: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(6-hydroxypyrazolo)[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 19

[0553] Compound 19i (30 mg, 0.037 mmol) was dissolved in trifluoroacetic acid (1 mL), heated to 50°C for 18 h, and the reaction solution was concentrated under reduced pressure. The crude product was separated by HPLC (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 30% to 50% acetonitrile, 30 mL / min) to give compound 19 (3.8 mg) as a white solid.

[0554] MS (ESI) m / z 727 [M+H] + .

[0555] 1 H NMR (400MHz, DMSO-d6) δ10.39(s,1H),8.34(s,1H),8.04(d,J=8.6Hz,1H),7.9 4(d,J=8.6Hz,1H),7.90(s,1H),7.69(d,J=8.6Hz,1H),6.82(s,1H),6.57(s,1 H),5.30(s,2H),4.52(d,J=12.4Hz,1H),4.24(m,2H),3.79(m,3H),3.49(m,3H ),3.17(m,2H),2.99(m,3H),2.81(m,1H),2.63(m,1H),1.17(t,J=7.6Hz,3H).

[0556] Example 20 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(5-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0557] Step 1: 5-(2,2,2-trifluoroacetyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 20a

[0558] 4,5,6,7-Tetrahydrothieno[3,2-c]pyridine hydrochloride (44a-1) (500 mg, 2.84 mmol) was dissolved in dry dichloromethane (5 mL). N,N-diisopropylethylamine (1.1 g, 8.54 mmol) was added, and the temperature was lowered to 0°C. Trifluoroacetic anhydride (1.2 g, 5.70 mmol) was added dropwise, and the mixture was allowed to react at 0°C for 1 h. After completion of the reaction, water (5 mL) was added to quench the reaction, followed by extraction with dichloromethane. The combined organic phases were dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford the crude product, compound 20a (700 mg), as a yellow oil.

[0559] MS (ESI) m / z 235.9 [M+H] + .

[0560] Step 2 5-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 20b

[0561] Compound 20a (300 mg, 1.28 mmol) was dissolved in dry tetrahydrofuran (3 mL), and a 1M borane-tetrahydrofuran solution (1.5 mL, 1.53 mmol) was added dropwise. The mixture was heated to 70°C with stirring for 4 h. TLC confirmed the reaction was complete. The reaction solution was then slowly added dropwise to water to quench the reaction. The mixture was extracted with ethyl acetate, and the combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography (PE:EA = 5:1) to afford compound 20b (130 mg) as a clear oil.

[0562] MS (ESI) m / z 222.0 [M+H] + .

[0563] Step 3 2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-5-(2,2,2-trifluoroacetyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine 20c

[0564] Compound 20b (130 mg, 0.59 mmol) was dissolved in dry tetrahydrofuran (2 mL), cooled to -70°C, and 2.5 M n-butyllithium (0.4 ml, 0.88 mmol) was slowly added dropwise. The temperature was maintained at -70°C and stirred for 30 min. Isopropyl pinacol borate (164.2 mg, 0.88 mmol) was added dropwise at -70°C. After addition, the temperature was raised to room temperature and stirred for 2 h. After completion of the reaction, water was added to quench the reaction, and the mixture was extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford the crude product, compound 20c (200 mg), as a reddish-brown oil.

[0565] MS (ESI) m / z 348.1 [M+H] + .

[0566] Step 4: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(5-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 20d

[0567] Compound 20c (200 mg, 0.58 mmol) and 43a (362.8 mg, 0.46 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). 1,1-Bis(diphenylphosphino)ferrocenepalladium dichloride (74.1 mg, 0.06 mmol) and sodium carbonate (183.15 mg, 1.73 mmol) were added. The atmosphere was purged with nitrogen three times and the reaction was carried out at 100°C for 4 h. After completion of the reaction, the mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography to afford compound 20d (160 mg) as a yellow solid.

[0568] MS (ESI) m / z 929.2 [M+H] + .

[0569] Step 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(5-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 20

[0570] Compound 20d (160 mg, 0.17 mmol) was dissolved in trifluoroacetic acid (3 mL), heated to 50°C for 3 h, and the reaction solution was concentrated under reduced pressure. The crude product was purified by preparative HPLC to give compound 20 (23.3 mg) as a yellow solid.

[0571] MS (ESI) m / z 839.2 [M+H] + .

[0572] 1 H NMR (400MHz, DMSO-d6) δ10.40(s,1H),10.24(s,1H),8.59(s,1H),8.06(d,J=8.0 Hz,1H),7.98(d,J=8.0Hz,1H),7.73(dd,J=8.0Hz,1H),7.61(s,1H),5.35(s,2H) ,4.53(d,J=8Hz,1H),3.51(d,J=8Hz,3H),3.35~3.42(m,4H),3.32~3.30(m,4H), 3.01(s,2H),2.84(d,J=8Hz,1H),2.65(s,1H),2.45(s,3H),1.20(t,J=8Hz,3H).

[0573] Example 21 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-((R)-4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0574] First step (2R)-4-(1-methoxy-1,3-dioxolan-2-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 21b

[0575] Compound 21a (4.74 g, 22.7 mmol) was dissolved in acetonitrile (40 mL), and (R)-tert-butyl 2-methylpiperazine-1-carboxylate (5 g, 24.97 mmol) and potassium carbonate (6.27 g, 45.39 mmol) were added. The mixture was reacted at room temperature for 4 h. Water was added and the mixture was extracted with ethyl acetate (100 mL × 3). The organic phases were combined and concentrated under reduced pressure to give the crude product, which was purified by silica gel column chromatography to give compound 21b (6.69 g).

[0576] MS (ESI) m / z 328 [M+H] + .

[0577] Step 2 (R)-4-(2-bromo-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 21d

[0578] The synthesis of compound 21c was described in WO2022249060A1. Compound 21b (3.5 g, 10.66 mmol), compound 21c (1.58 g, 9.69 mmol), and anhydrous phosphoric acid (1.04 g, 10.66 mmol) were added to a reaction flask, dissolved in ethanol (20 mL), heated to 80°C, and reacted for 16 h. The reaction solution was concentrated, water was added, and extraction was performed with ethyl acetate (50 mL x 3). The organic phases were combined and concentrated under reduced pressure to obtain a crude product, which was then purified by silica gel column chromatography to obtain compound 21d (1.65 g).

[0579] MS (ESI) m / z 441 [M+H] + .

[0580] Step 3 (R)-4-(2-bromo-4-(2-(2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 21e

[0581] Compound 21d (1.65 g, 3.74 mmol) was dissolved in acetonitrile (10 mL), and 2-bromo-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide (1.3 g, 4.11 mmol) and DIPEA (1.45 g, 11.22 mmol) were added. The mixture was reacted at room temperature for 3 h. Water was added and the mixture was extracted with ethyl acetate (30 mL×3). The organic phases were combined and concentrated under reduced pressure to give a crude product. The crude product was purified by silica gel column chromatography to give compound 21e (1.13 g).

[0582] MS (ESI) m / z 677 [M+H] + .

[0583] Step 4: (2R)-4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 21f

[0584] Compound 21e (0.50 g, 0.74 mmol) was dissolved in 1,4-dioxane (10 mL) and water (2 mL), and compound 2a (0.23 g, 0.89 mmol), potassium phosphate (0.47 g, 2.22 mmol) and Pd(dppf)Cl2 (54.0 mg, 0.074 mmol) were added. The mixture was heated to 85°C under N2 protection for 16 h. Water was added and the mixture was extracted with ethyl acetate (30 mL × 3). The organic phases were combined and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography to obtain compound 21f (243 mg).

[0585] MS (ESI) m / z 734 [M+H] + .

[0586] Step 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-((R)-3-methylpiperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 21g

[0587] Compound 21f (243 mg, 0.38 mmol) was dissolved in dichloromethane (3 mL), and trifluoroacetic acid (3 mL) was added. The mixture was reacted at room temperature for 1 h and then concentrated under reduced pressure to give a crude product 21 g (210.0 mg). The crude product was used directly in the next reaction without purification.

[0588] MS (ESI) m / z 634 [M+H] + .

[0589] Step 6: 2-(6-((R)-4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide for 21 hours

[0590] Compound 21g (210 mg, 0.33 mmol) was dissolved in THF (10 mL), and compound 1e (97 mg, 0.39 mmol), triethylamine (330 mg, 3.3 mmol) and T3P (300 mg, 0.49 mmol, 50% wt in EA) were added. The reaction solution was reacted at room temperature for 3 h, and then quenched by adding saturated sodium bicarbonate solution. The mixture was extracted with ethyl acetate (50 mL × 3). The organic phases were combined and concentrated under reduced pressure to give the crude product. The crude product was purified by silica gel column chromatography (PE / EtOAc = 1 / 1) to give compound 21h (103 mg).

[0591] MS (ESI) m / z 860.5 [M+H] + .

[0592] Step 7: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-((R)-4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 21

[0593] Compound 21h (103 mg, 0.15 mmol) was dissolved in trifluoroacetic acid (3 mL), heated to 50°C for 3 h, and then concentrated under reduced pressure to obtain a crude product. The crude product was purified by preparative liquid chromatography (preparative column: YMCC18, mobile phase A: 0.1% ammonia in H2O, mobile phase B: acetonitrile, 30%-50% acetonitrile, 30 mL / min) to obtain compound 21 (53 mg) as a white solid.

[0594] MS (ESI) m / z 770.4 [M+H] + .

[0595] 1 H NMR(400MHz,DMSO-d6)δ10.35(s,1H),10.25(br s,1H),8.56(d,J=4.4Hz,1H),8.06(d,J=8.6Hz,1H),7.96(d,J=2.1Hz,1H ),7.71(dd,J=8.7,2.1Hz,1H),6.74(s,1H),5.31(s,2H),4.78(m,0.5H),4 .39(d,J=12.4Hz,0.5H),3.74(m,3.5H),3.49(m,1.5H),3.12(m,1H),2.92 (m,1H),2.80(d,J=12.0Hz,0.5H),2.68(d,J=12.0Hz,0.5H),2.58(m,2H), 2.44(d,J=3.8Hz,2H),2.27(m,2H),1.89(m,2H),1.78-1.61(m,4H),1.40(m,3H),1.23(s,3H),1.20(t,J=7.5Hz,3H).

[0596] Example 22 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0597] Step 1 6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 22e

[0598] 4,5,6,7-Tetrahydrothieno[2,3-c]pyridine hydrochloride (43b-1) (1.0 g, 5.69 mmol) was dissolved in dry dichloromethane (10 mL). N,N-diisopropylethylamine (2.2 g, 17.07 mmol) was added, and the temperature was cooled to 0°C. Trifluoroacetic anhydride (2.4 g, 11.38 mmol) was added dropwise, and the reaction was maintained at 0°C for 1 h. After completion, the reaction was quenched with water and extracted with dichloromethane. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product, compound 22e (1.5 g), as a yellow oil.

[0599] MS (ESI) m / z 235.9 [M+H] + .

[0600] Step 2 6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 22f

[0601] Compound 22e (1.5 g, 6.62 mmol) was dissolved in dry tetrahydrofuran (10 mL), and a 1M borane-tetrahydrofuran solution (13.2 ml, 13.24 mmol) was added dropwise. The mixture was then heated to 70°C and allowed to react for 4 h. After completion, the reaction mixture was slowly added dropwise to water to quench the reaction. The mixture was extracted with ethyl acetate, and the combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography (PE:EA = 4:1) to afford compound 22f (970 mg) as a clear oil.

[0602] MS (ESI) m / z 222.0 [M+H] + .

[0603] Step 3 2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine 22g

[0604] Compound 22f (970 mg, 4.39 mmol) was dissolved in dry tetrahydrofuran (10 mL) and cooled to -70°C. 2.5 M n-butyllithium (2.6 ml, 6.58 mmol) was slowly added dropwise, and the mixture was maintained at -70°C with stirring for 30 min. Isopropyl pinacol borate (1.3 g, 6.58 mmol) was added dropwise at -70°C. The mixture was allowed to warm to room temperature and allowed to react for 2 h. After completion, the reaction was quenched with water and extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to yield the crude product, compound 22g (880 mg), as a reddish-brown oil.

[0605] MS (ESI) m / z 348.1 [M+H] + .

[0606] Step 4: 2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide for 22 hours

[0607] Compound 22g (300 mg, 0.86 mmol) and 43a (544.1 mg, 0.69 mmol) were dissolved in 1,4-dioxane (3 mL) and water (0.6 mL), and 1,1-bis(diphenylphosphino)ferrocenepalladium dichloride dichloromethane complex (70.6 mg, 0.09 mmol) and sodium carbonate (274.7 mg, 2.592 mmol) were added. The atmosphere was replaced with nitrogen three times and the reaction was carried out at 100°C overnight. After completion of the reaction, the mixture was cooled to room temperature, diluted with water, extracted with ethyl acetate, and the organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 22h (320 mg) as a reddish-brown solid.

[0608] MS (ESI) m / z 929.3 [M+H] + .

[0609] Step 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(6-(2,2,2-trifluoroethyl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 22

[0610] Compound 22h (320 mg, 0.35 mmol) was dissolved in trifluoroacetic acid (3 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure and the crude product was separated by preparative HPLC to obtain compound 22 (38.4 mg) as a yellow solid.

[0611] MS (ESI) m / z 839.2 [M+H] + .

[0612] 1 H NMR (400MHz, DMSO-d6) δ10.40(s,1H),10.24(s,1H),8.59(s,1H),8.06(d,J= 8.0Hz,1H),7.98(d,J=8.0Hz,1H),7.73(dd,J=8.0Hz,1H),7.61(s,1H),5.35(s,2H),4.53(d,J=8Hz,1H),3.51(d,J=8Hz,3 H),3.35~3.42(m,4H),3.32~3.30(m,4H),3.01(s,2H),2.84(d,J=8Hz,1H),2.65(s,1H),2.45(s,3H),1.20(t,J=8Hz,3H).

