MC4R antagonist compound, pharmaceutical composition and application of MC4R antagonist compound in medicine

Abstract

The invention relates to a compound as shown in a formula I, and a racemate, a stereoisomer, a tautomer, an isotope label, a nitrogen oxide, a solvate, a polymorphic substance, a metabolite, ester, a prodrug or pharmaceutically acceptable salt thereof. The compound has good MC4R antagonistic activity and good safety, the transmembrane activity and the drug bioavailability are improved, and the compound can be used as a drug for diagnosing, preventing and / or treating MC4R receptor mediated diseases or symptoms. # imgabs0 #

Description

MC4R antagonist compounds, pharmaceutical compositions and their medical applications

[0001] This application claims priority from the following patent applications:

[0002] A prior application, patent application number 202211461838.1, filed with the State Intellectual Property Office of China on November 17, 2022, entitled “MC4R antagonist compounds, pharmaceutical compositions and their use in medicine”;

[0003] A prior application, patent application number 202310223815.5, filed with the State Intellectual Property Office of China on March 9, 2023, entitled “MC4R antagonist compounds, pharmaceutical compositions and their use in medicine”;

[0004] A prior application, patent application number 202310388373.X, filed with the State Intellectual Property Office of China on April 12, 2023, entitled “MC4R antagonist compounds, pharmaceutical compositions, and their use in medicine”;

[0005] A prior application, filed with the State Intellectual Property Office of China on May 22, 2023, with patent application number 202310578810.4, entitled “MC4R antagonist compounds, pharmaceutical compositions, and their use in medicine”;

[0006] A prior application, patent application number 202310895827.2, filed with the State Intellectual Property Office of China on July 20, 2023, entitled “MC4R antagonist compounds, pharmaceutical compositions and their use in medicine”;

[0007] The prior application, filed with the State Intellectual Property Office of China on September 8, 2023, with patent application number 202311158862.2, entitled “MC4R antagonist compounds, pharmaceutical compositions and their use in medicine”,

[0008] The entire contents of said prior application are incorporated into the present application by reference. Technical Field

[0009] The present invention belongs to the field of pharmaceutical compounds, and specifically relates to MC4R antagonist compounds, pharmaceutical compositions and their applications in medicine. Background Art

[0010] The melanocortin-4 receptor (MC4R) is a member of the melanocortin receptor family (MCRs), belonging to the class A G protein-coupled receptor (GPCR) subfamily. Composed of five members (MC1R-MC5R), it mediates a variety of physiological functions in the human body. MC4R is a seven-transmembrane GPCR expressed primarily in the hypothalamus, hippocampus, and thalamus, and is a central regulator of body weight and energy homeostasis. MC1R, MC2R, MC3R, MC4R, and MC5R have been identified in mammals and are expressed in various tissues. MC1R is specifically expressed in melanocytes and melanomas. MC2R is an ACTH receptor expressed primarily in adrenal tissue. MC3R is primarily expressed in the brain and limbic system. MC4R is widely expressed in the brain and spinal cord. MC5R is expressed in the brain and many peripheral tissues, including skin, adipose tissue, skeletal muscle, and lymphoid tissue.

[0011] The MC4R is an unusual GPCR in that it possesses both endogenous agonists and antagonists. Melanocortin (α-MSH), derived from the hydrolysis product of proopiomelanocortin (POMC), acts as an endogenous ligand to activate the MC4R, promoting appetite and leading to weight loss. AgRP, secreted by AgRP neurons, inhibits MC4R signaling, thereby promoting appetite and increasing weight. Loss-of-function mutations in the MC4R cause obesity in mice: MC4R knockout mice gain weight at 5 weeks of age; at 15 weeks, homozygous females weigh, on average, twice as much as their wild-type littermates, while homozygous males are 50% heavier. Heterozygous MC4R knockout mice weigh an intermediate amount between their wild-type and homozygous littermates, demonstrating a gene dosage effect of MC4R knockout on body weight. Transgenic mice overexpressing AgRP display obesity, increased food intake, and hyperinsulinemia. Loss-of-function mutations in MC4R are associated with 6%-8% of cases of early-stage severe obesity, making it the most common form of monogenic obesity; whereas gain-of-function mutations are associated with a low body mass index (BMI). In addition to its key role in maintaining food intake and energy homeostasis, MC4R also plays a role in other areas of the central nervous system, such as pain perception, sexual function, anhedonia, and blood pressure, which also have clinical significance.

[0012] Based on the important functions of the MC4R signaling pathway in regulating food intake, energy balance and growth, MC4R has become a target for the treatment of obesity. The MC4R agonist Setmelanotide has been approved by the FDA for use in patients with POMC deficiency, leptin receptor deficiency and other forms of severe hereditary obesity.

[0013] In recent years, several small molecule MC4R antagonists have been reported in the literature and patent applications. These MC4R antagonists can treat and / or prevent diseases associated with MC4R, including cachexia (cancer-related cachexia, acquired immune deficiency syndrome (AIDS)-related cachexia, congestive heart failure (CHF)-related cachexia, chronic kidney disease (CKD)-related cachexia, and cachexia associated with the treatment of other chronic diseases); anorexia or anorexia nervosa (senile anorexia, anorexia associated with chemotherapy and / or radiotherapy); nausea and vomiting; weight loss (involuntary weight loss); growth retardation; sarcopenia; muscle atrophy; muscle weakness; frailty; osteoporosis; bone disease (bone loss); pain (neuropathic pain); anxiety (post-traumatic stress disorder or PTSD); depression; hypertension; malnutrition-related obesity (such as sarcopenia caused by chronic obesity); sexual dysfunction; and inflammatory diseases (inflammatory diseases associated with anorexia or cachexia, sarcopenia or muscle atrophy), etc.

[0014] Therefore, there is still an unmet medical need for the development of novel small molecule MC4R antagonists. We aim to obtain small molecule MC4R antagonists that are more effective, more selective, less toxic, more stable in physicochemical properties, better in oral bioavailability, and more metabolically stable for the treatment or prevention of MC4R-related diseases.

[0015] Summary of the Invention

[0016] To solve the problems existing in the prior art, the present invention provides a compound represented by Formula I and its racemate, stereoisomer, tautomer, isotope-labeled form, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt:

[0017] wherein X is selected from NR', O or S; R' is selected from H, C 1-12 Alkyl or C 3-12 Cycloalkyl;

[0018] Y is selected from CH or N;

[0019] to exist or not to exist;

[0020] R is selected from C 1-12 Alkyl, deuterated C 1-12 Alkyl, halogenated C 1-12 alkyl;

[0021] A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl, 3-14 membered heterocyclic group, C 3-12 Cycloalkyl; each Ra are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: C 1-12 Alkyl, C 1-12 Alkoxy; each R a1 the same or different, independently selected from deuterium, halogen, CN, C 1-12 Alkyl, C 1-12 alkoxy;

[0022] E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted 3-14 membered N-containing heterocyclic group; each R e the same or different, independently selected from H, deuterium, halogen, CN, OH, oxo (=O), C 1-12 Alkyl, C 1-12 Alkoxy; or two R e The atoms connected to it form C 3-12 Cycloalkyl or 3-14 membered heterocyclic group; N in E is Connected, C in E is connected to X;

[0023] G is selected from unsubstituted or optionally substituted with one, two or more R g Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl; each R g are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R g1 Substituted with the following groups: amino, C 1-12 Alkyl, C 1-12 Alkoxy, C 3-12 Cycloalkyl, C 6-14 Aryl, 5-14 membered heteroaryl; each R g1 the same or different, independently selected from deuterium, halogen, CN, C 1-12 Alkyl, C 1-12 alkoxy;

[0024] M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl, 3-14 membered heterocyclic group, C 3-12 Cycloalkyl; each R m are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R m1 Substituted with the following groups: C 1-12 Alkyl, C 1-12 Alkoxy, C 3-12Cycloalkyl; each R m1 the same or different, independently selected from deuterium, halogen, CN, OH, C 1-12 Alkyl, C 1-12 Alkoxy.

[0025] According to some embodiments, X is selected from NR', O or S; R' is selected from H, C 1-6 Alkyl or C 3-8 cycloalkyl, such as methyl, ethyl or cyclopropyl;

[0026] According to some embodiments, R is selected from C 1-6 Alkyl, deuterated C 1-6 Alkyl, halogenated C 1-6 An alkyl group; for example, a methyl group.

[0027] According to some embodiments, A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl, 3-10 membered heterocyclic group, C 3-8 Cycloalkyl; each R a are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: C 1-6 Alkyl, C 1-6 Alkoxy; each R a1 the same or different, independently selected from deuterium, halogen, CN, C 1-6 Alkyl, C 1-6 alkoxy;

[0028] According to some embodiments, A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: Each R a The same or different, independently selected from H, F, Cl, Br, I, CN, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl, halogenated C 1-6 alkoxy;

[0029] According to some embodiments, each R a are the same or different and are independently selected from H, F, Cl, Br, CN, methyl, methoxy, trifluoromethyl, difluoromethoxy;

[0030] According to some embodiments, A is selected from

[0031] According to some embodiments, E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted 3-8 membered N-containing heterocyclic group; each R e the same or different, independently selected from H, deuterium, halogen, CN, OH, oxo (=O), C 1-6 Alkyl, C 1-6 Alkoxy; or two R e The atoms connected to it form C 3-8 Cycloalkyl; N in E and Connected, C in E is connected to X;

[0032] According to some embodiments, E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted with the following groups: Each R e the same or different, independently selected from H, F, Cl, Br, I, CN, OH, oxo (=O), C 1-6 Alkyl; or two R e The atoms connected to it form C 3-8 Cycloalkyl;

[0033] According to some embodiments, each R e are the same or different and are independently selected from H, F, CN, OH, oxo (=O), methyl; or two R e The atom to which it is attached forms a cyclopropyl group;

[0034] According to some embodiments, E is selected from

[0035] According to some embodiments, G is selected from unsubstituted or optionally substituted with one, two or more R g Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl; each R g are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R g1 Substituted with the following groups: amino, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-8 Cycloalkyl, C 6-10 Aryl, 5-10 membered heteroaryl; each R g1 the same or different, independently selected from deuterium, C 1-6 Alkyl, C 1-6 alkoxy;

[0036] According to some embodiments, G is selected from unsubstituted or optionally substituted with one, two or more R gSubstituted with the following groups: Each R g The same or different, independently selected from H, halogen, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-8 Cycloalkyl, amino, C 1-6 Alkylamino, (C 1-6 Alkyl)2amino, C 6-10 Aryl, C 1-6 Alkyl 5-8 membered heteroaryl;

[0037] According to some embodiments, each R g are the same or different and are independently selected from H, F, methyl, ethyl, cyclopropyl, cyclobutyl, methoxy, methylamino (CH3NH-), dimethylamino ((CH3)2N-), phenyl,

[0038] According to some embodiments, G is selected from

[0039] According to some embodiments, M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl, 3-8 membered heterocyclic group; each R m are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R m1 Substituted with the following groups: C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl; each R m1 the same or different, independently selected from deuterium, halogen, CN, OH, C 1-6 Alkyl, C 1-6 alkoxy;

[0040] According to some embodiments, M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted groups: pyridyl, pyrimidinyl, phenyl, oxazolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, tetrazolyl, thiazolyl, tetrahydropyrrolyl, piperidinyl, morpholinyl, piperazinyl, naphthyl, quinolinyl, azepanyl, imidazolyl, Each R m The same or different, independently selected from H, deuterium, F, Cl, Br, I, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C3-6 Cycloalkyl, halogenated C 1-6 Alkyl, halogenated C 1-6 Alkoxy, hydroxy-C 1-6 alkyl;

[0041] According to some embodiments, each R m are the same or different and are independently selected from H, F, CN, methyl, methoxy, difluoromethyl, difluoromethoxy, trifluoromethyl, cyclopropyl,

[0042] According to some embodiments, each R m are the same or different and are independently selected from H, F, CN, methyl, methoxy, difluoromethyl, difluoromethoxy, trifluoromethyl, and cyclopropyl;

[0043] According to some embodiments, R m Selected from CN;

[0044] According to some embodiments, M is absent or selected from

[0045] According to some embodiments, M is selected from one, two or more R m Substituted 5-6 membered heteroaryl, wherein at least one R m CN, other R m is present or absent, and when present, is identical or different and is independently selected from halogen, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, halogenated C 1-6 Alkyl, halogenated C 1-6 Alkoxy.

