A pyridine derivative, its preparation method and uses

Abstract

A compound of Formula I, its stereoisomer or a pharmaceutically acceptable salt thereof, a pharmaceutical composition comprising the compound, a method of preparation thereof, and its use in the preparation of a medicament for treating diseases related to LSD1 are provided.

Description

Technical Field

[0001] This application relates to the field of medicinal chemistry, specifically to a pyridine derivative or a pharmaceutically acceptable salt thereof as an inhibitor of lysine-specific demethylase 1 (LSD1), its preparation method and use. Background Technology

[0002] Histone lysine-specific demethylase 1 (LSD1) is the first reported histone demethylase. With the assistance of flavin adenine dinucleotide (FAD), it can specifically recognize H3K4 and H3K9 substrates and remove their monomethyl or dimethyl modifications. Simultaneously, LSD1 can form transcriptional regulatory complexes with various factors, reaching the promoter regions of specific genes under the recruitment of transcription factors, thereby regulating gene expression.

[0003] Studies have shown that LSD1 is associated with a variety of diseases, is highly expressed in various tumor cells, and is closely related to poor tumor prognosis. Downregulating LSD1 expression or inhibiting its activity can significantly suppress tumor cell growth. Therefore, developing a highly effective and specific LSD1 inhibitor is of great significance for the treatment of diseases such as cancer. Summary of the Invention

[0004] This application relates to a pyridine derivative as an LSD1 inhibitor, and more particularly to a pyridine derivative thereof, its preparation method and its pharmaceutical application, especially the use of the pyridine derivative shown in Formula I below in the preparation of a drug for treating LSD1-mediated diseases, and more specifically, in the preparation of a drug suitable for oncology.

[0005] One aspect of this application provides pyridine derivatives, stereoisomers thereof, or pharmaceutically acceptable salts thereof with the structure shown in Formula I:

[0006]

[0007] in,

[0008] Ring A represents a saturated cycloalkyl group, a saturated heterocycloalkyl group, an unsaturated cycloalkyl group, or an unsaturated heterocycloalkyl group;

[0009] Ring B represents a monocyclic or bicyclic unsaturated hydrocarbon group, or a monocyclic or bicyclic unsaturated heterocyclic hydrocarbon group;

[0010] R 1 R 2 Each is independently selected from hydrogen, fluorine, cyano, And R 1 R 2 R can be either hydrogen or fluorine at the same time. 1 When it is hydrogen, R 2 Not fluorine, R 1When it is fluorine, R 2 Not hydrogen; or R 1 With R 2 It forms a ring with the attached carbon atom.

[0011] R 3 The group is selected from halogen, cyano, oxo, hydroxyl, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxyl, amino, C1-C6 alkyl, C1-C6 alkoxy, or C1-C6 alkylamide; preferably, when m is 2-5, R 3 Different functional groups can be selected;

[0012] R 4 The group is selected from halogen, cyano, oxo, hydroxy, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or (C3-C8) cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxy, amino, C1-C6 alkyl, C1-C6 alkoxy, or C1-C6 alkylamide; preferably, when n is 2-5, R 4 Different functional groups can be selected;

[0013] R 5 The group is selected from halogen, hydroxyl, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, (C3-C8)cycloalkyl or (C3-C8)cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxyl, amino, C1-C6 alkyl or C1-C6 alkoxy;

[0014] R 6 R 7 Each is independently selected from hydrogen, C1-C6 alkyl, or C3-C8 cycloalkyl;

[0015] m is selected from 0, 1, 2, 3, 4 or 5;

[0016] n is selected from 0, 1, 2, 3, 4 or 5;

[0017] q is selected from 0, 1, 2, 3 or 4.

[0018] In some implementations, ring A is selected from C4-C13 Cycloalkyl or 3- to 13-membered heterocycloalkyl, wherein the heterocycloalkyl contains 1 to 3 heteroatoms selected from nitrogen, oxygen or sulfur atoms;

[0019] Preferably, ring A is selected from C4-C8 monocycloalkyl, C7-C... 13 Spirocycloalkyl, 3- to 10-membered monoheterocycloalkyl, or 7- to 13-membered spiroheterocycloalkyl, wherein the heterocycloalkyl contains 1 to 3 heteroatoms selected from nitrogen or oxygen atoms;

[0020] More preferably, ring A is selected from C5-C7 monocycloalkyl, C8-C... 12 Spirocycloalkyl, 3- to 8-membered monoheterocycloalkyl, or 8- to 12-membered spiroheterocycloalkyl, wherein the heterocycloalkyl contains 1 to 3 heteroatoms selected from nitrogen or oxygen atoms; or, ring A is selected from C5-C7 monocycloalkyl (e.g., C6 monocycloalkyl), 3- to 8-membered monoheterocycloalkyl (e.g., 4- to 7-membered monoheterocycloalkyl), or 8- to 12-membered spiroheterocycloalkyl (e.g., 9- to 10-membered spiroheterocycloalkyl), wherein the heterocycloalkyl contains 1 to 3 heteroatoms selected from nitrogen or oxygen atoms;

[0021] Alternatively, preferably, ring A is selected from piperidinyl, tetrahydropyridinyl, piperazinyl, azircyclobutyl, azircyclopentyl, azircyclohexyl, azircycloheptyl, high-piperazinyl, cyclohexyl, cyclobutyl, cyclopentyl, cycloheptyl, diazaspirononyl, diazaspirodecyl, oxa-azaspirononyl, and oxa-azaspirodecyl.

[0022] In some implementations, ring B is selected from C5-C. 10 Cycloalkenyl, 5- to 10-membered heterocyclic alkenyl, C5-C 10 Aryl, 5- to 10-membered heteroaryl, wherein the heterocyclic alkenyl or heteroaryl contains 1 to 3 heteroatoms selected from nitrogen, oxygen or sulfur atoms;

[0023] Preferably, ring B is selected from C5-C8 aryl, 8-membered to 10-membered heteroaryl, and the heterocyclic alkenyl or heteroaryl contains 1-3 heteroatoms selected from nitrogen, oxygen or sulfur atoms;

[0024] Alternatively, preferably, ring B is selected from phenyl, pyridinyl, dihydropyridinyl, pyrazinyl, pyridazinyl, pyrimidinyl, benzo5- to 6-membered heterocyclic alkenyl groups containing 1 to 3 heteroatoms selected from nitrogen or oxygen atoms (e.g., indolyl, isoyindolyl, isoyindolinyl, dihydroindolyl, indazole, dihydroindazole, benzoxazolyl, benzoisoxazolyl, quinolinyl, isoquinolinyl, quinazolinyl, quinoxolinyl, benzodioxonelyl, benzofuranyl, benzoimidazolyl, benzodihydrooxazinyl).

[0025] In some implementations, R 1 R2 Each is independently selected from hydrogen, cyano, Or R 1 With R 2 It forms a ring with the attached carbon atom. R 6 R 7 Each is independently selected from hydrogen, C1-C6 alkyl, or C3-C6 cycloalkyl;

[0026] Preferably, R 1 Selected from hydrogen, cyano, (e.g., hydrogen, cyano or) ), R 2 Selected from hydrogen, cyano, (e.g., hydrogen, cyano, ), or R 1 With R 2 It forms a ring with the attached carbon atom. (For example ); R 6 R 7 Each is independently selected from hydrogen, C1-C5 alkyl (e.g., C1-C4 alkyl, C1-C3 alkyl) or C3-C5 cycloalkyl (e.g., C3-C4 cycloalkyl, cyclopropyl).

[0027] In some implementations, m is 0, 1, 2, 3, 4, or 5, R 3 The group is selected from halogens (including fluorine, chlorine, bromine, or iodine), oxo, hydroxyl, substituted or unsubstituted amino groups, C1-C6 alkyl, or C1-C6 alkoxy groups, wherein the C1-C6 alkyl or C1-C6 alkoxy group is unsubstituted or substituted with one or more substituents selected from: halogen, cyano, hydroxyl, amino. Preferably, when m is 2-5, the plurality of R groups... 3 It can be selected from different groups;

[0028] Alternatively, in some implementations, m is 0, 1, 2, or 3 (preferably 0, 1, or 2), R 3 Selected from fluorine, chlorine, bromine, oxo, hydroxyl, amino, or C1-C6 alkyl (e.g., C1-C3 alkyl), wherein the C1-C6 alkyl (e.g., C1-C3 alkyl) is unsubstituted or substituted with an amino group; preferably, when m is 2, the two R groups are... 3 They can be the same or different.

[0029] In some implementations, n is selected from 0, 1, 2, 3, 4, or 5, R 4The group is selected from halogens (including fluorine, chlorine, bromine, or iodine), oxo, hydroxyl, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, or C3-C6 cycloalkyloxy groups, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, or C3-C6 cycloalkyloxy groups are unsubstituted or substituted with one or more substituents selected from: halogen, hydroxyl, amino; preferably, when n is 2-5, the plurality of R groups... 4 It can be selected from different groups;

[0030] Alternatively, in some implementations, n is selected from 0, 1, 2, 3, or 4, R 4 The group is selected from fluorine, chlorine, bromine, oxo, hydroxyl, C1-C6 alkyl, and C1-C6 alkoxy groups, wherein the C1-C6 alkyl or C1-C6 alkoxy group is unsubstituted or substituted with one or more substituents selected from halogens or hydroxyl groups; preferably, when n is 2-4, the plurality of R groups... 4 It can be selected from different groups.

[0031] In some implementations, q is selected from 0, 1, or 2, R 5 The group is selected from halogens, hydroxyl groups, amino groups, C1-C6 alkyl groups, and C1-C6 alkoxy groups, wherein the C1-C6 alkyl or C1-C6 alkoxy group is unsubstituted or substituted by one or more substituents selected from halogens or hydroxyl groups;

[0032] Or in some implementations, q is 1 or 2, R 5 Selected from halogens, preferably R 5 Selected from fluorine, chlorine, or bromine.

[0033] Preferably, in a compound of formula I, its stereoisomer, or a pharmaceutically acceptable salt thereof,

[0034] Ring A is selected from 5- to 8-membered saturated monocyclic alkyl groups, 4- to 10-membered saturated monocyclic or spirocyclic heterocyclic alkyl groups, wherein the heterocyclic alkyl group contains 1-3 nitrogen atoms as heteroatoms;

[0035] Ring B is selected from 5- to 8-membered unsaturated monocyclic hydrocarbon groups, 5- to 14-membered unsaturated monocyclic or bicyclic heterocyclic hydrocarbon groups, wherein the heterocyclic hydrocarbon group contains 0-4 nitrogen atoms and 0-2 oxygen atoms as heteroatoms and has at least one of nitrogen and oxygen.

[0036] R 1 R 2 Each is independently selected from hydrogen, fluorine, cyano, And R 1 R 2 R can be either hydrogen or fluorine at the same time. 1 When it is hydrogen, R 2 Not fluorine, R 1 When it is fluorine, R 2Not hydrogen; or R 1 With R 2 It forms a ring with the attached carbon atom.

[0037] R 3 The group is selected from halogen, cyano, oxo, hydroxy, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more of the following substituents: halogen, cyano, hydroxy, amino, C1-C6 alkyl or C1-C6 alkoxy;

[0038] R 4 The group is selected from halogen, cyano, oxo, hydroxy, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or (C3-C8) cycloalkyloxy, wherein the (C1-C6) alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more of the following substituents: halogen, cyano, hydroxy, amino, C1-C6 alkyl or C1-C6 alkoxy;

[0039] R 5 The group is selected from halogen, hydroxyl, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxyl, amino, C1-C6 alkyl or C1-C6 alkoxy;

[0040] R 6 R 7 Each is independently selected from hydrogen, C1-C6 alkyl, or C3-C8 cycloalkyl;

[0041] m is selected from 0, 1, 2, 3, 4 or 5;

[0042] n is selected from 0, 1, 2, 3, 4 or 5;

[0043] q is selected from 0, 1, 2, 3 or 4.

[0044] Preferably, in a compound of formula I, its stereoisomer, or a pharmaceutically acceptable salt thereof,

[0045] Ring A is selected from 4- to 10-membered saturated monocyclic or spirocyclic heterocyclic alkyl groups, wherein the heterocyclic alkyl group contains 1-3 nitrogen atoms as heteroatoms; preferably, it contains 1-2 nitrogen atoms as heteroatoms; or, ring A is selected from piperidinyl, piperazine, azirrobutyl, azirroheptyl, homopiperazine, diazaspirononyl, diazaspirodecyl, or monooxaazaspirodecyl.

[0046] Ring B is selected from a 6-membered unsaturated monocyclic hydrocarbon group or a 6- to 10-membered unsaturated bicyclic heterocyclic hydrocarbon group, wherein the heterocyclic hydrocarbon group contains 0-4 nitrogen atoms and 0-2 oxygen atoms as heteroatoms and has at least one of nitrogen and oxygen; preferably, ring B is selected from a 6- to 10-membered aryl or heterocyclic aryl group, wherein the heterocyclic aryl group contains 1-2 nitrogen atoms and 0-2 oxygen atoms as heteroatoms and has at least one of nitrogen and oxygen; more preferably, ring B is selected from phenyl, pyridinyl, indolyl, dihydroindolyl, indazole, benzoisoxazolyl, benzoxazolyl, benzodioxonel, or benzodihydrooxazinyl.

[0047] R 1 R 2 Each is independently selected from hydrogen, fluorine, cyano, And R 1 R 2 R can be either hydrogen or fluorine at the same time. 1 When it is hydrogen, R 2 Not fluorine, R 1 When it is fluorine, R 2 Not hydrogen; or R 1 With R 2 It forms a ring with the attached carbon atom. Preferably, R 1 R 2 Each is independently selected from hydrogen, fluorine, cyano, (e.g. R) 1 R 2 Each is independently selected from hydrogen, fluorine, cyano, ), and R 1 R 2 R can be either hydrogen or fluorine at the same time. 1 When it is hydrogen, R 2 Not fluorine, R 1 When it is fluorine, R 2 Not hydrogen; or R 1 With R 2 It forms a ring with the attached carbon atom.

[0048] R 3The group is selected from halogen, cyano, oxo, hydroxyl, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxyl, amino, C1-C6 alkyl, or C1-C6 alkoxy; preferably, R 3 The group is selected from halogen, cyano, oxo, hydroxyl, substituted or unsubstituted amino, C1-C3 alkyl, C1-C3 alkoxy, C3-C5 cycloalkyl, or C3-C5 cycloalkyloxy, wherein the C1-C3 alkyl, C1-C3 alkoxy, C3-C5 cycloalkyl, or C3-C5 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxyl, amino, C1-C6 alkyl, or C1-C6 alkoxy; preferably, R 3 It is selected from fluorine, chlorine, oxo, hydroxyl, amino or C1-C3 alkyl (e.g. methyl), wherein the C1-C3 alkyl (e.g. methyl) is unsubstituted or substituted with amino;

[0049] R 4 The group is selected from halogen, cyano, oxo, hydroxy, substituted or unsubstituted amino, C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy, wherein the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxy, amino, C1-C6 alkyl, or C1-C6 alkoxy; preferably, the C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl, or C3-C8 cycloalkyloxy is C1-C3 alkyl, C1-C3 alkoxy, C3-C5 cycloalkyl, or C3-C5 cycloalkyloxy, and is unsubstituted or substituted by one or more substituents selected from: halogen, cyano, hydroxy, amino, C1-C6 alkyl, or C1-C6 alkoxy; preferably, R 4 Selected from fluorine, chlorine, oxo, methyl, ethyl, propyl, methoxy, ethoxy, propoxy, (For example, R) 4 Selected from fluorine, chlorine, oxo, methyl, methoxy, );

[0050] R 5Selected from halogens, hydroxyl groups, substituted or unsubstituted amino groups, C1-C6 alkyl groups, C1-C6 alkoxy groups, C3-C8 cycloalkyl groups, or C3-C8 cycloalkyloxy groups, wherein the C1-C6 alkyl group, C1-C6 alkoxy group, C3-C8 cycloalkyl group, or C3-C8 cycloalkyloxy group is unsubstituted or substituted by one or more substituents selected from: halogens, cyano groups, hydroxyl groups, amino groups, C1-C6 alkyl groups, or C1-C6 alkoxy groups; preferably, R 5 Selected from fluorine, chlorine, bromine, methyl, methoxy, trifluoromethyl, or trifluoromethoxy, for example, R 5 Selected from fluorine, chlorine, or bromine;

[0051] R 6 R 7 Each of the following is independently selected from hydrogen, C1-C6 alkyl, or C3-C8 cycloalkyl, preferably, R 6 R 7 Each is independently selected from hydrogen, C1-C6 alkyl, or C3-C6 cycloalkyl;

[0052] m is selected from 0, 1, 2, 3, 4 or 5; preferably, m is selected from 0, 1, 2 or 3;

[0053] n is selected from 0, 1, 2, 3, 4 or 5; preferably, n is selected from 0, 1, 2, 3 or 4;

[0054] q is selected from 0, 1, 2, 3 or 4; preferably, q is selected from 1 or 2.

[0055] In some embodiments, in a compound of formula I, its stereoisomers, or a pharmaceutically acceptable salt thereof,

[0056] Ring A is selected from piperidinyl, piperazinyl, aziridine, aziridine-heptyl, homopiperazinyl, diazaspirononyl, diazaspirodealkyl, or oxa-azaspirodealkyl;

[0057] Ring B is selected from phenyl, pyridyl, indolyl, dihydroindolyl, indazole, benzoisoxazolyl, benzoxazolyl, benzodioxonyl, or benzodihydrooxazinyl;

[0058] R 1 R 2 Each is independently selected from hydrogen, cyano, Or R 1 With R 2 It forms a ring with the attached carbon atom.

[0059] R 3 It is selected from fluorine, chlorine, bromine, oxo, hydroxyl, amino, or C1-C3 alkyl, wherein the C1-C3 alkyl is unsubstituted or substituted with amino;

[0060] R 4 The group is selected from fluorine, chlorine, bromine, oxo, hydroxyl, C1-C6 alkyl, and C1-C6 alkoxy, wherein the C1-C6 alkyl group is unsubstituted or substituted by one or more substituents selected from halogen or hydroxyl.

[0061] R 5 Selected from fluorine, chlorine, or bromine;

[0062] m is selected from 0, 1, 2, or 3;

[0063] n is selected from 0, 1, 2, 3 or 4;

[0064] q is selected from 1 or 2.

[0065] Preferably, the compound of formula I, its stereoisomer, or a pharmaceutically acceptable salt thereof further has the structure shown in formula II.

[0066]

[0067] in,

[0068] Ring A is selected from cyclohexyl, piperidinyl, piperazine, aziridine-heptyl, or homopiperazine; preferably, the piperidinyl, piperazine, aziridine-heptyl, or homopiperazine is attached to the pyridine ring via an N atom;

[0069] Ring B is selected from phenyl, pyridyl, indolyl, dihydroindolyl, or indazole;

[0070] R 2 Selected from hydrogen or fluorine;

[0071] R 3 Selected from fluorine, chlorine, oxo, hydroxyl, or amino groups;

[0072] R 4 Selected from fluorine, chlorine, oxo, methyl, methoxy,

[0073] m is selected from 0, 1, 2, or 3;

[0074] n is selected from 0, 1, 2, 3 or 4.

[0075] Alternatively, in some embodiments, in a compound of formula II, its stereoisomers, or a pharmaceutically acceptable salt thereof

[0076] Ring A is selected from cyclohexyl, piperidinyl, piperazinyl, aziridine, aziridine-heptyl, homopiperazinyl, diazaspironyl or diazaspironyldecyl; preferably, the piperidinyl, piperazinyl, aziridine-heptyl, homopiperazinyl, diazaspironyl or diazaspironyl is attached to the pyridine ring by an N atom;

[0077] Ring B is selected from phenyl, pyridyl, indolyl, dihydroindolyl, indazole, benzoisoxazolyl, benzoxazolyl, or benzodioxonel;

[0078] R 2 Selected from hydrogen, cyano, R 6 R 7 Each is independently selected from hydrogen, C1-C5 alkyl, or C3-C5 cycloalkyl;

[0079] R 3 It is selected from fluorine, chlorine, bromine, oxo, hydroxyl, amino, or C1-C6 alkyl, wherein the C1-C6 alkyl is unsubstituted or substituted with amino.

[0080] R 4 The group is selected from fluorine, chlorine, bromine, oxo, C1-C6 alkyl, and C1-C6 alkoxy, wherein the C1-C6 alkyl group is unsubstituted or substituted by one or more substituents selected from halogens or hydroxyl groups;

[0081] m is selected from 0, 1, 2, or 3;

[0082] n is selected from 0, 1, 2, 3 or 4.

[0083] More preferably, the heterocyclic derivatives of the structure shown in Formula I, their stereoisomers, or pharmaceutically acceptable salts thereof are selected from the following compounds, their stereoisomers, or pharmaceutically acceptable salts thereof:

[0084]

[0085]

[0086] This application also covers solutions obtained by any combination, deletion or substitution of the above-described embodiments and preferred solutions.