[0613] Example 23 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]-2'-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0614] Step 1-(Thiophen-3-yl)cyclopropane-1-carbonitrile 23b

[0615] Compound 23a (5 g, 40.6 mmol) was dissolved in DMF (100 mL), and NaH (4.06 g, 101 mmol) was added at 0°C. The mixture was reacted at 0°C for 30 min. 2-Bromo-1-chloroethane (6.96 g, 48.7 mmol) was then added and the reaction was continued at 25°C for 1 h. After completion of the reaction, water was slowly added to quench the reaction mixture, and the mixture was extracted with ethyl acetate. The organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was separated and purified by column chromatography to afford compound 23b (4.3 g) as a white solid.

[0616] Step 2 (1-(Thien-3-yl)cyclopropyl)methanamine 23c

[0617] 23b (4.2 g, 28.2 mmol) was dissolved in THF (80 mL), and LiAlH (56 mL, 1 M, 56.4 mmol) was added at 0°C. The mixture was allowed to react at 0°C for 2 h. After completion of the reaction, H2O (50 mL), 10% aqueous NaOH (50 mL), and H2O (50 mL) were slowly added sequentially. The mixture was stirred for 30 min, filtered, and extracted three times with ethyl acetate. The collected organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford compound 23c (3.9 g) as a white solid.

[0618] MS (ESI) m / z 154.0 [M+H] +

[0619] Step 3 6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]23d

[0620] 23c (3.9 g, 25.3 mmol) was dissolved in EtOH (60 mL), and HCHO (6.15 g, 37%, 75.9 mmol) and HCl (12 M, 2 mL) were added. The mixture was reacted at 80°C for 2 h. After completion of the reaction, water and ethyl acetate were added to the reaction solution, and the mixture was stirred for 10 min. The layers were allowed to stand for stratification. After separation, the pH of the aqueous phase was adjusted to 9 with 10% aqueous NaOH solution. The product was extracted three times with ethyl acetate. The collected organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford compound 23d (1.1 g) as a white solid.

[0621] MS (ESI) m / z 166.0 [M+H] +

[0622] 1 H NMR(400MHz, CDCl3)δ7.06(t,J=5.1Hz,1H),6.49–6.41(m,1H),4.05(d,J=69.8Hz,2H) ,3.43(s,1H),2.86(d,J=39.9Hz,2H),0.92(dt,J=11.3,4.5Hz,2H),0.85–0.76(m,2H)

[0623] Step 4: 6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]23e

[0624] 23d (1.6 g, 9.63 mmol) was dissolved in THF (30 mL), trideuterium methyl 4-methylbenzenesulfonate (1.09 g, 5.78 mmol), and TEA (1.94 g, 19.26 mmol) and reacted at 25°C for 6 h. After completion of the reaction, the reaction solution was filtered and dried, and the crude product was purified by column chromatography (DCM:MeOH = 10:1) to afford compound 23e (220 mg) as a pale yellow solid.

[0625] MS (ESI) m / z 183.1 [M+H] +

[0626] Step 5 (6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]-2'-yl)boronic acid 23f

[0627] 23e (220 mg, 1.20 mmol) was dissolved in THF (10 mL), and n-BuLi (0.72 mL, 2.5 M, 1.80 mmol) was added at -78°C. After reacting at -78°C for 30 min, 2,6-dimethyl-4-(propan-2-yloxy)-4-bora-3,5-dioxaheptane (339 mg, 1.80 mmol) was added and the reaction was continued at -78°C for 1 h. After completion of the reaction, the reaction solution was diluted with water and extracted with ethyl acetate. The collected aqueous phase was concentrated, and the crude product was purified by reverse-phase column chromatography (C18, H2O:MeCN=100:0) to afford compound 23f (250 mg) as a pale yellow solid.

[0628] MS (ESI) m / z 227.1 [M+H] +

[0629] Step 6: 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-2-(6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]-2'-yl)-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 23g

[0630] Compound 23f (204 mg, 906 μmol), 1a (300 mg, 453 μmol), Pd(dppf)Cl2 (33.1 mg, 45.3 μmol), and K2CO3 (156 mg, 1.13 mmol) were dissolved in 1,4-dioxane (10 mL) and H2O (2 mL). The atmosphere was replaced with nitrogen three times and the mixture was reacted at 80°C for 1 h. After completion of the reaction, the reaction solution was diluted with water and extracted three times with ethyl acetate. The organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by column chromatography (DCM:MeOH = 10:1) to afford compound 23g (120 mg) as a pale yellow solid.

[0631] MS (ESI) m / z 764.3 [M+H] +

[0632] Step 7: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridin-2'-yl)-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide for 23 hours

[0633] Compound 23g (110 mg, 157 μmol) was dissolved in DCM (4 mL), and TFA (89.5 mg, 785 μmol) was added. The mixture was allowed to react at 25°C for 1 h. After completion of the reaction, the pH of the reaction solution was adjusted to 8 with saturated aqueous NaHCO₃. The mixture was extracted three times with ethyl acetate. The collected organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 23h (50 mg) as a light yellow liquid.

[0634] MS (ESI) m / z 664.2 [M+H] +

[0635] Step 8 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]-2'-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 23i

[0636] 23h (50 mg, 90 μmol) and 1e (21.9 mg, 90 μmol) were dissolved in DMF (2 mL), and T3P (114 mg, 180 μmol) and DIEA (46.4 mg, 360 μmol) were added. The mixture was reacted at 25°C for 1 h. After completion of the reaction, water was added to the reaction solution, and the mixture was extracted three times with ethyl acetate. The collected organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by column chromatography to afford compound 23i (25 mg) as a light yellow solid.

[0637] MS (ESI) m / z 890.4 [M+H] +

[0638] Step 9: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6'-(methyl-d3)-6',7'-dihydro-5'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyridine]-2'-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 23

[0639] 23i (25 mg, 28 μmol) was dissolved in TFA (1 mL) and reacted at 50°C for 1 h. After completion of the reaction, the pH of the reaction solution was adjusted to 8 with saturated aqueous NaHCO₃ solution and extracted with ethyl acetate. The collected organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain the crude product. The crude product was separated by preparative HPLC to afford compound 23 (6.6 mg) as a white solid.

[0640] MS (ESI) m / z 800.2 [M+H] +

[0641] 1 H NMR (400MHz, DMSO-d6) δ10.38(s,1H),8.33(d,J=44.3Hz,2H),7.97(d,J=8.5Hz,1H) ,7.90(s,1H),7.65(d,J=7.2Hz,1H),5.26(s,2H),4.46(d,J=11.5Hz,1H),3.62(s,2 H),3.45(s,2H),3.21–3.15(m,4H),2.93(d,J=7.4Hz,4H),2.75(d,J=9.4Hz,1H),2. 58(d,J=12.0Hz,1H),2.34(s,3H),1.13(t,J=7.4Hz,3H),0.85(s,2H),0.75(s,2H).

[0642] Example 24 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(6-hydroxy-7-methylpyrazolo[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0643] Step 1: 2-(6-(4-(6-(benzyloxy)-7-methylpyrazolo[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 24a

[0644] Compound 22d (150 mg, 0.255 mmol) was dissolved in DMF (2 mL), and compound 23a (74.69 mg, 0.306 mmol) and DIEA (165.09 mg, 1.277 mmol) were added. The reaction solution was cooled to 0°C, and a 50% ethyl acetate solution of 1-propylphosphoric acid cyclic anhydride (121.7 mg, 0.3825 mmol) was added dropwise. The reaction was allowed to react at room temperature for 2 h. Water (5 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (5 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 24a (120 mg) as a reddish-brown solid.

[0645] MS (ESI) m / z 887.2 [M+H] + .

[0646] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(6-hydroxy-7-methylpyrazolo[1,5-a]pyrimidine-5-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 24

[0647] Compound 24a (120 mg, 0.135 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure and the crude product was purified by preparative liquid chromatography to give compound 24 (6.3 mg) as a light yellow solid.

[0648] MS (ESI) m / z 797.2 [M+H] + .

[0649] 1H NMR (400MHz, DMSO-d6) δ10.38(s,1H),8.12(s,1H),8.05(d,J=8.0Hz,1H),7.96(s, 1H),7.71(d,J=8.0Hz,1H),7.52(s,1H),6.69(d,J=8Hz,1H),5.31(s,2H),4.77(2, 2H),4.54(d,J=8Hz,2H),3.38(t,J=8Hz,2H),3.53-3.42(m,3H),3.03-2.98(m,2H),2 .85(d,J=8Hz,1H),2.71(s,1H),2.69(s,3H),2.62(d,J=8Hz,2H),1.19(t,J=8Hz,3H).

[0650] Example 25 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-2-(6-(methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0651] Step 1: (R)-4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-2-(6-(methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 25a

[0652] Compound 47a (100 mg, 0.5 mmol) and 21e (239.8 mg, 0.42 mmol) were dissolved in 1,4-dioxane (4 mL) and water (1 mL), and 1,1-bis(diphenylphosphino)ferrocenepalladium dichloride dichloromethane complex (34.1 mg, 0.04 mmol) and sodium carbonate (132.6 mg, 1.25 mmol) were added. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 100°C for 4 h. The mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 25a (180 mg) as a reddish-brown solid.

[0653] MS (ESI) m / z 752.4 [M+H] + .

[0654] Step 2 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(6-(methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-6-(3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 25b

[0655] Compound 25a (180 mg, 0.24 mmol) was dissolved in dry dichloromethane (1 mL), and trifluoroacetic acid (1 mL) was added. The mixture was allowed to react at room temperature for 1 h. The reaction solution was concentrated under reduced pressure, and a saturated aqueous sodium bicarbonate solution was added to the residue. The mixture was extracted with dichloromethane / isopropanol (volume ratio = 10 / 1). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 25b (160 mg) as a reddish-brown solid.

[0656] MS (ESI) m / z 652.3 [M+H] + .

[0657] Step 3 (R)-2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-5-ethyl-2-(6-methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-trifluoromethylphenyl)acetamide 25c

[0658] Compound 25b (160 mg, 0.24 mmol) was dissolved in DMF (1 mL), followed by the addition of compound 1e (56.1 mg, 0.23 mmol) and DPEA (99.2 mg, 0.77 mmol). The reaction mixture was cooled to approximately 0°C, and a 50% ethyl acetate solution of T3P (146.4 mg, 0.084 mmol) was added dropwise. After addition, the mixture was heated to 60°C and stirred overnight. After completion, the reaction was cooled to room temperature and quenched with water. The mixture was extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated by column chromatography to afford compound 25c (160 mg) as a reddish-brown solid.

[0659] MS (ESI) m / z 878.4 [M+H] + .

[0660] Step 4 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-2-(6-(methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 25

[0661] Compound 25c (160 mg, 0.18 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. The reaction solution was concentrated and the crude product was separated by preparative HPLC to obtain compound 25 (13.8 mg) as a light yellow solid.

[0662] MS (ESI) m / z 788.2 [M+H] + .

[0663] Example 26 N-(2,4-dichlorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0664] Step 1: 2-Bromo-N-(2,4-dichlorophenyl)acetamide 26b

[0665] 2,4-Dichloroaniline (26a) (1 g, 6.212 mmol) and DMAP (758.9 mg, 6.212 mmol) were dissolved in dry dichloromethane (10 mL). Bromoacetyl bromide (1.49 g, 7.454 mmol) was added dropwise at 0°C. The mixture was reacted at room temperature for 18 h. The reaction solution was concentrated and the crude product was purified by column chromatography (PE / EA = 20 / 1) to give compound 26b (980 mg) as a white solid.

[0666] MS (ESI) m / z 282.1 [M+H] + .

[0667] Step 2: tert-Butyl 4-(4-(2-((2,4-dichlorophenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 26d

[0668] Compound 26c (300 mg, 0.617 mmol) and compound 26b (260 mg, 0.9256 mmol) were dissolved in dry acetonitrile (3 mL), and DIEA (239 mg, 1.851 mmol) was added. The mixture was reacted at room temperature for 18 h. The reaction solution was concentrated, and the crude product was separated by column chromatography (PE / EA = 2 / 1) to give compound 26d (230 mg) as a light yellow solid.

[0669] MS (ESI) m / z 688.1 [M+H] + .

[0670] Step 3: N-(2,4-dichlorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 26e

[0671] Compound 26d (230 mg, 0.3347 mmol) was dissolved in dry dichloromethane (2 mL), and trifluoroacetic acid (2 mL) was added. The mixture was allowed to react at room temperature for 2 h. The reaction solution was concentrated under reduced pressure, and saturated aqueous sodium bicarbonate (3 mL) was added to the residue. The mixture was extracted with dichloromethane (5 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to afford compound 26e (200 mg) as a reddish-brown solid.

[0672] MS (ESI) m / z 588.1 [M+H] + .

[0673] Step 4: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2,4-dichlorophenyl)acetamide 26f

[0674] Compound 26e (150 mg, 0.255 mmol) was dissolved in DMF (2 mL), and compound 1e (74.69 mg, 0.306 mmol) and DIEA (164.78 mg, 1.275 mmol) were added. The reaction solution was cooled to 0°C, and T3P (243.4 mg, 0.3825 mmol, 50% in EA) was added dropwise. The reaction was allowed to react at room temperature for 2 h. Water (5 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (5 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 26f (130 mg) as a reddish-brown solid.

[0675] MS (ESI) m / z 814.2 [M+H]+ .

[0676] Step 5: N-(2,4-dichlorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 26

[0677] Compound 26f (130 mg, 0.1599 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure and the crude product was separated by preparative liquid chromatography to obtain compound 26 (34.2 mg) as a white solid.

[0678] MS (ESI) m / z 724.2 [M+H] + .

[0679] 1 H NMR (400MHz, DMSO-d6) δ10.25(s,1H),8.45(s,1H),7.72(d,J=8.0Hz,1H),7.7 1(s,1H),7.53(s,1H),7.42(dd,J=8.0Hz,1H),6.67(s,1H),5.26(s,2H),4.77 (s,2H),4.54(d,J=8.0Hz,1H),3.88(t,J=8Hz,2H),3.61-3.45(m,4H),3.01-2 .97(m,3H),2.83(m,1H),2.71(t,J=8Hz,2H),2.35(s,3H),1.20(t,J=8Hz,3H).