[0046] According to some embodiments, M is selected from 5-6 membered heteroaryl substituted with CN.

[0047] According to some embodiments, the compound represented by Formula I is selected from the structures shown below:

[0048] Among them, X, R, A, E, G, M, R a 、R e 、R g 、R m has the definitions described above; p1, p2, p3, p4 are independently selected from 0, 1, 2, 3, 4 or 5.

[0049] According to some embodiments, the compound represented by Formula I has a structure represented by Formula II below:

[0050] Among them, R, M, G, R a 、R e , p1, p2 have the definitions described above.

[0051] According to some embodiments, the compound represented by Formula I has a structure represented by Formula II-1, II-2 or II-3:

[0052] Among them, R, M, G, R a , p1 has the definition described above.

[0053] According to some embodiments, the compound represented by Formula I has a structure represented by Formula III below:

[0054] Among them, M, R, R a 、R e 、R g , p1, p2, p3 have the definitions described above.

[0055] According to some embodiments, the compound represented by Formula I is selected from the structures shown below:

[0056] Among them, M, R, R a 、R e 、R g , p1, p2, p3 have the definitions described above.

[0057] According to some embodiments, the compound represented by Formula I has a structure represented by Formula IV below:

[0058] Among them, R, G, R a 、R e 、R m , p1, p2, p4 have the definitions described above.

[0059] According to some embodiments, the compound represented by Formula I has a structure represented by Formula V below:

[0060] Among them, R, E, R a 、R g 、R m , p1, p3, p4 have the definitions described above.

[0061] According to some embodiments, the compound represented by Formula I has a structure represented by Formula VI below:

[0062] Among them, A, R, R e 、R g 、R m , p2, p3, and p4 have the definitions described above.

[0063] According to some embodiments, the compound represented by Formula I has a structure represented by Formula VII below:

[0064] Among them, R, R a 、R e 、R g 、R m , p1, p2, p3, p4 have the definitions described above.

[0065] According to some embodiments, the compound represented by Formula I has the structure represented by Formula VIII or VIII-A below:

[0066] Wherein, A and M have the definitions described above.

[0067] According to some embodiments, the compound represented by Formula I has the structure represented by the following Formula VIII-1 or VIII-2:

[0068] Among them, R a , p1, M have the definitions described above, and Y1 is selected from CH or N.

[0069] According to some embodiments, the compound represented by Formula I has a structure represented by Formula IX or Formula IX-1:

[0070] Among them, R a , p1 have the above definitions, W1, W2 are the same or different and are independently selected from CH or N.

[0071] According to some embodiments, the compound represented by formula I is selected from the following structures:

[0072] According to some embodiments, the compound represented by formula I is selected from the following structures:

[0073] The present invention also provides a method for preparing the compound represented by formula I, comprising the following steps:

[0074] Wherein, X, Y, R, A, E, G, and M have the definitions described above; L is selected from a leaving group, such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl, and methylsulfone.

[0075] According to some embodiments, the exemplary preparation method of the present invention comprises the following steps:

[0076] Wherein, X, Y, R, A, E, G, and M have the definitions described herein; L, L1, L2, and L3 are selected from leaving groups, such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl, and methylsulfone; and PG is selected from amino protecting groups, such as Cbz (benzyloxycarbonyl), Boc (tert-butyloxycarbonyl), Fmoc (tert-methyloxycarbonyl), Bn (benzyl), and PMB (p-methoxybenzyl).

[0077] According to some embodiments, the exemplary preparation method of the present invention comprises the following steps:

[0078] Wherein, X, Y, R, A, E, G, and M have the definitions described herein; L, L1, L2, and L3 are selected from leaving groups, such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl, and methylsulfonyl; and PG is selected from amino protecting groups, such as Cbz (benzyloxycarbonyl), Boc (tert-butyloxycarbonyl), Fmoc (tert-methyloxycarbonyl), Bn (benzyl), and PMB (p-methoxybenzyl).

[0079] According to some embodiments, the exemplary preparation method of the present invention comprises the following steps:

[0080] Among them, Y, R, A, R e 、R g 、Rm , PG, L, L1, L2, L3 have the definitions described in this article.

[0081] The present invention further provides a pharmaceutical composition comprising the compound of formula I described in the present invention and its racemate, stereoisomer, tautomer, isotope-labeled product, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt.

[0082] In some embodiments, the pharmaceutical composition described in the present invention further comprises a therapeutically effective amount of the compound of formula I described in the present invention and its racemate, stereoisomer, tautomer, isotope-labeled substance, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0083] The carrier in the pharmaceutical composition is "acceptable" in that it is compatible with the active ingredient of the composition (and preferably, capable of stabilizing the active ingredient) and not deleterious to the subject being treated. One or more pharmaceutical excipients may be used for delivery of the active compound.

[0084] The present invention further provides the use of the compound of formula I and its racemate, stereoisomer, tautomer, isotope-labeled product, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt or the pharmaceutical composition in the preparation of drugs.

[0085] According to some embodiments, the medicament is a medicament for diagnosing, preventing and / or treating a disease or disorder mediated by the MC4R receptor.

[0086] According to some embodiments, the drug is a MC4R antagonist.

[0087] According to some embodiments, the disease or condition is cachexia (cachexia associated with cancer, cachexia associated with acquired immune deficiency syndrome (AIDS), cachexia associated with congestive heart failure (CHF); cachexia associated with chronic kidney disease (CKD); cachexia associated with treatment of other chronic diseases); anorexia or anorexia nervosa (anorexia nervosa in the elderly, anorexia associated with chemotherapy and / or radiation therapy); nausea and vomiting; weight loss (involuntary weight loss); growth retardation; sarcopenia; muscle atrophy; muscle weakness; frailty; osteoporosis; bone disease (bone loss); pain (neuropathic pain); anxiety (post-traumatic stress disorder or PTSD); depression; hypertension; malnutrition obesity (such as sarcopenia caused by chronic obesity); sexual dysfunction; and inflammatory diseases (inflammatory diseases associated with anorexia or cachexia, sarcopenia or muscle atrophy).

[0088] The present invention also provides a method for diagnosing, preventing and / or treating a disease or condition mediated by the MC4R receptor, which method comprises administering to a patient in need of such treatment a therapeutically effective amount of at least one compound of the present invention, alone or, optionally, in combination with another compound of the present invention and / or at least one other type of therapeutic agent.

[0089] According to the present invention, the disease or condition is cachexia (cachexia associated with cancer, cachexia associated with acquired immunodeficiency syndrome (AIDS), cachexia associated with congestive heart failure (CHF); cachexia associated with chronic kidney disease (CKD); cachexia associated with treatment of other chronic diseases); anorexia or anorexia nervosa (senile anorexia, anorexia associated with chemotherapy and / or radiotherapy); nausea and vomiting; weight loss (involuntary weight loss); growth retardation; sarcopenia; muscle atrophy; muscle weakness; fragility; osteoporosis; bone disease (bone loss); pain (neuropathic pain); anxiety (post-traumatic stress disorder or PTSD); depression; hypertension; malnutrition obesity (such as sarcopenia caused by chronic obesity); sexual dysfunction; and inflammatory diseases (inflammatory diseases associated with anorexia or cachexia, sarcopenia or muscle atrophy), etc.

[0090] In some embodiments, the compound is used as an MC4R antagonist, including but not limited to: cachexia (cachexia associated with cancer, cachexia associated with acquired immunodeficiency syndrome (AIDS), cachexia associated with congestive heart failure (CHF); cachexia associated with chronic kidney disease (CKD); cachexia associated with treatment of other chronic diseases); anorexia or anorexia nervosa (anorexia senile, anorexia associated with chemotherapy and / or radiotherapy).

[0091] The compounds of the present invention may be used in combination with additional therapeutic agents. Beneficial effects

[0092] The compounds provided by the present invention have good MC4R antagonist activity; the compounds of the present invention not only have good biological activity and good safety, but also have improved transmembrane activity and drug bioavailability;

[0093] The present invention significantly improves the in vivo pharmacokinetic properties of the compound through structural optimization, while reducing the compound's inhibition on CYP3A4M, reducing the risk of drug interactions, and improving the drugability of the compound.

[0094] Definitions and Explanations of Terms

[0095] Unless otherwise indicated, the definitions of groups and terms in this specification and claims, including definitions used as examples, exemplary definitions, preferred definitions, definitions in tables, and definitions of specific compounds in the Examples, may be arbitrarily combined and coupled with one another. The group definitions and compound structures resulting from such combinations and couplings should be understood to be within the scope of this specification and / or claims.

[0096] The term "optional" (or "optionally", "optionally") in the general formula definitions of this application means the situation of being substituted by zero, one or more substituents, for example, "optionally substituted by one, two or more R" means that it may not be substituted by R (unsubstituted) or may be optionally substituted by one, two or more R.

[0097] "More" means three or more.

[0098] Unless otherwise indicated, numerical ranges recited in this specification and claims are equivalent to reciting at least each specific integer value therein. For example, the numerical range "1-12" is equivalent to reciting each integer value in the numerical range "1-12", namely, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12.

[0099] The term "C 1-12 "Alkyl" is understood to mean straight-chain and branched alkyl groups having 1 to 12 carbon atoms, "C 1-8 "Alkyl" means straight chain and branched chain alkyl groups having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms, "C 1-6 The term "alkyl" refers to straight-chain and branched alkyl groups having 1, 2, 3, 4, 5 or 6 carbon atoms. The alkyl group is, for example, methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, 2-methylbutyl, 1-methylbutyl, 1-ethylpropyl, 1,2-dimethylpropyl, neopentyl, 1,1-dimethylpropyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 2-ethylbutyl, 1-ethylbutyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbutyl, 2,3-dimethylbutyl, 1,3-dimethylbutyl or 1,2-dimethylbutyl, or the like or isomers thereof.