[0087] Another aspect of this application provides a method for preparing pyridine derivatives with the structure shown in Formula I above, the method comprising the following steps:

[0088] Method 1:

[0089]

[0090] Wherein, LG represents a leaving group, which includes, but is not limited to, halogen atoms, methanesulfonyloxy groups, p-toluenesulfonyloxy groups, etc. R 1 R 2 R 3 R 4 R 5 Ring A, ring B, m, n, and q are defined above for pyridine derivatives with the structure shown in Formula I;

[0091] (1) React compound I-1 with compound I-2 to obtain compound I-3.

[0092] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, N,N-dimethylformamide, N,N-dimethylacetamide, 1,4-dioxane, and any combination thereof, with N,N-dimethylformamide being preferred. The reaction is preferably carried out in the presence of a suitable base; and / or in the presence of a transition metal catalyst and a ligand. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, or potassium tert-butoxide, with potassium carbonate being preferred. The transition metal catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, or RuPhos-Pd-G3. The reaction is preferably carried out at a suitable temperature, preferably 50-80°C. The reaction is preferably carried out for a suitable time, for example, 2-8 hours.

[0093] (2) Compound I-3 was subjected to a substitution reaction to obtain compound I-4.

[0094] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from acetonitrile, toluene, chloroform, tetrahydrofuran, and any combination thereof, with acetonitrile being preferred. The reaction is preferably carried out in the presence of a suitable brominating agent. The brominating agent may be selected from N-bromosuccinimide, liquid bromine, or pyridine bromide, with N-bromosuccinimide being preferred. The reaction is preferably carried out at a suitable temperature, preferably 20-50°C. The reaction is preferably carried out for a suitable time, for example, 2-6 hours.

[0095] (3) Compound I-6 is obtained by coupling compound I-4 with boric acid or borate ester of compound I-5.

[0096] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0097] (4) Compound I-6 and compound I-7 were coupled together to obtain the compound with the structure shown in Formula I.

[0098] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0099] Method 2:

[0100]

[0101] Wherein, LG represents a leaving group, which includes, but is not limited to, halogen atoms, methanesulfonyloxy groups, p-toluenesulfonyloxy groups, etc. R 1 R 2 R 3 R 4 R 5 Ring A, ring B, m, n, and q are defined above for pyridine derivatives with the structure shown in Formula I;

[0102] (1) Compound I-1 was substituted to obtain compound I-8.

[0103] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from acetonitrile, toluene, chloroform, tetrahydrofuran, and any combination thereof, with acetonitrile being preferred. The reaction is preferably carried out in the presence of a suitable brominating agent. The brominating agent may be selected from N-bromosuccinimide, liquid bromine, or pyridine bromide, with N-bromosuccinimide being preferred. The reaction is preferably carried out at a suitable temperature, preferably 20-50°C. The reaction is preferably carried out for a suitable time, for example, 2-6 hours.

[0104] (2) Compound I-8 is coupled with boric acid or borate ester of compound I-5 to obtain compound I-9.

[0105] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0106] (3) Compound I-9 and compound I-7 were coupled together to obtain compound I-10.

[0107] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0108] (4) React compound I-10 with compound I-2 to obtain the compound with the structure shown in formula I.

[0109] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, N,N-dimethylformamide, N,N-dimethylacetamide, 1,4-dioxane, and any combination thereof, with N,N-dimethylformamide being preferred. The reaction is preferably carried out in the presence of a suitable base; and / or in the presence of a transition metal catalyst and a ligand. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, or potassium tert-butoxide, with potassium carbonate being preferred. The transition metal catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, or RuPhos-Pd-G3. The reaction is preferably carried out at a suitable temperature, preferably 50-80°C. The reaction is preferably carried out for a suitable time, for example, 2-8 hours.

[0110] Method 3:

[0111]

[0112] Wherein, LG represents a leaving group, which includes, but is not limited to, halogen atoms, methanesulfonyloxy groups, p-toluenesulfonyloxy groups, etc. R 1 R 2 R 3 R 4 R 5 Ring A, ring B, m, n, and q are defined above for pyridine derivatives with the structure shown in Formula I;

[0113] (1) Compound I-3 and compound I-7 were coupled together to obtain compound I-11.

[0114] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0115] (2) Compound I-11 was subjected to a substitution reaction to obtain compound I-12.

[0116] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from acetonitrile, toluene, chloroform, tetrahydrofuran, and any combination thereof, with acetonitrile being preferred. The reaction is preferably carried out in the presence of a suitable brominating agent. The brominating agent may be selected from N-bromosuccinimide, liquid bromine, or pyridine bromide, with N-bromosuccinimide being preferred. The reaction is preferably carried out at a suitable temperature, preferably 20-50°C. The reaction is preferably carried out for a suitable time, for example, 2-6 hours.

[0117] (3) Compound I-12 is coupled with boric acid or borate ester of compound I-5 to obtain compound I.

[0118] The reaction is preferably carried out in a suitable organic solvent. The organic solvent may be selected from tetrahydrofuran, 1,4-dioxane, toluene, acetonitrile, ethanol, water, and any combination thereof, preferably a combination of 1,4-dioxane and water. The reaction is preferably carried out in the presence of a suitable catalyst. The catalyst may be selected from Pd(dppf)Cl2, Pd(OAc)2, Pd2(dba)3, Pd(PPh3)4, preferably Pd(dppf)Cl2. The reaction is preferably carried out in the presence of a suitable base. The base may be selected from triethylamine, pyridine, 4-dimethylaminopyridine, diisopropylethylamine, potassium carbonate, cesium carbonate, sodium carbonate, preferably potassium carbonate. The reaction is preferably carried out at a suitable temperature, preferably 80-120°C. The reaction is preferably carried out for a suitable time, for example, 8-12 hours.

[0119] The specific conditions for each of the above reaction steps are well known in the art and are not specifically limited thereto in this application. If the teachings of this application are combined with common knowledge in the art, those skilled in the art can selectively replace the substituents in the general formula to prepare different compounds, and such selections and substitutions are all within the scope of protection of this application.

[0120] This application also relates to a pharmaceutical composition comprising the above-described compound of formula I, its stereoisomer or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0121] This application also relates to the use of the above-described Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical compositions in the preparation of a medicament for the prevention or treatment of LSD1-related diseases. Alternatively, this application also relates to a Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical compositions for the prevention or treatment of LSD1-related diseases. Alternatively, this application also relates to a method for the prevention or treatment of LSD1-related diseases, comprising administering to a subject in need a Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical compositions.

[0122] In some embodiments, the LSD1-related diseases are selected from tumors or cancers, such as acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), lymphoma, malignant sarcoma, breast cancer, cervical cancer, colon cancer, lung cancer, oral cancer, brain cancer, gastric cancer, liver cancer, colorectal cancer, pancreatic cancer, skin cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, fallopian tube tumors, peritoneal tumors, melanoma, glioma, glioblastoma, papillary malignancy, head and neck tumors, or myeloma.

[0123] This application also relates to the use of the above-described Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical composition in the preparation of an LSD1 inhibitor. Alternatively, this application also relates to the use of the above-described Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical composition as an LSD1 inhibitor. Alternatively, this application also relates to a method of inhibiting LSD1, comprising administering to a subject in need a above-described Formula I compound, its stereoisomers or pharmaceutically acceptable salts thereof, or the above-described pharmaceutical composition.

[0124] This application discovers a novel class of LSD1 inhibitors with the structure shown in Formula I, which exhibits good inhibitory activity. Attached Figure Description

[0125] Figure 1 The changes in tumor volume in mice in the test compound group and the solvent group are shown, where Cpd represents the compound.

[0126] Figure 2 The changes in tumor weight between the test compound group and the solvent group are shown.

[0127] Figure 3 The rate of change in body weight of mice in the test compound group and the solvent group is shown.

[0128] Figure 4 The changes in tumor volume in mice in the test compound group and the solvent group are shown.

[0129] Figure 5 The rate of change in body weight of mice in the test compound group and the solvent group is shown. Detailed Implementation

[0130] To make the aspects and technical solutions of this application clearer, the following describes the application in further detail with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the application. Furthermore, any specific experimental methods not mentioned in the following embodiments are performed according to conventional experimental methods.

[0131] Definitions and Explanations

[0132] Unless otherwise stated, the terms used in this application have the following meanings. A particular term should not be considered uncertain or unclear unless specifically defined, but should be understood in accordance with its ordinary meaning in the art.

[0133] When the covalent bonds in certain structural units or groups in this application are not connected to specific atoms, it means that the covalent bonds can be connected to any atom in the structural unit or group, as long as the rules of valence bond connection are not violated.

[0134] In this document, unless otherwise stated, "alkoxy" refers to -O-alkyl.

[0135] In this document, unless otherwise stated, "halogen" means fluorine, chlorine, bromine or iodine.

[0136] In this document, unless otherwise stated, “cycloalkyloxy” refers to -O-cycloalkyl.

[0137] In this document, unless otherwise stated, "saturated cycloalkyl" encompasses monocyclic, bicyclic bridged rings, bicyclic spirocyclic rings, etc., and refers to fully saturated alicyclic hydrocarbons. For example, cycloalkyl in this document may be C3-C6. 10 Single ring, C5-C 15 Spiral ring or C4-C 12 Cycloalkyl groups with bridged rings. Non-limiting examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, spiropentyl, spirohexyl, spiroheptyl, spiroheptyl, spiroctyl, spirononyl, spirdecyl, spirundecyl, etc.

[0138] In this document, unless otherwise stated, “saturated heterocyclic alkyl” means a cycloalkyl group in which one or more (e.g., 1-6, 1-5, 1-3, 1 or 2) carbon atoms are replaced by heteroatoms selected from N, O or S.

[0139] In this document, unless otherwise stated, "cyclic hydrocarbon group" refers to a hydrocarbon group in which carbon atoms form a ring, including saturated cyclic hydrocarbon groups and unsaturated cyclic hydrocarbon groups (e.g., aromatic groups). In this document, unsaturated cyclic hydrocarbon groups are sometimes also referred to as unsaturated cycloalkyl groups. In this document, cyclic hydrocarbon groups can be monocyclic, bicyclic bridged rings, bicyclic spirocyclic, or bicyclic fused rings, for example, C3-C... 10 Single ring, C5-C 15 Spiral ring, C4-C 12 Bridged ring, cyclic hydrocarbon group.

[0140] In this document, unless otherwise stated, “substituted or unsubstituted amino” refers to an unsubstituted amino group or an amino group substituted with a group selected from C1-C6 alkyl, C1-C6 alkoxy, C3-C8 cycloalkyl or halogen.

[0141] In this document, unless otherwise stated, the term "Cm-Cn" as used means that the portion modified by the term has mn carbon atoms (n is greater than m, and both are integers). For example, C1-C6 means that the portion it modifies has 1-6 carbon atoms, such as 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms.

[0142] The term “subject” is equivalent to “patient” and “individual” and refers to either a human or a non-human animal (mammal, such as primates, rodents, etc.). “Mammal” includes humans and livestock (such as laboratory mammals and domestic pets, such as cats, dogs, pigs, sheep, cattle, sheep, goats, horses, and rabbits), as well as non-domesticated mammals, such as wild mammals.

[0143] The term "treatment" means administering the compound or preparation described in this application to improve or eliminate a disease or one or more symptoms associated with the disease, including inhibiting the progression of the disease or condition and alleviating the disease or condition.

[0144] The term "pharmaceutically acceptable" means that a carrier, delivery substance, diluent, excipient, and / or the salt formed therefrom is generally chemically or physically compatible with the other components constituting a drug dosage form and physiologically compatible with the receptor, without excessive toxicity, irritation, allergic reactions or other problems or complications, and is commensurate with a reasonable benefit / risk ratio.

[0145] In this application, the terms “comprising,” “including,” and “containing,” and their equivalents, are to be understood in an open, non-exclusive sense, meaning “including but not limited to,” implying that in addition to the listed elements, components, and steps, other unspecified elements, components, and steps may also be included. In this document, unless the context clearly indicates otherwise, singular terms encompass plural referents, and vice versa. Similarly, unless the context clearly indicates otherwise, the word “or” is intended to include “and.”

[0146] Unless otherwise stated, in this document, parameter values ​​representing the amount of an ingredient, its physicochemical properties, or reaction conditions, etc., should be understood to be modified by the term "about" in all cases. When the term "about" is used to describe this application, the term "about" indicates an existing error value, such as a variation within ±5%, for example ±1%, or ±0.1% of a particular value.

[0147] In this application, when the chemical name and structural formula are inconsistent, the structural formula shall prevail, unless the chemical name rather than the structural formula can be inferred to be correct from the context.

[0148] The abbreviations used in this article have the following meanings:

[0149] <![CDATA[DMSO-d6]]> Hexadecimalized dimethyl sulfoxide q Four Peaks TMS Tetramethylsilane dd Double peak <![CDATA[ 1 H NMR]]> Hydrogen spectrum m multiplets MS mass spectrometry br Broad Peak s Single peak J Coupling constant d Double peak Hz hertz t Triple Peak NBS N-bromosuccinimide

[0150] The structure of the compound was determined by mass spectrometry (MS) or nuclear magnetic resonance (NMR). 1 It was determined by (H NMR). 1 HNMR shift (δ) is given in parts per million (ppm); nuclear magnetic resonance (NMR) 1 The 1H NMR (hydrogen spectroscopy) measurements were performed using a Bruker AVANCE-400 NMR spectrometer. The solvent was deuterated dimethyl sulfoxide (DMSO-d6), and the internal standard was tetramethylsilane (TMS). Chemical shifts were expressed in terms of 10⁻⁶ ppm. -6 (ppm) is given as the unit. Mass spectrometry (MS) determinations were performed using a FINNIGAN LCQAd (ESI) mass spectrometer (manufacturer: Therm, model: Finnigan LCQ advantage MAX). Thin-layer chromatography used Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates. Column chromatography generally used Yantai Huanghai 200-300 mesh silica gel as the support.

[0151] Unless otherwise specified in this application, all reactions mentioned herein are carried out under a nitrogen atmosphere. In this application, the term "nitrogen atmosphere" refers to, for example, connecting a reaction flask to a 1L nitrogen balloon.

[0152] In this application, the term "hydrogen atmosphere" refers to, for example, connecting a reaction vessel to a 1L hydrogen balloon.

[0153] Unless otherwise specified in this application, the solutions mentioned in the reactions described herein are aqueous solutions.

[0154] In this application, the term "room temperature" refers to a temperature between 10°C and 25°C.

[0155] For purposes of description and disclosure, all patents, patent applications, and other identified publications are expressly incorporated herein by reference. These publications are provided solely because their publication predates the filing date of this application. All statements regarding the dates of these documents or representations of their contents are based on information available to the applicant and do not constitute any acknowledgment of the accuracy of the dates or contents of these documents. Furthermore, in any country, any reference to these publications herein does not constitute an endorsement that such publication is part of the general knowledge in the art.

[0156] The present application will be described in detail below through embodiments; however, those skilled in the art will understand that the scope of protection of the present application is not limited thereto. Those skilled in the art can make various modifications, changes, combinations, etc., to the implementation methods and embodiments of the present application without departing from the spirit or scope of the present application, and the resulting adjusted solutions also fall within the scope of protection of the present application.

[0157] Example 1 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (1)

[0158]

[0159] Step 1: Preparation of tert-butyl (1-(6-chloro-4-cyanopyridin-2-yl)piperidin-4-yl)carbamate (1b)

[0160] Compound 1a (550 mg, 3.18 mmol), 4-tert-butoxycarbonylaminopiperidine (643 mg, 3.21 mmol), and K₂CO₃ (658 mg, 4.76 mmol) were dissolved in N,N-dimethylformamide (10 mL). The system was heated to 80 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 856 mg of the title compound (yield: 80%).

[0161] Step 2: Preparation of (1-(5-bromo-6-chloro-4-cyanopyridin-2-yl)piperidin-4-yl)carbamate tert-butyl ester (1c)

[0162] Compound 1b (538 mg, 1.60 mmol) was dissolved in acetonitrile (15 mL). NBS (288 mg, 1.62 mmol) was slowly added to the system under ice bath conditions. The reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography to give 542 mg of the title compound, yield: 82%.

[0163] Step 3: Preparation of [1-(6-chloro-4-cyano-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)piperidin-4-yl]tert-butyl carbamate (1d)

[0164] Compound 1c (231 mg, 0.56 mmol), 3-fluoro-4-methoxyphenylboronic acid (97 mg, 0.57 mmol), and K₂CO₃ (116 mg, 0.84 mmol) were dissolved in 1,4-dioxane (5 mL) and water (1 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (44 mg, 0.06 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 126 mg of the title compound (yield: 49%).

[0165] Step 4: Preparation of (1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (1e)

[0166] Compound 1d (47 mg, 0.10 mmol), 3-fluoro-4-cyanobenzonic acid (25 mg, 0.15 mmol), and K₂CO₃ (41 mg, 0.30 mmol) were dissolved in 1,4-dioxane (1 mL) and water (0.2 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (7 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 27 mg of the title compound (yield: 49%).

[0167] Step 5: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (1)

[0168] Compound 1e (27 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 16 mg of the title compound, yield: 73%.

[0169] LC-MS(ESI)m / z(M+H) + : 446.2

[0170] 1 H NMR (400MHz, DMSO-d6) δ7.83-7.77(m,1H),7.53(s,1H),7.37(dd,J=10.8,1.2Hz,1H),7.21-7.10(m,3H),6.98-6.93 (m,1H),4.34-4.25(m,2H),3.84(s,3H),3.08-2.98(m,2H),2.89-2.79(m,1H),1.82-1.73(m,2H),1.25-1.14(m,2H).

[0171] Example 2 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinamide (2)

[0172]

[0173] Step 1: Preparation of tert-butyl carbamate (2a)

[0174] Compound 1d (125 mg, 0.27 mmol) was dissolved in methanol (1 mL), followed by the addition of dimethyl sulfoxide (1 mL) and NaOH (32 mg, 0.80 mmol). H₂O₂ (153 mg, 1.35 mmol, 30% aqueous solution) was slowly added dropwise while stirring. The reaction mixture was kept at 50 °C and stirred for 2 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 90 mg of the title compound (yield: 69%).

[0175] Step 2: Preparation of tert-butyl carbamate (2b)

[0176] Compound 2a (55 mg, 0.11 mmol), 3-fluoro-4-cyanobenzonic acid (28 mg, 0.17 mmol), and K₂CO₃ (46 mg, 0.33 mmol) were dissolved in 1,4-dioxane (1 mL) and water (0.2 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (8 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 48 mg of the title compound (yield: 77%).

[0177] Step 3: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinamide (2)

[0178] Compound 2b (48 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 31 mg of the title compound, yield: 79%.

[0179] LC-MS(ESI)m / z(M+H) + : 464.2

[0180] 1 H NMR (400MHz, DMSO-d6) δ7.80-7.72(m,2H),7.45-7.40(m,1H),7.31(dd,J=10. 8,1.2Hz,1H),7.14(dd,J=8.0,1.2Hz,1H),7.02(t,J=8.4Hz,1H),6.95(dd,J=1 2.4,2.4Hz,1H),6.88(s,1H),6.88-6.72(m,1H),4.32-4.23(m,2H),3.80(s,3H) ),3.02-2.92(m,2H),2.86-2.77(m,1H),1.85-1.73(m,2H),1.26-1.13(m,2H).

[0181] Example 3 Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(3-fluoro-4-methoxyphenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (3)

[0182]

[0183] Step 1: Preparation of 2,6-dichloro-4-iodonicotinic acid (3b)

[0184] Compound 3a (10.0 g, 36.51 mmol) was dissolved in dry tetrahydrofuran (110 mL), and nitrogen gas was introduced. The system was placed at -50 °C, and LDA (20 mL, 40.00 mmol, 2 mol / L in THF / Heptane / Ethylbenzene) was slowly added dropwise. After the addition was complete, the system was kept at -50 °C and the reaction mixture was stirred for 2 hours. The reaction mixture was then quickly poured into a measuring cup containing crushed dry ice and stirred. The mixture was slowly brought to room temperature, and the reaction mixture was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was used directly in the next reaction without further purification.

[0185] Step 2: Preparation of methyl 2,6-dichloro-4-iodonicotinate (3c)

[0186] Compound 3b (12.0 g, 37.75 mmol) was dissolved in N,N-dimethylformamide (130 mL), followed by the sequential addition of K₂CO₃ (7.9 g, 57.16 mmol) and methyl iodoforme (8.1 g, 57.04 mmol). The reaction mixture was stirred at room temperature for 1 hour. The reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 8.3 g of the title compound. The combined yield of the two steps was 68%.