[0680] Example 27 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0681] Step 1: (R)-4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-3-methylpiperazine-1-carboxylic acid tert-butyl ester 27b

[0682] The synthesis of compound 27a refers to patent WO2022249060. Compound 27a (147 mg, 0.22 mmol), compound 22b (69 mg, 0.26 mmol), Pd(dppf)Cl2 (16 mg, 0.02 mmol), sodium carbonate (69 mg, 0.65 mmol), 1,4-dioxane (5 mL), and H2O (1 mL) were added to a reaction flask. The atmosphere was replaced with argon three times and the reaction was carried out at 100°C for 4 h. Ethyl acetate (50 mL) and saturated brine (10 mL) were added and extracted. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 27b (155 mg) as a light yellow solid.

[0683] MS (ESI) m / z 736.5 [M+H] + .

[0684] Step 2 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 27c

[0685] Compound 27b (155 mg, 0.21 mmol) was dissolved in DCM (2 mL) and TFA (2 mL) was added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated, ethyl acetate (50 mL) was added, and the pH was adjusted to 8 with saturated sodium bicarbonate. Extraction was performed, and the organic phase was washed with saturated brine and concentrated under reduced pressure to afford crude compound 27c (123 mg).

[0686] MS (ESI) m / z 636.4 [M+H] + .

[0687] Step 3 (R)-2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 27d

[0688] Compound 27c (123 mg, 0.19 mmol) was dissolved in THF (2 mL), and compound 1e (51 mg, 0.21 mmol), DIPEA (129 mg, 1.0 mmol), and T3P (191 mg, 0.3 mmol, 50% in EA) were added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, saturated sodium bicarbonate solution was added to quench the reaction. The mixture was extracted with ethyl acetate (50 mL). The organic phase was washed three times with saturated brine and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (DCM / MeOH = 30 / 1) to afford compound 27d (144 mg) as a pale yellow solid.

[0689] MS (ESI) m / z 862.6 [M+H] + .

[0690] Step 4 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 27

[0691] Compound 27d (144 mg, 0.17 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. After completion of the reaction, as monitored by TLC, the reaction mixture was concentrated under reduced pressure, ethyl acetate (50 mL) was added, and saturated aqueous sodium bicarbonate solution was adjusted to pH 8 for extraction. The organic phase was washed with saturated brine and concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography to afford compound 27 (51 mg).

[0692] MS (ESI) m / z 772.6 [M+H] + .

[0693] 1 H NMR (400MHz, CDCl3) δ11.78(s,1H),8.96(s,1H),8.58(s,1H),8.45(d,J=8.8Hz,1H), 7.61(s,1H),7.59(s,1H),7.52(d,J=8.8Hz,1H),5.65-5.62(m,1H),5.24(d,J=16.0Hz ,1H),5.13(d,J=16.0Hz,1H),4.82(s,2H),4.76(m,1H),3.98-3.84(m,3H),3.73(m,1H ),3.55-3.35(m,2H),3.16-2.95(m,2H),2.75(m,3H),2.56(s,3H),1.34-1.27(m,6H).

[0694] Example 28 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0695] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-2-(4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 28b

[0696] Compound 28a (160 mg, 0.55 mmol) and compound 1a (300 mg, 0.45 mmol) were dissolved in 1,4-dioxane (10 mL) and water (2 mL), and (1,1'-bis(diphenylphosphino)ferrocene)palladium dichloride (33 mg, 0.045 mmol) and sodium carbonate (144 mg, 1.36 mmol) were added. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 100°C for 4 h. The mixture was cooled to room temperature and diluted with water (20 mL). The mixture was extracted with ethyl acetate (30 mL x 3), and the organic phase was concentrated under reduced pressure. The crude product was purified by column chromatography (PE / EA = 2 / 1) to give compound 28b (180 mg) as a yellow solid.

[0697] MS (ESI) m / z 692 [M-C4H7] + .

[0698] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-7-oxo-6-(piperazin-1-yl)-2-(4'H,6'H-spiro[cyclopropane]-1,7'-thieno[3,2-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 28c

[0699] Compound 28b (180 mg, 0.24 mmol) was dissolved in dry dichloromethane (2 mL), and trifluoroacetic acid (2 mL) was added. The mixture was reacted at room temperature for 1 h, concentrated under reduced pressure, and saturated sodium bicarbonate solution (10 mL) was added. The mixture was extracted with dichloromethane (20 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to give compound 28c (150 mg) as a yellow solid.

[0700] MS (ESI) m / z 648 [M+H] + .

[0701] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(4'H,6'H-spiro)[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 28d

[0702] Compound 28c (100 mg, 0.154 mmol) and compound 1e (45 mg, 0.184 mmol) were dissolved in dry DMF (3 mL), and DIEA (99.5 mg, 0.77 mmol) was added. The mixture was cooled to 0°C, and a 50% ethyl acetate solution of 1-propylphosphoric acid cyclic anhydride (147 mg, 0.23 mmol) was added dropwise. The reaction was allowed to react at room temperature for 1 h, and water (10 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (20 mL x 3), and the organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by column chromatography (PE / EA = 1 / 3) to give compound 28d (100 mg) as a white solid.

[0703] MS (ESI) m / z 874 [M+H] + .

[0704] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(4'H,6'H-spiro[cyclopropane-1,7'-thieno[3,2-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 28

[0705] Compound 28d (100 mg, 0.11 mmol) was dissolved in trifluoroacetic acid (5 mL), reacted at 50°C for 18 h, and concentrated under reduced pressure. The crude product was dissolved in dichloromethane, and saturated aqueous sodium bicarbonate solution was added. The organic phase was concentrated under reduced pressure. The crude product was separated by preparative liquid chromatography to give compound 28 (33.0 mg) as a white solid.

[0706] MS (ESI) m / z 784 [M+H] + .

[0707] 1H NMR (400MHz, DMSO-d6) δ10.37(s,1H),8.57(s,1H),8.05(d,J=8.7Hz,1H),7.97(s,1H),7 .72(d,J=8.8Hz,1H),7.45(s,1H),5.32(s,2H),4.72(s,2H),4.52(d,J=11.4Hz,1H),3.67 (s,2H),3.55–3.45(m,3H),3.27–3.21(m,2H),2.99(s,3H),2.82(d,J=10.4Hz,1H),2.64( d,J=11.7Hz,1H),2.44(s,3H),1.19(t,J=7.4Hz,3H),1.08(d,J=6.7Hz,2H),0.93(s,2H).

[0708] Example 29 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0709] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 29b

[0710] Compound 1a (1.0 g, 1.51 mmol), compound 29a (485 mg, 1.81 mmol), Pd(dppf)Cl2 (110 mg, 150.85 μmol), and Na2CO3 (480 mg, 4.53 mmol) were dissolved in 1,4-dioxane (20 mL) and water (4 mL). The atmosphere was replaced with nitrogen three times and the reaction was carried out at 85°C for 3 h. The reaction solution was cooled to room temperature, and DCM (50 mL) was added. The reaction mixture was washed with saturated brine (50 mL). The organic phase was concentrated under reduced pressure, and the crude product was purified by column chromatography to afford compound 29b (1.0 g) as a yellow solid.

[0711] MS (ESI) m / z 668.1 [M-C4H7] +

[0712] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 29c

[0713] Compound 29b (500 mg, 690.42 μmol) was dissolved in DCM (2 mL) and TFA (2 mL) was added. The mixture was allowed to react at room temperature for 1.5 h. The mixture was concentrated under reduced pressure, and DCM (50 mL) and saturated aqueous NaHCO₃ were added. The organic phase was separated, dried over anhydrous Na₂SO₄, and concentrated under reduced pressure to afford compound 29c (420 mg) as a yellow solid.

[0714] MS (ESI) m / z 624.1 [M+H] +

[0715] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl-5,5-d2)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 29d

[0716] Compound 29c (400 mg, 640.95 μmol) and compound 1e (189 mg, 769.14 μmol) were dissolved in DMF (5.0 mL). DIPEA (414 mg, 3.20 mmol) and T3P (50% wt in EtOAc, 693 mg, 961.42 μmol) were added and allowed to react at room temperature for 1 h. After completion, the reaction was concentrated under reduced pressure, and DCM (20 mL) and saturated aqueous NaHCO3 (10 mL) were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography to afford compound 29d (520 mg) as a yellow solid.

[0717] MS (ESI) m / z 850.3 [M+H] +

[0718] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-(methoxymethyl)-piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 29

[0719] Compound 29d (520 mg, 611.54 μmol) was dissolved in TFA (5 mL) and reacted at 50°C for 4 h. The mixture was concentrated under reduced pressure, and DCM (10 mL) and saturated NaHCO3 solution (5 mL) were added. The organic phase was separated and concentrated under reduced pressure to give the crude product, which was purified by preparative liquid chromatography to afford compound 29 (15.1 mg) as a light yellow solid.

[0720] MS (ESI) m / z 760.2 [M+H] +

[0721] 1 H NMR (400MHz, DMSO-d6) δ10.40(s,1H),8.54(s,1H),8.05(d,J=8.8Hz,1H),7.97(d, J=1.2Hz,1H),7.71(dd,J=8.8,1.6Hz,1H),7.52(s,1H),5.34(s,2H),4.76(s,2H),4 .53(d,J=12.0Hz,1H),3.52-3.46(m,3H),3.26(m,1H),3.05-2.95(m,3H),2.83(d, J=10.8Hz,1H),2.69(s,2H),2.66-2.62(m,1H),2.43(s,3H),1.19(t,J=7.6Hz,3H).

[0722] Example 30 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-((3-methyloxetan-3-yl)ethynyl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0723] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-2-((3-methyloxetane-3-yl)ethynyl)-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 30b

[0724] Compound 1a (200 g, 301.71 μmol), compound 30a (203 mg, 1.21 mmol), Xphos-Pd-G2 (24 mg, 30.17 μmol), and CsF (275 mg, 301.71 μmol) were dissolved in DMF (2 mL) and TEA (0.4 mL). The atmosphere was replaced with nitrogen three times, and the reaction mixture was stirred at 90°C for 3 h. The reaction mixture was concentrated under reduced pressure, and EtOAc (15 mL) and saturated aqueous NH4Cl (10 mL) were added. The organic phase was separated and concentrated under reduced pressure to give the crude product, which was purified by column chromatography to afford compound 30b (200 mg) as a yellow solid.

[0725] MS (ESI) m / z 622.1 [M-C4H7] +

[0726] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-((3-methyloxetan-3-yl)ethynyl)-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4-(7H)-yl)acetamide 30c

[0727] Compound 30b (80 mg, 117.97 μmol) was dissolved in DCM (1.5 mL) and TFA (1.5 mL) was added. The mixture was allowed to react at room temperature for 1.5 h. After completion, the reaction was concentrated under reduced pressure, and DCM (15 mL) and saturated aqueous NaHCO₃ were added. The organic phase was separated and concentrated under reduced pressure to afford compound 30c (72 mg) as a yellow solid.

[0728] MS (ESI) m / z 578.1 [M+H] +

[0729] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-((3-methyloxetan-3-yl)ethynyl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4-(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 30d

[0730] Compound 30c (72 mg, 124.57 μmol) and compound 1g (37 mg, 149.48 μmol) were dissolved in DMF (1.0 mL). DIPEA (81 mg, 622.84 μmol) and T3P (50% wt in EtOAc, 134.63 mg, 186.85 μmol) were added and reacted at room temperature for 1 h. After completion, the reaction was concentrated under reduced pressure, and DCM (10 mL) and saturated aqueous NaHCO3 (5 mL) were added. The organic phase was separated and concentrated under reduced pressure to give the crude product, which was purified by column chromatography to afford compound 30d (86 mg) as a yellow solid.

[0731] MS (ESI) m / z 804.3 [M+H] +

[0732] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-((3-methyloxetane-3-yl)ethynyl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4-(7H)-yl)acetamide 30

[0733] Compound 30d (84 mg, 104.45 μmol) was dissolved in TFA (1 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure, and DCM (10 mL) and saturated aqueous NaHCO₃ (5 mL) were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product, which was purified by preparative liquid chromatography to afford compound 30 (27.3 mg) as a white solid.

[0734] MS (ESI) m / z 714.2 [M+H] +

[0735] 1 H NMR (400MHz, DMSO-d6) δ10.33(s,1H),8.52(s,1H),8.10(d,J=8.6Hz,1H),7.96(d,J=1.3 Hz,1H),7.71(dd,J=8.6,1.3Hz,1H),5.33(s,2H),4.75(d,J=5.6Hz,2H),4.52(d,J=12.0H z,1H),4.45(d,J=5.6Hz,2H),3.51-3.44(m,3H),3.22(m,1H),3.03-2.94(m,3H),2.82(d ,J=11.2Hz,1H),2.64(d,J=11.2Hz,1H),2.43(s,3H),1.64(s,3H),1.18(t,J=7.4Hz,3H).

[0736] Example 31 N-(2-chloro-4-cyanophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0737] Step 1: 2-Bromo-N-(2-chloro-4-cyanophenyl)acetamide 31b

[0738] Compound 31a (2.0 g, 13.11 mmol) and DMAP (2.5 g, 20.46 mmol) were dissolved in DCM (40 mL). The reaction mixture was cooled to 0°C, and bromoacetyl bromide (4.68 g, 23 mmol) was added. The mixture was allowed to react at room temperature for 1 h. After completion, the reaction mixture was quenched with water (50 mL), extracted with dichloromethane, and the organic phase was concentrated under reduced pressure. The resulting mixture was purified by silica gel column chromatography to afford compound 31b (3.0 g) as a light yellow solid.

[0739] MS (ESI) m / z 273.1 [M+H] +

[0740] Step 2 4-(2-bromo-4-(2-((2-chloro-4-cyanophenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 31c

[0741] Compound 5c (500 mg, 1.17 mmol) and compound 31b (400 mg, 1.46 mmol) were dissolved in acetonitrile (10 mL), and DIEA (0.42 mL, 2.40 mmol) was added. The mixture was allowed to react at room temperature for 3 h. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography to afford compound 31c (680 mg) as a yellow solid.

[0742] MS (ESI) m / z 563.3 [M–C4H7] +

[0743] Step 3 tert-Butyl 4-(4-(2-((2-chloro-4-cyanophenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 31d

[0744] Compound 31c (300 mg, 483.95 μmol) and compound 22b (160 mg, 601.14 μmol) were dissolved in 1,4-dioxane (5 mL) and water (1 mL). NaCO (160 mg, 1.51 mmol) and Pd(dppf)Cl (35.0 mg, 47.83 μmol) were added. The reaction mixture was purged with nitrogen three times and reacted at 85°C for 16 h. The reaction mixture was concentrated under reduced pressure, and the crude product was purified by column chromatography to afford compound 31d (70 mg) as a yellow solid.