[0100] The term "C 3-12 "Cycloalkyl" is understood to mean a saturated monovalent monocyclic, bicyclic (such as condensed, bridged, spiro) hydrocarbon ring or tricyclic alkane having 3 to 12 carbon atoms, preferably "C 3-10 Cycloalkyl", more preferably "C 3-8 Cycloalkyl". The term "C 3-12"Cycloalkyl" is understood to mean a saturated monovalent monocyclic, bicyclic (eg bridged, spiro) hydrocarbon ring or tricyclic alkane having 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. 3-12 The cycloalkyl group may be a monocyclic hydrocarbon group such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl or cyclodecyl, or a bicyclic hydrocarbon group such as borneol, indolyl, hexahydroindolyl, tetrahydronaphthyl, decahydronaphthyl, bicyclo[2.1.1]hexyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.1]heptenyl, 6,6-dimethylbicyclo[3.1.1]heptyl, 2,6,6-trimethylbicyclo[3.1.1]heptyl, bicyclo[2.2.2]octyl, 2,7-diazaspiro[3,5]nonyl, 2,6-diazaspiro[3,4]octyl, or a tricyclic hydrocarbon group such as adamantyl.

[0101] The term "C 6-14 "Aryl" should be understood to mean preferably a monovalent aromatic or partially aromatic monocyclic, bicyclic (such as fused, bridged, spiro) or tricyclic hydrocarbon ring having 6 to 14 carbon atoms, which can be a single aromatic ring or a polyaromatic ring fused together, preferably "C 6-10 Aryl". The term "C 6-14 "Aryl" is understood to mean preferably a monovalent aromatic or partially aromatic monocyclic, bicyclic or tricyclic hydrocarbon ring ("C 6-14 or a ring having 9 carbon atoms ("C9 aryl"), for example indanyl or indenyl, or a ring having 10 carbon atoms ("C 10 aryl) such as tetrahydronaphthyl, dihydronaphthyl or naphthyl, or a ring having 13 carbon atoms ("C 13 aryl), such as fluorenyl, or a ring having 14 carbon atoms ("C 14 aryl”), such as anthracenyl. When the C 6-20 When the aryl group is substituted, it may be monosubstituted or polysubstituted. Furthermore, there is no limitation on the position of substitution, and for example, substitution may be at the ortho, para or meta position.

[0102] The term "5-14 membered heteroaryl" is understood to include monovalent monocyclic, bicyclic (e.g. fused, bridged, spiro) or tricyclic aromatic ring systems having 5 to 14 ring atoms and containing 1 to 5 heteroatoms independently selected from N, O and S, for example "5-10 membered heteroaryl". The term "5-14 membered heteroaryl" is understood to include monovalent monocyclic, bicyclic or tricyclic aromatic ring systems having 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 ring atoms, in particular 5 or 6 or 9 or 10 carbon atoms, and containing 1 to 5, preferably 1 to 3, heteroatoms each independently selected from N, O and S and, in each case, may additionally be benzofused. "Heteroaryl" also refers to a radical in which a heteroaromatic ring is fused to one or more aryl, alicyclic or heterocyclyl rings, wherein the radical or point of attachment is on the heteroaromatic ring. Non-limiting examples include 1-, 2-, 3-, 5-, 6-, 7-, or 8-indolizinyl, 1-, 3-, 4-, 5-, 6-, or 7-isoindolyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl, 2-, 3-, 4-, 5-, 6-, or 7-indazolyl, 2-, 4-, 5-, 6-, 7-, or 8-purinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, or 9-quinolizinyl, 2-, 3-, 4-, 5-, 6-, 7-, or 8-quinolyl, 1-, 3-, 4-, 5-, 6-, 7-, or 8-isoquinolyl, 1-, 4-, 5-, 6-, 7-, or 8-phthalazinyl (pht) halazinyl), 2-, 3-, 4-, 5-, or 6-naphthyridinyl, 2-, 3-, 5-, 6-, 7-, or 8-quinazolinyl, 3-, 4-, 5-, 6-, 7-, or 8-cinnolinyl, 2-, 4-, 6-, or 7-pteridinyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, or 8-4aHcarbazolyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, or 8-carbazolylcarbazolyl, 1-, 3-, 4-, 5-, 6-, 7-, 8-, or 9-carbolinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, 9-, or 10-phenanthridinyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, 8- or 9-acridinyl, 1-, 2-, 4-, 5-, 6-, 7-, 8- or 9-piperidinyl, 2-, 3-, 4-, 5-, 6-, 8-, 9- or 10-phenanthrolinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8- or 9-phenazinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8- or 9-phenothiazinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, 9- or 10-phenothiazinyl, 1-, 3-, 4-, 6-, 7-, 8-, 9-, or 10-phenazinyl, 2-, 3-, 4-, 5-, 6-, or 1-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, or 10-benzoisoquinolinyl, 2-, 3-, 4-, or thieno[2,3-b]furanyl, 2-, 3-, 5-, 6-, 7-, 8-, 9-, 10-, or 11-7H-pyrazino[2,3-c]carbazolyl, 2-, 3-, 5-, 6- or 7-2H-furo[3,2-b]-pyranyl, 2-, 3-, 4-, 5-, 7- or 8-5H-pyrido[2,3-d]-o-oxazinyl, 1-, 3- or 5-1H-pyrazolo[4,3-d]oxazolyl, 2-, 4- or 54H-imidazo[4,5-d]thiazolyl, 3-, 5- or 8-pyrazino[2,3-d]pyridazinyl, 2-, 3-, 5- or 6-imidazo[2,1-b]thiazolyl, 1-, 3-, 6-, 7-, 8- or 9-furo[3,4-c]cinnolinyl, 1-, 2-, 3-, 4-, 5-, 6-, 8-, 9-, 10 or 11-4H-pyrido[2,3

[0015] In some embodiments, the present invention further comprises carbazolyl, 2-, 3-, 6-, or 7-imidazo[1,2-b][1,2,4]triazinyl, 7-benzo[b]thienyl, 2-, 4-, 5-, 6-, or 7-benzoxazolyl, 2-, 4-, 5-, 6-, or 7-benzimidazolyl, 2-, 4-, 4-, 5-, 6-, or 7-benzothiazolyl, 1-, 2-, 4-, 5-, 6-, 7-, 8-, or 9-benzoxapinyl, 2-, 4-, 5-, 6-, 7-, or 8-benzoxazinyl, 1-, 2-, 3-, 5-, 6-, 7-, 8-, 9-, 10-, or 11-4H-pyrrolo[1,2-b][2]benzazepinyl. Typical fused heteroaryl groups include, but are not limited to, 2-, 3-, 4-, 5-, 6-, 7-, or 8-quinolyl, 1-, 3-, 4-, 5-, 6-, 7-, or 8-isoquinolyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl, 2-, 3-, 4-, 5-, 6-, or 7-benzo[b]thienyl, 2-, 4-, 5-, 6-, or 7-benzoxazolyl, 2-, 4-, 5-, 6-, or 7-benzimidazolyl, and 2-, 4-, 5-, 6-, or 7-benzothiazolyl. When the 5- to 14-membered heteroaryl group is linked to other groups to form a compound of the present invention, the carbon atoms on the 5- to 14-membered heteroaryl ring may be linked to the other groups, or heteroatoms on the 5- to 14-membered heteroaryl ring may be linked to the other groups. When the 5- to 14-membered heteroaryl group is substituted, it may be monosubstituted or polysubstituted. Furthermore, there is no limitation on the substitution site, for example, a hydrogen atom connected to a carbon atom on a heteroaryl ring may be substituted, or a hydrogen atom connected to a heteroatom on a heteroaryl ring may be substituted.

[0103] Unless otherwise defined, the term "3-14 membered heterocyclyl" refers to a saturated or unsaturated non-aromatic ring or ring system, for example, a 4-, 5-, 6- or 7-membered monocyclic ring, a 7-, 8-, 9-, 10-, 11- or 12-membered bicyclic ring (such as a fused ring, a bridged ring, a spirocyclic ring) or a 10-, 11-, 12-, 13- or 14-membered tricyclic ring system, and contains at least one, for example 1, 2, 3, 4, 5 or more heteroatoms selected from O, S and N, wherein N and S may also be optionally oxidized to various oxidation states to form nitrogen oxides, -S(O)- or -S(O)2- states. For example, the "3-14 membered heterocyclyl" may be a 3-14 membered N-containing heterocyclyl (containing at least one N). Preferably, the heterocyclyl may be selected from a "3-10 membered heterocyclyl". The term "3-10 membered heterocyclyl" means a saturated or unsaturated non-aromatic ring or ring system, and contains at least one heteroatom selected from O, S and N. The heterocyclyl can be connected to the rest of the molecule through any one of the carbon atoms or the nitrogen atom (if present). The heterocyclyl can include fused or bridged rings and spirocyclic rings. In particular, the heterocyclyl can include, but is not limited to, 4-membered rings such as azetidinyl, oxetane; 5-membered rings such as tetrahydrofuranyl, dioxolyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, pyrrolinyl; or 6-membered rings such as tetrahydropyranyl, piperidinyl, morpholinyl, dithianyl, thiomorpholinyl, piperazinyl or trithianyl; or 7-membered rings such as diazepanyl. Optionally, the heterocyclyl can be benzo-fused. The heterocyclic group may be bicyclic, such as, but not limited to, a 5,5-membered ring, such as a hexahydrocyclopenta[c]pyrrol-2(1H)-yl ring, or a 5,6-membered bicyclic ring, such as a hexahydropyrrolo[1,2-a]pyrazin-2(1H)-yl ring. The heterocyclic group may be partially unsaturated, i.e., it may contain one or more double bonds, such as, but not limited to, dihydrofuranyl, dihydropyranyl, 2,5-dihydro-1H-pyrrolyl, 4H-[1,3,4]thiadiazinyl, 1,2,3,5-tetrahydrooxazolyl, or 4H-[1,4]thiazinyl, or it may be benzo-fused, such as, but not limited to, dihydroisoquinolinyl. When the 3-14-membered heterocyclic group is linked to other groups to form the compounds of the present invention, the linking may be to a carbon atom of the 3-14-membered heterocyclic group or to a heteroatom on the 3-14-membered heterocyclic group ring. For example, when the 3-14 membered heterocyclic group is selected from piperazinyl, the nitrogen atom on the piperazinyl group may be connected to the other group. Or when the 3-14 membered heterocyclic group is selected from piperidinyl, the nitrogen atom on the piperidinyl ring and the carbon atom at the para position thereof may be connected to the other group.

[0104] The term "spirocyclic" refers to a ring system in which two rings share one ring atom.

[0105] The term "fused ring" refers to a ring system in which two rings share two ring atoms.

[0106] The term "bridged ring" refers to a ring system in which two rings share three or more ring atoms.

[0107] The term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0108] "Halo" means substituted with one or more halogens.

[0109] Wavy lines intersecting chemical bonds Used to indicate the connection position of a group to other atoms in the molecular structure.

[0110] It will be appreciated by those skilled in the art that the compounds of formula (I) may exist in the form of various pharmaceutically acceptable salts. If these compounds have a basic center, they may form acid addition salts; if these compounds have an acidic center, they may form base addition salts; if these compounds contain both an acidic center (e.g., a carboxyl group) and a basic center (e.g., an amino group), they may also form internal salts.

[0111] The compounds of the present invention may exist in the form of solvates (e.g., hydrates), wherein the compounds of the present invention contain a polar solvent as a structural element of the crystal lattice of the compound, in particular water, methanol or ethanol. The amount of polar solvent, in particular water, may be present in a stoichiometric or non-stoichiometric ratio.