[0187] Step 3: Preparation of methyl 2,6-dichloro-4-cyanonicotinate (3d)

[0188] Compound 3c (4.3 g, 12.95 mmol) and Zn(CN)₂ (4.56 g, 38.85 mmol) were dissolved in N,N-dimethylformamide (40 mL), and the mixture was purged with nitrogen. Pd(PPh₃)₄ (750 mg, 0.65 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was stirred overnight at 100 °C. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.0 g of the title compound (yield: 33%).

[0189] Step 4: Preparation of methyl 2-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-6-chloro-4-cyanonicotinic acid (3e)

[0190] Compound 3d (1.0 g, 4.33 mmol), 4-tert-butoxycarbonylaminopiperidine (1.3 g, 6.49 mmol), and triethylamine (900 mg, 8.89 mmol) were dissolved in tetrahydrofuran (10 mL). The system was heated to 50 °C and stirred for 3 hours. The reaction system was cooled to room temperature, the reaction solution was poured into water, and the mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 647 mg of the title compound, yield: 38%.

[0191] Step 5: Preparation of (1-(6-chloro-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (3f)

[0192] Compound 3e (647 mg, 1.64 mmol) dissolved in methanol (10 mL) was added to a reaction flask, followed by the addition of ammonia (10 drops) and Raney-Ni (267 mg), purging the hydrogen atmosphere. The reaction system was then incubated overnight at room temperature under a hydrogen atmosphere. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 218 mg of the title compound, yield: 36%.

[0193] Step 6: Preparation of tert-butyl carbamate (3g) of 1-(7-bromo-6-chloro-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)carbamate

[0194] Compound 3f (175 mg, 0.48 mmol) was dissolved in acetonitrile (4 mL). NBS (128 mg, 0.72 mmol) was slowly added to the system under ice bath conditions, and the reaction mixture was stirred for 1 hour while maintaining the ice bath conditions. The reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 210 mg of the title compound, yield: 99%.

[0195] Step 7: Preparation of (1-(6-chloro-7-(3-fluoro-4-methoxyphenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (3h)

[0196] 3 g (120 mg, 0.27 mmol) of the compound, 46 mg (0.27 mmol) of 3-fluoro-4-methoxyphenylboronic acid, and 112 mg (0.81 mmol) of K₂CO₃ were dissolved in 2 mL of 1,4-dioxane and 0.2 mL of water. Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (36 mg, 0.05 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 70 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 33 mg of the title compound (yield: 25%).

[0197] Step 8: Preparation of (1-(6-(4-cyano-3-fluorophenyl)-7-(3-fluoro-4-methoxyphenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (3i)

[0198] Compound 3h (33 mg, 0.07 mmol), 3-fluoro-4-cyanobenzonic acid (16 mg, 0.01 mmol), and K₂CO₃ (56 mg, 0.40 mmol) were dissolved in 1,4-dioxane (1 mL) and water (0.1 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (22 mg, 0.03 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 34 mg of the title compound (yield: 88%).

[0199] Step 9: Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(3-fluoro-4-methoxyphenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (3)

[0200] Compound 3i (34 mg, 0.06 mmol) was dissolved in 1,4-dioxane (2 mL), and then 1,4-dioxane hydrochloride solution (2 mL, 4 M in 1,4-dioxane) was added. The reaction system was stirred at room temperature for 2 hours. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 12 mg of the title compound, yield: 43%.

[0201] LC-MS(ESI)m / z(M+H) + : 476.2

[0202] 1 H NMR (400MHz, DMSO-d6) δ8.54 (s, 1H), 7.84-7.78 (m, 1H), 7.45 (dd, J = 10.8, 1.2Hz, 1H),7.24(dd,J=8.0,1.6Hz,1H),7.18(dd,J=12.0,2.0Hz,1H),7.12(t,J=8.8Hz,1 H),6.95-6.90(m,1H),4.49-4.37(m,2H),4.15(s,2H),3.83(s,3H),3.06-2.95(m ,2H),2.87-2.76(m,1H),2.03-1.93(m,1H),1.86-1.74(m,2H),1.39-1.33(m,1H).

[0203] Example 4 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (4)

[0204]

[0205] Step 1: Preparation of tert-butyl carbamate (4a)

[0206] Compound 1c (130 mg, 0.31 mmol), 1-methylindazole-5-boric acid (56 mg, 0.32 mmol), and K₂CO₃ (64 mg, 0.46 mmol) were dissolved in 1,4-dioxane (3 mL) and water (0.6 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (22 mg, 0.03 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 120 mg of the title compound (yield: 82%).

[0207] Step 2: Preparation of tert-butyl carbamate (4b)

[0208] Compound 4a (50 mg, 0.11 mmol), 3-fluoro-4-cyanobenzonic acid (28 mg, 0.17 mmol), and K₂CO₃ (46 mg, 0.33 mmol) dissolved in 1,4-dioxane (1 mL) and water (0.2 mL) were added to a reaction flask. Nitrogen gas was then introduced, followed by the addition of Pd(dppf)Cl₂ (8 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction system was heated to 100 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction mixture was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 20 mg of the title compound (yield: 34%).

[0209] Step 3: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (4)

[0210] Compound 4b (20 mg, 0.04 mmol) was dissolved in ethyl acetate (0.5 mL), and then hydrochloric acid-ethyl acetate solution (0.5 mL, 3 M) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 15 mg of the title compound, yield: 92%.

[0211] LC-MS(ESI)m / z(M+H) + : 452.2

[0212] 1 H NMR (400MHz, DMSO-d6) δ8.06(d,J=0.8Hz,1H),7.75-7.70(m,1H),7.69-7.67(m,1H),7.63-7.59(m,1H),7.57(s,1H),7.38(dd,J=10.8,1. 2Hz,1H),7.18-7.12(m,2H),4.38-4.28(m,2H),4.05(s,3H),3.10-3.00(m,2H),2.91-2.81(m,1H),1.85-1.74(m,2H),1.26-1.16(m,2H).

[0213] Example 5 Preparation of 6-(4-amino-3-hydroxypiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (5)

[0214]

[0215] Step 1: Preparation of 4-amino-3-hydroxypiperidine-1-carboxylic acid benzyl ester (5b)

[0216] Compound 5a (1.0 g, 4.29 mmol) was dissolved in ethanol (5 mL), followed by the addition of ammonia (5 mL). The reaction mixture was sealed and heated to 70 °C with stirring overnight. The reaction mixture was cooled to room temperature, extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was used directly in the next reaction step without purification.

[0217] Step 2: Preparation of 4-((tert-Butoxycarbonyl)amino)-3-hydroxypiperidine-1-carboxylic acid benzyl ester (5c)

[0218] Compound 5b (700 mg, 2.80 mmol) was dissolved in dichloromethane (10 mL). Triethylamine (424 mg, 4.19 mmol) and di-tert-butyl dicarbonate (610 mg, 2.80 mmol) were added sequentially to the system under ice bath conditions. The reaction mixture was stirred overnight at room temperature. The reaction was quenched with water, and the mixture was extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 500 mg of the title compound. The combined yield for both steps was 33%.

[0219] Step 3: Preparation of (3-hydroxypiperidin-4-yl)carbamate tert-butyl ester (5d)

[0220] Compound 5c (500 mg, 1.43 mmol) was dissolved in isopropanol (5 mL), and then Pd / C (50 mg, 10%) was added. The air in the system was replaced with hydrogen, and the system was stirred overnight at room temperature for 4 hours under a hydrogen atmosphere. The reaction system was filtered and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 200 mg of the title compound, yield: 65%.

[0221] Step 4: Preparation of 2-chloro-6-((4-methoxybenzyl)amino)isonicotinonitrile (5e)

[0222] Compound 1a (7.0 g, 40.46 mmol) was dissolved in DMSO (70 mL), followed by the addition of DIEA (5.3 g, 41.09 mmol) and p-methoxybenzylamine (5.7 g, 41.55 mmol). The reaction mixture was stirred overnight at 100 °C. After cooling to room temperature, the reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was used directly in the next step without purification.

[0223] Step 5: Preparation of 2-amino-6-chloroisocyanuric acid nitrile (5f)

[0224] Compound 5e (9.0 g, 32.88 mmol) was dissolved in dichloromethane (50 mL), and then trifluoroacetic acid (50 mL) was added. The reaction mixture was stirred at room temperature for 6 hours. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 5.0 g of the title compound. The total yield of the two steps was 81%.

[0225] Step 6: Preparation of 6-amino-3-bromo-2-chloroisocyanuric acid nitrile (5g)

[0226] Compound 5f (5.0 g, 32.56 mmol) was dissolved in acetonitrile (50 mL). Bromine (5.3 g, 33.13 mmol) was slowly added dropwise to the system under ice bath conditions. The reaction system was stirred at room temperature for 6 hours. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The system was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 4.0 g of the title compound, yield: 53%.

[0227] Step 7: Preparation of 6-amino-2-chloro-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (5h)

[0228] 5 g (2.0 g, 8.60 mmol) of the compound was dissolved in 20 mL of 1,4-dioxane and 4 mL of water. Then, 1.5 g (8.83 mmol) of 3-fluoro-4-methoxyphenylboronic acid and 3.6 g (26.05 mmol) were added sequentially, and nitrogen was introduced to purge the atmosphere. Pd(dppf)Cl2 (628 mg, 0.86 mmol) was added, and nitrogen was introduced again. The reaction mixture was stirred at 80 °C for 6 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.5 g of the title compound, yield: 63%.

[0229] Step 8: Preparation of 6-amino-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (5i)

[0230] Compound 5h (1.5 g, 5.40 mmol) was dissolved in 1,4-dioxane (15 mL) and water (3 mL), followed by the addition of 3-fluoro-4-cyanobenzonic acid (2.7 g, 16.37 mmol) and K₂CO₃ (2.3 g, 16.64 mmol) to purge with nitrogen. Pd(dppf)Cl₂ (395 mg, 0.54 mmol) was then added, followed by another purge with nitrogen. The reaction mixture was stirred at 80 °C for 9 hours. After cooling to room temperature, the reaction solution was concentrated under reduced pressure. The solution was diluted with water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.5 g of the title compound (yield: 77%).

[0231] Step 9: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-iodoisocyanate (5j)

[0232] Compound 5i (1.5 g, 4.14 mmol) was dissolved in acetonitrile (15 mL), followed by the addition of potassium iodide (4.1 g, 24.70 mmol) and isoamyl nitrite (2.9 g, 24.75 mmol). The reaction mixture was stirred at 75 °C for 36 hours. After cooling to room temperature, the reaction solution was concentrated under reduced pressure. The solution was diluted with water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 700 mg of the title compound (yield: 36%).

[0233] Step 10: Preparation of tert-butyl carbamate (1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-3-hydroxypiperidin-4-yl)carbamate (5k)

[0234] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by compound 5d (48 mg, 0.22 mmol) and Cs₂CO₃ (108 mg, 0.33 mmol). Nitrogen gas was then added, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was purged again. The reaction mixture was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 20 mg of the title compound (yield: 34%).

[0235] Step 11: Preparation of 6-(4-amino-3-hydroxypiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (5)

[0236] Compound 5k (20 mg, 0.04 mmol) was dissolved in 2 mL of 1,4-dioxane solution, followed by the addition of 2 mL of 1,4-dioxane hydrochloride solution (4 M in 1,4-dioxane). The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 8 mg of the title compound, yield: 49%.

[0237] LC-MS(ESI)m / z(M+H) + : 462.2

[0238] Example 6 Preparation of 6-(4-(aminomethyl)-4-hydroxypiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (6)

[0239]

[0240] Step 1: Preparation of 4-(aminomethyl)-1-benzylpiperidine-4-ol (6b)

[0241] Compound 6-1 (500 mg, 2.46 mmol) was dissolved in methanol (2 mL), and then ammonia (5 mL) was added. The system was sealed and stirred at room temperature for 4 hours. The reaction system was cooled to room temperature, extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was used directly in the next reaction without purification.

[0242] Step 2: Preparation of ((1-benzyl-4-hydroxypiperidin-4-yl)methyl)tert-butyl carbamate (6c)

[0243] Compound 6b (500 mg, 2.27 mmol) was dissolved in dichloromethane (10 mL). Di-tert-butyl dicarbonate (495 mg, 2.27 mmol) was added to the system under ice bath conditions, and the reaction mixture was stirred overnight at room temperature. The reaction was quenched with water, and the mixture was extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 600 mg of the title compound. The combined yield for both steps was 76%.

[0244] Step 3: Preparation of ((4-hydroxypiperidin-4-yl)methyl)tert-butyl carbamate (6d)

[0245] Compound 6c (300 mg, 0.94 mmol) was dissolved in isopropanol (5 mL), and then palladium hydroxide on carbon (30 mg, 10%) was added at room temperature. The air in the system was replaced with hydrogen, and the system was then stirred overnight at room temperature under a hydrogen atmosphere. The reaction system was filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 100 mg of the title compound, yield: 46%.

[0246] Step 4: Preparation of tert-butyl carbamate (6e) of 1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-4-hydroxypiperidin-4-yl)methyl)carbamate

[0247] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by compound 6d (51 mg, 0.22 mmol) and Cs₂CO₃ (108 mg, 0.33 mmol) in sequence. Nitrogen gas was then added, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was purged again. The reaction mixture was stirred at 80 °C for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 22 mg of the title compound (yield: 36%).

[0248] Step 5: Preparation of 6-(4-(aminomethyl)-4-hydroxypiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (6)

[0249] Compound 6e (22 mg, 0.04 mmol) was dissolved in 2 mL of 1,4-dioxane solution, followed by the addition of 2 mL of 1,4-dioxane hydrochloride solution (4 M in 1,4-dioxane). The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 10 mg of the title compound, yield: 55%.

[0250] LC-MS(ESI)m / z(M+H) + : 476.2

[0251] 1H NMR (400MHz, DMSO-d6) δ7.86-7.77(m,1H),7.60-7.53(m,1H),7.43-7.36(m,1H),7.23-7.10(m,3H),7.00-6. 92(m,1H),4.60(brs,1H),4.21-4.07(m,2H),3.85(s,3H),3.52-3.36(m,2H),2.54(s,2H),1.58-1.39(m,4H).

[0252] Example 7 Preparation of 6-(3-aminoazacyclobutane-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (7)

[0253]

[0254] Step 1: Preparation of (1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)azacyclobutane-3-yl)tert-butyl carbamate (7a)

[0255] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of tert-butylazacyclobutane-3-ylcarbamate (38 mg, 0.22 mmol) and Cs₂CO₃ (108 mg, 0.33 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was stirred at 80 °C for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 22 mg of the title compound (yield: 40%).

[0256] Step 2: Preparation of 6-(3-aminoazacyclobutane-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (7)

[0257] Compound 7a (22 mg, 0.04 mmol) was dissolved in dichloromethane (2 mL), and then trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 15 mg of the title compound, yield: 85%.

[0258] LC-MS(ESI)m / z(M+H) + : 418.2

[0259] 1 H NMR(400MHz,DMSO-d6)δ7.81(dd,J=8.0,6.8Hz,1H),7.37(dd,J=10.4,1.2Hz,1H),7.19-7.10(m ,3H),7.09(s,1H),6.97-6.91(m,1H),4.27-4.19(m,2H),3.89-3.81(m,4H),3.70-3.63(m,2H).

[0260] Example 8 Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(1-methyl-1H-indazol-5-yl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (8)

[0261]

[0262] Step 1: Preparation of tert-butyl (1-(6-(4-cyano-3-fluorophenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-ylcarbamate (8a)

[0263] Compound 3f (287 mg, 0.78 mmol), 3-fluoro-4-cyanobenzonic acid (193 mg, 1.17 mmol), and K₂CO₃ (647 mg, 4.68 mmol) were dissolved in 1,4-dioxane (4 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (29 mg, 0.04 mmol), and nitrogen gas was introduced again. The reaction mixture was stirred at 100 °C for 2 hours. After cooling to room temperature, the reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 315 mg of the title compound (yield: 89%).

[0264] Step 2: Preparation of (1-(7-bromo-6-(4-cyano-3-fluorophenyl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (8b)

[0265] Compound 8a (310 mg, 0.69 mmol) was dissolved in acetonitrile (12 mL). NBS (246 mg, 1.38 mmol) was slowly added to the system under ice bath conditions, and the reaction mixture was stirred for 1 hour while maintaining the ice bath conditions. The reaction solution was poured into water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 364 mg of the title compound, yield: 100%.

[0266] Step 3: Preparation of (1-(6-(4-cyano-3-fluorophenyl)-7-(1-methyl-1H-indazol-5-yl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (8c)

[0267] Compound 8b (70 mg, 0.13 mmol), 1-methylindazole-5-boric acid (35 mg, 0.20 mmol), and K₂CO₃ (108 mg, 0.78 mmol) were dissolved in 1,4-dioxane (4 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was stirred at 100 °C for 1 hour. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 57 mg of the title compound (yield: 74%).

[0268] Step 4: Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(1-methyl-1H-indazol-5-yl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (8)

[0269] Compound 8c (57 mg, 0.10 mmol) was dissolved in dichloromethane (3 mL), and then trifluoroacetic acid (3 mL) was added at 0 °C. The reaction system was stirred at 0 °C for 2 h. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 15 mg of the title compound, yield: 32%.

[0270] LC-MS(ESI)m / z(M+H) + : 482.2

[0271] 1H NMR (400MHz, DMSO-d6) δ8.60(s,1H),8.02(s,1H),7.77-7.70(m,1H),7.68(s,1H),7.62(d,J=8.4Hz,1H),7.49(d,J=10.4Hz,1H),7.20(d,J=8.0Hz,1 H),7.16(d,J=8.4Hz,1H),4.61-4.51(m,2H),4.15(s,2H),4.06(s,3H),3. 30-3.17(m,1H),3.07-2.95(m,2H),2.04-1.92(m,2H),1.72-1.58(m,2H).

[0272] Example 9 Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (9)

[0273]

[0274] Step 1: Preparation of 1-(5-bromo-6-fluoro-1H-indazol-1-yl)-2-methylpropane-2-ol (9b)

[0275] Compound 9a (430 mg, 2.00 mmol) was dissolved in N,N-dimethylformamide (10 mL), followed by the sequential addition of Cs₂CO₃ (2.0 g, 6.14 mmol) and 1,1-dimethylethylene oxide (216 mg, 3.00 mmol). The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 488 mg of the title compound (yield: 85%).

[0276] Step 2: Preparation of 1-(6-fluoro-5-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)-1H-indazol-1-yl)-2-methylpropane-2-ol (9c)

[0277] Compound 9b (286 mg, 1.00 mmol), pinacol diborate (381 mg, 1.50 mmol), and potassium acetate (196 mg, 0.10 mmol) were dissolved in N,N-dimethylformamide (5 mL), and the mixture was purged with nitrogen. Triphenylphosphine (52 mg, 0.20 mmol) and Pd(AcO)₂ (23 mg, 0.10 mmol) were added sequentially, and the mixture was purged with nitrogen again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 296 mg of the title compound (yield: 89%).

[0278] Step 3: Preparation of (1-(6-(4-cyano-3-fluorophenyl)-7-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)-3-oxo)-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-4-yl)piperidin-4-yl)tert-butyl carbamate (9d)

[0279] Compound 8b (110 mg, 0.21 mmol), compound 9c (107 mg, 0.32 mmol), and K₂CO₃ (175 mg, 1.27 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.2 mL). Nitrogen gas was purged, and Pd(dppf)Cl₂ (15 mg, 0.02 mmol) was added. Nitrogen gas was purged again, and the reaction mixture was heated to 100 °C and stirred for 5 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 12 mg of the title compound (yield: 9%).

[0280] Step 4: Preparation of 4-(4-(4-aminopiperidin-1-yl)-7-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)-3-oxo-2,3-dihydro-1H-pyrrolo[3,4-c]pyridin-6-yl)-2-fluorobenzonitrile (9)

[0281] Compound 9d (12 mg, 0.02 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 6 mg of the title compound, yield: 59%.

[0282] LC-MS(ESI)m / z(M+H) + 558.2

[0283] 1 H NMR (400MHz, DMSO-d6) δ8.55 (s, 1H), 8.08 (s, 1H), 7.86 (d, J = 7.2Hz, 1H), 7.80-7.72 (m,1H),7.54(d,J=10.8Hz,1H),7.44(d,J=10.4Hz,1H),7.27(d,J=8.4Hz,1H),4.65 (s,1H),4.55-4.42(m,2H),4.40-4.22(m,3H),3.96-3.87(m,1H),3.11-2.98(m,2H) ,2.87-2.75(m,1H),1.89-1.75(m,2H),1.44-1.32(m,2H),1.16(s,3H),1.10(s,3H).