[0745] MS (ESI) m / z 623.3 [M–C4H7] +

[0746] Step 4: N-(2-chloro-4-cyanophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 31e

[0747] Compound 31d (70 mg, 103.06 μmol) was dissolved in DCM (2 mL) and TFA (1 mL) and allowed to react at room temperature for 1 h. The reaction mixture was concentrated under reduced pressure, and DCM (10 mL) and saturated aqueous NaHCO₃ were added. The organic phase was concentrated under reduced pressure and purified by column chromatography to afford compound 31e (77 mg) as a yellow solid.

[0748] MS (ESI) m / z 579.4 [M+H] +

[0749] Step 5 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-cyanophenyl)acetamide 31f

[0750] Compound 31e (77 mg, 132.97 μmol) and compound 1e (42 mg, 171.95 μmol) were dissolved in DMF (1.5 mL). DIPEA (86 mg, 665.39 μmol) and T3P (143 mg, 396.93 μmol) were added and reacted at room temperature for 2 h. The reaction solution was concentrated under reduced pressure, and the crude product was purified by column chromatography to afford compound 31f (55 mg) as a yellow solid.

[0751] MS (ESI) m / z 805.5 [M+H] +

[0752] Step 6: N-(2-chloro-4-cyanophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 31

[0753] Compound 31f (53 mg, 65.81 μmol) was dissolved in TFA (2.0 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated, and DCM (15 mL) and saturated aqueous NaHCO3 (2 mL) were added. The organic phase was separated and concentrated under reduced pressure. The crude product was purified by preparative liquid chromatography to afford compound 31 (12.7 mg) as a white solid.

[0754] MS (ESI) m / z 715.5 [M+H] +

[0755] 1 H NMR(400MHz,DMSO-d6)δ10.29(br s,1H),8.54(s,1H),8.15(d,J=1.8Hz,1H),8.05(d,J=8.4Hz,1H),7.80(dd,J =8.4,1.8Hz,1H),7.51(s,1H),5.34(s,2H),5.32(m,1H),4.76(s,2H),4.53( d,J=12.0Hz,1H),3.87(t,J=5.6Hz,2H),3.50(m,2H),2.99(m,2H),2.82(m,1 H), 2.71-2.63 (m, 3H), 2.43 (s, 3H), 2.04-1.95 (m, 2H), 1.18 (t, J = 7.2Hz, 3H).

[0756] Example 32 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(1-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-1,2,3,6-tetrahydropyridin-4-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0757] Step 1: tert-Butyl 4-(2-bromo-4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylate 32c

[0758] The synthesis of compounds 32a and 32b is described in WO2022249060. Compound 32a (500 mg, 1.18 mmol) was dissolved in acetonitrile (8 mL), and 2-bromo-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide (32b) (392 mg, 1.24 mmol) and DIEA (456 mg, 3.54 mmol) were added. The mixture was reacted at 70°C for 18 h. The reaction mixture was concentrated under reduced pressure, and the crude product was purified by column chromatography (PE / EA = 2 / 1) to afford compound 32c (420 mg) as a pale yellow solid.

[0759] MS (ESI) m / z 604 [M-C4H7] + .

[0760] Step 2: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylate 32d

[0761] Compound 32c (320 mg, 0.486 mmol) was dissolved in DMF (5 mL), and compound 22b (190 mg, 0.714 mmol), X-phosPdG3 (20 mg, 0.02 mmol), potassium phosphate (200 mg, 0.94 mmol), and water (0.5 mL) were added. The mixture was reacted at 80°C under nitrogen for 1 h. The reaction solution was concentrated under reduced pressure, and the crude product was purified by column chromatography (PE / EA = 1 / 1) to afford compound 32d (400 mg) as a white solid.

[0762] MS (ESI) m / z 663.1 [M-C4H7] + .

[0763] Step 3: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-6-(1,2,3,6-tetrahydropyridin-4-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 32e

[0764] Compound 32d (400 mg, 0.55 mmol) was dissolved in DCM (8 mL) and trifluoroacetic acid (4 mL) and allowed to react at room temperature for 1 h. The mixture was concentrated under reduced pressure, saturated sodium bicarbonate solution was added, and extraction was performed with ethyl acetate (20 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford compound 32e (250 mg) as a white solid.

[0765] MS (ESI) m / z 619.1 [M+1] + .

[0766] Step 4: 2-(6-(1-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)-1,2,3,6-tetrahydropyridin-4-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 32f

[0767] Compound 32e (250 mg, 0.405 mmol) was dissolved in DMF (10 mL), and compound 1e (125 mg, 0.526 mmol), 1-propylphosphonic anhydride in ethyl acetate (250 mg, 0.607 mmol), and DIEA (150 mg, 1.16 mmol) were added. The mixture was allowed to react at room temperature for 1 h. Water (50 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (30 mL x 2). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford compound 32f (300 mg) as a white solid.

[0768] MS (ESI) m / z 845.2 [M+1] + .

[0769] Step 5: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(1-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-1,2,3,6-tetrahydropyridin-4-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 32

[0770] Compound 32f (30 mg, 0.036 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 1.5 h. The reaction mixture was adjusted to neutrality by adding saturated sodium bicarbonate solution, extracted with dichloromethane, and concentrated under reduced pressure to obtain a crude product, which was purified by column chromatography to afford compound 32 (13 mg) as a white solid.

[0771] MS (ESI) m / z 755.2 [M+1] + .

[0772] 1H NMR(400MHz,DMSO-d6)δ8.05(dd,J=8.0,4.0Hz,1H),7.88(s,1H),7.82(d,J=8.0H z,1H),7.64(d,J=8.0Hz,1H),7.53(s,1H),6.02–5.63(m,2H),5.41–5.10(m,3H), 4.77(s,2H),4.01(d,J=4.0Hz,1H),3.87(t,J=5.5Hz,2H),2.78–2.64(m,4H),2 .36–2.29(m,1H),2.21(s,3H),1.99(s,1H),1.76(d,J=4.0Hz,1H),1.23(s,3H).

[0773] Example 33 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0774] Step 1: (S)-tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylate 33b

[0775] The synthesis of compound 33a refers to patent WO2022249060. Compound 33a (490 mg, 0.72 mmol), compound 22b (193 mg, 0.72 mmol), Pd(dppf)Cl2 (32 mg, 0.04 mmol), and sodium carbonate (228 mg, 2.16 mmol) were dissolved in 1,4-dioxane (20 mL) and H2O (4 mL). The atmosphere was replaced with nitrogen three times and the mixture was reacted at 100°C for 4 h. Ethyl acetate (150 mL) and saturated brine (30 mL) were added, and the organic phase was separated, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 33b (220 mg) as a light yellow solid.

[0776] MS (ESI) m / z 736.5 [M+H] + .

[0777] Step 2 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 33c

[0778] Compound 33b (220 mg, 0.65 mmol) was dissolved in DCM (10 mL), and ethanolic hydrogen chloride (2 mL, 4 mol / L) was added. The mixture was allowed to react at room temperature for 3 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to afford crude compound 33c (160 mg).

[0779] MS (ESI) m / z 636.4 [M+H] + .

[0780] Step 3 (S)-2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 33d

[0781] Compound 33c (100 mg, 0.17 mmol) was dissolved in DMF (2 mL), and compound 1e (41 mg, 0.17 mmol), triethylamine (52 mg, 0.51 mmol), EDCI (33 mg, 0.17 mmol), and HOBt (23 mg, 0.17 mmol) were added. The mixture was allowed to react at room temperature for 16 h. After TLC monitoring, ethyl acetate (50 mL) was added, and the organic phase was washed three times with saturated brine (20 mL). The organic phase was concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography (DCM / MeOH = 30 / 1) to obtain compound 33d (76 mg) as a light yellow solid.

[0782] MS (ESI) m / z 862.6 [M+H] + .

[0783] Step 4 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 33

[0784] Compound 33d (76 mg, 0.1 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 3 h. After completion of the reaction, as monitored by TLC, the mixture was concentrated under reduced pressure. Ethyl acetate (50 mL) and saturated aqueous sodium bicarbonate were added, and the organic phase was separated and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (DCM / MeOH = 10 / 1) to obtain compound 33 (2.5 mg).

[0785] MS (ESI) m / z 772.2 [M+H] + .

[0786] 1 H NMR (400MHz, CDCl3) δ11.77(s,0.5H),11.70(s,0.5H),9.04(s,1H),8.60(d,J=6.6Hz,1H),8.49(d,J=8.6Hz,1H),7.63(m,2H),7 .53(m,1H),5.75(m,0.5H),5.48(m,0.5H),5.13(s,2H),5.06(m,0.5H),4.84(s,2H),4.68(m,0.5H),3.98(m,3.5H),3.81(m,1H), 3.69(m,0.5H),3.36-3.16(m,2H),2.86-2.66(m,4H),2.57(s,1.5H),2.56(s,1.5H),1.40-1.32(m,6H).

[0787] Example 34 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0788] Step 1: (R)-4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-methylpiperazine-1-carboxylic acid tert-butyl ester 34a

[0789] Compound 21e (500 mg, 0.74 mmol), compound 22b (236 mg, 0.89 mmol), Pd(dppf)Cl2 (32 mg, 0.04 mmol), and sodium carbonate (229 mg, 2.22 mmol) were dissolved in 1,4-dioxane (20 mL) and H2O (4 mL). The atmosphere was replaced with nitrogen three times and the mixture was reacted at 100°C for 4 h. Ethyl acetate (150 mL) and saturated brine (30 mL) were added, the organic phase was separated, and the mixture was concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 34a (480 mg) as a pale yellow solid.

[0790] MS (ESI) m / z 736.5 [M+H] + .

[0791] Step 2 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 34b

[0792] Compound 34a (480 mg, 0.65 mmol) was dissolved in DCM (10 mL), and HCl-EtOH (2 mL, 4 M) was added. The mixture was allowed to react at room temperature for 3 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to give crude compound 34b (180 mg).

[0793] MS (ESI) m / z 636.4 [M+H] + .

[0794] Step 3 (R)-2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidine-4(7H)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 34c

[0795] Compound 34b (110 mg, 0.17 mmol) was dissolved in DMF (2 mL), and compound 1e (42 mg, 0.17 mmol), triethylamine (52 mg, 0.51 mmol), EDCI (33 mg, 0.17 mmol), and HOBt (23 mg, 0.17 mmol) were added. The mixture was allowed to react at room temperature for 16 h. After completion of the reaction, as monitored by TLC, ethyl acetate (50 mL) was added, and the organic phase was washed three times with saturated brine (20 mL x 3). The organic phase was concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (DCM / MeOH = 30 / 1) to afford compound 34c (86 mg) as a pale yellow solid.

[0796] MS (ESI) m / z 862.6 [M+H] + .

[0797] Step 4 (R)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 34

[0798] Compound 34c (86 mg, 0.1 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure, and ethyl acetate (50 mL) and saturated aqueous sodium bicarbonate were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography (DCM / MeOH = 10 / 1) to obtain compound 34 (25.2 mg).

[0799] MS (ESI) m / z 772.5 [M+H] + .

[0800] 1 H NMR (400MHz, DMSO-d6) δ10.39(s,1H),8.53(s,1H),8.15(s,1H),8.05(d,J=8.6Hz,1H),7.96(s,1H),7.71(d,J=8.0Hz,1H),7.52(s,1H ),5.33(s,2H),4.76(s,2H),3.87(m,2H),3.69(m,2H),3.53(m,3H),3.20(m,3H),2.96(m,2H),2.70(m,2H),2.42(m,2H),1.23(m,6H).

[0801] Example 35 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0802] Step 1: (S)-tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-3-methylpiperazine-1-carboxylate 35b

[0803] Compound 35a was prepared according to patent WO2022249060. Compound 35a (235 mg, 0.35 mmol), compound 22b (111.1 mg, 0.42 mmol), Pd(dppf)Cl2 (25.5 mg, 0.03 mmol), sodium carbonate (110.7 mg, 1.05 mmol), 1,4-dioxane (5 mL), and H2O (1 mL) were added to a reaction flask. The atmosphere was replaced with argon three times and the reaction was carried out at 100°C for 4 h. Ethyl acetate (50 mL) and saturated brine (10 mL) were added, and the mixture was extracted. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 35b (100 mg) as a light yellow solid.

[0804] MS (ESI) m / z 680.2 [M–C4H7] + .

[0805] Step 2 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 35c

[0806] Compound 35b (100 mg, 0.13 mmol) was dissolved in DCM (4 mL) and TFA (3 mL) was added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated, ethyl acetate (50 mL) was added, and the mixture was adjusted to alkaline with saturated sodium bicarbonate. Extraction was performed, and the organic phase was washed with saturated brine and concentrated under reduced pressure to afford crude compound 35c (60 mg).

[0807] MS (ESI) m / z 636.1 [M+H] + .

[0808] Step 3 (S)-2-(6-(4-(5-benzyloxy)-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 35d

[0809] Compound 35c (60 mg, 0.09 mmol) was dissolved in THF (2 mL), and compound 1e (25.32 mg, 0.10 mmol), DIPEA (61 mg, 0.47 mmol), and T3P (45 mg, 0.14 mmol, 50% in EA) were added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, saturated sodium bicarbonate solution was added to quench the reaction. The mixture was extracted with ethyl acetate (50 mL). The organic phase was washed three times with saturated brine and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (DCM / MeOH = 30 / 1) to afford compound 35d (68 mg) as a pale yellow solid.

[0810] MS (ESI) m / z 862.3 [M+H] + .

[0811] Step 4 (S)-N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-2-methylpiperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 35

[0812] Compound 35d (68 mg, 0.07 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure, ethyl acetate (50 mL) was added, and the pH was adjusted to 8 with saturated aqueous sodium bicarbonate solution. Extraction was performed, and the organic phase was washed with saturated brine and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography (EtOAc) to obtain compound 35 (27.3 mg).

[0813] MS (ESI) m / z 772.2 [M+H] + .