[0112] Depending on their molecular structure, the compounds of the present invention may be chiral and therefore may exist in various enantiomeric forms. These compounds may thus exist in racemic or optically active forms. The compounds of the present invention encompass isomers or mixtures thereof, racemates, in which each chiral carbon is in the R or S configuration. The compounds of the present invention or their intermediates can be separated into enantiomeric compounds by chemical or physical methods well known to those skilled in the art, or used in this form for synthesis. In the case of racemic amines, diastereomers are prepared from the mixture by reaction with an optically active resolving agent. Examples of suitable resolving agents are optically active acids, such as R and S forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid, suitable N-protected amino acids (e.g., N-benzoylproline or N-phenylsulfonylproline) or various optically active camphorsulfonic acids. Chromatographic enantiomer resolution can also be advantageously performed with the aid of optically active resolving agents (e.g., dinitrobenzoylphenylglycine, cellulose triacetate or other carbohydrate derivatives or chirally derivatized methacrylate polymers immobilized on silica gel). Suitable eluents for this purpose are aqueous or alcoholic solvent mixtures, e.g., hexane / isopropanol / acetonitrile.

[0113] The corresponding stable isomers can be separated according to known methods, for example by extraction, filtration or column chromatography.

[0114] The term "patient" refers to any animal including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, pigs, cows, sheep, horses or primates, and most preferably humans.

[0115] The term "therapeutically effective amount" refers to that amount of an active compound or drug that will elicit the biological or medical response that a researcher, veterinarian, physician, or other clinician is seeking in a tissue, system, animal, individual, or human, and includes one or more of the following: (1) prevents disease, e.g., prevents a disease, disorder, or condition in an individual who is susceptible to the disease, disorder, or condition but who is not yet experiencing or developing the pathology or symptoms of the disease. (2) inhibits disease, e.g., inhibits the disease, disorder, or condition (i.e., prevents further development of the pathology and / or symptoms) in an individual who is experiencing or developing the pathology or symptoms of the disease, disorder, or condition. (3) alleviates disease, e.g., alleviates the disease, disorder, or condition (i.e., reverses the pathology and / or symptoms) in an individual who is experiencing or developing the pathology or symptoms of the disease, disorder, or condition. DETAILED DESCRIPTION

[0116] The technical solutions of the present invention will be described in further detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanations of the present invention and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are encompassed within the scope of protection that the present invention is intended to protect.

[0117] The structures of the compounds of the present invention are determined by nuclear magnetic resonance (NMR) and / or liquid chromatography-mass spectrometry (LC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer, using deuterated dimethyl sulfoxide (DMSO-d6), deuterated methanol (CD3OD), and deuterated chloroform (CDCl3) as the solvents, with tetramethylsilane (TMS) as the internal standard.

[0118] Liquid chromatography-mass spectrometry (LC-MS) was performed using an Agilent 1200 Infinity Series mass spectrometer. HPLC was performed using an Agilent 1200DAD high-pressure liquid chromatograph (Sunfire C18 150×4.6 mm column) and a Waters 2695-2996 high-pressure liquid chromatograph (Gimini C18 150×4.6 mm column).

[0119] Thin layer chromatography silica gel plates use Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plates. The specifications used for TLC are 0.15mm-0.20mm, and the specifications used for thin layer chromatography separation and purification products are 0.4mm-0.5mm. Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

[0120] Unless otherwise stated, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods. Unless otherwise stated, all reactions of the present invention were carried out under a dry nitrogen or argon atmosphere with continuous magnetic stirring, with dry solvents, and reaction temperatures are reported in degrees Celsius.

[0121] Example 1

[0122] Step 1: tert-Butyl 3-((6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate 1b

[0123] At room temperature, 2-bromo-6-methylpyridine (1.71 g), tert-butyl 3-aminopyrrolidine-1-carboxylate (2.23 g), palladium acetate (0.2 g), (R)-(-)-1-[(S)-2-(dicyclohexylphosphino)ferrocenyl]ethyl-di-tert-butylphosphine (54.8 mg), and sodium tert-butoxide (1.33 g) were added to a 40 mL sample bottle. Ethylene glycol dimethyl ether (15 mL) was then added to the above system. Under a nitrogen atmosphere, the system was stirred at 90° C. overnight. The reaction was monitored by LCMS for completion, the solvent was removed under reduced pressure, and the resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-50%) as the eluent to obtain compound 1b (2.32 g).

[0124] LC-MS (ESI): m / z = 278.05 [M+H] +

[0125] Step 2 tert-Butyl 3-((5-bromo-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate 1c

[0126] To a solution of compound 1b (1.4 g) in tetrahydrofuran / dichloromethane (1:1, 10 mL) was added dropwise a solution of N-bromosuccinimide (0.90 g) in acetonitrile (5 mL) at 0°C. After complete addition, the reaction was allowed to stand at room temperature for 2 hours. Following completion of the reaction as monitored by LCMS, the solvent was removed under reduced pressure, and the resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-50%) as the eluent to afford compound 1c (1.41 g).

[0127] LC-MS (ESI): m / z = 355.95 [M+H] +

[0128] Step 3 tert-Butyl 3-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)pyrrolidine-1-carboxylate 1d

[0129] At room temperature, compound 1c (700 mg), 2-(tributyltin)pyrimidine (1087.95 mg), tetrakis(triphenylphosphine)palladium (113.53 mg), cuprous iodide (18.71 mg), and anhydrous lithium chloride (83.29 mg) were added to a 40 mL sample bottle. The atmosphere was purged with nitrogen three times, and then anhydrous toluene (15 mL) was added to the sample bottle. The reaction system was stirred at 120°C overnight. LCMS monitored the reaction completion, and the solvent was removed under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (20%-80%) as the eluent to obtain compound 1d (500 mg).

[0130] LC-MS (ESI): m / z = 355.90 [M+H] +

[0131] Step 4: 6-methyl-5-(pyrimidin-2-yl)-N-(pyrrolidin-3-yl)pyridin-2-amine 1e

[0132] To a solution of compound 1d (135 mg) in dichloromethane (1 mL) was added trifluoroacetic acid (0.2 mL) at room temperature, and the mixture was stirred at room temperature for 2 hours. The reaction was complete as monitored by LCMS, and the resulting residue was concentrated under reduced pressure to afford the trifluoroacetic acid salt of compound 1e (97 mg).

[0133] LC-MS (ESI): m / z = 256.05 [M+H] +

[0134] Step 5 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-(3-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)pyrrolidin-1-yl)propyl-1-one 001

[0135] Under nitrogen protection, to a solution of the trifluoroacetate of compound 1e (97 mg) in N,N-dimethylformamide (1 mL) were added (R)-2-(5-fluoro-2-methoxypyridin-4-yl)propanoic acid (37.61 mg, prepared by reference to "WO2021250541A1"), N,N-diisopropylethylamine (245.51 mg), and 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (216.68 mg) at room temperature. The reaction was stirred for 2 hours, and the product was generated as monitored by LCMS. The crude product was purified by preparative HPLC (column specifications: Kinetex EVO C18 Column, 30 mm x 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; elution gradient: 10% B to 45% B) to obtain compound 001.

[0136] LC-MS (ESI): m / z = 436.85 [M+H] +

[0137] 1 H NMR(400MHz,DMSO-d6)δ8.85–8.80(m,2H),8.14–7.93(m,2H),7.33–7.28(m,1H),7 .17–6.92(m,1H),6.74–6.61(m,1H),6.48–6.32(m,1H),4.49–4.31(m,1H),4.20–4 .02(m,1H),3.91–3.73(m,4H),3.70–3.53(m,1H),3.45–3.36(m,1H),3.28–3.14(m ,1H),2.65–2.58(m,3H),2.28–2.04(m,1H),1.99–1.73(m,1H),1.37–1.28(m,3H).

[0138] Example 2 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((3S,4S)-3-methyl-4-(6-methyl-5-(pyrimidin-2-yl)pyridin-2-ylamino)pyrrolidin-1-yl)propan-1-one (051-1)

[0139] The first step: (3S,4S)-3-methyl-4-(6-methyl-5-(pyrimidin-2-yl)pyridin-2-ylamino)pyrrolidine-1-carboxylic acid tert-butyl ester (051-1b)

[0140] Under nitrogen protection, tris(dibenzylideneacetone)dipalladium (209 mg), sodium tert-butoxide (439 mg), and 2-dicyclohexylphosphine-2',6'-dimethoxybiphenyl (187 mg) were added to a toluene (5 mL) solution of 051-1a (549 mg) and 083-1c (470 mg) at room temperature. The resulting mixture was stirred at 80°C for 2 hours. The system was cooled to room temperature, diluted with water (50 mL), and the reaction mixture was extracted with ethyl acetate (3×50 mL). The organic phases were combined, backwashed with saturated brine (1×20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1:1) to obtain the title compound 051-1b (500 mg).

[0141] LC-MS: (ESI, m / z) = 369.95 [M+H] +

[0142] Step 2: 6-methyl-N-((3S,4S)-4-methylpyrrolidin-3-yl)-5-(pyrimidin-2-yl)pyridin-2-amine (051-1c)

[0143] A solution of 051-1b (120 mg) in 1,4-dioxane (1 mL) was stirred at room temperature for 30 minutes and then concentrated to obtain the hydrochloride salt of the title compound 051-1c (100 mg, crude product). The crude product was not further purified and was directly used in the next step.

[0144] LC-MS: (ESI, m / z) = 270.10 [M+H] +

[0145] Step 3 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((3S,4S)-3-methyl-4-(6-methyl-5-(pyrimidin-2-yl)pyridin-2-ylamino)pyrrolidin-1-yl)propan-1-one (051-1)

[0146] Under nitrogen protection, N,N-diisopropylethylamine (336 mg) was added to a solution of the hydrochloride salt of 051-1c (35 mg) and 1f (26 mg) in N,N-dimethylacetamide (1 mL) at room temperature. After stirring for 5 minutes, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (74.11 mg) was added portionwise at room temperature. The resulting mixture was stirred at room temperature for 30 minutes. The crude product was purified by high performance liquid chromatography (chromatographic column specifications: YMC Triart C18 ExRs, 30 mm × 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate aqueous solution), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 60% B) to obtain compound 051-1 (9.17 mg).

[0147] LC-MS: (ESI, m / z) = 450.95 [M+H] +

[0148] 1H NMR(400MHz,DMSO-d6)δ8.84-8.80(m,2H),8.12-8.09(m,1H),8.05-8.00(m,1H), 7.32-7.27(m,1H),7.02-6.89(m,1H),6.75-6.69(m,1H),6.55-6.46(m,1H),4.63-4.47(m,1H),4.19-4.03(m,1H),3.8 2(s,3H),3.69-3.47(m,2H),3.32-3.09(m,2H),2.61(d,3H),2.59-2.38(m,1H),1.38-1.28(m,3H),0.97-0.79(m,3H).

[0149] Example 3 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-3-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)pyrrolidin-1-yl)propan-1-one (083-1)

[0150] The first step is 6-chloro-2-methyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (083-1b)

[0151] To a solution of 083-1a (50 g) and pinacol diboron (122.99 g) in 1,4-dioxane was added [1,1-bis(diphenylphosphino)ferrocene]palladium dichloride (3.54 g) and potassium acetate (52.29 g) at room temperature. The nitrogen atmosphere was replaced, and the mixture was stirred at 100°C for 2 hours. The reaction solution was then concentrated under reduced pressure. The residue was dissolved in N,N-dimethylformamide (30 mL), and purified by reverse-phase column chromatography using the following conditions: column specifications (C18, 330 g), mobile phase: water and acetonitrile, 0% to 100%, UV 254 nm detector. The residue was concentrated under reduced pressure to yield the title compound 083-1b (28 g).