[0284] Example 10 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinamide (10)

[0285]

[0286] Step 1: Preparation of tert-butyl carbamate (10a)

[0287] Compound 4a (190 mg, 0.41 mmol) was dissolved in methanol (4 mL), followed by the addition of dimethyl sulfoxide (2 mL), NaOH (49 mg, 1.23 mmol), and H₂O₂ (232 mg, 2.05 mmol, 30% aqueous solution). The reaction mixture was stirred at 50 °C for 2 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 126 mg of the title compound (yield: 64%).

[0288] Step 2: Preparation of tert-butyl carbamate (10b)

[0289] Compound 10a (100 mg, 0.21 mmol), 3-fluoro-4-cyanobenzonic acid (53 mg, 0.32 mmol), and K₂CO₃ (87 mg, 0.63 mmol) were dissolved in 1,4-dioxane (4 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 95 mg of the title compound (yield: 81%).

[0290] Step 3: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinamide (10)

[0291] Compound 10b (95 mg, 0.17 mmol) was dissolved in ethyl acetate (2 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 31 mg of the title compound, yield: 40%.

[0292] LC-MS(ESI)m / z(M+H) + : 470.2

[0293] 1 H NMR (400MHz, DMSO-d6) δ7.98 (d, J = 0.4Hz, 1H), 7.73-7.64 (m, 2H), 7.51-7.46 ( m,2H),7.34-7.31(m,1H),7.31(dd,J=9.2,1.2Hz,1H),7.10(dd,J=8.0,1.2Hz ,1H),7.10(dd,J=8.4,1.6Hz,1H),6.91(s,1H),4.35-4.25(m,2H),4.02(s,3H ),3.03-2.93(m,2H),2.89-2.79(m,1H),1.85-1.74(m,2H),1.29-1.16(m,2H).

[0294] Example 11 Preparation of 2-(4-aminopiperidin-1-yl)-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (11)

[0295]

[0296] Step 1: Preparation of 2,6-dichloronicotinamide (11b)

[0297] Compound 11a (3.0 g, 15.63 mmol) was dissolved in chloroform (30 mL). Thionyl chloride (18.6 g, 156.30 mmol) was slowly added to the system under ice bath conditions. The system was heated to 70 °C and stirred for 3 hours. The reaction mixture was concentrated under reduced pressure and dissolved in tetrahydrofuran (10 mL). This solution was then added dropwise to a 10 mL solution of tetrahydrofuran in ammonia water (30 mL) cooled in an ice bath, and the mixture was stirred for 30 minutes. Water was added to the reaction mixture, and the mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 2.8 g of the title compound (yield: 94%).

[0298] Step 2: Preparation of (1-(3-carbamoyl-6-chloropyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (11c)

[0299] Compound 11b (2.8 g, 14.66 mmol), 4-tert-butoxycarbonylaminopiperidine (4.9 g, 24.47 mmol), and triethylamine (3.0 g, 29.65 mmol) were dissolved in acetonitrile (30 mL). The system was heated to 70 °C and stirred for 3 hours. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 4.4 g of the title compound, yield: 88%.

[0300] Step 3: Preparation of (1-(3-carbamoyl-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (11d)

[0301] Compound 11c (400 mg, 1.13 mmol), 3-fluoro-4-cyanobenzonic acid (280 mg, 1.70 mmol), and K₂CO₃ (937 mg, 6.78 mmol) were dissolved in 1,4-dioxane (5 mL) and water (0.5 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (41 mg, 0.06 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred for 4 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 448 mg of the title compound (yield: 90%).

[0302] Step 4: Preparation of tert-butyl carbamate (11e)

[0303] Compound 11d (448 mg, 1.02 mmol) was dissolved in N,N-dimethylformamide (5 mL). NBS (200 mg, 1.12 mmol) was slowly added to the system under ice bath conditions, and the reaction mixture was stirred at 0 °C for 2 hours. The reaction solution was poured into water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 528 mg of the title compound (yield: 100%).

[0304] Step 5: Preparation of tert-butyl carbamate (11f)

[0305] Compound 11e (518 mg, 1.00 mmol), 1-methylindazole-5-boronic acid (263 mg, 1.49 mmol), and K₂CO₃ (829 mg, 6.00 mmol) were dissolved in 1,4-dioxane (6 mL) and water (0.6 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (37 mg, 0.05 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 423 mg of the title compound (yield: 74%).

[0306] Step 6: Preparation of 2-(4-aminopiperidin-1-yl)-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (11)

[0307] Compound 11f (30 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 6 mg of the title compound, yield: 24%.

[0308] LC-MS(ESI)m / z(M+H) + : 470.2

[0309] 1 H NMR(400MHz,DMSO-d6)δ8.02(s,1H),8.00-7.95(m,1H),7.79(s,1H),7.79-7. 73(m,1H),7.68-7.62(m,2H),7.58-7.54(m,1H),7.50-7.45(m,1H),7.22(dd, J=8.4,1.6Hz,1H),7.22(dd,J=8.8,1.6Hz,1H),4.04(s,3H),3.91-3.80(m,2H ),3.02-2.90(m,2H),2.84-2.73(m,1H),1.86-1.74(m,2H),1.43-1.32(m,2H).

[0310] Example 12 Preparation of 2-(4-aminopiperidin-1-yl)-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)nicotinonitrile (12)

[0311]

[0312] Step 1: Preparation of tert-butyl carbamate (12a)

[0313] Compound 11f (200 mg, 0.35 mmol) was dissolved in tetrahydrofuran (6 mL), and nitrogen gas was introduced. Under ice bath conditions, a solution of pyridine (194 mg, 2.45 mmol) and trifluoroacetic anhydride (147 mg, 0.70 mmol) in dichloromethane (1 mL) was added sequentially. The reaction mixture was slowly brought to room temperature and stirred overnight. The reaction solution was poured into water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 10 mg of the title compound, yield: 5%.

[0314] Step 2: Preparation of 2-(4-aminopiperidin-1-yl)-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)nicotinonitrile (12)

[0315] Compound 12a (10 mg, 0.02 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 1 mg of the title compound, yield: 12%.

[0316] LC-MS(ESI)m / z(M+H) + : 452.2

[0317] Example 13 Preparation of 6-(4-amino-3-fluoropiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (13)

[0318]

[0319] Step 1: Preparation of tert-butyl 4-amino-3-fluoropiperidine-1-carboxylic acid (13b)

[0320] Compound 13-1 (1.0 g, 4.60 mmol) was dissolved in methanol (10 mL), and ammonium acetate (2.5 g, 32.43 mmol) and sodium cyanoborohydride (375 mg, 5.97 mmol) were added sequentially. The reaction mixture was stirred overnight at room temperature. The mixture was diluted with water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 500 mg of the title compound, yield: 50%.

[0321] Step 2: Preparation of 4-(((benzyloxy)carbonyl)amino)-3-fluoropiperidine-1-carboxylic acid tert-butyl ester (13c)

[0322] Compound 13b (500 mg, 2.29 mmol) was dissolved in ethyl acetate (5 mL), followed by the addition of saturated sodium bicarbonate solution (10 mL). Benzyl chloroformate (391 mg, 2.29 mmol) was added to the system under ice bath conditions, and the reaction mixture was stirred overnight at room temperature. The reaction solution was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 420 mg of the title compound (yield: 52%).

[0323] Step 3: Preparation of (3-fluoropiperidin-4-yl)carbamate benzyl hydrochloride (13d)

[0324] Compound 13c (200 mg, 0.57 mmol) was dissolved in ethyl acetate (10 mL), and then hydrochloric acid-ethyl acetate solution (4 mL, 3 M in EA) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, and the crude product was used directly in the next step without purification.

[0325] Step 4: Preparation of (1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-3-fluoropiperidin-4-yl)carbamate (13e)

[0326] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of compound 13d (64 mg, 0.22 mmol) and t-BuONa (53 mg, 0.55 mmol). Nitrogen was then introduced, followed by the addition of RuPhos-Pd-G3 (9 mg, 0.01 mmol), and the reaction mixture was stirred at 80 °C for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 21 mg of the title compound (yield: 33%).

[0327] Step 5: Preparation of 6-(4-amino-3-fluoropiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (13)

[0328] Compound 13e (21 mg, 0.04 mmol) was dissolved in ethyl acetate (2 mL), and then Pd / C (2 mg, 10%) was added at room temperature. The air in the system was replaced with hydrogen, and the system was then heated to 35 °C and stirred for 4 hours under a hydrogen atmosphere. The reaction system was filtered and concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to give 10 mg of the title compound, yield: 61%.

[0329] LC-MS(ESI)m / z(M+H) + : 464.2

[0330] 1 H NMR(400MHz, DMSO-d6)δ7.95-7.78(m,1H),7.64(s,1H),7.41(dd,J=10.8,1.6Hz,1H),7.23-7.11(m,3H),7.00-6.94(m,1H),4 .22-4.15(m,2H),4.10-4.00(m,1H),3.85(s,3H),3.45-3.33(m,2H),3.03-2.92(m,1H),1.93-1.82(m,1H),1.41-1.30(m,1H).

[0331] Example 14 Preparation of 6-(4-aminocyclohexyl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (14)

[0332]

[0333] Step 1: Preparation of (4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)cyclohex-3-en-1-yl)tert-butyl carbamate (14a)

[0334] Compound 5j (80 mg, 0.17 mmol) was dissolved in 1,4-dioxane (3 mL) and water (0.5 mL). Then, (4-(4,4,5,5-tetramethyl-1,3,2-dioxaboran-2-yl)cyclohex-3-en-1-yl)tert-butyl carbamate (165 mg, 0.51 mmol) and K₂CO₃ (70 mg, 0.51 mmol) were added sequentially, purging with nitrogen. Pd(dppf)Cl₂ (15 mg, 0.02 mmol) was added, and nitrogen was purged again. The system was heated to 80 °C and stirred for 2 hours. The reaction mixture was cooled to room temperature, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 79 mg of the title compound (yield: 86%).

[0335] Step 2: Preparation of (4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)cyclohexyl)tert-butyl carbamate (14b)

[0336] Compound 14a (40 mg, 0.07 mmol) was dissolved in ethyl acetate (2 mL), and then Pd / C (4 mg, 10%) was added at room temperature. The air in the system was replaced with hydrogen, and the system was stirred overnight at room temperature under a hydrogen atmosphere. The reaction system was filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to give 23 mg of the title compound, yield: 57%.

[0337] Step 3: Preparation of 6-(4-aminocyclohexyl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (14)

[0338] Compound 14b (23 mg, 0.04 mmol) was dissolved in ethyl acetate (5 mL), and then hydrochloric acid-ethyl acetate solution (4 mL, 3 M in EA) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 11 mg of the title compound, yield: 59%.

[0339] LC-MS(ESI)m / z(M+H) + : 445.2

[0340] 1 H NMR(400MHz,DMSO-d6)δ8.08(s,0.48H),8.01(s,0.47H),7.89-7.82(m,1 H),7.48-7.41(m,1H),7.55-7.28(m,1H),7.25-7.16(m,2H),7.11-7.05( m,1H),3.88-3.84(m,3H),3.16-3.10(m,0.5H),2.95-2.86(m,0.5H),2.8 6-2.68(m,1H),2.15-1.89(m,3H),1.75-1.60(m,4H),1.33-1.21(m,1H).

[0341] Example 15 Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazacycloheptane-1-yl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (15)

[0342]

[0343] Step 1: Preparation of tert-butyl 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-1,4-diazacycloheptane-1-carboxylic acid (15a)

[0344] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the addition of tert-butyl 1,4-diazacycloheptan-1-carboxylate (44 mg, 0.22 mmol) and Cs₂CO₃ (108 mg, 0.33 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure to remove the solvent, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 24 mg of the title compound (yield: 42%).

[0345] Step 2: Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazacycloheptane-1-yl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (15)

[0346] Compound 15a (24 mg, 0.04 mmol) was dissolved in ethyl acetate (5 mL), and then hydrochloric acid-ethyl acetate solution (4 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 10 mg of the title compound, yield: 51%.

[0347] LC-MS(ESI)m / z(M+H) + : 446.2

[0348] 1 H NMR(400MHz,DMSO-d6)δ7.85-7.79(m,1H),7.42-7.34(m,2H),7.23-7.11(m,3H),7.00-6.93(m ,1H),3.85(s,3H),3.80-3.68(m,4H),2.93-2.86(m,2H),2.76-2.69(m,2H),1.84-1.73(m,2H).

[0349] Example 16 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (16)

[0350]

[0351] Step 1: Preparation of (1-(4-cyano-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (16a)

[0352] Compound 1b (1.7 g, 5.05 mmol), 3-fluoro-4-cyanobenzoic acid (1.2 g, 7.28 mmol), and K₂CO₃ (2.1 g, 15.20 mmol) were dissolved in acetonitrile (20 mL) and water (10 mL). Nitrogen gas was introduced, followed by the addition of Pd(PPh₃)₄ (289 mg, 0.25 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.8 g of the title compound (yield: 85%).

[0353] Step 2: Preparation of (1-(5-bromo-4-cyano-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (16b)

[0354] Compound 16a (421 mg, 1.00 mmol) was dissolved in acetonitrile (15 mL). NBS (182 mg, 1.02 mmol) was slowly added to the system under ice bath conditions. The reaction system was stirred at room temperature for 3 hours. The reaction system was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography to give 450 mg of the title compound, yield: 90%.

[0355] Step 3: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (16c)

[0356] Compound 16b (100 mg, 0.20 mmol), compound 9c (100 mg, 0.30 mmol), and K₂CO₃ (83 mg, 0.60 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.2 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 30 mg of the title compound (yield: 24%).

[0357] Step 4: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (16)

[0358] Compound 16c (30 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 24 mg of the title compound, yield: 95%.

[0359] LC-MS(ESI)m / z(M+H) + 528.2

[0360] 1 H NMR (400MHz, DMSO-d6) δ8.11(s,1H),7.80-7.72(m,2H),7.61(s,1H),7.57(d,J=10.8Hz,1H),7.40-7.34(m,1H),7.18(dd,J=8.0,1.2Hz,1 H),4.68(s,1H),4.40-4.24(m,4H),3.12-3.01(m,2H),2.92-2.82(m,1H),1.85-1.75(m,2H),1.27-1.19(m,2H),1.14(s,3H),1.10(s,3H).

[0361] Example 17 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinamide (17)

[0362]

[0363] Step 1: Preparation of tert-butyl(1-(6-chloro-4-cyano-5-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (17a)

[0364] Compound 1c (229 mg, 0.55 mmol), compound 9c (368 mg, 1.10 mmol), and K₂CO₃ (228 mg, 1.65 mmol) were dissolved in 1,4-dioxane (5 mL) and water (1 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (44 mg, 0.06 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 138 mg of the title compound (yield: 46%).

[0365] Step 2: Preparation of tert-butyl (1-(4-carbamoyl-6-chloro-5-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (17b)

[0366] Compound 17a (138 mg, 0.25 mmol) was dissolved in methanol (1 mL), followed by the sequential addition of dimethyl sulfoxide (1 mL), NaOH (30 mg, 0.75 mmol), and H₂O₂ (170 mg, 1.50 mmol, 30% aqueous solution). The reaction mixture was stirred at 50 °C for 2 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 126 mg of the title compound (yield: 88%).

[0367] Step 3: Preparation of tert-butyl (1-(4-carbamoyl-6-(4-cyano-3-fluorophenyl)-5-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (17c)

[0368] Compound 17b (126 mg, 0.22 mmol), 3-fluoro-4-cyanobenzonic acid (54 mg, 0.33 mmol), and K₂CO₃ (91 mg, 0.66 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 74 mg of the title compound (yield: 51%).

[0369] Step 4: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinamide (17)

[0370] Compound 17c (74 mg, 0.11 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 24 mg of the title compound, yield: 38%.

[0371] LC-MS(ESI)m / z(M+H) + 546.2

[0372] 1 H NMR (400MHz, DMSO-d6) δ8.03 (d, J = 0.8, 1H), 7.85 (s, 1H), 7.25-7.67 (m, 1H), 7.5 9(d,J=7.2,1H),7.42-7.34(m,2H),7.27(dd,J=10.4,1.2Hz,1H),7.16(dd,J=8.0 ,1.6Hz,1H),6.96(s,1H),4.66(s,1H),4.37-4.19(m,4H),3.04-2.94(m,2H),2. 88-2.48(m,1H),1.85-1.75(m,2H),1.27-1.19(m,2H),1.13(s,3H),1.07(s,3H).

[0373] Example 18 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-2-oxoindol-5-yl)isonicotinonitrile (18)

[0374]

[0375] Step 1: Preparation of tert-butyl carbamate (18a)

[0376] Compound 18b (75 mg, 0.15 mmol), 1-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxoboronyl-2-yl)dihydroindole-2-one (63 mg, 0.23 mmol), and K₂CO₃ (62 mg, 0.46 mmol) were dissolved in 1,4-dioxane (1 mL) and water (0.2 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 36 mg of the title compound (yield: 42%).

[0377] Step 2: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-2-oxoindol-5-yl)isonicotinonitrile (18)

[0378] Compound 18a (36 mg, 0.06 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to obtain 18 mg of the title compound, yield: 61%.

[0379] LC-MS(ESI)m / z(M+H) + : 467.2

[0380] 1H NMR (400MHz, DMSO-d6) δ7.77(dd,J=8.0,7.2Hz,1H),7.53(s,1H),7.39(dd,J=10.8,1.2Hz,1H),7.19-7.12(m,2H),7.09-7.07(m,1H),7.0 0-6.96(m,1H),4.34-4.26(m,2H),3.51(s,2H),3.12(s,3H),3.08-2.98(m,2H),2.89-2.80(m,1H),1.82-1.74(m,2H),1.26-1.17(m,2H).

[0381] Example 19 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(2-fluoro-4-(2-hydroxy-2-methylpropyl)phenyl)isonicotinonitrile (19)

[0382]

[0383] Step 1: Preparation of methyl 2-(4-bromo-3-fluorophenyl)acetate (19b)

[0384] Compound 19a (1.0 g, 4.29 mmol) was dissolved in acetonitrile (40 mL), followed by the sequential addition of K₂CO₃ (1.8 g, 13.02 mmol) and methyl iodoform (1.5 g, 10.57 mmol). The reaction mixture was heated to 60 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.0 g of the title compound (yield: 94%).

[0385] Step 2: Preparation of 1-(4-bromo-3-fluorophenyl)-2-methylpropane-2-ol (19c)

[0386] Compound 19b (815 mg, 3.30 mmol) was dissolved in dry tetrahydrofuran (30 mL), and then methylmagnesium bromide (5.5 mL, 3 M in THF) was slowly added dropwise under an ice-water bath. After the addition was complete, the reaction system was slowly warmed to room temperature and stirred overnight. The reaction system was cooled to 0 °C, and an appropriate amount of water was slowly added to quench the reaction. The mixture was filtered, and the filtrate was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 366 mg of the title compound, yield: 45%.

[0387] Step 3: Preparation of 1-(3-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)phenyl)-2-methylpropane-2-ol (19d)

[0388] Compound 19c (360 mg, 1.46 mmol), pinacol diborate (741 mg, 2.92 mmol), and potassium acetate (429 mg, 4.37 mmol) were dissolved in 1,4-dioxane (15 mL), and the mixture was purged with nitrogen. Pd(dppf)Cl2 (107 mg, 0.15 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was heated to 100 °C and stirred for 24 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 177 mg of the title compound (yield: 41%).

[0389] Step 4: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(2-fluoro-4-(2-hydroxy-2-methylpropyl)phenyl)pyridin-2-yl)piperidin-4-yl)carbamate (19e)

[0390] Compound 16b (100 mg, 0.20 mmol), compound 19d (118 mg, 0.40 mmol), and K₂CO₃ (83 mg, 0.60 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.2 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 50 mg of the title compound (yield: 43%).

[0391] Step 5: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(2-fluoro-4-(2-hydroxy-2-methylpropyl)phenyl)isonicotinonitrile (19)

[0392] Compound 19e (50 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 36 mg of the title compound as a yellow solid, yield: 87%.

[0393] LC-MS(ESI)m / z(M+H) + : 488.2

[0394] 1 H NMR(400MHz,DMSO-d6)δ7.82-7.76(m,1H),7.59(s,1H),7.27-7.17(m,3H),7.08(s,1H),7.07-7.04(m,1H),4.56(s,1H),4.37-4 .27(m,2H),3.10-3.00(m,2H),2.91-2.80(m,1H),2.68(s,2H),1.85-1.74(m,2H),1.27-1.17(m,2H),1.04(s,3H),1.03(s,3H).

[0395] Example 20 Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazaaniline-1-yl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (20)

[0396]

[0397] Step 1: Preparation of tert-butyl 4-(6-chloro-4-cyanopyridin-2-yl)-1,4-diazane-1-carboxylic acid (20a)

[0398] Compound 1a (1.5 g, 8.67 mmol), tert-butyl 1,4-diazacycloheptan-1-carboxylate (1.7 g, 8.49 mmol), and K₂CO₃ (2.4 g, 17.36 mmol) were dissolved in acetonitrile (20 mL). The system was heated to 80 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 2.1 g of the title compound, yield: 72%.