[0814] 1H NMR (400MHz, CDCl3) δ11.77(s,1H),9.02(s,1H),8.58(d,J=9.4Hz,1H),8.44(d,J=8.8Hz, 1H),7.61(s,1H),7.57(s,1H),7.51(dd,J=8.8,2.0Hz,1H),5.65-5.56(m,1H),5.27(d,J= 16.0Hz,1H),5.15(d,J=16.0Hz,1H),4.81(s,2H),4.75(m,1H),3.97-3.85(m,3H),3.72(m ,1H),3.55-3.35(m,2H),3.09-2.92(m,2H),2.74(m,3H),2.56(s,3H),1.34-1.27(m,6H).

[0815] Example 36 N-(2-chloro-4-fluorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0816] Step 1: 2-Bromo-N-(2-chloro-4-fluorophenyl)acetamide 36b

[0817] Compound 36a (2 g, 13.7 mmol), 2-bromoacetyl bromide (3.0 g, 15 mmol), and DMAP (2.5 g, 20.4 mmol) were dissolved in DCM (40 mL). The atmosphere was replaced with argon three times, and the reaction mixture was stirred in an ice bath for 4 h. Ethyl acetate (150 mL) and saturated brine (50 mL) were added, followed by extraction. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (PE / EtOAc = 5 / 1) to afford compound 36b (2.87 g) as a white solid.

[0818] MS (ESI) m / z 266.1 [M+H] + .

[0819] Step 2 4-(2-bromo-4-(2-(2-chloro-4-fluorophenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 36c

[0820] Compound 36b (2.05 g, 4.8 mmol), compound 5c (1.4 g, 5.28 mmol), DIEA (1.86 g, 14.4 mmol), and acetonitrile (100 mL) were added to a reaction flask. The atmosphere was replaced with argon three times and the reaction was allowed to proceed at room temperature for 3 h. After completion of the reaction, as monitored by TLC, ethyl acetate (150 mL) and saturated brine (50 mL) were added and extracted. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to afford compound 36c (1.1 g) as a white solid.

[0821] MS (ESI) m / z 556.2 [M–C4H7] + .

[0822] Step 3 4-(2-bromo-4-(2-(2-chloro-4-fluorophenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 36d

[0823] Compound 36c (1 g, 3.7 mmol), compound 22b (326 mg, 1.2 mmol), Pd(dppf)Cl2 (99 mg, 0.13 mmol), and sodium carbonate (433 mg, 4 mmol) were dissolved in 1,4-dioxane (50 mL) and H2O (10 mL). The atmosphere was replaced with argon three times and the mixture was reacted at 100°C for 4 h. Ethyl acetate (150 mL) and saturated brine (50 mL) were added, and the mixture was extracted. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (PE / EtOAc = 1 / 1) to obtain compound 36d (408 mg) as a white solid.

[0824] MS (ESI) m / z 616.4 [M–C4H7] + .

[0825] Step 4: N-(2-chloro-4-fluorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 36e

[0826] Compound 36d (200 mg, 0.29 mmol) was dissolved in DCM (4 mL) and TFA (2.5 mL) was added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, the mixture was concentrated under reduced pressure. Ethyl acetate (50 mL) and saturated aqueous sodium bicarbonate were added, and the mixture was extracted. The organic phase was washed with saturated brine and concentrated under reduced pressure to afford crude compound 36e (170 mg).

[0827] MS (ESI) m / z 572.4 [M+H] + .

[0828] Step 5 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-fluorophenyl)acetamide 36f

[0829] Compound 36e (120 mg, 0.20 mmol) was dissolved in THF (10 mL), and compound 1e (56.3 mg, 0.23 mmol), DIPEA (135 mg, 1.04 mmol), and T3P (100 mg, 0.31 mmol, 50% in EA) were added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, saturated sodium bicarbonate solution was added to quench the reaction. The mixture was extracted with ethyl acetate (50 mL). The organic phase was washed three times with saturated brine and concentrated under reduced pressure to afford the crude product. The crude product was purified by column chromatography (DCM / MeOH = 30 / 1) to afford compound 36f (30 mg) as a pale yellow solid.

[0830] MS (ESI) m / z 798.6 [M+H] + .

[0831] Step 6: N-(2-chloro-4-fluorophenyl)-2-(2-(4,7-dihydro-5H-thieno[2,3-c]pyran-2-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 36

[0832] Compound 36f (30 mg, 0.03 mmol) was dissolved in trifluoroacetic acid (2 mL) and reacted at 50°C for 4 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure, ethyl acetate (50 mL) was added, and the pH was adjusted to 8 with saturated aqueous sodium bicarbonate solution. Extraction was performed, and the organic phase was washed with saturated brine and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography (EtOAc) to obtain compound 36 (8.8 mg).

[0833] MS (ESI) m / z 708.5 [M+H] +

[0834] 1H NMR (400MHz, CDCl3) δ11.80(s,1H),8.74(s,1H),8.59(s,1H),8.23(m,1H),7.62(s ,1H),7.11(d,J=7.6Hz,1H),7.00(m,1H),5.66(m,1H),5.35(m,1H),5.10(s,2H),4 .84(s,2H),3.98(m,2H),3.82(m,2H),3.49(m,1H),3.25-3.15(m,2H),3.07(m,1H) ,2.85(m,1H),2.78(m,2H),2.56(s,3H),2.24-2.19(m,1H),1.33(t,J=7.6Hz,3H).

[0835] Example 37 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0836] Step 1: tert-Butyl 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-5-ethyl-7-oxo-2-(5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 37b

[0837] Compound 37a (0.5 g, 1.71 mmol), 1g (0.94 g, 1.42 mmol), Pd(dppf)Cl2.DCM (0.09 g, 0.11 mmol) and sodium carbonate (0.35 g, 3.3 mmol) were dissolved in 1,4-dioxane (20 mL) and water (4 mL), and the atmosphere was replaced with nitrogen three times. The reaction was carried out at 100°C for 6 hours, and water (20 mL) was added to dilute the mixture. The mixture was extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 37b (0.64 g) as a white solid.

[0838] MS (ESI) m / z 748.2 [M+H] +

[0839] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-7-oxo-6-(piperazin-1-yl)-2-(5'H,7'H-spiro[cyclopropane]-1,4'-thieno[2,3-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 37c

[0840] Compound 37b (0.2 g, 0.26 mmol) was dissolved in dry dichloromethane (5 mL), and trifluoroacetic acid (5 mL) was added. The mixture was reacted at room temperature for 3 h, and then concentrated under reduced pressure. Ethyl acetate and saturated sodium bicarbonate were added to adjust the aqueous solution. The mixture was extracted with ethyl acetate three times, and the organic phases were combined and concentrated to obtain compound 37c (173 mg).

[0841] MS (ESI) m / z 648.1 [M+H] +

[0842] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(5'H,7'H-spiro)[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 37d

[0843] Compound 37c (173 mg, 0.26 mmol) was dissolved in THF (10 mL), and compound 1e (78 mg, 0.32 mmol), triethylamine (270 mg, 2.67 mmol), and T3P (250 mg, 0.39 mmol, 50% wt in EA) were added. The reaction was allowed to react at room temperature for 3 h. The reaction was quenched by the addition of saturated sodium bicarbonate solution, and the mixture was extracted with ethyl acetate (50 mL x 3). The organic phases were combined and concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography to obtain compound 37d (39.8 mg).

[0844] MS (ESI) m / z 874.2 [M+H] +

[0845] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(5'H,7'H-spiro[cyclopropane-1,4'-thieno[2,3-c]pyran]-2'-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 37

[0846] Compound 37d (39.8 mg, 0.045 mmol) was dissolved in trifluoroacetic acid (10 mL) and reacted at 50°C for 4 h. The mixture was concentrated under reduced pressure to obtain a crude product. Ethyl acetate and saturated aqueous sodium bicarbonate solution were added, and the product was extracted three times with ethyl acetate (20 mL). The organic phases were combined, concentrated under reduced pressure, and purified by column chromatography to obtain compound 37 (10 mg).

[0847] MS (ESI) m / z 784.2 [M+H] +

[0848] 1 H NMR (400MHz, CDCl3) δ11.79(s,1H),9.04(s,1H),8.59(s,1H),8.48(d,J=8.6Hz,1H),7. 62(s,1H),7.53(d,J=8.6Hz,1H),7.32(s,1H),5.67(d,J=13.2Hz,1H),5.13(s,2H),4.9 3(s,2H),4.79(d,J=13.2Hz,1H),3.81(t,J=11.7Hz,2H),3.73(s,2H),3.49(m,1H),3.2 0(m,2H),3.07(m,1H),2.80(m,2H),2.56(s,3H),1.34(m,3H),1.01(m,2H),0.91(m,2H).

[0849] Example 38 2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-fluoro-4-(trifluoromethyl)phenyl)acetamide

[0850] Step 1: tert-Butyl 4-(5-ethyl-4-(2-((2-fluoro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylate 38b

[0851] Compound 38a (200 mg, 0.31 mmol) and compound 2a (123 mg, 0.465 mmol) were dissolved in 1,4-dioxane (4 mL) and water (0.4 mL), and 1,1-bis(diphenylphosphino)ferrocenepalladium dichloride dichloromethane complex (24.5 mg, 0.03 mmol) and sodium carbonate (98.5 mg, 0.93 mmol) were added. The atmosphere was replaced with nitrogen three times, and the reaction was carried out at 100°C for 16 h. The mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (10 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography to obtain compound 38b (53 mg) as a yellow-brown solid.

[0852] MS (ESI) m / z 647.3 [M–C4H7] + .

[0853] Step 2: 2-(5-ethyl-7-oxo-6-(piperazin-1-yl)-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-fluoro-4-(trifluoromethyl)phenyl)acetamide 38c

[0854] Compound 38b (53 mg, 0.075 mmol) was dissolved in dry dichloromethane (1 mL), trifluoroacetic acid (1 mL) was added, and the mixture was reacted at room temperature for 1 h. The mixture was concentrated under reduced pressure, and the residue was dissolved in dichloromethane (5 mL) and concentrated under reduced pressure. This process was repeated three times to obtain compound 38c (50 mg) as a yellow-brown solid.

[0855] MS (ESI) m / z 604.4 [M+H] + .

[0856] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-fluoro-4-(trifluoromethyl)phenyl)acetamide 38d

[0857] Compound 38c (50 mg, 0.082 mmol) was dissolved in DMF (2 mL), and compound 1e (23.92 mg, 0.098 mmol) and triethylamine (52.99 mg, 0.41 mmol) were added. The reaction solution was cooled to 0°C, and a 50% ethyl acetate solution of 1-propylphosphoric acid cyclic anhydride (78.27 mg, 0.123 mmol) was added dropwise. The reaction was allowed to react at room temperature for 2 h. Water (10 mL) was added to quench the reaction, and the mixture was extracted with dichloromethane (10 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified on a silica gel plate (PE / EA = 1 / 2) to afford compound 38d (30 mg) as a pale yellow solid.

[0858] MS (ESI) m / z 830.6 [M+H] + .

[0859] Step 4: 2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-2-(1-oxaspiro[4.5]dec-7-en-8-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-fluoro-4-(trifluoromethyl)phenyl)acetamide

[0860] Compound 38d (30 mg, 0.036 mmol) was dissolved in trifluoroacetic acid (1 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure and purified by preparative liquid chromatography to give compound 38 (15.7 mg) as a white solid.

[0861] MS (ESI) m / z 740.5 [M+H] + .

[0862] 1 H NMR (400MHz, DMSO-d6) δ10.68(s,1H),8.43(s,1H),8.22(s,1H),7.79(dd,J=8.0Hz,1 H),7.56(d,J=8.0Hz,1H),6.71(s,1H),5.29(s,2H),4.52(d,J=8Hz,1H),3.73(t,J=8 Hz,2H),3.60-3.43(m,4H),3.02-2.93(m,3H),2.82-2.79(m,2H),2.68-2.59(m,2H), 2.40(s,3H),2.27(s,2H),1.92-1.85(m,2H),1.79-1.63(m,4H),1.17(t,J=8Hz,3H).

[0863] Example 39 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-((3aR,6aS)-5-(5-hydroxy-6-methylpyrimidine-4-carbonyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0864] Step 1: (3aR,6aS)-5-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran)-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)hexahydropyrrolo[3,4-c]pyrrole-2(1H)-carboxylic acid tert-butyl ester 39a

[0865] Compound 4a (95 mg, 0.17 mmol) was dissolved in DMSO (1 mL), and cis-2-Boc-hexahydropyrrolo[3,4-c]pyrrole (360 mg, 1.7 mmol) and AgBF4 (33 mg, 0.17 mmol) were added. The mixture was reacted at 130°C for 3 hours under N2 protection. After completion of the reaction, the organic phase was concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (DCM / MeOH = 10 / 1) to afford compound 39a (35 mg).

[0866] MS (ESI) m / z 692.2 [M+H] + .

[0867] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-((3aR,6aS)-hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 39b

[0868] Compound 39a (35 mg, 0.05 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The mixture was allowed to react at room temperature for 4 h. After completion of the reaction, as monitored by TLC, the mixture was concentrated under reduced pressure to afford compound 39b (35 mg).

[0869] MS (ESI) m / z 592.2 [M+H] + .

[0870] Step 3: 2-(6-((3aR,6aS)-5-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 39c

[0871] Compound 1e (13.4 mg, 0.55 mmol), EDCI (10.6 mg, 0.55 mmol), HOBt (2.9 mg, 0.55 mmol), and DIEA (19 mg, 150 mmol) were dissolved in DMF (3 mL). Compound 39b (35 mg, 0.5 mmol) was added and allowed to react at room temperature for 16 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain the crude product, which was then purified by column chromatography (DCM / MeOH = 10:1) to afford compound 39c (22 mg).

[0872] MS (ESI) m / z 818.3 [M+H] + .

[0873] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-((3aR,6aS)-5-(5-hydroxy-6-methylpyrimidine-4-carbonyl)hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 39

[0874] Compound 39c (22 mg, 0.03 mmol) was dissolved in TFA (2 mL) and reacted at 50°C for 16 h. After completion of the reaction as monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain a crude product, which was then purified by column chromatography (DCM / MeOH = 10:1) to afford compound 39 (6.4 mg).

[0875] MS (ESI) m / z 728.4 [M+H] + .