[0152] LC-MS: (ESI, m / z) = 254.25 [M+H] +

[0153] Step 2: 2-(6-chloro-2-methylpyridin-3-yl)pyrimidine (083-1c)

[0154] To a solution of compound 083-1b (23 g) and 2-bromopyrimidine (17.31 g) in 1,4-dioxane (400 mL) was added 1,1'-bis(diphenylphosphino)ferrocene-palladium(II) chloride (6.64 g) and aqueous sodium carbonate solution (68 mL, 4 M). The nitrogen atmosphere was replaced and the mixture was reacted at 100°C for 2 hours. The mixture was cooled to room temperature, diluted with water (200 mL), and extracted with ethyl acetate (300 mL x 3). The resulting organic phase was washed once with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (4:1) to obtain the title compound 083-1c (18 g).

[0155] LC-MS: (ESI, m / z) = 206.05 [M+H] +

[0156] Step 3: (S)-tert-Butyl 3-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)pyrrolidine-1-carboxylate (083-1d)

[0157] Under nitrogen protection at room temperature, palladium acetate (1.09 g), 2-dicyclohexylphosphino-2',6'-diisopropoxybiphenyl (2.27 g), and sodium tert-butoxide (9.35 g) were added to a toluene solution (150 mL) of 083-1c (10 g) and (S)-3-aminopyrrolidine-1-carboxylic acid tert-butyl ester (13.59 g). The nitrogen atmosphere was replaced, and the reaction was carried out at 90°C for 3 hours. The desired product was found in the liquid. The resulting residue was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (2:1) to obtain the title compound 083-1d (12 g).

[0158] LC-MS: (ESI, m / z) = 356.15 [M+H] +

[0159] Step 4 (S)-6-methyl-5-(pyrimidin-2-yl)-N-(pyrrolidin-3-yl)pyridin-2-amine (083-1e)

[0160] To 083-1d (1.0 g) was added a 4 M solution of hydrogen chloride in 1,4-dioxane (10 mL) at room temperature. After the addition was complete, the system was stirred at room temperature for 30 minutes. The desired product was found in the liquid phase. The reaction mixture was directly concentrated under reduced pressure to afford the hydrochloride salt of the title compound 083-1e (850 mg). The crude product was used in the next step without further purification.

[0161] LC-MS: (ESI, m / z): 255.95 [M+H] +

[0162] Step 5 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-3-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)pyrrolidin-1-yl)propan-1-one (083-1)

[0163] To a solution of the hydrochloride salt of compound 083-1e (850 mg) and 083-1f (700 mg, synthesized according to the method of patent WO2021250541A1) in N,N-dimethylacetamide (10 mL) was added N,N-diisopropylethylamine (2.10 g) at room temperature. After stirring at room temperature for 5 minutes, 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (1.85 g) was added portionwise at room temperature. After the addition was complete, the system was stirred at room temperature for 2 hours. The desired product was found in the liquid phase. The crude product was purified by HPLC (Ultimate@XB-C18 column, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile, gradient: 25% B to 55% B) to obtain the title compound 083-1.

[0164] LC-MS (ESI): m / z = 452.95 [M+H] +

[0165] 1H NMR(400MHz,DMSO-d6)δ8.84-8.81(m,2H),8.23(d,1H),8.05-8.00(m,1H),7 .33-7.27(m,1H),7.18-7.00(m,1H),6.72(d,1H),6.46-6.38(m,1H),4.41-4 .28(m,1H),4.20-4.05(m,1H),3.84(d,3H),3.74-3.50(m,2H),3.43-3.24(m ,2H),2.61(d,3H),2.28-2.04(m,1H),1.95-1.81(m,1H),1.34-1.27(m,3H).

[0166] Example 4 2-(6-(((S)-1-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-5-carbonitrile (110)

[0167] Step 1: (S)-tert-Butyl 3-((5-bromo-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate (110b)

[0168] To a solution of 110a (10 g) and cesium carbonate (34.99 g) in N,N-dimethylacetamide (54 mL) was added 3-bromo-6-fluoro-2-methylpyridine (9.18 g) at room temperature. After the addition was complete, the atmosphere was replaced with nitrogen and the system was stirred at 120°C overnight. The desired product was found in the liquid. The reaction mixture was diluted with water (300 ml). The reaction mixture was extracted with ethyl acetate (3 × 200 mL). The organic phases were combined, backwashed with saturated brine (2 × 100 mL), and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (0% to 100%) to obtain the title compound 110b (4.7 g).

[0169] LC-MS (ESI): m / z = 355.70 [M+H] +

[0170] Step 2: (S)-tert-Butyl 3-((6-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)amino)pyrrolidine-1-carboxylate (110c)

[0171] To a solution of 110b (970 mg) and bis(pinacol boronate) (1.38 g) in 1,4-dioxane (10 mL) at room temperature were added [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (39.84 mg) and potassium acetate (587.86 mg). After the addition was complete, the system was purged with nitrogen, the temperature was raised to 100°C, and stirring was continued for 2 hours. The desired product was observed in the liquid phase. The reaction mixture was cooled to room temperature. The resulting residue was purified by reverse-phase column chromatography (C18 column, size 330 g), mobile phase: A: water (10 mmol / L ammonium bicarbonate) and B: acetonitrile, gradient: B: 0% to 100%) to afford the title compound 110c (850 mg).

[0172] LC-MS (ESI): m / z = 403.75 [M+H] +

[0173] Step 3: (S)-tert-Butyl 3-((5-(5-chloropyrimidin-2-yl)-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate (110d)

[0174] To a solution of 110c (200 mg) and 2-bromo-5-chloropyrimidine (287.75 mg) in 1,4-dioxane (4 mL) at room temperature were added [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (72.57 mg) and aqueous sodium carbonate (0.38 mL, 4 M). After the addition was complete, the atmosphere was replaced with nitrogen and the mixture was stirred at 100°C for 2 hours. The desired product was found in the liquid. The reaction mixture was diluted with water (10 mL) and then extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated brine (2 × 10 mL), and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate = 7:1 to obtain the title compound 110d (150 mg).

[0175] LC-MS (ESI): m / z = 390.05 [M+H] +

[0176] Step 4: (S)-tert-Butyl 3-((5-(5-cyanopyrimidin-2-yl)-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate (110e)

[0177] To a solution of 110d (90 mg) and zinc cyanide (54.21 mg) in N,N-dimethylacetamide (2 mL) at room temperature were added tris(dibenzylideneacetone)dipalladium (21.14 mg) and 2-(dicyclohexylphosphino)-2',4',6'-triisopropyl-1,1'-biphenyl (22.01 mg). After the addition was complete, the atmosphere was replaced with nitrogen and stirring was continued at 120°C for 2 hours. The desired product was found in the liquid. The reaction mixture was diluted with water (10 mL) and then extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated brine (2 × 10 mL), and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether:ethyl acetate = 10:1 to obtain the title compound 110e (80 mg).

[0178] LC-MS (ESI): m / z = 381.05 [M+H] +

[0179] Step 5: (6-(((S)-1-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-5-carbonitrile (110f)

[0180] Trifluoroacetic acid (1 mL) was added dropwise to a solution of 110e (50 mg) in dichloromethane (3 mL) at room temperature. After the addition was complete, the system was stirred at room temperature for 30 minutes. The desired product was found in the liquid. The reaction mixture was directly concentrated under reduced pressure to afford the trifluoroacetate salt of the title compound 110f (35 mg, crude product). The resulting mixture was used in the next step without further purification.

[0181] LC-MS (ESI): m / z = 281.05 [M+H] +

[0182] Step 6: 2-(6-(((S)-1-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-5-carbonitrile (110)

[0183] To a solution of the trifluoroacetate salt of 110f (35 mg) and 1f (25 mg) in N,N-dimethylacetamide (2 mL) at room temperature was added N,N-diisopropylethylamine (165 mg). The mixture was stirred at room temperature for 5 minutes, followed by the addition of 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (71.59 mg). After the addition was complete, the system was stirred at room temperature for 2 hours. The desired product was found in the liquid phase. The reaction mixture was directly purified by HPLC (Kinetex EVO C18 column, 30 mm x 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 55% B) to obtain the title compound 110 (24.24 mg).

[0184] LC-MS (ESI): m / z = 461.95 [M+H] +

[0185] 1H NMR(400MHz,DMSO-d6)δ9.24(d,2H),8.23-8.17(m,1H),8.13-8.09(m,1H),7. 52-7.35(m,1H),6.73-6.67(m,1H),6.51-6.43(m,1H),4.47-4.36(m,1H),4.1 9-4.02(m,1H),3.82(d,3H),3.77-3.68(m,1H),3.65-3.49(m,1H),3.46-3.21 (m,2H),2.67(d,3H),2.30-2.04(m,1H),1.96-1.81(m,1H),1.37-1.28(m,3H).

[0186] Example 5 2-(6-(((S)-1-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (111)

[0187] The first step: tert-Butyl (3S)-3-((5-(4-chloropyrimidin-2-yl)-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate (111b)

[0188] At room temperature, 111a (220 mg) and 110c (366.99 mg) were dissolved in 1,4-dioxane (5 mL). Aqueous sodium carbonate (1.7 mL, 2 mol / L) and (1,1'-bis(diphenylphosphino)ferrocene)palladium dichloride (83 mg) were added, respectively. The atmosphere was replaced with nitrogen three times, and the reaction was stirred at 100°C for 2 hours. Liquid chromatography-mass spectrometry indicated the formation of product. The mixture was cooled to room temperature and diluted with water (20 mL). The reaction mixture was extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated brine (1 × 20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1:1) to give the title compound 111b (160 mg).

[0189] LC-MS: (ESI, m / z) = 390.10 [M+H] +

[0190] Step 2: tert-Butyl (3S)-3-((5-(4-cyanopyrimidin-2-yl)-6-methylpyridin-2-yl)amino)pyrrolidine-1-carboxylate (111c)

[0191] At room temperature, 111b (160 mg) and zinc cyanide (96.37 mg) were dissolved in N,N-dimethylacetamide (2 mL). Tris(dibenzylideneacetone)dipalladium (37.58 mg) and 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (39.13 mg) were added, respectively. The atmosphere was replaced with nitrogen three times, and the reaction was stirred at 120°C for 2 hours. After cooling to room temperature, the mixture was diluted with water (20 mL), and extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated brine (1 × 10 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1:1) to obtain the title compound 111c (80 mg).

[0192] LC-MS: (ESI, m / z) = 381.20 [M+H] +

[0193] Step 3 (S)-2-(2-methyl-6-(pyrrolidin-3-ylamino)pyridin-3-yl)pyrimidine-4-carbonitrile (111d)

[0194] At room temperature, 111c (55 mg), dichloromethane (1 mL), and trifluoroacetic acid (0.5 mL) were added to the reaction flask. The reaction was stirred for 30 minutes. Liquid chromatography-mass spectrometry indicated the formation of product. The precipitate was concentrated to afford the trifluoroacetic acid salt of the title compound 111d (50 mg, crude). The crude product was used in the next step without further purification.