[0399] Step 2: Preparation of tert-butyl 4-(4-cyano-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (20b)

[0400] Compound 20a (1.0 g, 2.97 mmol), 3-fluoro-4-cyanobenzonic acid (735 mg, 4.46 mmol), and Na₂CO₃ (630 mg, 5.94 mmol) were dissolved in acetonitrile (10 mL) and water (5 mL). Nitrogen gas was introduced, followed by the addition of Pd(PPh₃)₄ (173 mg, 0.15 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 85 °C and stirred for 6 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 800 mg of the title compound (yield: 64%).

[0401] Step 3: Preparation of tert-butyl 4-(5-bromo-4-cyano-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (20c)

[0402] Compound 20b (500 mg, 1.19 mmol) was dissolved in acetonitrile (5 mL). NBS (211 mg, 1.19 mmol) was slowly added to the system under ice bath conditions. The reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography to give 420 mg of the title compound, yield: 71%.

[0403] Step 4: Preparation of tert-butyl 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (20d)

[0404] Compound 20c (50 mg, 0.10 mmol), 1-methylindazole-5-boric acid (53 mg, 0.30 mmol), and K₂CO₃ (41 mg, 0.30 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (7 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 80 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 30 mg of the title compound (yield: 54%).

[0405] Step 5: Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazaaniline-1-yl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (20)

[0406] Compound 20d (30 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to obtain 20 mg of the title compound, yield: 82%.

[0407] LC-MS(ESI)m / z(M+H) + : 452.2

[0408] 1 H NMR (400MHz, DMSO-d6) δ8.07-8.05(m,1H),7.75-7.70(m,1H),7.69-7.66(m,1H),7.63-7.59(m,1H),7.40-7.35(m ,2H),7.18-7.13(m,2H),4.06(s,3H),3.83-3.67(m,4H),2.93-2.87(m,2H),2.76-2.69(m,2H),1.85-1.74(m,2H).

[0409] Example 21 Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazaaniline-1-yl)-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (21)

[0410]

[0411] Step 1: Preparation of tert-butyl 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid ester (21a)

[0412] Compound 20c (50 mg, 0.10 mmol), compound 9c (100 mg, 0.30 mmol), and K₂CO₃ (41 mg, 0.30 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (7 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 80 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 10 mg of the title compound (yield: 16%).

[0413] Step 2: Preparation of 2-(4-cyano-3-fluorophenyl)-6-(1,4-diazaaniline-1-yl)-3-(6-fluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (21)

[0414] Compound 21a (10 mg, 0.02 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 4 mg of the title compound, yield: 48%.

[0415] LC-MS(ESI)m / z(M+H) + 528.2

[0416] 1H NMR(400MHz, DMSO-d6)δ8.11(s,1H),7.79-7.73(m,2H),7.58(d,J=10.4Hz,1H),7.41(s,1H),7.39-7.33(m,1H),7.21-7.16(m,1H),4 .67(s,1H),4.32-4.23(m,2H),3.88-3.67(m,4H),2.94-2.85(m,2H),2.76-2.68(m,2H),1.86-1.72(m,2H),1.15(s,3H),1.10(s,3H).

[0417] Example 22 Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(2,7-diazaspiro[3.5]non-7-yl)isonicotinonitrile (22)

[0418]

[0419] Step 1: Preparation of 7-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-2,7-diazaspirocyclic[3.5]nonane-2-carboxylic acid tert-butyl ester (22a)

[0420] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of 2-tert-butoxycarbonyl-2,7-diazaspiro[3.5]nonane (50 mg, 0.22 mmol) and Cs₂CO₃ (107 mg, 0.33 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 30 mg of the title compound (yield: 50%).

[0421] Step 2: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(2,7-diazaspiro[3.5]non-7-yl)isonicotinonitrile (22)

[0422] Compound 22a (30 mg, 0.05 mmol) was dissolved in dichloromethane (2 mL), and then trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 15 mg of the title compound, yield: 61%.

[0423] LC-MS(ESI)m / z(M+H) + : 472.2

[0424] 1 H NMR(400MHz,DMSO-d6)δ7.85-7.78(m,1H),7.59(s,1H),7.44-7.36(m,1H),7.22-7 .11(m,3H),6.99-6.93(m,1H),3.85(s,3H),3.72-3.56(m,8H),1.85-1.67(m,4H).

[0425] Example 23 Preparation of 6-(4-aminocyclohexyl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (23-1,23-2)

[0426]

[0427] Step 1: Preparation of 6-amino-2-chloro-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (23a)

[0428] 5 g (1.5 g, 6.45 mmol) of the compound was dissolved in 20 mL of 1,4-dioxane and 3 mL of water. Then, 1.1 g (6.25 mmol) of 1-methylindazole-5-boric acid and 1.8 g (13.02 mmol) of K₂CO₃ were added sequentially, and nitrogen was purged. Pd(dppf)Cl₂ (472 mg, 0.64 mmol) was added, and nitrogen was purged again. The reaction mixture was stirred at 80 °C for 3 hours. The reaction mixture was cooled to room temperature, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.1 g of the title compound, yield: 60%.

[0429] Step 2: Preparation of 6-amino-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (23b)

[0430] Compound 23a (1.1 g, 3.88 mmol) was dissolved in 1,4-dioxane (30 mL) and water (3 mL), followed by the addition of 3-fluoro-4-cyanobenzonic acid (1.9 g, 11.52 mmol) and K₂CO₃ (1.6 g, 11.58 mmol) to purge with nitrogen. Pd(dppf)Cl₂ (278 mg, 0.38 mmol) was then added, followed by another purge with nitrogen. The reaction mixture was stirred overnight at 80 °C. The reaction mixture was cooled to room temperature, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.1 g of the title compound (yield: 77%).

[0431] Step 3: Preparation of 2-(4-cyano-3-fluorophenyl)-6-iodo-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (23c)

[0432] Compound 23b (500 mg, 1.36 mmol) was dissolved in acetonitrile (10 mL), followed by the addition of potassium iodide (451 mg, 2.72 mmol) and amyl nitrite (319 mg, 2.72 mmol). The reaction mixture was stirred overnight at 80 °C. After cooling to room temperature, the reaction solution was concentrated under reduced pressure. The solution was diluted with water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 90 mg of the title compound (yield: 14%).

[0433] Step 4: Preparation of tert-butyl carbamate (23d)

[0434] Compound 23c (90 mg, 0.19 mmol) was dissolved in 1,4-dioxane (3 mL) and water (0.6 mL). Then, (4-(4,4,5,5-tetramethyl-1,3,2-dioxaboran-2-yl)cyclohex-3-en-1-yl)tert-butyl carbamate (184 mg, 0.57 mmol) and K₂CO₃ (79 mg, 0.57 mmol) were added sequentially, and nitrogen was purged. Pd(dppf)Cl₂ (14 mg, 0.02 mmol) was added, and nitrogen was purged again. The reaction system was stirred at 80 °C for 2 hours. The reaction system was cooled to room temperature, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 80 mg of the title compound, yield: 78%.

[0435] Step 5: Preparation of (4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)pyridin-2-yl)cyclohexyl)tert-butyl carbamate (23e)

[0436] Compound 23d (80 mg, 0.14 mmol) was dissolved in ethyl acetate (3 mL), and then Pd / C (8 mg, 10%) was added at room temperature. The air in the system was replaced with hydrogen, and the system was stirred overnight at room temperature under a hydrogen atmosphere. The reaction system was filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to give 67 mg of the title compound, yield: 83%.

[0437] Step 6: Preparation of 6-(4-aminocyclohexyl)-2-(4-cyano-3-fluorophenyl)-3-(1-methyl-1H-indazol-5-yl)isonicotinonitrile (23-1,23-2)

[0438] Compound 23e (67 mg, 0.12 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (3 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography (DCM / MeOH = 10:1) to give 10 mg of the title compound 23-1 (top point: R). f =0.3), yield: 18%, and 10 mg of title compound 23-2 (bottom point: R) f =0.25), yield: 18%.

[0439] Top point:

[0440] LC-MS(ESI)m / z(M+H) + : 451.2

[0441] 1 H NMR (400MHz, DMSO-d6) δ8.13-8.08(m,2H),7.83-7.80(m,1H),7.79-7.73(m,1H),7.69-7.64(m,1H),7.49-7.44(m,1H),7.26-7.22(m ,1H),7.21-7.16(m,1H),4.07(s,3H),3.24-3.18(m,1H),3.03-2.92(m,1H),2.15-2.02(m,2H),1.80-1.60(m,4H),1.25-1.10(m,2H).

[0442] Next point:

[0443] LC-MS(ESI)m / z(M+H) + : 451.2

[0444] 1 H NMR (400MHz, DMSO-d6) δ8.12-8.10(m,1H),8.03(s,1H),7.82-7.80(m,1H),7.79-7.73(m,1H),7.68-7.64(m,1H),7.46-7.41(m,1H ),7.26-7.22(m,1H),7.20-7.16(m,1H),4.07(s,3H),2.88-2.72(m,2H),2.03-1.91(m,4H),1.75-1.62(m,2H),1.36-1.25(m,2H).

[0445] Example 24 Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(7-oxo-1,4-diaza-1-yl)isonicotinonitrile (24)

[0446]

[0447] Preparation of step one: 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-5-oxy-1,4-diazane-1-carboxylic acid tert-butyl ester (24a)

[0448] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of 1-tert-butoxycarbonyl-1,4-diaza-5-cycloheptanone (47 mg, 0.22 mmol) and Cs₂CO₃ (107 mg, 0.33 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 40 mg of the title compound (yield: 68%).

[0449] Step 2: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(7-oxo-1,4-diaza-1-yl)isonicotinonitrile (24)

[0450] Compound 24a (40 mg, 0.07 mmol) was dissolved in dichloromethane (2 mL), and then trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 16 mg of the title compound, yield: 49%.

[0451] LC-MS(ESI)m / z(M+H) + 460.2

[0452] 1H NMR (400MHz, DMSO-d6) δ8.29(s,1H),7.89-7.82(m,1H),7.49(dd,J=10.4,1.2Hz,1H),7.31(dd,J=12.0 ,2.0Hz,1H),7.25-7.18(m,2H),7.11-7.05(m,1H),4.29-4.19(m,2H),3.85(s,3H),3.03-2.94(m,4H), 2.91-2.84(m,2H).

[0453] Example 25 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(2,2-difluorobenzo[1,3-d]dioxane-5-yl)isonicotinonitrile (25)

[0454]

[0455] Step 1: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(2,2-difluorobenzo[1,3-d]dioxane-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (25a)

[0456] Compound 16b (75 mg, 0.15 mmol), (2,2-difluorobenzo[1,3-d]dioxacyclopenten-5-yl)boronic acid (46 mg, 0.23 mmol), and K₂CO₃ (62 mg, 0.45 mmol) were dissolved in 1,4-dioxane (1.5 mL) and water (0.3 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 53 mg of the title compound (yield: 61%).

[0457] Step 2: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(2,2-difluorobenzo[1,3-d]dioxane-5-yl)isonicotinonitrile (25)

[0458] Compound 25a (53 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 34 mg of the title compound, yield: 78%.

[0459] LC-MS(ESI)m / z(M+H) + : 478.2

[0460] 1 H NMR (400MHz, DMSO-d6) δ7.80(dd,J=8.0,6.8Hz,1H),7.57(s,1H),7.44(d,J=1.6Hz,1H),7.43-7.38(m,2H),7.15(dd,J=8.0,1.2Hz ,1H),7.04(dd,J=8.0,1.2Hz,1H),4.36-4.26(m,2H),3.10-3.00(m,2H),2.90-2.80(m,1H),1.83-1.73(m,2H),1.26-1.14(m,2H).

[0461] Example 26 Preparation of 6-(4-cyano-3-fluorophenyl)-2-(1,4-diaza-1-yl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (26)

[0462]

[0463] Step 1: Preparation of tert-butyl 4-(3-carbamoyl-6-chloropyridin-2-yl)-1,4-diazane-1-carboxylic acid (26a)

[0464] Compound 11b (550 mg, 2.88 mmol), tert-butyl 1,4-diazacycloheptan-1-carboxylate (572 mg, 2.86 mmol), and K₂CO₃ (792 mg, 5.73 mmol) were dissolved in acetonitrile (7 mL). The system was heated to 80 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 620 mg of the title compound, yield: 61%.

[0465] Step 2: Preparation of tert-butyl 4-(3-carbamoyl-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (26b)

[0466] Compound 26a (620 mg, 1.75 mmol), 3-fluoro-4-cyanobenzonic acid (433 mg, 2.63 mmol), and K₂CO₃ (723 mg, 5.23 mmol) were dissolved in 1,4-dioxane (7 mL) and water (0.7 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (64 mg, 0.09 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 754 mg of the title compound (yield: 98%).

[0467] Step 3: Preparation of tert-butyl 4-(5-bromo-3-carbamoyl-6-(4-cyano-3-fluorophenyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (26c)

[0468] Compound 26b (754 mg, 1.72 mmol) was dissolved in N,N-dimethylformamide (9 mL). NBS (609 mg, 3.42 mmol) was slowly added to the system under ice bath conditions. The reaction mixture was stirred at room temperature for 2 hours. The reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 315 mg of the title compound, yield: 35%.

[0469] Step 4: Preparation of tert-butyl 4-(3-carbamoyl-6-(4-cyano-3-fluorophenyl)-5-(1-methyl-1H-indazol-5-yl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (26d)

[0470] Compound 26c (160 mg, 0.31 mmol), 1-methylindazole-5-boric acid (82 mg, 0.47 mmol), and K₂CO₃ (128 mg, 0.93 mmol) were dissolved in 1,4-dioxane (3 mL) and water (0.3 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (12 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 145 mg of the title compound (yield: 82%).

[0471] Step 5: Preparation of 6-(4-cyano-3-fluorophenyl)-2-(1,4-diaza-1-yl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (26)

[0472] Compound 26d (72 mg, 0.13 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 2 hours. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 49 mg of the title compound, yield: 83%.

[0473] LC-MS(ESI)m / z(M+H) + : 470.2

[0474] 1 H NMR(400MHz,DMSO-d6)δ8.02(s,1H),8.02-7.98(m,1H),7.79-7.74(m,1H) ,7.73-7.69(m,1H),7.68-7.64(m,1H),7.58-7.53(m,1H),7.51-7.41(m,2H ),7.26-7.21(m,1H),7.14-7.08(m,1H),4.04(s,3H),3.76-3.66(m,2H),3 .56-3.49(m,2H),2.98-2.91(m,2H),2.77-2.70(m,2H),1.91-1.78(m,2H).

[0475] Example 27 Preparation of 6-(4-cyano-3-fluorophenyl)-2-(1,4-diazaaniline-1-yl)-N,N-dimethyl-5-(1-methyl-1H-indazol-5-yl)nicotinamide (27)

[0476]

[0477] Step 1: Preparation of 2,6-dichloro-N,N-dimethylnicotinamide (27a)

[0478] Compound 11a (500 mg, 2.60 mmol) was dissolved in tetrahydrofuran (5 mL), followed by the addition of dimethylamine hydrochloride (636 mg, 7.80 mmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (1.2 g, 3.16 mmol), and N,N-diisopropylethylamine (1.0 g, 7.74 mmol). The mixture was stirred overnight at room temperature. The reaction solution was poured into water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give 570 mg of the title compound, which was used directly in the next reaction without purification.

[0479] Step 2: Preparation of tert-butyl 4-(6-chloro-3-(dimethylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (27b)

[0480] Compound 27a (570 mg, 2.60 mmol), tert-butyl 1,4-diazacycloheptan-1-carboxylate (518 mg, 2.59 mmol), and K₂CO₃ (708 mg, 5.12 mmol) were dissolved in acetonitrile (7 mL). The system was heated to 80 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 653 mg of the title compound. The overall yield of the two steps was 65%.

[0481] Step 3: Preparation of tert-butyl 4-(6-(4-cyano-3-fluorophenyl)-3-(dimethylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (27c)

[0482] Compound 27b (520 mg, 1.36 mmol), 3-fluoro-4-cyanobenzonic acid (335 mg, 2.03 mmol), and K₂CO₃ (559 mg, 4.04 mmol) were dissolved in 1,4-dioxane (7 mL) and water (0.7 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (50 mg, 0.07 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 399 mg of the title compound (yield: 63%).

[0483] Step 4: Preparation of tert-butyl 4-(5-bromo-6-(4-cyano-3-fluorophenyl)-3-(dimethylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (27d)

[0484] Compound 27c (389 mg, 0.83 mmol) was dissolved in N,N-dimethylformamide (4 mL). NBS (295 mg, 1.66 mmol) was slowly added to the system under ice bath conditions, and the reaction mixture was stirred at room temperature for 2 hours. The reaction solution was poured into water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 311 mg of the title compound (yield: 68%).

[0485] Step 5: Preparation of tert-butyl 4-(6-(4-cyano-3-fluorophenyl)-3-(dimethylcarbamoyl)-5-(1-methyl-1H-indazol-5-yl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid ester (27e)

[0486] Compound 27d (160 mg, 0.29 mmol), 1-methylindazole-5-boric acid (76 mg, 0.43 mmol), and K₂CO₃ (120 mg, 0.87 mmol) were dissolved in 1,4-dioxane (3 mL) and water (0.3 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (11 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 53 mg of the title compound (yield: 30%).

[0487] Step 6: Preparation of 6-(4-cyano-3-fluorophenyl)-2-(1,4-diazaaniline-1-yl)-N,N-dimethyl-5-(1-methyl-1H-indazol-5-yl)nicotinamide (27)

[0488] Compound 27e (53 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 3 hours. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 31 mg of the title compound, yield: 70%.

[0489] LC-MS(ESI)m / z(M+H) + 498.3

[0490] 1H NMR(400MHz,DMSO-d6)δ8.01(s,1H),7.79-7.73(m,1H),7.65(s,1H),7.57-7.52(m,2H),7.49-7.43(m,1H),7.26-7.21(m,1H),7.1 2-7.06(m,1H),4.03(s,3H),3.73-3.40(m,4H),3.00(s,3H),2.95(s,3H),2.94-2.89(m,2H),2.79-2.71(m,2H),1.90-1.74(m,2H).

[0491] Example 28 Preparation of 6-(4-cyano-3-fluorophenyl)-N-cyclopropyl-2-((1,4-diazaaniline-1-yl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (28)

[0492]

[0493] Step 1: Preparation of 2,6-dichloro-N-cyclopropylnicotinamide (28a)

[0494] Compound 11a (500 mg, 2.60 mmol) was dissolved in chloroform (10 mL), and nitrogen was purged. Thionyl chloride (3.1 g, 26.00 mmol) was slowly added to the system under ice bath conditions, and nitrogen was purged again. The system was heated to 70 °C and stirred for 2 hours. The reaction mixture was concentrated under reduced pressure and dissolved in dichloromethane (1 mL), then added dropwise to a 1 mL solution of cyclopropylamine (327 mg, 5.73 mmol) in dichloromethane, cooled in an ice bath. The mixture was stirred for 1 hour. Cold water was added to the reaction mixture, and a solid precipitated. After filtration, 600 mg of the title compound was obtained and used directly in the next reaction without purification.

[0495] Step 2: Preparation of tert-butyl 4-(6-chloro-3-(cyclopropylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (28b)

[0496] Compound 28a (600 mg, 2.60 mmol), tert-butyl 1,4-diazacycloheptan-1-carboxylate (520 mg, 2.60 mmol), and K₂CO₃ (718 mg, 5.20 mmol) were dissolved in acetonitrile (8 mL). The system was heated to 80 °C and stirred overnight. The reaction system was cooled to room temperature, and the reaction solution was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 913 mg of the title compound. The overall yield of the two steps was 89%.

[0497] Step 3: Preparation of 4-(6-(4-cyano-3-fluorophenyl)-3-(cyclopropylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid tert-butyl ester (28c)

[0498] Compound 28b (520 mg, 1.32 mmol), 3-fluoro-4-cyanobenzonic acid (327 mg, 1.98 mmol), and K₂CO₃ (547 mg, 3.96 mmol) were dissolved in 1,4-dioxane (5 mL) and water (0.5 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (48 mg, 0.07 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 630 mg of the title compound (yield: 100%).

[0499] Step 4: Preparation of tert-butyl 4-(5-bromo-6-(4-cyano-3-fluorophenyl)-3-(cyclopropylcarbamoyl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid (28d)

[0500] Compound 28c (630 mg, 1.31 mmol) was dissolved in N,N-dimethylformamide (8 mL). NBS (480 mg, 2.70 mmol) was slowly added to the system under ice bath conditions, and the reaction mixture was stirred at room temperature for 2 hours. The reaction solution was poured into water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 315 mg of the title compound (yield: 43%).