[0876] 1H NMR (400MHz, CDCl3) δ12.73(s,1H),8.98(s,1H),8.61(s,1H),8.47(d,J=8.8Hz,1H),7.6 3(s,1H),7.53(d,J=8.4Hz,1H),6.96(s,1H),5.35(m,1H),5.07(s,2H),4.53(m,2H),4.4 0(m,1H),4.36(m,2H),4.06-4.00(m,1H),3.91(t,J=5.2Hz,2H),3.68(m,1H),3.18(m,1H ),3.04(m,3H),2.72(m,2H),2.55(s,3H),2.22(t,J=7.6Hz,1H),2.02(m,1H),1.31(t,3H)

[0877] Example 40 2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)acetamide

[0878] Step 1: N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)-1,1-diphenylmethanamine 40b

[0879] The synthesis of compound 40a is described in reference to WO2023000834 A1. Compound 40a (1.1 g, 3.957 mmol) and diphenylmethaneimine (788.89 mg, 4.353 mmol) were dissolved in toluene (10 mL). 1,1'-binaphthyl-2,2'-bisdiphenylphosphine (108.97 mg, 0.119 mmol), tris(dibenzylideneacetone)dipalladium (147.57 mg, 0.237 mmol), and sodium tert-butoxide (1.14 g, 11.871 mmol) were added. The atmosphere was purged with nitrogen three times, and the mixture was reacted at 110°C for 18 h. The mixture was cooled to room temperature and filtered through celite. The filter cake was washed with ethyl acetate (10 mL x 3). The filtrates were combined and concentrated under reduced pressure to give crude compound 40b (1.3 g) as a yellow-brown oil.

[0880] MS (ESI) m / z 380.1 [M+H] +

[0881] Step 2: 2-Methyl-4-(trifluoromethyl)benzofuran-7-amine 40c

[0882] Compound 40b (1.3 g, 3.428 mmol) was dissolved in 2 M hydrochloric acid-ethanol solution (20 mL), reacted at 50°C for 3 h, cooled to room temperature, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE / EA = 1 / 1) to give crude compound 40c (420 mg) as a yellow oil.

[0883] MS (ESI) m / z 216.1 [M+H] +

[0884] Step 3 2-Bromo-N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)acetamide 40d

[0885] Compound 40c (420 mg, 1.953 mmol) and DMAP (238.60 mg, 1.95 mmol) were dissolved in dry dichloromethane (5 mL), cooled to about 0°C, and bromoacetyl bromide (473.11 mg, 2.344 mmol) was added dropwise. The temperature was returned to room temperature and the reaction was allowed to react for 2 h. Water (5 mL) was added to dilute the mixture, and the mixture was extracted with dichloromethane (5 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated on a silica gel plate (PE / EA = 20 / 1) to give compound 40d (96 mg) as a light yellow solid.

[0886] MS (ESI) m / z 336.8 [M+H] +

[0887] Step 4: 4-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-4-(2-((2-methyl-4-(trifluoromethyl)benzofuran-7-yl)amino)-2-oxoethyl)-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)piperazine-1-carboxylic acid tert-butyl ester 40e

[0888] Compound 40d (98.65 mg, 0.2293 mmol) and compound 5c (96 mg, 0.2866 mmol) were dissolved in dry acetonitrile (3 mL), and DIEA (88.83 mg, 0.6879 mmol) was added. The mixture was reacted at room temperature for 18 h. The reaction solution was concentrated under reduced pressure, and the crude product was separated on a silica gel plate (PE / EA = 1 / 1) to give compound 40e (80 mg) as a light yellow solid.

[0889] MS (ESI) m / z 686.3 [M+H] +

[0890] Step 5: 2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-6-(piperazin-1-yl)-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)acetamide 40f

[0891] Compound 40e (80 mg, 0.1167 mmol) was dissolved in dry dichloromethane (1 mL), and trifluoroacetic acid (1 mL) was added. The mixture was allowed to react at room temperature for 2 h. The reaction solution was concentrated under reduced pressure, and saturated sodium bicarbonate (2 mL) was added to the residue. The mixture was extracted with dichloromethane (3 mL x 3). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford crude compound 40f (70 mg) as a reddish-brown solid.

[0892] MS (ESI) m / z 586.3 [M+H] + .

[0893] Step 6: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazine-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)acetamide 40g

[0894] Compound 40f (70 mg, 0.1196 mmol), compound 1e (43.79 mg, 0.1794 mmol), and DIEA (77.29 mg, 0.598 mmol) were dissolved in DMF (1 mL). The reaction solution was cooled to 0°C, and a 50% ethyl acetate solution of 1-propylphosphoric acid cyclic anhydride (76.11 mg, 0.1196 mmol) was added dropwise. The mixture was allowed to warm to room temperature and react for 2 h. Water (2 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (3 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to afford compound 40g (70 mg) as a reddish-brown solid, which was used directly in the next step.

[0895] MS (ESI) m / z 812.3 [M+H] + .

[0896] Step 9: 2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-methyl-4-(trifluoromethyl)benzofuran-7-yl)acetamide 40

[0897] Compound 40g (70 mg, 0.025 mmol) was dissolved in trifluoroacetic acid (1 mL), and the mixture was reacted at 50°C for 1 h. The reaction solution was concentrated under reduced pressure, and the crude product was separated by preparative liquid chromatography to obtain compound 40 (10.7 mg) as a white solid.

[0898] MS (ESI) m / z 722.3 [M+H] + .

[0899] 1 H NMR (400MHz, DMSO-d6) δ11.02(s,1H),8.40(s,1H),7.96(d,J=8.0Hz,1H),7.51(d,J= 8.0Hz,1H),6.81(s,1H),6.78(s,1H),5.36(s,2H),4.53(d,J=8.0Hz,1H),4.23(d,J=8 .0Hz,1H),3.79(t,J=8Hz,2H),3.56-3.42(m,4H),3.26-3.21(m,3H),3.02-2.95(m,3 H),2.82(d,J=8.0Hz,1H),2.63(s,1H),2.56(s,3H),2.39(s,3H),1.19(t,J=8Hz,3H).

[0900] Example 41 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-(methoxymethyl)-piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0901] Step 1: 4-(4-(2-((2-chloro-4-(trifluoromethyl)phenyl)amino)-2-oxoethyl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidin-6-yl)-2-(methoxymethyl)piperazine-1-carboxylic acid tert-butyl ester 41b

[0902] Compound 4a (400 mg, 713.32 μmol) and compound 41a (1.6 g, 7.13 mmol) were dissolved in NMP (0.03 mL), and AgO (330 mg, 1.43 mmol) was added. The mixture was deoxygenated three times and reacted at 140°C for 2 h. The reaction solution was cooled to room temperature, dissolved in EtOAc (30 mL), and washed with saturated aqueous NH4Cl (20 mL). The organic phase was separated, and the crude product was separated by preparative liquid chromatography to afford compound 41b (42 mg) as a white solid.

[0903] MS (ESI) m / z 710.2 [M+H] +

[0904] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(3-(methoxymethyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4-(7H)-yl)acetamide 41c

[0905] Compound 41b (41 mg, 57.73 μmol) was dissolved in DCM (1 mL) and TFA (1 mL) was added. The mixture was allowed to react at room temperature for 0.5 h. The reaction mixture was concentrated under reduced pressure, and DCM (20 mL) and saturated aqueous NaHCO₃ (5 mL) were added. The organic phase was separated and concentrated under reduced pressure to afford compound 41c (42 mg) as a yellow solid.

[0906] MS (ESI) m / z 610.3 [M+H] +

[0907] Step 3: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)-3-(methoxymethyl)piperazin-1-yl)-2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4-(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 41d

[0908] Compound 41c (42 mg, 68.85 μmol) and compound 1e (20 mg, 82.62 μmol) were dissolved in DMF (1 mL). DIPEA (44 mg, 344.24 μmol) and T3P (50% wt in EtOAc, 74 mg, 103.27 μmol) were added and reacted at room temperature for 0.5 h. The reaction mixture was concentrated under reduced pressure, and DCM (15 mL) and saturated aqueous NaHCO3 (10 mL) were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product, which was purified by column chromatography to afford compound 41d (58 mg) as a yellow solid.

[0909] MS (ESI) m / z 836.3 [M+H] +

[0910] Step 4: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)-3-(methoxymethyl)-piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 41

[0911] Compound 41d (58 mg, 69.36 μmol) was dissolved in TFA (2 mL) and reacted at 50°C for 3 h. The reaction solution was concentrated under reduced pressure, and DCM (10 mL) and saturated aqueous NaHCO3 (5 mL) were added. The organic phase was separated and concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative liquid chromatography to afford compound 41 (15.1 mg) as a light yellow solid.

[0912] MS (ESI) m / z 746.2 [M+H] +

[0913] 1 H NMR(400MHz,DMSO-d6)δ10.36(br s,2H),8.54(d,J=4.4Hz,1H),8.05(d,J=8.8Hz,1H),7.97(d,J=1.6Hz,1H),7.72(d,J=8.8Hz,1H),6.83(s,1H),5.37-5.26(m,2H), 4.78-4.73(m,0.5H),4.44-4.37(m,0.5H),4.26(s,1H),4.25(s,1H),3.92(dd,J=17.6,8.8Hz,1H),3.80(t,J=5.2Hz,2H),3.66(dt ,J=11.6,3.6Hz,1H),3.59-3.48(m,2H),3.43-3.37(m,1H),3.32(s,3H),3.17(m,0.5H),3.11-3.05(m,1H),2.93(d,J=11.8Hz,0.5 H),2.88-2.77(m,2H),2.62(d,J=10.8Hz,0.5H),2.52(m,2H),2.44(s,1.5H),2.43(s,1.5H),1.99(m,0.5H),1.20(t,J=7.6Hz,3H)

[0914] Example 42 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(2-(3,6-dihydro-2H-pyran-4-yl)-5-ethyl-6-(4-(4-hydroxy-1,5,6-trimethyl-2-oxo-1,2-dihydropyridine-3-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0915] Compound 42a (190 mg, 0.96 mmol) was dissolved in DMF (5 mL). EDCI (76 mg, 0.40 mmol), HOBt (54 mg, 0.40 mmol), and DIEA (154 mg, 1.20 mmol) were added at room temperature, followed by compound 6f. The reaction was allowed to react at 50°C for 16 h. After completion of the reaction, as monitored by TLC, the reaction solution was concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative liquid chromatography to afford compound 42 (1.4 mg).

[0916] MS (ESI) m / z 745.2 [M+H] + .

[0917] 1 H NMR (400MHz, CDCl3) δ8.99 (s, 1H), 8.50 (d, J = 8.6Hz, 1H), 7.63 (s, 1H), 7.52 (d ,J=8.6Hz,1H),7.52(s,1H),6.96(s,1H),5.35(m,2H),5.08(s,2H),4.36(d,J =2.7Hz,2H),3.91(t,J=5.4Hz,2H),3.65(m,2H),3.51(s,3H),3.16(m,2H),2. 73(m,4H),2.34(s,3H),2.22(t,J=7.6Hz,2H),2.07(s,3H),1.35-1.31(m,3H)

[0918] Example 43 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0919] Step 1: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 43c

[0920] Compound 43b (80 mg, 0.29 mmol) was dissolved in 1,4-dioxane (4 mL) and water (0.5 mL). Compound 43a (190 mg, 0.24 mmol), potassium phosphate (153 mg, 0.72 mmol), and Pd(dppf)Cl2 (22 mg, 0.03 mmol) were added. The atmosphere was purged with nitrogen three times, and the mixture was reacted at 100°C for 3 hours. After cooling, the reaction solution was diluted with ethyl acetate (30 mL) and washed three times with saturated brine. The organic phase was concentrated under reduced pressure to obtain a crude product. The crude product was separated and purified by column chromatography (DCM / MeOH = 10:1) to obtain compound 43c (90 mg).

[0921] MS (ESI) m / z 861.5 [M+H] + .

[0922] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 43

[0923] Compound 43c (90 mg, 0.10 mmol) was dissolved in TFA (5 mL) and reacted at 50°C for 3 h. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative HPLC to afford compound 43 (38.2 mg) as a white solid.

[0924] MS (ESI) m / z 771.3 [M+H] + .

[0925] 1H NMR(400MHz,MeOD-d4)δ8.55(s,1H),8.16(d,J=8.5Hz,1H),7.82(s,1H),7.64-7.5 9(m,2H),5.42(s,2H),4.72(d,J=11.9Hz,2H),4.23(s,2H),4.11(d,J=13.0Hz,1H), 3.74(br.,2H),3.47(d,J=13.9Hz,1H),3.17-3.06(m,4H),3.04-2.95(m,3H),2.86(s,3H),2.80(m,1H),2.52(s,3H),1.36-1.31(m,3H).

[0926] Example 44 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(5-methyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0927] Step 1: 2-(6-(4-(5-(benzyloxy)-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-5-ethyl-2-(5-methyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)-N-(2-chloro-4-(trifluoromethyl)phenyl)acetamide 44b

[0928] Compound 44a (50 mg, 0.2537 mmol) and compound 43a (139.77 mg, 0.2114 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL), and Pd(dppf)Cl2.CH2Cl2 (17.31 mg, 0.0212 mmol) and sodium carbonate (67.23 mg, 0.6343 mmol) were added. The atmosphere was replaced with nitrogen three times, and the temperature was raised to 100°C for 18 h. The mixture was cooled to room temperature, diluted with water (5 mL), and extracted with ethyl acetate. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by column chromatography (DCM / MeOH = 10:1) to give compound 44b (42 mg) as a light yellow solid.

[0929] MS (ESI) m / z 861.3 [M+H] + .

[0930] Step 2: N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(5-methyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide 44

[0931] Compound 44b (42 mg, 0.0488 mmol) was dissolved in trifluoroacetic acid (1 mL), heated to 50°C for 3 h, and the reaction solution was concentrated under reduced pressure. The crude product was separated by preparative HPLC to obtain compound 44 (15.7 mg) as a light yellow solid.

[0932] MS (ESI) m / z 771.2 [M+H] + .