[0195] LC-MS: (ESI, m / z) = 281.05 [M+H] +

[0196] Step 4: 2-(6-(((S)-1-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (111)

[0197] The trifluoroacetate salt of 111d (50 mg, crude) and N,N-dimethylacetamide (2 mL) were dissolved in N,N-diisopropylethylamine (454 mg) at room temperature. 1f (35 mg) was added and the mixture was stirred for 5 minutes. Then, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (100 mg) was added at room temperature. The resulting residue was stirred for 30 minutes, and product was observed in the liquid phase. The crude product was purified by HPLC (column size: Kinetex 5μm EVO C18, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 58% B) to afford the title compound 111 (9.17 mg).

[0198] LC-MS: (ESI, m / z) = 462.00 [M+H] +

[0199] 1H NMR(400MHz,DMSO-d6)δ9.14-9.10(m,1H),8.15-8.06(m,2H),7.90-7.86(m,1H ),7.40-7.24(m,1H),6.73-6.68(m,1H),6.50-6.43(m,1H),4.44-4.34(m,1H), 4.19-4.03(m,1H),3.82(d,3H),3.76-3.68(m,1H),3.65-3.50(m,1H),3.45-3. 20(m,2H),2.63(d,3H),2.29-2.03(m,1H),1.95-1.80(m,1H),1.37-1.27(3H).

[0200] Example 6 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((3S,4S)-3-methyl-4-(6-methyl-5-(pyrimidin-2-yl)pyridin-2-ylamino)pyrrolidin-1-yl)propan-1-one (112)

[0201] Under nitrogen, N,N-diisopropylethylamine (335 mg) was added to a solution of the hydrochloride salt of 051-1c (35 mg) and 083-1f (28 mg) in N,N-dimethylacetamide (1 mL) at room temperature. After stirring for 5 minutes, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (74.11 mg) was added portionwise at room temperature. The resulting mixture was stirred at room temperature for 30 minutes. The crude product was purified by HPLC (Kinetex EVO C18 Column, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 30% B to 55% B) to afford the title compound 112 (16.19 mg).

[0202] LC-MS: (ESI, m / z) = 466.95 [M+H] +

[0203] 1H NMR(400MHz,DMSO-d6)δ8.84-8.80(m,2H),8.25-8.20(m,1H),8.02(d,1H),7 .32-7.27(m,1H),7.03-6.86(m,1H),6.74(d,1H),6.56-6.47(m,1H),4.62-4 .47(m,1H),4.19-4.04(m,1H),3.84(s,3H),3.68-3.47(m,2H),3.26-3.11(m ,2H),2.61(d,3H),2.58-2.37(m,1H),1.36-1.27(m,3H),0.97-0.80(m,3H).

[0204] Example 7 2-(6-(((S)-1-((R)-2-(5-chloro-2-methoxypyridin-4-yl)propionyl)pyrrolidin-3-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (113)

[0205] To a solution of the trifluoroacetate salt of 111d (35 mg) and 083-1f (27 mg) in N,N-dimethylacetamide (1 mL) at room temperature was added N,N-diisopropylethylamine (322 mg). After stirring for 5 minutes, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (71 mg) was added at room temperature. After 2 hours of reaction at room temperature, the reaction was complete as determined by HPLC. The crude product was purified by HPLC (column size: Kinetex 5μm EVO C18, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate in water), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 58% B) to afford the title compound 113 (5 mg).

[0206] LC-MS: (ESI, m / z) = 477.95 [M+H] +

[0207] 1H NMR(400MHz,DMSO-d6)δ9.15-9.10(m,1H),8.23(d,1H),8.11-8.05(m,1H),7.90- 7.86(m,1H),7.41-7.21(m,1H),6.72(d,1H),6.50-6.42(m,1H),4.43-4.29(m,1H ),4.21-4.05(m,1H),3.84(d,3H),3.74-3.65(m,1H),3.63-3.49(m,1H),3.46-3. 21(m,2H),2.63(d,3H),2.30-2.03(m,1H),1.96-1.81(m,1H),1.36-1.25(m,3H).

[0208] Example 8 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (114)

[0209] Step 1: (S)-7-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (114-a)

[0210] To a 40 mL vial were added compound 083-1c (500 mg), (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (567.77 mg), tris(dibenzylideneacetone)dipalladium (222.64 mg), 2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl (199.63 mg), and sodium tert-butoxide (467.32 mg). The atmosphere was replaced with nitrogen three times, and toluene (5 mL) was added. The mixture was stirred at 100°C for 2 hours, and the desired product was observed in the liquid. The mixture was cooled to room temperature, diluted with water (20 mL), and extracted with ethyl acetate (3 x 20 mL). The organic phases were combined and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-50%) to obtain compound 114-a (850 mg).

[0211] LC-MS: (ES, m / z) = 382.05 [M+H] +

[0212] Step 2 (S)-N-(6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)-5-azaspiro[2.4]heptane-7-amine (114-b)

[0213] Compound 114-a (500 mg) was added to a 4 M solution of hydrogen chloride in 1,4-dioxane (5 mL) and stirred at room temperature for 30 minutes. LCMS analysis indicated product formation. The reaction solution was concentrated under reduced pressure to afford the hydrochloride salt of compound 114-b (420 mg, crude product). The crude product was used directly in the next step without further purification.

[0214] LC-MS: (ES, m / z) = 282.40 [M+H] +

[0215] Step 3: (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (114)

[0216] The hydrochloride salt of compound 114-b (130 mg) was dissolved in N,N-dimethylacetamide (4 mL), followed by the addition of compound 083-1f (153.28 mg), N,N-diisopropylethylamine (275.62 mg), and 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (405.42 mg). The reaction mixture was stirred at room temperature for 2 hours, and the reaction was monitored for completion by LCMS. The reaction mixture was directly purified by HPLC (column: Kinetex 5μm EVO C18, 30 mm x 150 mm; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 22% B to 57% B over 8 min) to obtain the title compound 114 (195.78 mg).

[0217] LC-MS: (ES, m / z) = 478.95 [M+H] +

[0218] 1 H NMR(400MHz, DMSO-d6)δ8.84–8.81(m,2H),8.22(d,1H),8.03–7.98(m,1H),7.33-7.28(m,1H),7.12–7.00(m,1H),6.73(d,1H),6.53–6.42(m,1 H),4.20–4.05(m,2H),3.84(d,3H),3.74–3.53(m,3H),3.53–3.48(m,0. 5H),3.25(d,0.5H),2.60(d,3H),1.37–1.25(m,3H),0.90–0.48(m,4H).

[0219] Example 9 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[6-methyl-5-(pyrimidin-2-yl)pyridin-2-yl]amino}-5-azaspiro[2.4]heptane-5-yl]propan-1-one (056-1)

[0220] At room temperature, dissolve the hydrochloride salt of compound 114-b (35 mg) and compound 1f (25 mg) in N,N-dimethylacetamide (3 ml). Add N,N-diisopropylethylamine (243.23 mg) and then 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (71 mg). After the addition is complete, stir the system at room temperature for 2 hours. Filter the reaction mixture, rinse the filter with N,N-dimethylacetamide (0.5 mL x 2), and combine the filtrates. The crude product was purified by high performance liquid chromatography (chromatographic column specifications: Kinetex EVO C18 column, 30 mm*150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 ml / min; gradient: 20% B to 55% B in 8 minutes) to obtain the title compound 056-1 (27.7 mg).

[0221] LC-MS (ESI, m / z): =463.00 [M+H] +

[0222] 1 H NMR(400MHz,DMSO-d6)δ8.84–8.80(m,2H),8.13–8.09(m,1H),8.03–7.98( m,1H),7.32–7.27(m,1H),7.10–6.97(m,1H),6.73–6.68(m,1H),6.52–6.4 2(m,1H),4.26–4.04(m,2H),3.82(d,3H),3.79–3.69(m,2H),3.57–3.46(m ,1.5H),3.24(d,0.5H),2.60(d,3H),1.37–1.27(m,3H),0.89–0.46(m,4H).

[0223] Example 10 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methoxy-4-methylquinazolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (115)

[0224] Step 1: (S)-7-((6-methoxy-4-methylquinazolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (115b)

[0225] Under nitrogen, compound 115a (100 mg, prepared with reference to WO2012083165A1) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (122 mg) were added to the reaction flask. Tris(dibenzylideneacetone)dipalladium (43 mg), sodium tert-butoxide (92 mg), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (39 mg) were then added. The nitrogen atmosphere was replaced, and finally, ultra-dry toluene (2 mL) was added. The reaction was continued at 120°C. Liquid chromatography-mass spectrometry indicated the formation of product. The reaction mixture was diluted with water (50 mL) and extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride aqueous solution (1 × 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (50-100%) as eluent to give the title compound 115b (100 mg).

[0226] LC-MS: (ESI, m / z) = 385.10 [M+H] +

[0227] Step 2 (S)-6-methoxy-4-methyl-N-(5-azaspiro[2.4]heptane-7-yl)quinazolin-2-amine (115c)

[0228] Compound 115b (40 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (1 mL) were added to a reaction flask at room temperature. The reaction was allowed to react for 30 minutes. Liquid chromatography-mass spectrometry indicated the formation of product. The reaction solution was concentrated under reduced pressure to afford the hydrochloride salt of the title compound 115c (35 mg, crude). The crude product was used in the next step without further purification.

[0229] LC-MS: (ESI, m / z) = 285.00 [M+H] +

[0230] Step 3 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methoxy-4-methylquinazolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (115)

[0231] At room temperature, the hydrochloride salt of compound 115c (35 mg) and compound 1f (25 mg) were added to a reaction flask. The mixture was dissolved in N,N-dimethylacetamide (1 mL), followed by N,N-diisopropylethylamine (94 mg). After stirring for five minutes, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (70 mg) was added. The mixture was stirred at room temperature for 1 hour. LC / MS monitoring indicated the formation of the product. The crude product was purified by HPLC (Kinetex EVO C18 column, 30 mm x 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 40% B to 64% B over 10 minutes) to afford the title compound 115 (17.5 mg).

[0232] LC-MS: (ESI, m / z) = 466.00 [M+H] +

[0233] 1 H NMR (400MHz, DMSO-d6) δ8.12–8.09(m,1H),7.41–7.24(m,4H),6.73–6.69(m,1H),4.27–4.04(m,2H),3.85(d,6 H),3.82(d,3H),3.78–3.47(m,3.5H),3.25–3.19(m,0.5H),2.70(d,3H),1.37–1.26(m,3H),0.93–0.41(m,4H).

[0234] Example 11 2-(6-(((S)-5-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propanoyl)-5-azaspiro[2.4]heptane-7-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (116)

[0235] Step 1: 6-Fluoro-2-methyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (116b)

[0236] Compound 116a (5 g) and pinacol diboronate (13.44 g) were dissolved in 1,4-dioxane (50 mL). [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex (432 mg) and potassium acetate (5.7 g) were then added. The atmosphere was purged with nitrogen three times, the temperature was raised to 100°C, and stirring was continued for 2 hours. The reaction mixture was cooled to room temperature and filtered. The filter cake was washed with 1,4-dioxane (20 mL), and the filtrate was concentrated under reduced pressure. The resulting residue was purified by reverse-phase column chromatography (C18 column, mobile phase: acetonitrile and water, gradient from 0% to 75% over 25 minutes) to obtain the title compound 116b (4.8 g).