[0501] Step 5: Preparation of tert-butyl 4-(6-(4-cyano-3-fluorophenyl)-3-(cyclopropylcarbamoyl)-5-(1-methyl-1H-indoleazol-5-yl)pyridin-2-yl)-1,4-diazane-1-carboxylic acid ester (28e)

[0502] Compound 28d (160 mg, 0.29 mmol), 1-methylindazole-5-boric acid (76 mg, 0.43 mmol), and K₂CO₃ (120 mg, 0.87 mmol) were dissolved in 1,4-dioxane (3 mL) and water (0.3 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (11 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 52 mg of the title compound (yield: 30%).

[0503] Step 6: Preparation of 6-(4-cyano-3-fluorophenyl)-N-cyclopropyl-2-((1,4-diazaaniline-1-yl)-5-(1-methyl-1H-indazol-5-yl)nicotinamide (28)

[0504] Compound 28e (52 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 5 hours. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 20 mg of the title compound, yield: 46%.

[0505] LC-MS(ESI)m / z(M+H) + 510.3

[0506] 1 H NMR(400MHz,DMSO-d6)δ8.60-8.57(m,1H),8.01(s,1H),7.79-7.73(m,1H),7.66(s,1H) ),7.60(s,1H),7.57-7.52(m,1H),7.47-7.41(m,1H),7.25-7.20(m,1H),7.11-7.06(m ,1H),4.01(s,3H),3.71-3.64(m,2H),3.54-3.47(m,2H),2.96-2.88(m,2H),2.84-2.7 6(m,1H),2.75-2.68(m,2H),1.89-1.77(m,2H),0.72-0.65(m,2H),0.54-0.48(m,2H).

[0507] Example 29 Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(2,8-diazaspiro[4.5]dec-8-yl)isonicotinonitrile (29)

[0508]

[0509] Step 1: Preparation of tert-butyl 8-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-2,8-diazaspirocyclic[4.5]decane-2-carboxylic acid (29a)

[0510] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of 2,8-diazaspirocyclic [4.5]decane-2-carboxylic acid tert-butyl ester (53 mg, 0.22 mmol) and Cs₂CO₃ (107 mg, 0.33 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 32 mg of the title compound, yield: 52%.

[0511] Step 2: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(2,8-diazaspiro[4.5]dec-8-yl)isonicotinonitrile (29)

[0512] Compound 29a (32 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 16 mg of the title compound, yield: 60%.

[0513] LC-MS(ESI)m / z(M+H) + 486.3

[0514] 1H NMR(400MHz,DMSO-d6)δ7.85-7.77(m,1H),7.57(s,1H),7.46-7.36(m,1H),7.24-7.11(m,3H),7.01-6.92(m,1H),3.85(s ,3H),3.81-3.57(m,4H),3.50-3.24(m,3H),2.96-2.88(m,1H),1.81-1.72(m,1H),1.65-1.58(m,1H),1.58-1.48(m,4H).

[0515] Example 30 Preparation of 6-(aza-4-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (30)

[0516]

[0517] Step 1: Preparation of 4-((((trifluoromethyl)sulfonyl)oxy)-2,3,6,7-tetrahydro-1H-aza-1-carboxylic acid tert-butyl ester (30b)

[0518] Compound 30a (3.0 g, 14.07 mmol) was dissolved in tetrahydrofuran (50 mL), purged with nitrogen, and cooled to -78 °C. Bistrimethylsilylaminolithium (19 mL, 1 M in THF) was added dropwise, and the reaction was continued at -78 °C for 1 hour. Then, N-phenylbis(trifluoromethanesulfonyl)imide (6.5 g, 18.19 mmol) was added, and the reaction was continued for another 0.5 hours. The system was then allowed to return to room temperature and reacted for 14 hours. The mixture was diluted with water, extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 2.0 g of the title compound (yield: 41%).

[0519] Step 2: Preparation of 4-(4,4,5,5-tetramethyl-1,3,2-dioxobenzaldehyde-2-yl)-2,3,6,7-tetrahydro-1H-azacyclic-1-carboxylic acid tert-butyl ester (30c)

[0520] Compound 30b (500 mg, 1.45 mmol), pinacol diborate (1.1 g, 4.33 mmol), and potassium acetate (426 mg, 4.34 mmol) were dissolved in 1,4-dioxane (5 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl2 (106 mg, 0.15 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 90 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain 300 mg of the title compound (yield: 64%).

[0521] Step 3: Preparation of 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)-2,3,6,7-tetrahydro-1H-azapyridine-1-carboxylic acid tert-butyl ester (30d)

[0522] Compound 5j (50 mg, 0.11 mmol), compound 30c (177 mg, 0.55 mmol), and K₂CO₃ (45 mg, 0.33 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (7 mg, 0.01 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 80 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 20 mg of the title compound (yield: 35%).

[0523] Step 4: Preparation of 4-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3-fluoro-4-methoxyphenyl)pyridin-2-yl)azacycloheptan-1-carboxylic acid tert-butyl ester (30e)

[0524] Compound 30d (20 mg, 0.04 mmol) was dissolved in ethyl acetate (2 mL), and then Pd / C (2 mg, 10%) was added at room temperature. The air in the system was replaced with hydrogen, and the system was stirred overnight at room temperature under a hydrogen atmosphere. The reaction system was filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel filtration to give 10 mg of the title compound, yield: 50%.

[0525] Step 5: Preparation of 6-(aza-4-yl)-2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (30)

[0526] Compound 30e (10 mg, 0.02 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 6 mg of the title compound, yield: 74%.

[0527] LC-MS(ESI)m / z(M+H) + : 445.2

[0528] 1 H NMR(400MHz,DMSO-d6)δ8.03-7.98(m,1H),7.88-7.82(m,1H),7.46-7.41(m,1H),7.33-7.28(m,1H),7.24-7.16(m,2H),7.1 0-7.04(m,1H),3.86(s,3H),3.66-3.45(m,1H),3.22-3.10(m,1H),3.08-2.88(m,2H),2.87-2.74(m,1H),2.05-1.58(m,6H).

[0529] Example 31 Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(6-hydroxy-1,4-diaza-1-yl)isonicotinonitrile (31)

[0530]

[0531] Preparation of N,N'-(ethane-1,2-diyl)bis(4-methylbenzenesulfonamide) (31b) in the first step

[0532] Compound 31a (10.0 g, 75.18 mmol) was dissolved in pyridine (200 mL), and then p-toluenesulfonyl chloride (28.0 g, 146.87 mmol) was added. The system was heated to 100 °C and stirred for 16 hours. After cooling to room temperature, the reaction mixture was poured into water, and a solid precipitated. The solid was filtered and dried under vacuum to give 18.0 g of the title compound, yield: 65%.

[0533] Step 2: Preparation of 1,4-di-p-toluenesulfonyl-1,4-diaza-6-ol (31c)

[0534] Compound 31b (5.8 g, 15.74 mmol) was dissolved in acetonitrile (100 mL), and potassium hydroxide (2.2 g, 39.21 mmol) was added. The reaction mixture was heated to 80 °C and reacted for 0.5 h. After cooling to room temperature, 1,3-dibromo-2-propanol (4.1 g, 18.82 mmol) was added, and the reaction mixture was heated to 80 °C again and reacted for 16 h. After cooling to room temperature, the mixture was concentrated under reduced pressure, diluted with water, and extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 5.3 g of the title compound (yield: 79%).

[0535] Step 3: Preparation of 1,4-diaza-6-ol hydrogen bromide (31d)

[0536] Compound 31c (2.0 g, 4.71 mmol) was dissolved in an aqueous solution of hydrogen bromide (20 mL, 40%), and the reaction mixture was heated to 100 °C and reacted for 22 hours. After cooling to room temperature, the mixture was concentrated under reduced pressure and then slurried with methyl tert-butyl ether to give 600 mg of crude product, yield: 46%.

[0537] Step 4: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(6-hydroxy-1,4-diaza-1-yl)isonicotinonitrile (31)

[0538] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of compound 31d (214 mg, 0.77 mmol) and Cs₂CO₃ (358 mg, 1.10 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 5 mg of the title compound (yield: 10%).

[0539] LC-MS(ESI)m / z(M+H) + : 462.2

[0540] Example 32 Preparation of 2-(4-cyano-3-fluorophenyl)-6-(6-fluoro-1,4-diaza-1-yl)-3-(3-fluoro-4-methoxyphenyl)isonicotinonitrile (32-1, 32-2)

[0541]

[0542] Step 1: Preparation of 6-fluoro-1,4-di-p-toluenesulfonyl-1,4-diazane (32a)

[0543] Compound 31c (2.5 g, 5.89 mmol) was dissolved in dichloromethane (30 mL), purged with nitrogen, and cooled to 0 °C. Diethylaminosulfur trifluoride (1.9 g, 11.79 mmol) was then added, and the reaction mixture was heated to room temperature for 16 hours. The reaction was quenched with water, and the mixture was extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.6 g of the title compound (yield: 64%).

[0544] Step 2: Preparation of 6-fluoro-1,4-diazacycloheptane (32b)

[0545] Compound 32a (1.6 g, 3.75 mmol) was dissolved in an aqueous solution of hydrogen bromide (20 mL, 40%), and the reaction mixture was heated to 120 °C and reacted for 22 hours. The reaction mixture was then cooled to room temperature and concentrated under reduced pressure. The concentrate was then slurried with methyl tert-butyl ether to give 700 mg of crude product, yield: 67%.

[0546] Step 3: Preparation of 2-(4-cyano-3-fluorophenyl)-3-(3-fluoro-4-methoxyphenyl)-6-(6-hydroxy-1,4-diaza-1-yl)isonicotinonitrile (32-1, 32-2)

[0547] Compound 5j (50 mg, 0.11 mmol) was dissolved in 1,4-dioxane (3 mL), followed by the sequential addition of compound 32b (216 mg, 0.77 mmol) and Cs₂CO₃ (358 mg, 1.10 mmol). Nitrogen gas was then introduced, followed by the addition of RuPhos-Pd-G₃ (9 mg, 0.01 mmol), and nitrogen gas was introduced again. The system was heated to 80 °C and stirred for 3 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain the title compounds 32-1 (3 mg, yield: 6%) and 32-2 (4 mg, yield: 8%).

[0548] 32-1:LC-MS(ESI)m / z(M+H) + : 464.2

[0549] 1H NMR (400MHz, DMSO-d6) δ7.85-7.78(m,1H),7.56(s,1H),7.43-7.38(m,1H),7.24-7.12(m,3H),7.00-6.94(m,1H),4.53-4 .44(m,1H),4.41-4.28(m,2H),4.26-4.18(m,1H),3.85(s,3H),3.03-2.88(m,2H),2.78-2.65(m,2H),2.03-1.95(m,1H).

[0550] 32-2:LC-MS(ESI)m / z(M+H) + : 464.2

[0551] 1 H NMR(400MHz,DMSO-d6)δ7.85-7.79(m,1H),7.47(s,1H),7.40(dd,J=10.8,1.2Hz,1H),7.23-7.12(m,3H),6 .99-6.93(m,1H),4.20-3.91(m,2H),3.85(s,3H),3.80-3.69(m,2H),3.00-2.75(m,4H),2.03-1.94(m,1H).

[0552] Example 33 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(5-fluoro-3-toluidine[d]isoxazol-6-yl)isonicotinonitrile (33)

[0553]

[0554] Step 1: Preparation of 3-bromo-4-fluorophenylacetic acid ester (33b)

[0555] Compound 33a (2.0 g, 10.47 mmol) was dissolved in dichloromethane (20 mL). Triethylamine (1.6 g, 15.81 mmol) was slowly added to the system under ice bath conditions, followed by the slow addition of acetyl chloride (1.2 g, 15.29 mmol). The reaction system was then brought to room temperature and stirred for 1 hour. The reaction solution was poured into water and extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 2.4 g of the title compound, yield: 98%.

[0556] Step 2: Preparation of 1-(4-bromo-5-fluoro-2-hydroxyphenyl)ethane-1-one (33c)

[0557] Compound 33b (2.4 g, 10.30 mmol) was dissolved in boron trifluoride acetic acid solution (26 mL, 33% BF3), and nitrogen gas was introduced. The reaction system was heated to 150 °C and stirred for 8 hours. The reaction system was cooled to room temperature, and the reaction solution was poured into water, precipitating a solid. After filtration, 1.6 g of the crude title compound was obtained and used directly in the next step without purification.

[0558] Step 3: Preparation of 6-bromo-5-fluoro-3-toluidine[d]isoxazole (33d)

[0559] Compound 33c (1.6 g, 6.87 mmol) was dissolved in methanol (16 mL), followed by the addition of hydroxylamine hydrochloride (963 mg, 13.86 mmol) and sodium acetate (853 mg, 10.40 mmol). The reaction mixture was heated to 60 °C and stirred for 1 hour. The mixture was cooled to room temperature, poured into cold water, and a solid precipitated. This solid was filtered, and the filter cake was dried to obtain a brown solid. The obtained solid was dissolved in acetic anhydride (6 mL), and the reaction mixture was heated to 60 °C and stirred for 2 hours. The mixture was cooled to room temperature, poured into cold water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was dissolved in N,N-dimethylformamide (10 mL), followed by the addition of K₂CO₃ (1.9 g, 13.75 mmol). The reaction mixture was stirred overnight at room temperature. The reaction solution was poured into water and extracted three times with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 693 mg of the title compound. Two-step yield: 29%.

[0560] Step 4: Preparation of 5-fluoro-3-methyl-6-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)benzo[d]isoxazole (33e)

[0561] Compound 33d (693 mg, 3.01 mmol), pinacol diborate (914 mg, 3.60 mmol), and potassium acetate (588 mg, 5.99 mmol) were dissolved in 1,4-dioxane (7 mL), and the mixture was purged with nitrogen. Pd(dppf)Cl2 (66 mg, 0.09 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 757 mg of the title compound (yield: 91%).

[0562] Step 5: Preparation of (1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(5-fluoro-3-methylbenzo[d]isoxazol-6-yl)pyridin-2-yl)piperidin-4-yl)tert-butyl carbamate (33f)

[0563] Compound 16b (150 mg, 0.30 mmol), compound 33e (166 mg, 0.60 mmol), and K₂CO₃ (124 mg, 0.90 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.2 mL). Nitrogen gas was purged, and Pd(dppf)Cl₂ (15 mg, 0.02 mmol) was added. Nitrogen gas was purged again, and the reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 41 mg of the title compound (yield: 24%).

[0564] Step 6: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(5-fluoro-3-toluidine[d]isoxazol-6-yl)isonicotinonitrile (33)

[0565] Compound 33f (41 mg, 0.07 mmol) was dissolved in ethyl acetate (1 mL), and then ethyl hydrochloride solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 12 mg of the title compound as a yellow solid, yield: 35%.

[0566] LC-MS(ESI)m / z(M+H) + : 471.2

[0567] 1 H NMR (400MHz, DMSO-d6) δ7.90-7.87(m,1H),7.82-7.74(m,2H),7.66(s,1H),7.47-7.42(m,1H),7.16(dd,J=8.0,1.6H z,1H),4.42-4.26(m,2H),3.14-3.03(m,2H),2.92-2.82(m,1H),2.55(s,3H),1.84-1.74(m,2H),1.26-1.16(m,2H).

[0568] Example 34 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3,3-difluoro-1-methyl-2-oxoindol-5-yl)isonicotinonitrile (34)

[0569]

[0570] Step 1: Preparation of 5-bromo-3,3-difluoroindole-2-one (34b)

[0571] Compound 34a (2.0 g, 8.85 mmol) was dissolved in dichloromethane (30 mL), purged with nitrogen, and cooled to 0 °C. Then, diethylaminosulfur trifluoride (4.3 g, 26.68 mmol) was added, and the reaction mixture was allowed to react at room temperature for 16 hours. The reaction was quenched with water, and the mixture was extracted three times with dichloromethane. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 1.8 g of the title compound (yield: 82%).

[0572] Step 2: Preparation of 5-bromo-3,3-difluoro-1-methylindole-2-one (34c)

[0573] Compound 34b (1.0 g, 4.03 mmol) was dissolved in acetonitrile (15 mL), followed by the addition of iodomethane (1.1 g, 7.75 mmol) and Cs₂CO₃ (2.0 g, 6.14 mmol). The reaction mixture was stirred at room temperature for 16 hours. The reaction solution was concentrated under reduced pressure, diluted with water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 900 mg of the title compound, yield: 85%.

[0574] Step 3: Preparation of 3,3-difluoro-1-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)indol-2-one (34d)

[0575] Compound 34c (500 mg, 1.91 mmol), pinacol diborate (1.5 g, 5.91 mmol), and potassium acetate (562 mg, 5.73 mmol) were dissolved in 1,4-dioxane (5 mL), and the mixture was purged with nitrogen. Pd(dppf)Cl2 (139 mg, 0.19 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was heated to 90 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 420 mg of the title compound (yield: 71%).

[0576] Step 4: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3,3-difluoro-1-methyl-2-oxoindol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (34e)

[0577] Compound 16b (80 mg, 0.16 mmol), compound 34d (247 mg, 0.80 mmol), and K₂CO₃ (66 mg, 0.48 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 80 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 30 mg of the title compound (yield: 31%).

[0578] Step 5: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3,3-difluoro-1-methyl-2-oxoindol-5-yl)isonicotinonitrile (34)

[0579] Compound 34e (30 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 22 mg of the title compound, yield: 88%.

[0580] LC-MS(ESI)m / z(M+H) + 503.2

[0581] 1 H NMR(400MHz,DMSO-d6)δ7.85-7.78(m,1H),7.63-7.54(m,2H),7.51-7.45(m,1H),7.44-7.38(m,1H),7.26-7.20(m,1H),7.1 9-7.13(m,1H),4.40-4.26(m,2H),3.18(s,3H),3.11-2.98(m,2H),2.95-2.82(m,1H),1.86-1.73(m,2H),1.27-1.15(m,2H).

[0582] Example 35 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(4-(1,1-difluoro-2-hydroxy-2-methylpropyl)-2-fluorophenyl)isonicotinonitrile (35)

[0583]

[0584] Step 1: Preparation of ethyl 2-(4-bromo-3-fluorophenyl)-2,2-difluoroacetate (35b)

[0585] Compound 35a (1.3 g, 4.32 mmol) was dissolved in dimethyl sulfoxide (20 mL), purged with nitrogen, and copper powder (707 mg, 11.13 mmol) was added. The mixture was stirred at room temperature for 1 hour, followed by the addition of ethyl 2-bromo-2,2-difluoroacetate (2.3 g, 11.33 mmol). The reaction mixture was heated to 80 °C and stirred for 2 hours. The reaction mixture was cooled to room temperature, filtered, and the filtrate was poured into water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 719 mg of the title compound (yield: 56%).

[0586] Step 2: Preparation of 1-(4-bromo-3-fluorophenyl)-1,1-difluoro-2-methylpropane-2-ol (35c)

[0587] Compound 35b (719 mg, 2.42 mmol) was dissolved in dry tetrahydrofuran (20 mL), and then methylmagnesium bromide reagent (4.1 mL, 3 M in THF) was slowly added dropwise under an ice-water bath. After the addition was complete, the reaction system was slowly brought to room temperature and stirred overnight. The reaction system was cooled to 0 °C, and the reaction was quenched by slowly adding an appropriate amount of water. The mixture was filtered, and the filtrate was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 577 mg of the title compound, yield: 84%.

[0588] Step 3: Preparation of 1,1-difluoro-1-(3-fluoro-4-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)phenyl)-2-methylpropane-2-ol (35d)

[0589] Compound 35c (577 mg, 2.04 mmol), pinacol diborate (1.0 g, 3.94 mmol), and potassium acetate (600 mg, 6.12 mmol) were dissolved in 1,4-dioxane (20 mL), and the mixture was purged with nitrogen. Pd(dppf)Cl2 (146 mg, 0.20 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was heated to 100 °C and stirred for 24 hours. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 364 mg of the title compound (yield: 54%).

[0590] Step 4: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(4-(1,1-difluoro-2-hydroxy-2-methylpropyl)-2-fluorophenyl)pyridin-2-yl)piperidin-4-yl)carbamate (35e)

[0591] Compound 16b (100 mg, 0.20 mmol), compound 35d (132 mg, 0.40 mmol), and K₂CO₃ (83 mg, 0.60 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.2 mL). Nitrogen gas was purged, and Pd(dppf)Cl₂ (15 mg, 0.02 mmol) was added. Nitrogen gas was purged again, and the reaction mixture was heated to 100 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 57 mg of the title compound (yield: 46%).

[0592] Step 5: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(4-(1,1-difluoro-2-hydroxy-2-methylpropyl)-2-fluorophenyl)isonicotinonitrile (35)

[0593] Compound 35e (57 mg, 0.09 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (1 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to give 37 mg of the title compound as a yellow solid, yield: 77%.