[0933] 1 H NMR (400MHz, DMSO-d6) δ10.40(s,1H),8.59(s,1H),8.15(s,1H),8.06(d,J=8.0H z,1H),7.98(d,J=8.0Hz,1H),7.72(dd,J=8.0Hz,1H),7.48(s,1H),5.35(s,2H),4 .53(d,J=8Hz,1H),3.60(s,2H),3.54-3.46(m,4H),3.05-2.96(m,4H),2.92(d,J =8.0Hz,2H),2.83(d,J=12Hz,3H),2.48(s,3H),2.45(s,3H),1.20(t,J=8Hz,3H).

[0934] Example 45 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(5-(methyl-d3)-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0935] The synthesis of intermediate 45a was based on intermediate 43b, except that methyl p-toluenesulfonate was replaced by D3-p-toluenesulfonylmethyl ester and 4,5,6,7-tetrahydrothieno[2,3-c]pyridine hydrochloride (43b-1) was replaced by 4,5,6,7-tetrahydrothieno[3,2-c]pyridine hydrochloride (44a-1); MS (ESI) m / z 283.2 [M+H] + .

[0936] Example 45 was synthesized by referring to Example 44, except that intermediate 44a was replaced with 5-(methyl-d3)-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine (45a).

[0937] MS (ESI) m / z 774.3 [M+H] + .

[0938] 1H NMR (400MHz, CDCl3) δ9.06(s,1H),8.59(s,1H),8.45(d,J=8.7Hz,1H),7.62(d,J=2.0Hz,1H),7.52(d,J=8.8Hz,1H),7.49(s,1H),5.67(d,J=13.2Hz ,1H),5.16(s,2H),4.78(s,1H),4.01(s,2H),3.80(m,2H),3.49(m,2H),3 .27(s,2H),3.16(s,4H),2.81(m,2H),2.57(s,3H),1.33(t,J=7.5Hz,3H).

[0939] Example 46 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-2-(5-ethyl-4,5,6,7-tetrahydrothieno[3,2-c]pyridin-2-yl))-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0940] The synthesis of intermediate 46a was based on intermediate 43b; methyl p-toluenesulfonate was replaced by ethyl p-toluenesulfonate and 4,5,6,7-tetrahydrothieno[2,3-c]pyridine hydrochloride (43b-1) was replaced by 4,5,6,7-tetrahydrothieno[3,2-c]pyridine hydrochloride (44a-1); MS (ESI) m / z 294.2 [M+H] + .

[0941] Synthesis of Example 46 Referring to Example 44, intermediate 44a was replaced with 5-ethyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine (46a).

[0942] MS (ESI) m / z 785.3 [M+H] + .

[0943] 1H NMR(400MHz, CDCl3)δ9.07(s,1H),8.60(s,1H),8.48(d,J=8.63Hz,1H),8.20(br.,1H),7.63(s,1H),7.57-7.50(m,2H),5.66(br.,1 H),5.15(s,2H),4.81(m,1H),3.94-3.75(m,4H),3.26-3.01(m,8H),2.92(m,2H),2.84-2.78(m,2H),2.58(s,3H),1.39-1.28(m,6H).

[0944] Example 47 N-(2-chloro-4-(trifluoromethyl)phenyl)-2-(5-ethyl-6-(4-(5-hydroxy-6-methylpyrimidine-4-carbonyl)piperazin-1-yl)-2-(6-(methyl-d3)-4,5,6,7-tetrahydrothieno[2,3-c]pyridin-2-yl)-7-oxo-[1,2,4]triazolo[1,5-a]pyrimidin-4(7H)-yl)acetamide

[0945] The synthesis of intermediate 47a was based on intermediate 43b, except that methyl p-toluenesulfonate was replaced with p-D3-toluenesulfonylmethyl ester; MS (ESI) m / z 283.2 [M+H] + .

[0946] The synthesis of Example 47 was carried out with reference to Example 44; intermediate 44a was replaced with 6-(methyl-d3)-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine (47a).

[0947] MS (ESI) m / z 774.3 [M+H] + .

[0948] 1 H NMR (400MHz, DMSO-d6) δ10.40(s,1H),10.24(s,1H),8.59(s,1H),8.06(d,J=8.0 Hz,1H),7.98(d,J=8.0Hz,1H),7.73(dd,J=8.0Hz,1H),7.61(s,1H),5.35(s,2H) ,4.53(d,J=8Hz,1H),3.51(d,J=8Hz,3H),3.35~3.42(m,4H),3.32~3.30(m,4H), 3.01(s,2H),2.84(d,J=8Hz,1H),2.65(s,1H),2.45(s,3H),1.20(t,J=8Hz,3H).

[0949] NSC617145 was purchased from a commercial manufacturer, Bidex Pharmaceuticals, catalog number: BD628938

[0950] The synthesis of control compound 1 refers to patent WO2022249060A1.

[0951] Test Example 1: Test of compound activity against WRN (517-1238) enzyme

[0952] This experiment studies the inhibitory effect of compounds on WRN (517-1238) enzyme activity by detecting the effects of compounds on WRN (517-1238) enzyme activity in vitro. NSC617145 is the internal control compound in this project.

[0953] Experimental reagents and consumables

[0954] Compounds were serially diluted in DMSO using a 384 PP Plate. Using an Echo, 0.15 μL of compound was transferred to a 384-well microplate (Corning 4514) to ensure a final DMSO content of 1% (in duplicate). 5 μL of WRN (517-1238) enzyme solution was added to each well of the 384-well microplate and incubated at 25°C for 10 minutes. Wells containing 1% DMSO and enzyme served as high controls, while wells containing the same amount of DMSO and buffer served as low controls. 5 μL of dsDNA solution and 5 μL of ATP solution were added to each well and incubated at 25°C for 20 minutes (final concentrations: 1 nM WRN, 5 nM dsDNA, 50 μM ATP). Fluorescence signals were read on a BMG (CLARIO Star Plusacu) microplate reader (Ex: 620 nm, Em: 685 nm). The percentage inhibition of compound-treated wells was normalized between high control and low control. % inhibition = ((signal value high control - signal value 化合物处理 ) / (signal value high control-signal value low control)*100). Then, the four-parameter IC was fitted by XLfit 5.5.0. 50 Curve and analyze.

[0955] Table 1 Inhibition results of test compounds on WRN(517-1238)_FI

[0956] As can be seen from the above table, the compounds of the present invention have a good inhibitory effect on WRN(517-1238)_FI.

[0957] Test Example 2: Testing of the compound's inhibitory activity on SW48 cell proliferation

[0958] Complete medium (L-15 + 10% FBS + 1% P / S) was prepared to resuscitate SW48 cells (ATCC). After two passages, cells in the logarithmic growth phase were collected by centrifugation and counted. The cells were resuspended to an appropriate concentration and inoculated into a 96-well plate with 100 μL of cell suspension per well at a seeding density of 5000 cells / well. The 96-well plate was placed in a CO2 incubator (0% CO2) overnight, 50 μL of the upper medium was aspirated, and 40 μL of fresh medium was added. The test compound was prepared as a stock solution in DMSO and serially diluted four-fold with DMSO, starting at 2.5 mM, to create nine concentration gradients. The test compound was diluted 25-fold with culture medium, and 10 μL of each was added to 90 μL of cells in a 96-well plate. Cell-free culture medium (containing 0.4% DMSO) was added to the Min control wells, while 10 μL of a DMSO-cell culture medium mixture (final DMSO concentration of 0.4%) was added to the Max control wells. The cells were incubated for 120 hours at 37°C in a 0% CO2 incubator with a relative humidity of 90% or higher. The reaction was terminated by adding 50 μL / well of CellTiter Glo. The cells were incubated at room temperature in the dark for 10 minutes, gently shaken, and then detected using Envision. The fluorescence (RLU) value of each well was measured, and the cell inhibition rate was calculated using the formula: % cell growth inhibition rate = (1 - As / Ac) × 100. Where As: RLU sample (cells + CTG + test compound) - RLU Min (culture medium without cells), Ac: RLU normal growth cell control (cells + CTG + DMSO) - RLU Min (culture medium without cells). Input the inhibition rate Inh% (Y) corresponding to each concentration (X) in EXCEL, and use Graphpad Prism 8 software according to the built-in four-parameter fitting formula Y = Bottom + (Top-Bottom) / (1 + (IC 50 / X)*HillSlope) to calculate the half-maximal inhibitory concentration IC of each compound 50 value.

[0959] Table 2 Inhibition results of test compounds on SW48 cell proliferation

[0960] The experimental results show that compared with the control compound 1, the compound of the present invention has a good inhibitory effect on the proliferation of SW48 cells.

[0961] Test Example 3: Thermodynamic Solubility Test of Compounds

[0962] Simulated artificial gastrointestinal fluid (FaSSGF (pH = 1.6) and FaSSIF (pH = 6.5)) is a commercial reagent purchased from Biorelevant or other commonly used commercial companies. Accurately weigh approximately 2 mg of compound per pH and add appropriate volumes of simulated artificial gastrointestinal fluid (FaSSGF (pH = 1.6) and FaSSIF (pH = 6.5) to obtain a 2 mg / mL solution. After sonication for 10 minutes, place the tube on a shaker and shake overnight at room temperature (at least 8 hours). After shaking, sonicate for 10 minutes and centrifuge at 13,000 rpm for 15 minutes. Pipette 0.1 mL of the supernatant into a new tube, shake and rinse for 5 minutes, and then discard the liquid. Transfer another 0.5 mL of the supernatant to a new tube, centrifuge at 13,000 rpm for 15 minutes, and remove the supernatant (dilute with water if necessary) for analysis by LC-UV (or LC-MS if UV detection is unavailable). Sample concentrations are quantified using a fitted calibration line (3-5 points).

[0963] Table 3 Thermodynamic solubility test results of test compounds

[0964] The experimental results showed that compared with the control compound 1, the compound of the present invention had better solubility in FaSSGF (pH=1.6) and FaSSIF (pH=6.5) solvents.

[0965] Test Example 4: Pharmacokinetic Test

[0966] Mouse pharmacokinetic studies were conducted using male ICR mice weighing 20-25 g, provided by the Laboratory Animal Management Department of the Shanghai Institute of Family Planning Sciences. Three mice fasted overnight were administered the drug (10 mg / kg) orally by gavage. Blood was collected before dosing, and 15, 30 minutes, and 1, 2, 4, 8, and 24 hours after dosing. Three additional mice were administered the drug (1 mg / kg) intravenously, and blood was collected before dosing, and 15, 30 minutes, and 1, 2, 4, 8, and 24 hours after dosing. Blood samples were centrifuged at 6800 g for 6 minutes at 2-8°C, and plasma was collected and stored at -80°C. Plasma was collected at each time point and mixed with 10-20 times the volume of methanol or acetonitrile containing the internal standard. The mixture was vortexed for 1 minute and centrifuged at 18,000 rpm at 4°C for 10 minutes. The supernatant was directly analyzed by LC-MS / MS. Key pharmacokinetic parameters were analyzed using WinNonlin 7.0 software using a non-compartmental model.

[0967] Table 4 Pharmacokinetic test results of test compounds in mice

[0968] The results of the mouse pharmacokinetic experiment showed that the compounds 28 and 37 of the present invention had high oral exposure, good pharmacokinetic properties and good drugability.

[0969] Test Example 5: Manual patch clamp technique to detect the effect of the test substance on the current of stably overexpressed hERG channels

[0970] 5.1. Test Article Preparation Information

[0971] A small amount of compound 47 was weighed and prepared into a 31.346 mM stock solution in DMSO. The test sample stock solution was sequentially diluted with DMSO to produce 10 mM, 3.33 mM, 1 mM, 0.33 mM, and 0.1 mM dilutions. The test sample dilutions were then diluted sequentially with extracellular fluid containing 0.5% F68 to produce 30 μM, 10 μM, 3 μM, 1 μM, and 0.3 μM working solutions, ensuring a DMSO content of 0.3%. All concentrations were sonicated for 20 min. The solubility of the test sample was visually assessed; all concentrations dissolved completely without visible precipitation.

[0972] Weigh a small amount of control compound I and prepare a 31.311 mM stock solution in DMSO. Dilute the test compound stock solution sequentially with DMSO to 10 mM, 3.33 mM, 1 mM, 0.33 mM, and 0.1 mM dilutions. Dilute the test compound dilutions sequentially with extracellular fluid containing 0.5% F68 to prepare working solutions at 30 μM, 10 μM, 3 μM, 1 μM, and 0.3 μM, ensuring a DMSO concentration of 0.3%. Sonicate all concentrations for 20 minutes. Visually inspect the solubility of the test sample; all concentrations dissolve completely without visible precipitation.

[0973] 3.37mg of the positive control substance Cisapride was prepared into a 10mM stock solution with 682.41μL DMSO. The Cisapride stock solution was diluted with DMSO to 1mM, 333.33μM, 33.33μM, 3.33μM, 0.33μM and 0.033μM dilutions. The five concentrations of the Cisapride dilutions were then diluted with extracellular fluid to prepare working solutions of 1000nM, 100nM, 10nM, 1nM and 0.1nM, ensuring that the DMSO concentration was 0.3%. The solubility of Cisapride was visually inspected and all concentrations were completely dissolved without visible precipitation. 5.2 Cell Culture

[0974] HEK-293 cells stably expressing hERG potassium channel were used. hERG potassium channel cells were purchased from Creacell (Cat. No. A-0320). All operations followed the standard cell culture operating procedures of Beijing Aisiyipu Biotechnology Co., Ltd.

[0975] The cell culture method is as follows:

[0976] HEK-293 cells stably expressing hERG potassium channel were cultured in DMEM medium containing 10% fetal bovine serum and 0.8 mg / mL G418 at a temperature of 37° C. and a carbon dioxide concentration of 5%.

[0977] Cell passaging: Remove old culture medium and wash once with PBS, then add 1 mL of TrypLE TM Express solution, incubate at 37°C for about 0.5 min. When the cells detach from the bottom of the dish, add about 5 mL of complete culture medium preheated at 37°C. Gently pipette the cell suspension to separate the aggregated cells. Transfer the cell suspension to a sterile centrifuge tube and centrifuge at 1000 rpm for 5 min to collect the cells. For expansion or maintenance culture, seed the cells in a 6 cm cell culture dish with 2.5 x 10 cells seeded per dish. 5 cells (final volume: 5 mL). To maintain the electrophysiological activity of the cells, the cell density must not exceed 80%. Before patch clamp testing, the cells were TM Express separation, 4x 10 3 The cells were plated on coverslips and cultured in 24-well plates (final volume: 500 μL). After 18 hours, the assay was performed.