[0237] LC-MS: (ESI, m / z) = 238.10 [M+H] +

[0238] Step 2: 2-(6-Fluoro-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (116c)

[0239] Compound 116b (500 mg) and 2-bromopyrimidine-4-carbonitrile (773 mg) were dissolved in a mixture of 1,4-dioxane (5 mL) and water (1.25 mL). [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex (86 mg) and potassium carbonate (874 mg) were then added. The atmosphere was purged with nitrogen three times, and the temperature was raised to 100°C with stirring for 1 hour. The reaction mixture was cooled to room temperature, diluted with water (50 mL), and extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-20%) as the eluent to afford compound 116c (320 mg, 70.9%).

[0240] LC-MS: (ESI, m / z) = 215.00 [M+H] +

[0241] Step 3: (S)-7-((5-(4-cyanopyrimidin-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (116d)

[0242] Compound 116c (100 mg) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (150 mg) were dissolved in N,N-dimethylacetamide (2 mL). N,N-diisopropylethylamine (182 mg) was added at room temperature and the temperature was raised to 100°C with continued stirring overnight. The reaction mixture was cooled to room temperature, diluted with water (40 mL), and the mixture was extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-37%) as eluent to obtain the title compound 116d (120 mg).

[0243] LC-MS: (ESI, m / z) = 407.25 [M+H] +

[0244] Step 4 (S)-2-(6-((5-azaspiro[2.4]hept-7-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (116e)

[0245] Compound 116d (100 mg) was dissolved in dichloromethane (0.5 mL), trifluoroacetic acid (1 mL) was added at room temperature, and stirring was continued for 1 hour. The reaction mixture was concentrated under reduced pressure to afford the trifluoroacetic acid salt of title compound 116e (80 mg, crude product). The crude product was used in the next step without further purification.

[0246] LC-MS: (ESI, m / z) = 307.10 [M+H] +

[0247] Step 5: 2-(6-(((S)-5-((R)-2-(5-fluoro-2-methoxypyridin-4-yl)propionyl)-5-azaspiro[2.4]heptane-7-yl)amino)-2-methylpyridin-3-yl)pyrimidine-4-carbonitrile (116)

[0248] The trifluoroacetate salt of compound 116e (80 mg, crude) and compound 1f (49 mg) were dissolved in N,N-dimethylacetamide (1 mL). N,N-diisopropylethylamine (317 mg) was added to make the mixture alkaline. 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (187 mg) was then added and stirring continued at room temperature for 1 hour. The resulting mixture was filtered, and the filter cake was washed with N,N-dimethylacetamide (2 × 1 mL). The filtrate was purified by HPLC (Kinetex 5μm EVO C18 column, 30×150 mm, 5μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 30% B to 65% B over 8 minutes) to afford the title compound 116 (26.8 mg).

[0249] LC-MS: (ESI, m / z) = 488.00 [M+H] +

[0250] 1 H NMR(400MHz,DMSO-d6)δ9.14–9.10(m,1H),8.13–8.09(m,1H),8.09–8.04( m,1H),7.89–7.86(m,1H),7.34-7.27(m,1H),6.73–6.68(m,1H),6.56–6.4 5(m,1H),4.25–4.05(m,2H),3.82(d,3H),3.78–3.60(m,2H),3.56–3.47(m ,1.5H),3.23(d,0.5H),2.62(d,3H),1.36–1.28(m,3H),0.88–0.47(m,4H).

[0251] Example 12 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((7-fluoro-6-methoxyquinolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (117)

[0252] Step 1: Ethyl (E)-3-(2-amino-4-fluoro-5-methoxyphenyl)acrylate (117b)

[0253] Compound 117a (2 g) and ethyl acrylate (1.1 g) were dissolved in N,N-dimethylformamide (20 mL), followed by the addition of palladium acetate (204 mg), tri(o-tolyl)phosphine (305 mg), and triethylamine (3.69 g). The atmosphere was purged with nitrogen three times, and the temperature was raised to 105°C and stirred for 2 hours. The mixture was cooled to room temperature, diluted with water (100 mL), and extracted with ethyl acetate (3 × 200 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 200 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-25%) as the eluent to obtain the title compound 117b (1.4 g).

[0254] LC-MS: (ESI, m / z) = 239.95 [M+H] +

[0255] Step 2 7-Fluoro-6-methoxyquinolin-2(1H)-one (117c)

[0256] Compound 117b (940 mg) was dissolved in anhydrous ethanol (10 mL), followed by the addition of 21% sodium ethoxide-ethanol solution (2.4 g). The mixture was purged with nitrogen three times and heated to 90°C for 2 hours. The reaction mixture was concentrated under reduced pressure, and saturated ammonium chloride solution was added to adjust the pH to neutral and stirred for 10 minutes. The resulting mixture was filtered, the filter cake was collected, and concentrated under reduced pressure to give the title compound 117c (500 mg, crude product). The crude product was not further purified and was directly used in the next step.

[0257] LC-MS: (ESI, m / z) = 194.15 [M+H] +

[0258] Step 3: 2-chloro-7-fluoro-6-methoxyquinoline (117d)

[0259] Compound 117c (200 mg, crude product) was dissolved in phosphorus oxychloride (4 mL), replaced with nitrogen three times, and the temperature was raised to 80°C and stirred for 3 hours. After the reaction mixture was cooled to room temperature, it was added dropwise to ice water (40 mL) to quench the mixture, and then the mixture was extracted with ethyl acetate (3×50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2×50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-12%) as eluent to obtain the title compound 117d (118 mg).

[0260] LC-MS: (ESI, m / z) = 212.10 [M+H] +

[0261] Step 4: (S)-7-((7-fluoro-6-methoxyquinolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (117e)

[0262] Compound 117d (55 mg) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (66 mg) were dissolved in toluene (2 mL). Tris(dibenzylideneacetone)dipalladium (24 mg), 2-dicyclohexylphosphine-2′,6′-dimethoxy-biphenyl (21 mg), and sodium tert-butoxide (50 mg) were then added. The atmosphere was purged with nitrogen three times and the temperature was raised to 100°C with stirring for 2 hours. The reaction mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-13%) as the eluent to afford the title compound 117e (60 mg).

[0263] LC-MS: (ESI, m / z) = 388.15 [M+H] +

[0264] Step 5 (S)-7-Fluoro-6-methoxy-N-(5-azaspiro[2.4]heptane-7-yl)quinolin-2-amine (117f)

[0265] Compound 117e (48 mg) was placed in a reaction flask. A 4 M solution of hydrogen chloride in 1,4-dioxane (2 mL) was added at room temperature and stirred for 1 hour. The reaction mixture was concentrated under reduced pressure to afford the hydrochloride salt of title compound 117f (38 mg, crude product). The crude product was used in the next step without further purification.

[0266] LC-MS: (ESI, m / z) = 288.05 [M+H] +

[0267] Step 6 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((7-fluoro-6-methoxyquinolin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one (117)

[0268] The hydrochloride salt of compound 117f (38 mg, crude) and compound 1f (25 mg) were dissolved in N,N-dimethylacetamide (1 mL). N,N-diisopropylethylamine (161 mg) was added at room temperature to make the mixture alkaline. 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (95 mg) was then added and stirring continued at room temperature for 1 hour. The resulting mixture was filtered, and the filter cake was washed with N,N-dimethylacetamide (2 × 1 mL). The filtrate was purified by HPLC (XBridge BEH Shield RP 18 column, 30 x 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 35% B to 75% B over 8 minutes) to afford the title compound 117 (17.3 mg).

[0269] LC-MS: (ESI, m / z) = 469.00 [M+H] +

[0270] 1 H NMR(400MHz, DMSO-d6)δ8.13–8.08(m,1H),7.85–7.78(m,1H),7.34-7.28(m,1H),7.24–7.17(m,1H),7.15–7.00(m,1H),6.83–6.66(m,2H), 4.26–4.05(m,2H),3.87(d,3H),3.82(d,3H),3.77–3.61(m,2H),3.56 –3.50(m,1.5H),3.24(d,0.5H),1.37–1.25(m,3H),0.89–0.48(m,4H).

[0271] Example 13 2R-2-(5-chloro-2-methoxypyridin-4-yl)-1-[(7S)-7-[(6-methoxy-4-methylquinazolin-2-yl)amino]-5-azaspiro[2.4]heptan-5-yl]propan-1-one (118)

[0272] To a solution of the hydrochloride salt of compound 115c (30 mg, crude) and compound 083-1f (26.54 mg) in N,N-dimethylacetamide (2 mL) was added N,N-diisopropylethylamine (318.16 mg). After stirring for 5 minutes, 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (70.20 mg) was added. The reaction was stirred at room temperature for 1 hour. The crude product was purified by HPLC (Kinetex EVO C18 column, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 38% B to 53% B) to afford the title compound 118 (7.36 mg).

[0273] LC-MS (ESI): m / z = 481.90 [M+H] +

[0274] 1 H NMR(400MHz,DMSO-d6)δ8.21(d,1H),7.42–7.22(m,4H),6.73(d,1H),4.25–4.06(m,2H),3.89–3.8 0(m,6H),3.78–3.49(m,3.5H),3.23(d,0.5H),2.70(d,3H),1.35–1.24(m,3H),0.93–0.41(m,4H).

[0275] Example 14 (R)-1-((S)-7-((5-(5-(difluoromethyl)-1-methyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one (119)

[0276] Step 1: 6-chloro-3-(5-(difluoromethyl)-1-methyl-1H-1,2,4-triazol-3-yl)-2-methylpyridine (119b)

[0277] Compound 083-1b (500 mg) was dissolved in 1,4-dioxane (5 mL), followed by the addition of [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex (72.15 mg) and a 2M sodium carbonate solution (627.07 mg). The atmosphere was purged with nitrogen three times, followed by the addition of compound 119a (501.71 mg). The atmosphere was purged with nitrogen three more times, and the temperature was raised to 100°C with stirring for 2 hours. The reaction mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-20%) as the eluent to obtain the title compound 119b (400 mg).

[0278] LC-MS: (ESI, m / z) = 259.35 [M+H] +

[0279] Step 2: (S)-tert-Butyl 7-((5-(5-(difluoromethyl)-1-methyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate (119c)

[0280] Compound 119b (200 mg), tert-butyl (7S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylate (180.56 mg), tris(dibenzylideneacetone)dipalladium (70.81 mg), 2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl (63.49 mg), sodium tert-butoxide (148.62 mg), and toluene (2 mL) were added sequentially to a reaction flask. The atmosphere was purged with nitrogen three times and the mixture was stirred at 110°C for 2 hours. The reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (3 x 10 mL). The organic phases were combined and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-20%) to afford the title compound 119c (400 mg).

[0281] LC-MS: (ESI, m / z) = 435.50 [M+H] +

[0282] Step 3: (S)-N-(5-(5-(difluoromethyl)-1-methyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl)-5-azaspiro[2.4]heptane-7-amine (119d)

[0283] Compound 119c (60 mg) was added to a solution of hydrogen chloride in 1,4-dioxane (4 M, 2 mL) and stirred at room temperature for 30 minutes. The reaction solution was concentrated under reduced pressure to obtain the hydrochloride salt of compound 119d (45 mg, crude product). The crude product was used directly in the next step without purification.