[0594] LC-MS(ESI)m / z(M+H) +524.2

[0595] 1 H NMR(400MHz,DMSO-d6)δ7.84-7.78(m,1H),7.64(s,1H),7.50-7.43(m,1H),7.39-7.28(m,3H),7.24-7.19(m,1H),5.4 9(s,1H),4.40-4.28(m,2H),3.14-3.03(m,2H),2.91-2.82(m,1H),1.85-1.75(m,2H),1.28-1.18(m,2H),1.15(s,6H).

[0596] Example 36 Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3,6-difluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (36)

[0597]

[0598] Step 1: Preparation of 5-bromo-3,6-difluoro-1H-indazole (36a)

[0599] Compound 9a (500 mg, 2.33 mmol) was dissolved in acetonitrile (7.5 mL), and glacial acetic acid (0.75 mL) and 1-chloromethyl-4-fluoro-1,4-diazabicyclo[2.2.2]octane di(tetrafluoroborate) salt (1.6 g, 4.66 mmol) were added sequentially. The reaction system was heated to 80 °C and stirred for 5 hours. The reaction system was cooled to room temperature, and the reaction was quenched with water. The mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 300 mg of the title compound, yield: 55%.

[0600] Step 2: Preparation of 1-(5-bromo-3,6-difluoro-1H-indazol-1-yl)-2-methylpropane-2-ol (36b)

[0601] Compound 36a (300 mg, 1.29 mmol) was dissolved in acetonitrile (10 mL), and then Cs₂CO₃ (630 mg, 1.94 mmol) and 1,1-dimethylethylene oxide (185 mg, 2.57 mmol) were added sequentially. The reaction system was heated to 80 °C and stirred for 16 hours. The reaction system was cooled to room temperature, the reaction solution was poured into water, and the mixture was extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 150 mg of the title compound, yield: 38%.

[0602] Step 3: Preparation of 1-(3,6-difluoro-5-(4,4,5,5-tetramethyl-1,3,2-dioxane-2-yl)-1H-indazol-1-yl)-2-methylpropane-2-ol (36c)

[0603] Compound 36b (150 mg, 0.49 mmol), pinacol diborate (373 mg, 1.47 mmol), and potassium acetate (144 mg, 1.47 mmol) were dissolved in 1,4-dioxane (5 mL), and the mixture was purged with nitrogen. Pd(dppf)Cl2 (37 mg, 0.05 mmol) was added, and the mixture was purged with nitrogen again. The reaction mixture was heated to 90 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give 76 mg of the title compound (yield: 44%).

[0604] Step 4: Preparation of tert-butyl(1-(4-cyano-6-(4-cyano-3-fluorophenyl)-5-(3,6-difluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)pyridin-2-yl)piperidin-4-yl)carbamate (36d)

[0605] Compound 16b (107 mg, 0.21 mmol), compound 36c (74 mg, 0.21 mmol), and K₂CO₃ (87 mg, 0.63 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL). Nitrogen gas was introduced, followed by the addition of Pd(dppf)Cl₂ (15 mg, 0.02 mmol), and nitrogen gas was introduced again. The reaction mixture was heated to 80 °C and stirred overnight. The reaction mixture was cooled to room temperature, poured into water, and extracted three times with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer chromatography using silica gel to obtain 35 mg of the title compound (yield: 26%).

[0606] Step 5: Preparation of 6-(4-aminopiperidin-1-yl)-2-(4-cyano-3-fluorophenyl)-3-(3,6-difluoro-1-(2-hydroxy-2-methylpropyl)-1H-indazol-5-yl)isonicotinonitrile (36)

[0607] Compound 36d (35 mg, 0.05 mmol) was dissolved in ethyl acetate (1 mL), and then hydrochloric acid-ethyl acetate solution (2 mL, 3 M in EA) was added. The reaction system was stirred at room temperature for 1 hour. The reaction system was concentrated under reduced pressure, diluted with water, and the pH was adjusted to approximately 8 with saturated sodium bicarbonate solution. The mixture was extracted three times with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by thin-layer silica gel chromatography to obtain 23 mg of the title compound, yield: 78%.

[0608] LC-MS(ESI)m / z(M+H) + 546.3

[0609] 1 H NMR(400MHz, DMSO-d6)δ7.86-7.81(m,1H),7.80-7.74(m,1H),7.62(s,1H),7.60-7.54(m,1H), 7.44-7.38(m,1H),7.19(dd,J=8.0,1.2Hz,1H),4.67(brs,1H),4.40-4.28(m,2H),4.20-4.07(m,2H), 3.13-3.01(m,2H),2.94-2.82(m,1H),1.86-1.75(m,2H),1.30-1.18(m,2H),1.14(s,3H),1.10(s,3H).

[0610] Biological evaluation

[0611] Compounds 1-36 used in the following experimental examples were prepared by the synthetic methods of Examples 1-36, respectively.

[0612] Experimental Example 1: LSD1 Enzyme Activity Assay

[0613] 1. Purpose of the experiment

[0614] This study employed time-resolved homogeneous fluorescence (HTRF) technology in a two-step method to detect the inhibitory activity of compounds on LSD1 enzyme. The first step involved incubating LSD1 enzyme, different concentrations of the compound, and a biotin-labeled methylated polypeptide substrate (H3(1-21)K4me1-biotin) at room temperature for a fixed time. The second step involved adding an antibody against the demethylated polypeptide substrate (Anti-H3K4me0-Eu(K)) and a detection reagent containing streptomycin labeled with XL-665 (SA-XL665), followed by further incubation at room temperature for a fixed time. The concentration of the demethylated polypeptide substrate was determined using EnVision's Intensity (665 nM) / Intensity (615 nM) ratio to evaluate the effect of the test compound on LSD1 enzyme activity. Simultaneously, the IC50 of the test compound against LSD1 enzyme was calculated using the inhibition rate. 50 value.

[0615] 2. Test materials

[0616] 2.1. The compound to be tested:

[0617] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0618] Test drug: Compounds 1-36.

[0619] 2.2 Test Reagents and Instruments

[0620] LSD1, Active Motif;

[0621] DMSO, Sigma;

[0622] OptiPlate-384, PerkinElmer;

[0623] Labcyte ultrasonic pipette;

[0624] EnVision microplate reader, PerkinElmer;

[0625] Centrifuge, Eppendorf.

[0626] 3. Test methods

[0627] 3.1 Compound Preparation

[0628] Preparation of 10mM compound stock solution: Dissolve the compound powder in 100% DMSO to prepare 10mM compound stock solutions.

[0629] 3.2 Enzyme Reaction Process

[0630] (1) Prepare 1× buffer according to the LSD1 enzymology kit provided by Active Motif (catalog number: 31432).

[0631] (2) Preparation of compound solution: Compound IC 50 The test started at a final concentration of 500 nM, with 5-fold dilutions, resulting in 6 concentrations. Each concentration was tested in single or duplicate wells. Solutions were diluted to the corresponding final concentration of 1000 times in a 384-well Source plate, and then 10 nL was transferred to each well of the 384-well reaction plate using an Echo 550. 10 nL of 100% DMSO was transferred to each of the Min and Max wells.

[0632] (3) Prepare 2× enzyme solution using 1× reaction solution.

[0633] (4) Prepare a 2× substrate mixture solution using 1× reaction solution.

[0634] (5) Add 5 μL of 2× enzyme solution to each well, add 5 μL of 1× reaction solution to the Min well, centrifuge at 1000 rpm for 1 min, and incubate at room temperature for 15 min.

[0635] (6) Add 5 μL of 2× substrate mixture to each well of the reaction plate to start the reaction, centrifuge at 1000 rpm for 1 min, and incubate at room temperature for 60 min.

[0636] (7) Add 10 μL of detection solution to each well, centrifuge at 1000 rpm for 1 min, and incubate at room temperature for 60 min.

[0637] (8) Use EnVision to read the signal values ​​Intensity(665nM) / Intensity(615nM).

[0638] 3.3 Data Analysis

[0639] The inhibition percentage was calculated using the maximum, minimum, and signal values ​​(maximum value: fluorescence intensity of 5 μL 2× enzyme solution + 5 μL 2× substrate mixture + 10 nL 100% DMSO + 10 μL detection solution; minimum value: fluorescence intensity of 5 μL 1× reaction solution + 5 μL 2× substrate mixture + 10 nL 100% DMSO + 10 μL detection solution; signal value: fluorescence intensity of 5 μL 2× enzyme solution + 5 μL 2× substrate mixture + 10 nL diluted analyte + 10 μL detection solution), using the following formula:

[0640]

[0641] Plotting the logarithm of concentration on the X-axis and the percentage inhibition rate on the Y-axis, dose-response curves were fitted using the log(inhibitor) vs. response-variable slop(four parameters) formula in Graphpad Prism 5 software to derive the IC50 of each compound on enzyme activity. 50 value.

[0642] 4. Test Results

[0643] The inhibitory activities of the test compounds and positive controls against LSD1 enzyme in this application are shown in Table 1.

[0644] Table 1. Inhibitory activity of the test compound and positive control against LSD1 enzyme in this application.

[0645]

[0646] As can be seen from the data on the inhibitory activity of each compound against LSD1 enzyme shown in Table 1, the compounds of this application can have significant inhibitory activity against LSD1 enzyme.

[0647] Experimental Example 2: Kasumi-1 Cell Proliferation Inhibition Test

[0648] 1. Experimental Principle

[0649] Studies have confirmed that LSD1 plays a crucial role in the development of various cancer types, including small cell lung cancer (SCLC) and acute myeloid leukemia (AML). Therefore, using the human AML (Kasumi-1) cell line, the ATP content in cells at different drug concentrations was measured using the CelltiterGLO assay kit. Cell viability was assessed by measuring the luminescence intensity, and the IC50 of different compounds on Kasumi-1 cells was calculated using the inhibition rate. 50 Value. This study investigates the inhibitory effect of the compounds of this application on the proliferation of Kasumi-1 cells to evaluate the antitumor efficacy of the test compounds at the cellular level.

[0650] 2. Test materials

[0651] 2.1 Compounds:

[0652] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0653] Test drugs: compounds 4, 16, 18 and 21.

[0654] 2.2 Test reagents and instruments:

[0655] RPMI 1640, Gibico;

[0656] Penicillin-Streptomycin (Gibico);

[0657] Fetal bovine serum (FBS), Gibico;

[0658] Phosphate-Buffered Saline (PBS), Gibic;

[0659] Dimethyl sulfoxide: DMSO, Sigma;

[0660] CelltiterGlo assay Kit (CTG), Promega;

[0661] White 384-well cell culture plate, PerkinElmer;

[0662] Vibrating plate device, QILINBE;

[0663] Centrifuge, Eppendorf;

[0664] CO2 incubator, Thermo Scientific;

[0665] Microscope, OLYMMPUS;

[0666] Gibco fully automated cell counter;

[0667] 384-well Source plate, Echo Qualified 384-Well Polypropylene, LABCYTE;

[0668] Echo, LABCYTE;

[0669] Envision multi-functional microplate reader, PerkinElmer.

[0670] 2.3. Experimental cells:

[0671] Kasumi-1 cells were purchased from ATCC.

[0672] 3. Test methods

[0673] 3.1 Test Procedure

[0674] On Day 1, cells were seeded into 384-well cell culture plates and cultured overnight in a 37°C, 5% CO2 cell culture incubator.

[0675] Day 0: The test compound was prepared into a 10 mM stock solution using DMSO. Starting with 1 μM, it was diluted 3-fold, resulting in 10 concentration points in double replicates. The diluted compound was added to 384-well cell culture plates and incubated at 37°C in a 5% CO2 incubator for 7 days. The DMSO concentration in the cell culture system was 0.1 vol%.

[0676] Day 7: Following the culture system: Celltiter GLO = 1:1 volume ratio, Celltiter GLO was added to a 384-well cell culture plate. The plate was shaken in the dark for 3 minutes, then placed at room temperature in the dark for 30 minutes. The plate was then read using a multifunctional enzyme-linked chemiluminescence module.

[0677] 3.2 Data Analysis

[0678] The inhibition rate was calculated using the luminescence intensity of the DMSO group (with DMSO added), the experimental group (with the test compound or positive control solution added), and the blank group (without cell treatment). The calculation formula is as follows:

[0679]

[0680] Using the logarithm of concentration as the X-axis and the percentage inhibition rate as the Y-axis, dose-response curves were fitted using the log(inhibitor) vs. response-variable slop(four parameters) formula in Graphpad Prism 8 software to derive the IC50 of each compound's inhibitory activity against Kasumi-1 proliferation. 50 value.

[0681] 4. Test Results

[0682] The inhibitory activity of the test compound and positive control against Kasumi-1 cell growth in this application is shown in Table 2.

[0683] Table 2. Inhibitory activity of the test compound and positive control against Kasumi-1 cells in this application.

[0684] 4 3.07 16 0.59 18 8.79 21 1.03 CC-90011 3.18

[0685] As shown in Table 2, the compounds in this application have a strong inhibitory effect on the growth of Kasumi-1 cells.

[0686] Experimental Example 3: NCI-H1417 Cell Proliferation Inhibition Test

[0687] 1. Experimental Principle

[0688] Studies have confirmed that LSD1 plays a crucial role in the development of various cancer types, including small cell lung cancer (SCLC) and acute myeloid leukemia (AML). Therefore, using the human small cell lung cancer cell line (NCI-H1417), the ATP content in cells at different drug concentrations was measured using the Celltiter GLO assay kit, and cell viability was assessed by the fluorescence intensity. The IC50 of different compounds on NCI-H1417 cells was calculated using the inhibition rate. 50 The study investigated the inhibitory effect of the compounds in this application on the proliferation of NCI-H1417 cells to evaluate the antitumor efficacy of the test compounds at the cellular level.

[0689] 2. Test materials

[0690] 2.1 Compounds:

[0691] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0692] Test drugs: compounds 4, 16, 18 and 21.

[0693] 2.2 Test reagents and instruments:

[0694] RPMI 1640, Gibico;

[0695] Penicillin-Streptomycin (Gibico);

[0696] Fetal bovine serum (FBS), Gibico;

[0697] Phosphate-Buffered Saline (PBS), Gibic;

[0698] Dimethyl sulfoxide: DMSO, Sigma;

[0699] CelltiterGlo assay Kit (CTG), Promega;

[0700] White 384-well cell culture plate, PerkinElmer;

[0701] Vibrating plate device, QILINBE;

[0702] Centrifuge, Eppendorf;

[0703] CO2 incubator, Thermo Scientific;

[0704] Microscope, OLYMMPUS;

[0705] Gibco fully automated cell counter;

[0706] 384-well Source plate, Echo Qualified 384-Well Polypropylene, LABCYTE;

[0707] Echo, LABCYTE;

[0708] Envision multi-functional microplate reader, PerkinElmer.

[0709] 2.3. Experimental cells:

[0710] NCI-H1417 cells were purchased from ATCC.

[0711] 3. Test methods

[0712] 3.1 Test Procedure

[0713] On Day 1, cells were seeded into 384-well cell culture plates and cultured overnight in a 37°C, 5% CO2 cell culture incubator.

[0714] On Day 0, the test compound was prepared into a 10 mM stock solution using DMSO. Starting with 1 μM, it was diluted 3-fold, resulting in 10 concentration points in duplicate wells. The diluted compound was added to a 384-well cell culture plate and incubated at 37°C in a 5% CO2 incubator for 7 days. The DMSO concentration in the cell culture system was 0.1%.

[0715] Day 7: Following the culture system: Celltiter GLO = 1:1 volume ratio, Celltiter GLO was added to a 384-well cell culture plate. The plate was shaken in the dark for 3 minutes, then placed at room temperature in the dark for 30 minutes. The plate was then read using a multifunctional enzyme-linked chemiluminescence module.

[0716] 3.2 Data Analysis

[0717] The inhibition percentage was calculated using the luminescence intensity of the DMSO group (with DMSO added), the experimental group (with test compound and positive control solution added), and the blank group (no cell treatment). The calculation formula is as follows:

[0718]

[0719] Plotting the logarithm of concentration on the X-axis and the percentage inhibition rate on the Y-axis, dose-response curves were fitted using the log(inhibitor) vs. response-variable slop(four parameters) formula in Graphpad Prism 8 software to derive the IC50 of each compound's inhibitory activity against NCI-H1417 proliferation. 50 value.

[0720] 4. Test Results

[0721] The inhibitory activities of the test compound and positive control on the growth of NCI-H1417 cells are shown in Table 3.

[0722] Table 3. Inhibitory activity of the test compound and positive control against NCI-H1417 cells in this application.

[0723] 4 16.74 16 2.79 18 33.30 21 6.51 36 4.00 CC-90011 22.88

[0724] As shown in Table 3, the compounds in this application have a strong inhibitory effect on the growth of NCI-H1417 cells.

[0725] Experimental Example 4: Pharmacokinetic Study of Rats After Oral Administration

[0726] 1. Experimental Principle

[0727] Using SD rats as test animals, the plasma concentration of the compound of this application at different time points after oral administration of the compound in a clarifying cassette was determined by LC-MS / MS. The pharmacokinetic parameters of the compound of this application in rats were obtained to study its pharmacokinetic characteristics.

[0728] 2. Test materials

[0729] 2.1 Compounds:

[0730] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0731] Test drugs: compounds 1, 4, 14-16, 18, 20, 21 and 36.

[0732] 2.2. Test Instruments:

[0733] Shimadzu LC-30A AB API4500 tandem mass spectrometer, vacuum blood collection tubes, blood collection needles, filter paper, syringes, etc.

[0734] 2.3 Experimental Animals

[0735] Male SD rats, weighing 180-220g, were used in groups of 3. After purchase, the animals were housed in an animal facility for at least 3 days to acclimatize, and were used in the experiment only after passing quarantine.

[0736] 3. Test methods

[0737] 3.1 Grouping: The rats were randomly grouped according to Table 4. After grouping, there was no statistically significant difference in body weight among the different groups of SD rats.

[0738] Table 4. Trial Groups and Dosing Regimens

[0739]

[0740] 3.2 Blood sample collection and testing:

[0741] According to Table 4, rats in each group were administered the corresponding test drug or positive control drug by gavage. Before administration and at 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h, and 24 h after administration, a fixed volume of blood was collected through the orbital sinus and placed in an EDTA-K2 anticoagulant tube. The tube was centrifuged at 8000 rpm for 1 min to separate the plasma into a centrifuge tube and then frozen at -20°C.

[0742] 3.3 Analytical Methods

[0743] Plasma samples stored at -20℃ at various time points were collected, acetonitrile was added, the mixture was vortexed at 1500 rpm for 2 min, centrifuged for 15 min (3500 rpm), and a fixed volume of the supernatant was taken for LC-MS / MS analysis.

[0744] 4. Calculation of pharmacokinetic parameters

[0745] The pharmacokinetic behavior of the test compounds was fitted using a non-compartmental model, and the main pharmacokinetic parameters (T0, T ...) were calculated using DAS 3.31 software. 1 / 2 T max C max AUC last wait).

[0746] 5. Test Results

[0747] Table 5 Pharmacokinetic parameters of compounds in the examples

[0748]

[0749]

[0750] As can be seen from the experimental results in Table 5, compared with the positive group (CC-90011), the groups given with compounds 1, 4, 14-16, 18, 20-21, and 36 showed better plasma exposure than CC-90011. This indicates that the compounds prepared in Examples 1, 4, 14, 15, 16, 18, 20, 21, and 36 of this application exhibit significantly improved pharmacokinetic properties compared with CC-90011. Therefore, the compounds of this application can exhibit good pharmacokinetic properties.

[0751] Experimental Example 5: Pharmacokinetic Study of Rats After Oral and Intravenous Administration

[0752] 1. Experimental Principle

[0753] Using SD rats as test animals, the plasma concentrations of the compound of this application at different time points after oral and intravenous administration were determined by LC-MS / MS. The pharmacokinetic parameters and bioavailability of the compound of this application in rats were obtained, and its pharmacokinetic characteristics were studied.

[0754] 2. Test materials

[0755] 2.1 Compounds:

[0756] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0757] Test drug: Compound 16;

[0758] 2.2. Test Instruments:

[0759] Shimadzu LC-30A AB API4500 tandem mass spectrometer, vacuum blood collection tubes, blood collection needles, filter paper, syringes, etc.

[0760] 2.3 Experimental Animals

[0761] Male SD rats, weighing 180-220g, were used in groups of 3. After purchase, the animals were housed in an animal facility for at least 3 days to acclimatize, and were used in the experiment only after passing quarantine.

[0762] 3. Test methods

[0763] 3.1 Grouping: The rats were randomly grouped according to Table 6. After grouping, there was no statistically significant difference in body weight among the different groups of SD rats.