[0978] 5.3 Electrophysiological Recording

[0979] The liquids used were recorded as intracellular and extracellular fluids. The extracellular fluid was stored for 2 weeks. After the intracellular fluid was prepared, it was divided into 1 mL per tube and stored in a -20°C refrigerator. Freshly thawed intracellular fluid was used for each test. All intracellular fluids were used within three months. If the amount exceeded three months, the old intracellular fluid was discarded and re-prepared. The preparation methods of intracellular and extracellular fluids are as follows:

[0980] Extracellular solution: 140 mM NaCl, 3.5 mM KCl, 1 mM MgCl2·6H2O, 2 mM CaCl2·2H2O, 10 mM D-Glucose, 10 mM HEPES, 1.25 mM NaH2PO4·2H2O, pH adjusted to 7.4 with NaOH.

[0981] Intracellular solution: 20 mM KCl, 115 mM K-Aspartic acid, 1 mM MgCl2·6H2O, 5 mM EGTA, 10 mM HEPES, 2 mM Na2-ATP, pH adjusted to 7.2 with KOH.

[0982] 5.4. Patch clamp assay

[0983] The voltage stimulation protocol for whole-cell patch-clamp recording of hERG currents was as follows: after forming a whole-cell seal, the cell membrane voltage was clamped at -80 mV. The clamping voltage was depolarized from -80 mV to -50 mV for 0.5 s (to detect leakage current), then stepped to 30 mV for 2.5 s, and then quickly returned to -50 mV for 4 s to stimulate hERG tail current. Data were collected repeatedly every 10 s to observe the effects of drugs on hERG tail current. Experimental data were acquired using an IPA amplifier (Sutter Instrument) and stored in SutterPatch (with Igor Pro) software.

[0984] The patch clamp procedure begins by pulling a recording electrode from a glass capillary using a microelectrode puller. The electrode, filled with intracellular fluid, is then placed in a microelectrode holder. Under an inverted microscope, the microelectrode manipulator is used to immerse the electrode in the extracellular fluid and record the electrode resistance (Rpip). The electrode is then slowly brought into contact with the cell surface, and negative pressure is applied to create a GΩ seal. Fast capacitance compensation is then performed, and negative pressure is continued to rupture the cell membrane, establishing whole-cell recording mode. Finally, slow capacitance compensation is performed, and experimental parameters such as series resistance (Rs) are recorded. No leakage compensation is performed.

[0985] When the hERG current recorded by the whole cell is stable, the drug is administered. After each drug concentration acts for about 5 minutes (or the current stabilizes), the next concentration is detected. Multiple concentrations are detected for each test compound. The coverslip with cells is placed in a recording bath under an inverted microscope. The blank control external solution and the working solution of the test compound are flowed through the recording bath from low concentration to high concentration by gravity perfusion to act on the cells. A peristaltic pump is used for liquid exchange during recording. The current detected for each cell in the external solution without the compound serves as its own control group. At least two cells are used for each concentration and the test is repeated twice independently. All electrophysiological experiments are performed at room temperature.

[0986] 5.5 Data Analysis

[0987] First, the peak tail current after each drug concentration is compounai ) and blank control tail current (Peak tail current control ) normalized, and then calculated the inhibition rate corresponding to each drug concentration The mean (Mean), standard deviation (SD) and standard error (SE) of the inhibition rate at each concentration were calculated, and the data were expressed as Mean±SE.

[0988] Y=Bottom+(Top-Bottom) / (1+10^((LogIC 50-X)*HillSlope))

[0989] The IC of each compound was calculated using the above equation 50 The dose-effect curve was fitted nonlinearly, where IC 50 IC is the half inhibitory concentration. 50 5.6 Experimental results were calculated and curve fitting was completed using GraphPad Prism software

[0990] The inhibitory effects of compound 47 and control compound I hydrochloride on hERG channels were tested in two independent repeated experiments, and the IC values ​​of the test substances on hERG current were calculated by fitting the concentration-effect curves. 50 The test results are as follows:

[0991] Table 5. Inhibition rate of control compound I and compound 47 on hERG current

[0992] The experimental results show that compared with the control compound I, the compound of the present invention has weaker inhibition on hERG at concentrations of 10 μM and 30 μM and has good safety.

[0993] All documents mentioned in this application are incorporated herein by reference, just as if each document were incorporated herein by reference individually. It should also be understood that after reading the above teachings of the present invention, those skilled in the art may make various changes or modifications to the present invention, and that such equivalents also fall within the scope of the claims appended hereto.

Claims

1. A compound as represented by formula (I), its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, in, X is selected from CH or N; and when X is CH, the CH may be substituted by R5 or R6 (i.e., X is C-R5 or C-R6); m, n, e and q are each independently selected from: 0, 1, 2 or 3; R1 is selected from the following group: (1) R is selected from CH, CD, CR 1e or N; Z and W are independently selected from CH2, CR 1e R 1f NR 1e or O, and Z and W are not CH2 or CR at the same time 1e R 1f ; Among them, R 1e and R 1f Each independently selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl; Y and U are independently selected from CH2 or CR 1e R 1f , and Y and U are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; or Z and W are independently selected from CH2 or NR 1e , and Z and W are not CH2 at the same time; where R 1e Selected from the group consisting of deuterium, C 1-4 Deuterated alkyl; Both Y and U are CR 1e R 1f ; Among them, R 1e and R 1f are independently selected from the following groups: H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, or R 1e and R 1f Together with the carbon atoms to which they are connected, they form a 3-5 membered carbocyclic or heterocyclic ring. The heteroatom contained in the heterocycle is selected from O, N and S; (2) R is selected from CH, CD, CR 1e or N; F is selected from NR 1e or O; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl; K and G are independently selected from CH2, CR 1e R 1f , and K and G are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; R2 is selected from the following group: C 1-4 Alkyl, C 1-4 Deuterated alkyl or C 1-4 Haloalkyl; R3 and R4 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 3-6 Alkoxy, C 6-10 Alkylamino, C 1-4 R3 and R4 can form a 4-6 membered carbocyclic ring or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently substituted by 0, 1, 2 or 3 R7; R5 and R6 are each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Haloalkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, -SO2CF3, -SF5, C 3-6 Cycloalkyl, C 6-10 Alkylamino, C 1-4 Alkenyl, C 2-6 Alkynyl, C 1-4 Halogenated alkenyl, C 2-6 Alkynyl; R5 and R6 can form a 4-6 membered carbocyclic or heterocyclic ring together with the carbon atoms to which they are attached, wherein the heterocyclic ring contains heteroatoms selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above contains 0, 1, 2 or 3 unsaturated double bonds, which are optionally substituted by 0, 1, 2 or 3 R8, respectively and independently; R7 and R8 are each independently selected from the group consisting of hydroxy, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Haloalkoxy, or two R7 or R8 together form =O; Cy A Select from the following group: Among them, the Cy A Can be optionally replaced by one or more R 12 Substituents are substituted; d is selected from 0, 1, 2 or 3; each R 12 Each independently selected from the group consisting of hydroxyl, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Haloalkoxy, or two R 12 Together they form =O; B is selected from O or S; Cy B R3 and R4 are each independently selected from the following groups: H, D, hydroxyl, halogen, cyano, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 6-10 Alkylamino, C 1-4 R3 and R4 can form a 4-6 membered carbocyclic or heterocyclic ring with the carbon atoms to which they are attached, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; the 4-6 membered carbocyclic or heterocyclic ring formed above can be independently and optionally replaced by 0, 1, 2 or 3 R 42 replace; R 42 Each independently selected from the group consisting of hydroxyl, halogen, cyano, C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Haloalkoxy, or two R 42 co-form = O; Wherein, the heteroaryl or heterocyclic group includes 1, 2, 3, 4 or 5 heteroatoms selected from the following group as ring atoms: N, O, S or Se; the carbocyclic ring, heterocyclic ring, heterocyclic group may be a monocyclic ring, a fused ring, a spirocyclic ring or a bridged ring structure, which may be saturated or partially unsaturated, but not aromatic; R 1a and R 1b Each independently selected from the group consisting of H, D, oxo, halogen, cyano, hydroxyl, C 1-4 Alkyl, C 1-4 Haloalkyl, C 1-4 Alkoxy, C 1-4 Halogenated alkoxy, C 3-6 Alkynyl, C 3-6 Alkenyl, C 3-6 Halogenated alkenyl, C 1-6 Alkylamino; R 1b and R 1c Together with the carbon atoms to which they are attached, they can form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; R 1c and R 1d Each independently selected from the following group: H, D, hydroxyl, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Alkoxy, C 1-4 Haloalkoxy, sulfone, C 1-6 Alkylamino; R 1e and R 1f Each independently selected from the group consisting of H, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 3-6 Halogenated cycloalkyl, Methylamino-substituted C 1-4 Alkyl, methoxy substituted C 1-4 alkyl; Each R 1g Each independently selected from the group consisting of H, D, hydroxyl, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Alkoxy, C 1-4 Haloalkoxy; R i C 3-6 Cycloalkoxy; Each R h Each independently selected from the group consisting of H, D, hydroxy, halogen, cyano, C 1-4 Alkyl, C 3-6 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Halogenated alkoxy.

2. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: R1 is selected from the following group: (1) R is selected from CH, CD, CR 1e or N; Z and W are independently selected from CH2, CR 1e R 1f or O, and Z and W are not CH2 or CR at the same time 1e R 1f ; Among them, R 1e and R 1f Each independently selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl; Y and U are independently selected from CH2 or CR 1e R 1f , and Y and U are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; or Z and W are independently selected from CH2 or NR 1e , and Z and W are not CH2 at the same time; where R 1e Selected from the group consisting of deuterium, C 1-4 Deuterated alkyl; Both Y and U are CR 1e R 1f ; Among them, R 1e and R 1f are independently selected from the following groups: H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatoms contained in the heterocyclic ring are selected from O, N and S; (2) R is selected from CH, CD, CR 1e or N; F is selected from NR 1e or O; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl; K and G are independently selected from CH2, CR 1e R 1f , and K and G are not CH2 at the same time; where R 1e Selected from the group consisting of H, deuterium, halogen, hydroxyl, C 1-4 Alkyl, C 1-4 Deuterated alkyl, R 1f Selected from the group consisting of deuterium, C 1-4 deuterated alkyl; or R 1e and R 1f Together with the carbon atoms to which they are attached, they form a 3-5 membered carbocyclic or heterocyclic ring, wherein the heteroatom contained in the heterocyclic ring is selected from O, N and S.

3. The compound according to claim 2, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: R1 is selected from the following group:

4. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: Cy A Select from the following group: in represents a covalent bond to the parent nucleus, and represents a covalent bond to the carbonyl group; R 12 and d are as defined in claim 1.

5. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: Cy B Select from the following group: wherein h is selected from 0, 1, 2 or 3; R 44 , R 45 and R 46 Each independently selected from H, D, halogen, CN, OH, C 1-6 Alkyl, C 1-4 Deuterated alkyl, C 3-6 Cycloalkyl, C 3-6 Cycloalkoxy, C 1-6 Alkoxy, C 1-6 Haloalkyl, C 1-6 Halogenated alkoxy, C 1-6 Alkylamine Base, C 2-4 Alkenyl, C 1-4 Halogenated alkenyl, C 2-4 Alkynyl; R as described above 45 and R 46 can form a 4-6 membered carbocyclic ring or heterocyclic ring or heteroaromatic ring together with the carbon atoms to which they are connected, wherein the heteroatoms contained in the heterocyclic ring or heteroaromatic ring are selected from O, N and S; the 4-6 membered carbocyclic ring or heterocyclic ring formed above can be independently optionally replaced by 0, 1, 2 or 3 R 47 replace; R 47 Independently selected from the group consisting of H, D, halogen, hydroxyl, cyano, C 1-4 Alkyl, C 1-4 Cycloalkyl, C 1-4 Haloalkyl, C 1-4 Haloalkyl, C 1-4 Halogenated cycloalkyl, C 1-4 Halogenated alkoxy.

6. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: Cy B Select from the following group:

7. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: Select from the following group: Select from the following group: Among them, R 48 , R 49 and R 50 Each independently selected from the following group: C 1-4 Alkyl, C 1-4 Haloalkyl, cyano, fluorine, chlorine.

8. The compound according to claim 1, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, characterized in that: The compound is selected from:

9. A pharmaceutical composition comprising a compound as described in any one of claims 1 to 8, a stereoisomer thereof, an optical isomer thereof, a pharmaceutically acceptable salt thereof, a crystalline form thereof, an isotopic derivative thereof, a prodrug thereof, a metabolite thereof, a solvate thereof or a hydrate thereof; and a pharmaceutically acceptable carrier.

10. Use of the compound according to any one of claims 1 to 8, its stereoisomer, its optical isomer, its pharmaceutically acceptable salt, its crystal form, its isotopic derivative, its prodrug, its metabolite, its solvate or its hydrate, or the pharmaceutical composition according to claim 9 in the preparation of a drug for treating a disease mediated by WRN helicase; Preferably, the disease is a tumor; more preferably, the disease is a microsatellite unstable tumor; further preferably, the tumor is a tumor characterized by microsatellite instability-high (MSI-H), or a tumor with defective mismatch repair (dMMR) or "dMMR signature"; in, The microsatellite unstable tumors include, but are not limited to, uterine body tumors, endometrial cancer, colon adenocarcinoma, gastric adenocarcinoma, rectal adenocarcinoma, adrenal cortical carcinoma, uterine carcinosarcoma, cervical squamous cell carcinoma, endocervical adenocarcinoma, esophageal cancer, breast cancer, renal clear cell carcinoma, and ovarian serous cystic carcinoma; Among them, tumors with "defective mismatch repair" (dMMR) or "dMMR characteristics" include but are not limited to lung cancer, breast cancer, kidney cancer, colorectal cancer, ovarian cancer, prostate cancer, upper aerodigestive tract cancer, gastric cancer, endometrial cancer, liver cancer, pancreatic cancer, hematopoietic and lymphoid tissue cancer, skin cancer, thyroid cancer, pleural cancer, autonomic nervous system tumors, soft tissue tumors, rhabdoid sarcoma, melanoma and other tumors.

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