[0764] 3.2 Solvent: 0.5% sodium methylcellulose aqueous solution

[0765] iv: 5% DMSO+10% Solutol+85% Saline

[0766] Table 6. Trial Groups and Dosing Regimens

[0767]

[0768] 3.2 Blood sample collection and testing:

[0769] According to Table 6, rats in each group were administered the corresponding test drug or positive control drug by gavage. Before administration and at 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h, 10 h, and 24 h after administration, a fixed volume of blood was collected through the orbital sinus and placed in an EDTA-K2 anticoagulant tube. The tube was centrifuged at 8000 rpm for 1 min to separate the plasma into a centrifuge tube and then frozen at -20°C.

[0770] 3.3 Analytical Methods

[0771] Plasma samples stored at -20℃ at various time points were collected, acetonitrile was added, the mixture was vortexed at 1500 rpm for 2 min, centrifuged for 15 min (3500 rpm), and a fixed volume of the supernatant was taken for LC-MS / MS analysis.

[0772] 4. Calculation of pharmacokinetic parameters:

[0773] The pharmacokinetic behavior of the test compounds was fitted using a non-compartmental model, and the main pharmacokinetic parameters (T0, T ...) were calculated using DAS 3.31 software. 1 / 2 T max C max AUC last wait).

[0774] 5. Test Results:

[0775] Table 7 Pharmacokinetic parameters of compounds in the examples

[0776]

[0777] As can be seen from the experimental results in Table 7, at the same dose, compound 16 is superior to CC-90011 in terms of plasma exposure, maximum blood concentration, and bioavailability when administered orally and intravenously, indicating that the compound prepared in Example 16 of this application has significantly improved pharmacokinetic properties compared with CC-90011.

[0778] Experimental Example 6: Pharmacokinetic Study of Oral Administration in Mice

[0779] 1. Experimental Principle

[0780] Using Balb / c mice as test animals, the plasma concentrations of the compound of this application at different time points after drug administration were determined by LC-MS / MS under different solvent conditions. Pharmacokinetic parameters of the compound of this application in mice under different solvent conditions were obtained to study its pharmacokinetic characteristics.

[0781] 2. Test materials

[0782] 2.1 Compounds:

[0783] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0784] Test drug: Compound 16.

[0785] 2.2. Test Instruments:

[0786] Shimadzu LC-30A AB API4500 tandem mass spectrometer, vacuum blood collection tubes, blood collection needles, filter paper, syringes, etc.

[0787] 2.3 Experimental Animals

[0788] Balb / c mice, male, weighing 18-22g, 6 mice per group. After purchase, the animals were housed in the animal facility for at least 3 days to acclimatize, and were used for the experiment after passing quarantine.

[0789] 3. Test methods

[0790] 3.1 Grouping: The mice were randomly grouped according to Table 8. After grouping, there was no statistically significant difference in body weight among the Balb / c mouse groups.

[0791] Table 8. Trial Groups and Dosing Regimens

[0792]

[0793] 3.2 Blood sample collection and testing:

[0794] According to Table 8, mice in each group were administered the corresponding test drug or positive control drug by gavage. Before administration and at 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h, 10 h, and 24 h after administration, a fixed volume of blood was collected through the orbital sinus and placed in an EDTA-K2 anticoagulant tube. The tube was centrifuged at 8000 rpm for 1 min to separate the plasma into a centrifuge tube and then frozen at -20°C.

[0795] 3.3 Analytical Methods

[0796] Plasma samples stored at -20℃ at various time points were collected, acetonitrile was added, the mixture was vortexed at 1500 rpm for 2 min, centrifuged for 15 min (3500 rpm), and a fixed volume of the supernatant was taken for LC-MS / MS analysis.

[0797] 4. Calculation of pharmacokinetic parameters:

[0798] The pharmacokinetic behavior of the test compounds was fitted using a non-compartmental model, and the main pharmacokinetic parameters (T0, T ...) were calculated using DAS 3.31 software. 1 / 2 T max C max AUC last wait).

[0799] 5. Test Results:

[0800] Table 9. Pharmacokinetic parameters of compounds in the examples.

[0801]

[0802] As can be seen from the experimental results in Table 9, compared with the positive group (CC-90011), compound 16 is superior to CC-90011 in terms of plasma exposure and maximum blood drug concentration under different solvent conditions. This indicates that the compound prepared in Example 16 of this application has significantly improved pharmacokinetic properties compared with CC-90011, and has drug metabolism kinetic advantages under different solvent conditions.

[0803] Experiment Example 7: Liver Microsomal Stability Test Experiment

[0804] 1. Experimental Principle

[0805] The liver microsome in vitro incubation method utilizes liver microsomes, supplemented by an NADPH regeneration system, to conduct metabolic reactions in vitro under simulated physiological conditions. After a certain reaction time, LC-MS / MS is used to determine the parent drug and metabolites in the incubation solution, and a preliminary analysis of their content is performed. Studying the stability of drugs in liver microsomes across different species is crucial for understanding the changes in drugs within the body.

[0806] 2. Test materials

[0807] 2.1 Liver microsomes in different species of rats, mice, humans, dogs, and monkeys

[0808] 2.2 Compounds:

[0809] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0810] Test drugs: compounds 16 and 36.

[0811] Positive reference compounds: testosterone, diclofenac, propafenone

[0812] 2.3. Test Instruments:

[0813] Shimadzu LC-30A AB API4500 tandem mass spectrometer, electric thermostatic shaker, multi-tube vortex oscillator, and pipettes of various capacities.

[0814] 3. Test methods

[0815] Following the instructions of the Phase I metabolic stability kit, the liver microsome incubation system was prepared by adding PBS buffer, liver microsomes, NADPH regeneration system solution A, NADPH regeneration system solution B, and the analyte in that order, and incubating at 37°C. Each sample was in triplicate, with a sample not containing the NADPH generation system serving as a negative control. The reaction was terminated by adding an equal volume of pre-cooled acetonitrile at 0, 5, 15, 30, 45, and 60 min. The parent drug content in the incubation solution was determined by LC-MS / MS.

[0816] 4. Test Results

[0817] Table 10 Results of liver microsomal stability tests of compounds in different species.

[0818]

[0819]

[0820]

[0821]

[0822] The above experimental results indicate that compounds 16 and 36 exhibit high metabolic stability in human, rat, mouse, dog, and monkey liver microsomes. Therefore, the compounds of this application demonstrate good drug-like properties.

[0823] Experiment Example 8: In vitro hERG inhibitory activity assay

[0824] 1. Experimental Principle

[0825] The rapidly activated human delayed rectified outward potassium current is primarily mediated by hERG ion channels and participates in human cardiomyocyte repolarization. Drug blockade of this current is a major cause of clinical manifestations such as QT prolongation syndrome, acute arrhythmias, and even sudden death. Using whole-cell patch technology, we examined the blocking effect of a compound on hERG channels in HEK293 cells stably expressing hERG channels and determined the half-maximal inhibitory concentration (IC50) of the compound. 50 As part of the cardiac safety evaluation, it was used to conduct a preliminary evaluation of its safety in in vitro screening for cardiotoxicity.

[0826] 2. Test materials

[0827] 2.1 Experimental cell lines:

[0828] HEK293 cells with homeostatic expression of hERG ion channels

[0829] 2.2 Compounds:

[0830] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-177-77;

[0831] Test drug: Compound 16.

[0832] 2.3. Test Instruments:

[0833] Patch clamp instrument: PC-505B

[0834] Micro-manipulation instrument: MP-225

[0835] Electrode drawing instrument: PC-10 (Narishige, Japan)

[0836] 3. Test methods

[0837] Cells were transferred to a perfusion tank, and an extracellular solution prepared with 137 mM NaCl, 4 mM KCl, 1.8 mM CaCl2, 1 mM MgCl2, 10 mM HEPES, and 10 mM glucose was used for perfusion, adjusted to pH 7.4 with NaOH. Electrodes were fabricated using PC-10 (Narishige, Japan). Whole-cell patch-clamp recording was performed, with noise filtered at one-fifth of the sampling frequency. Cells were clamped at -80 mV, then depolarized to 40 mV with a 4-second square wave, followed by hyperpolarization to -40 mV with a 2-second square wave to obtain the hERG tail current. This procedure was repeated every 20 seconds. The hERG tail current was the pure hERG current. The maximum current induced by the second square wave was detected, and after stabilization, the test compound was perfused. Once the reaction stabilized, the blocking strength was calculated.

[0838] 4. Test Results

[0839] Table 11. IC of the compound on hERG current 50 value

[0840] Compound numbering <![CDATA[IC 50 (μM)]]>

[0841] CC-90011 6.48

[0842] The above experimental results indicate that, compared with the positive phase (CC-90011), compound 16 has an IC50 inhibitory effect on hERG. 50 A value greater than 10 μM indicates

[0843] The compound of this application has a lower risk of cardiotoxicity than the positive control.

[0844] Experimental Example 9: In vivo efficacy trial in a Kasumi-1 cell subcutaneous xenograft tumor model

[0845] 1. Test Methods

[0846] 1.1 Cell Culture

[0847] RPMI 1640 medium containing 20% ​​fetal bovine serum (FBS), 37°C, 5% CO2.

[0848] 1.2 Compounds:

[0849] Solvent: 0.5% methylcellulose aqueous solution;

[0850] Test drugs: compounds 16 and 36.

[0851] 1.3 Experimental Procedure

[0852] CB17 SCID mice, female, 6-8 weeks old, weighing approximately 18-22 grams, were subcutaneously injected with 0.1 mL (1×10⁻⁶) on the right side of each mouse. 7 Kasumi-1 cells (including matrix gel). When the average tumor volume reached 100-200 mm³, mice were grouped and administered the drug. The dosage and administration method are shown in Table 12. Tumor volume was measured twice a week, and the mouse's body weight, tumor length and width were recorded at each measurement. Tumor volume = length × width × width / 2 mm. 3 When the average tumor volume in the solvent group reached over 3000 mm³, administration was terminated, mice in each group were sacrificed, tumor masses were dissected, and the tumor weight and volume of each mouse were weighed. The difference in average tumor volume between the test compound group and the solvent group was compared. The tumor-inhibiting effect of the compound was evaluated using TGI (%), which reflects the tumor growth inhibition rate.

[0853] Calculation of TGI (%): TGI (%) = [1 - (average tumor volume of compound in the current test - average tumor volume of compound at the beginning) / (average tumor volume of solvent group in the current test - average tumor volume of solvent group at the beginning)] × 100%

[0854] Changes in tumor volume and weight in each group of mice Figure 1 and Figure 2 As shown in the image.

[0855] 2. Test Results

[0856] Table 12. In vivo tumor suppression experiment data

[0857] solvent group 5 qd,po / 20 / Compound 16 5 qd,po 3.5 20 78.28 Compound 16 5 qd,po 5 20 86.13 Compound 36 5 qd,po 5 20 84.87

[0858] 3. Experimental Conclusions

[0859] The above experimental results indicate that compounds 16 and 36 of this application exhibit good in vivo efficacy in a human acute promyelocytic leukemia Kasumi-1 cell subcutaneous xenograft tumor model, demonstrating significant tumor-suppressive activity (TGI>60%). Furthermore, the test animals showed good tolerance to compounds 16 and 36, and the mice did not experience a decrease in body weight after administration (e.g., Figure 3 (As shown). Therefore, the compounds of this application can exhibit good in vivo tumor-suppressing effects.

[0860] Experimental Example 10: In vivo efficacy trial in a subcutaneous xenograft tumor model of HL-60 cells

[0861] 1. Test Methods

[0862] 1.1 Cell Culture

[0863] DMEM medium containing 10% fetal bovine serum (FBS), 37°C, 5% CO2.

[0864] 1.2 Compounds:

[0865] Positive drug: CC-90011, Chengdu Haibowei Pharmaceutical Co., Ltd., batch number HBW-013-203-33;

[0866] Solvent: 0.5% methylcellulose aqueous solution;

[0867] Test drug: Compound 16.

[0868] 1.3 Experimental Procedure

[0869] Balb / C nu mice, female, 6-8 weeks old, weighing approximately 18-22 grams, were subcutaneously injected with 0.1 mL (1×10⁻⁶) on the right side of each mouse. 7 HL-60 cells (cells + matrix gel). When the average tumor volume reached 100-200 mm³, mice were divided into groups and administered the drug. The dosage and administration method are shown in Table 13 below. Tumor volume was measured twice a week, and the mouse's body weight, tumor length and width were recorded at each measurement. Tumor volume = length × width × width / 2 mm. 3 When the average tumor volume in the solvent group reached over 3000 mm³, administration was discontinued, and the difference in average tumor volume between the test compound group and the solvent group was compared. The tumor-inhibiting effect of the compounds was evaluated using TGI (%), which reflects the tumor growth inhibition rate.

[0870] Calculation of TGI (%): TGI (%) = [1 - (average tumor volume of compound in the current test - average tumor volume of compound at the beginning) / (average tumor volume of solvent group in the current test - average tumor volume of solvent group at the beginning)] × 100%

[0871] Changes in tumor volume in each group of mice Figure 4 As shown in the image.

[0872] 2. Test Results

[0873] Table 12. In vivo tumor suppression experiment data

[0874] solvent group 5 qd,po / 10 / CC-90011 5 qd,po 10 10 35.3 Compound 16 5 qd,po 10 10 73.2

[0875] 3. Experimental Conclusions

[0876] The above experimental results show that, compared with the positive compound CC-90011, the compound of this application exhibits good in vivo efficacy in a human promyelocytic leukemia HL-60 cell subcutaneous xenograft tumor model, with significant tumor-suppressive activity (TGI>60%). Furthermore, the test animals showed good tolerance to the compound of this application, and the body weight of mice did not decrease after administration (e.g., Figure 5 (As shown).

[0877] It should be understood that the detailed description and accompanying embodiments above are merely exemplary and should not be construed as limiting the scope of this application, which is defined only by the appended claims and their equivalents. Various changes and modifications to the disclosed embodiments will be readily apparent to those skilled in the art. Such changes and modifications may be made without departing from their spirit and scope, including but not limited to those relating to the chemical structure, substituents, derivatives, intermediates, synthetic methods, formulations, and / or methods of use of this application. All publications, patents, and patent applications cited herein are incorporated herein by reference in their entirety for various purposes.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt thereof, in, Ring A is selected from cyclohexyl, piperidinyl, or homopiperazinyl; Ring B can be phenyl, indolyl, dihydroindolyl, or indazole. R 1 R 2 Each is independently selected from: hydrogen, cyano or And R 1 R 2 Not both hydrogen; or R 1 With R 2 It forms a ring with the attached carbon atom. ; R 3 Selected from: hydroxyl or unsubstituted amino groups; R 4 Selected from: halogen, oxo, (C1-C6)alkyl, (C1-C6)alkoxy, wherein the (C1-C6)alkyl, (C1-C6)alkoxy is unsubstituted or substituted by one or more substituents selected from: halogen, hydroxyl, (C1-C6)alkyl or (C1-C6)alkoxy; R 5 It is a halogen; R 6 R 7 Each is independently selected from: hydrogen or (C1-C6) alkyl; m is selected from: 0 or 1; n is selected from: 1, 2, 3 or 4; q is set to 1.

2. The compound of claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, The piperidinyl or homopiperazinyl group is attached to the pyridine ring via an N atom.

3. The compound of claim 2 or a pharmaceutically acceptable salt thereof, characterized in that, The ring A is piperidinyl.

4. The compound or a pharmaceutically acceptable salt thereof as described in any one of claims 1-3, characterized in that, The ring B is selected from phenyl, dihydroindolyl, or indazole.

5. The compound of claim 4 or a pharmaceutically acceptable salt thereof, characterized in that, The ring B is an indazole group.

6. The compound of claim 4 or a pharmaceutically acceptable salt thereof, characterized in that, R 1 R 2 Each is independently selected from: hydrogen, cyano or And R 1 R 2 They are not both hydrogen.

7. The compound of claim 6 or a pharmaceutically acceptable salt thereof, characterized in that, R 1 For hydrogen, R 2 It is cyano; or R 1 It is cyano or R 2 It is hydrogen.

8. The compound of claim 4 or a pharmaceutically acceptable salt thereof, characterized in that, The R 3 It is an unsubstituted amino group.

9. The compound or a pharmaceutically acceptable salt thereof as described in any one of claims 5-7, characterized in that, The R 3 It is an unsubstituted amino group.

10. The compound of claim 9 or a pharmaceutically acceptable salt thereof, characterized in that, The R 4 Selected from: fluorine, chlorine, oxo, methyl, methoxy or .

11. The compound or a pharmaceutically acceptable salt thereof as described in any one of claims 5-7, characterized in that, The R 4 Selected from: fluorine, chlorine, oxo, methyl, methoxy or .

12. The compound of claim 11 or a pharmaceutically acceptable salt thereof, characterized in that, The R 4 Selected from methyl, methoxy or .

13. The compound of claim 11 or a pharmaceutically acceptable salt thereof, characterized in that, n is selected from 1 or 2.

14. The compound of claim 12 or a pharmaceutically acceptable salt thereof, characterized in that, n is selected from: 1 or 2.

15. The compound of claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound of formula I has the structure of formula II. in, Ring A is selected from cyclohexyl, piperidinyl, or homopiperazinyl; Ring B is selected from phenyl, indolyl, dihydroindolyl, and indazole. R 2 It is hydrogen; R 3 It is an amino group; R 4 Selected from: fluorine, chlorine, oxo, methyl, methoxy or ; m is selected from: 0 or 1; n is selected from: 1, 2, 3 or 4.

16. The compound of claim 15 or a pharmaceutically acceptable salt thereof, characterized in that, The piperidinyl or homopiperazinyl group is attached to the pyridine ring via an N atom.

17. The compound of claim 16 or a pharmaceutically acceptable salt thereof, characterized in that, The ring B is an indazole group.

18. The compound of claim 17 or a pharmaceutically acceptable salt thereof, characterized in that, R 4 Selected from: methyl, methoxy or .

19. The compound of claim 18 or a pharmaceutically acceptable salt thereof, characterized in that, n is selected from: 1 or 2.

20. A compound or a pharmaceutically acceptable salt thereof, said compound being selected from: , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , or .

21. A pharmaceutical composition, characterized in that, It comprises a compound of formula I as claimed in any one of claims 1-19 or a pharmaceutically acceptable salt thereof, or a compound of claim 20 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

22. Use of a compound of formula I as claimed in any one of claims 1-19 or a pharmaceutically acceptable salt thereof, or the compound of claim 20 or a pharmaceutically acceptable salt thereof, in the preparation of a medicament for the prevention or treatment of lysine-specific demethylase 1 (LSD1)-mediated diseases.

23. The use as described in claim 22, wherein the lysine-specific demethylase 1 (LSD1)-mediated disease is selected from tumors or cancer.

24. The use as described in claim 22 or 23, wherein the disease is selected from acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), lymphoma, malignant sarcoma, breast cancer, cervical cancer, colon cancer, lung cancer, oral cancer, brain cancer, gastric cancer, liver cancer, colorectal cancer, pancreatic cancer, skin cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, fallopian tube tumors, peritoneal tumors, melanoma, glioma, glioblastoma, papillary malignancy, head and neck tumors, or myeloma.

25. A method for preparing a pyridine derivative with the structure shown in Formula I as described in any one of claims 1-19, comprising the following steps: Where LG represents the leaving group.

26. The preparation method according to claim 25, characterized in that, The leaving group is selected from halogen atoms, methanesulfonyloxy groups, or p-toluenesulfonyloxy groups; (1) React compound I-1 with compound I-2 to obtain I-3; (2) Compound I-3 is substituted to give I-4; (3) I-6 is obtained by coupling compound I-4 with the boric acid or borate ester of compound I-5; (4) Compound I-6 and compound I-7 are coupled together to obtain pyridine derivatives with the structure shown in Formula I.

27. A method for preparing a pyridine derivative with the structure shown in Formula I as described in any one of claims 1-19, comprising the following steps: Where LG represents the leaving group.

28. The preparation method according to claim 27, characterized in that, The leaving group is selected from halogen atoms, methanesulfonyloxy groups, or p-toluenesulfonyloxy groups; (1) Compound I-1 is substituted to give I-8; (2) I-9 is obtained by coupling compound I-8 with the boric acid or borate ester of compound I-5; (3) Compound I-9 and compound I-7 are coupled together to obtain formula I-10; (4) React compound I-10 with compound I-2 to obtain pyridine derivatives with the structure shown in Formula I.

29. A method for preparing a pyridine derivative with the structure shown in Formula I as described in any one of claims 1-19, comprising the following steps: Where LG represents the leaving group.

30. The preparation method according to claim 29, characterized in that, The leaving group is selected from halogen atoms, methanesulfonyloxy groups, or p-toluenesulfonyloxy groups; (1) Compound I-3 and compound I-7 are coupled together to obtain formula I-11; (2) Compound I-11 is substituted to give I-12; (3) The pyridine derivatives with the structure shown in Formula I are obtained by coupling reaction of compound I-12 with boric acid or borate ester of compound I-5.

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