[0284] LC-MS: (ESI, m / z) = 335.50 [M+H] +

[0285] Step 4: (R)-1-((S)-7-((5-(5-(difluoromethyl)-1-methyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one (119)

[0286] The hydrochloride salt of compound 119d (45 mg, crude product) was dissolved in N,N-dimethylacetamide (1 mL). Compound 1f (29.78 mg), N,N-diisopropylethylamine (57.98 mg), and 2-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium hexafluorophosphate (85.29 mg) were added sequentially. The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was directly purified by HPLC (column: XSelect CSH Fluorophenyl 5μm, 30mm x 150mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 30% B to 55% B over 10 minutes) to obtain the title compound 119 (40 mg).

[0287] LC-MS: (ESI, m / z) = 515.95 [M+H] +

[0288] 1 H NMR(400MHz,DMSO-d6)δ8.13–8.08(m,1H),7.88–7.81(m,1H),7.56–7.26( m,1H),7.03–6.90(m,1H),6.73–6.68(m,1H),6.49–6.39(m,1H),4.21–4.0 4(m,2H),4.00(s,3H),3.82(d,3H),3.75–3.58(m,2H),3.55–3.44(m,1.5H ),3.26–3.20(m,0.5H),2.57(d,3H),1.35–1.28(m,3H),0.88–0.45(m,4H).

[0289] Synthesis of intermediate 119a

[0290] Step 1: 3-Bromo-1-methyl-1H-1,2,4-triazole-5-carbaldehyde (119f)

[0291] Compound 119e (14 g) and ultra-dry tetrahydrofuran (140 mL) were added to a 250 mL three-necked flask. The nitrogen atmosphere was replaced three times, and the mixture was cooled to -45°C. n-Butyl lithium (4.47 g) was added dropwise. The mixture was stirred at -45°C for 30 minutes, then cooled to -75°C. N,N-dimethylformamide (5.52 g) was added dropwise. After 15 minutes, the cooling bath was removed and the reaction mixture was allowed to warm to room temperature. The reaction mixture was allowed to react for 2 hours. The mixture was poured into water (200 mL), and the reaction mixture was extracted with ethyl acetate (4 x 150 mL). The organic phases were combined and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-20%) to obtain the title compound 119f (5.5 g).

[0292] LC-MS: (ESI, m / z)=208.05[M+H2O+H] +

[0293] Step 2: 3-Bromo-5-difluoromethyl-1-methyl-1H-1,2,4-triazole (119a)

[0294] Under ice bath, compound 119f (4 g) and diethylaminosulfur trifluoride (13.57 g) were added to a 100 ml three-necked flask. Under nitrogen protection, the obtained residue was stirred and reacted overnight at room temperature. The reaction mixture was quenched with 0°C saturated aqueous ammonium bicarbonate solution (50 mL). The reaction mixture was extracted with dichloromethane (3×50 mL). The organic phases were combined, backwashed with saturated brine (50 mL), and dried over anhydrous sodium sulfate. After the obtained mixture was filtered, the filtrate was concentrated under reduced pressure. Purification by reverse phase column chromatography (C18 chromatography column, mobile phase, water (0.5% formic acid) and acetonitrile, 10% to 50% gradient 10 minutes) gave intermediate 119a (1.6 g).

[0295] LC-MS: (ESI, m / z) = 212.30 [M+H] +

[0296] Example 15 (2R)-1-[(7S)-7-{[5-(1,5-dimethyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl]amino}-5-azaspiro[2.4]heptane-5-yl]-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 132

[0297] Step 1: 3-Bromo-1,5-dimethyl-1H-1,2,4-triazole 132b

[0298] To a solution of 132a (3 g) in tetrahydrofuran (30 mL) was added dropwise a solution of n-butyllithium in n-hexane (2.5 M, 6.00 mL) at -78 degrees Celsius. After stirring for 1 hour, iodomethane (2.30 g) was added dropwise at -40 degrees Celsius. The reaction was stirred for 2 hours. The mixture was then quenched with a saturated aqueous ammonium chloride solution (30 mL) at 0 degrees Celsius. The reaction mixture was extracted with ethyl acetate (50 mL*3). The organic phases were combined, backwashed with saturated brine (50 ml), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The title compound 132b (2 g, crude product) was obtained.

[0299] GC-MS: (EI, m / z) = 175.00 [M] +

[0300] Step 2 6-chloro-3-(1,5-dimethyl-1H-1,2,4-triazol-3-yl)-2-methylpyridine 132c

[0301] To a solution of 132b (2 g) and 083-1b (2.88 g) in 1,4-dioxane (30 mL) were added [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride (831.41 mg) and sodium carbonate solution (2 M, 10 mL). Under nitrogen protection, the reaction was stirred at 100 degrees Celsius for 2 hours. The reaction mixture was extracted with ethyl acetate (30 mL*3). The organic phases were combined, backwashed with saturated brine (20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (1:1) to obtain the title compound 132c (300 mg).

[0302] LC-MS: (ESI, m / z) = 223.40 [M+H] +

[0303] Step 3: (7S)-7-{[5-(1,5-dimethyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl]amino}-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester 132d

[0304] To a toluene (3 mL) solution of compound 132c (300 mg) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (286 mg) were added tris(dibenzylideneacetone)dipalladium (61.68 mg), 2-dicyclohexylphosphine-2',6'-dimethoxybiphenyl (55.31 mg), and sodium tert-butoxide (258.95 mg). The atmosphere was replaced with nitrogen, and the reaction was stirred at 100 degrees Celsius for 2 hours. The mixture was then cooled to room temperature, diluted with water (10 ml), and extracted with ethyl acetate (20 mL*3). The organic phases were combined, backwashed with saturated brine (20 ml), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with dichloromethane / methanol (1:1) to obtain the title compound 132d (330 mg).

[0305] LC-MS: (ESI, m / z) = 399.25 [M+H] +

[0306] Step 4 (7S)-N-[5-(1,5-dimethyl-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl]-5-azaspiro[2.4]hept-7-amine 132e

[0307] To the reaction flask were added 132d (330 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (4 mL). The resulting residue was stirred and reacted for 1 hour. The reaction mixture was directly spin-dried to afford the hydrochloride salt of the title compound 132e (270 mg, crude product). The crude product was not further purified and was used directly in the next step.

[0308] LC-MS: (ESI, m / z) = 299.10 [M+H] +

[0309] Step 5 (2R)-1-[(7S)-7-{[5-(1,5-dimethyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl]amino}-5-azaspiro[2.4]heptane-5-yl]-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 132

[0310] To a solution of the hydrochloride salt of 132e (250 mg) and 1f (166.88 mg) in N,N-dimethylacetamide (5 mL) was added N,N-diisopropylethylamine (2165.71 mg). After stirring for 5 minutes, 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (477.85 mg) was added. The resulting residue was stirred for 1 hour. The reaction solution was directly purified by HPLC (XBridge BEH Shield RP18 column, 30 mm × 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 50% B) to obtain the title compound 132 (131.5 mg).

[0311] LC-MS (ESI): m / z = 480.20 [M+H] +

[0312] 1 H NMR(400MHz,DMSO-d6)δ8.12–8.09(m,1H),7.84–7.79(m,1H),6.86–6.73(m,1H ),6.73–6.68(m,1H),6.45–6.34(m,1H),4.18–4.04(m,2H),3.82(d,3H),3.78(d ,3H),3.75–3.58(m,2H),3.53(s,1H),3.50–3.44(m,0.5H),3.23(d,0.5H),2.5 6(d,3H),2.40(d,3H),1.36–1.27(m,3H),0.89-0.78(m,1H),0.71–0.44(m,3H).

[0313] Example 16 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl]amino}-5-azaspiro[2.4]heptan-5-yl]propan-1-one 136

[0314] Step 1 6-chloro-2-methyl-3-(1-methyl-1H-1,2,4-triazol-3-yl)pyridine 136b

[0315] Under nitrogen, 136a (383 mg) and 083-1b (200 mg) were added to a reaction flask, followed by 1,1'-bis(diphenylphosphino)ferrocenepalladium dichloride (64 mg) and a 2M sodium carbonate aqueous solution (1.2 mL). The nitrogen atmosphere was replaced, and finally 1,4-dioxane (4 mL) was added. The reaction was allowed to react at 100°C overnight. The resulting mixture was diluted with water (50 mL) and extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride aqueous solution (1 × 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-50%) as the eluent to afford the title compound 136b (150 mg).

[0316] LC-MS: (ESI, m / z) = 208.95 [M+H] +

[0317] Step 2 (S)-tert-Butyl 7-((6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 136c

[0318] Under nitrogen, 136b (130 mg) and tert-butyl (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylate (132 mg) were added to the reaction flask. Tris(dibenzylideneacetone)dipalladium (57 mg), sodium tert-butoxide (119 mg), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (51 mg) were then added. The nitrogen atmosphere was replaced, and finally, ultra-dry toluene (4 mL) was added. The reaction was continued at 100°C for 2 hours. The reaction mixture was diluted with water (50 mL) and extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride (1 × 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-50%) as the eluent to afford the title compound 136c (120 mg, 50.05%).

[0319] LC-MS: (ESI, m / z) = 385.15 [M+H] +

[0320] Step 3 (S)-N-(6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 136d

[0321] Compound 136c (40 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (1 mL) were added to the reaction flask at room temperature for 30 minutes. The reaction solution was concentrated under reduced pressure to obtain the hydrochloride salt of the title compound 136d (35 mg, crude product). The crude product was used in the next step without further purification.

[0322] LC-MS: (ESI, m / z) = 285.05 [M+H] +

[0323] Step 4 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl]amino}-5-azaspiro[2.4]heptan-5-yl]propan-1-one 136

[0324] At room temperature, the hydrochloride salt of 136d (30 mg) and 1f (22 mg) were added to a reaction flask and dissolved in N,N-dimethylacetamide (2 mL). N,N-diisopropylethylamine (40 mg) was then added and stirred for several minutes. Finally, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (60 mg) was added. The resulting mixture was stirred at room temperature for 2 hours. The mixture was directly purified by HPLC (XBridge BEH C18 column, 30 mm x 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 45% B over 10 minutes) to afford the title compound 136 (26.9 mg).

[0325] LC-MS: (ESI, m / z) = 465.95 [M+H] +

[0326] 1 H NMR(400MHz,DMSO-d6)δ8.43(d,1H),8.13–8.08(m,1H),7.87–7.82(m,1H),6.9 0–6.76(m,1H),6.73–6.68(m,1H),6.47–6.37(m,1H),4.19–4.04(m,2H),3.88(d ,3H),3.82(d,3H),3.76–3.58(m,2H),3.53(s,1H),3.50–3.43(m,0.5H),3.23( d,0.5H),2.57(d,3H),1.37–1.26(m,3H),0.89-0.78(m,1H),0.72–0.43(m,3H).

[0327] Example 17 (2R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[5-(1,5-dimethyl-1H-1,2,4-triazol-3-yl)-6-methylpyridin-2-yl]amino}-5-azaspiro[2.4]heptan-5-yl]propan-1-one 141

[0328] To a solution of the hydrochloride salt of 132e (50 mg) and 083-1f (36.13 mg) in N,N-dimethylacetamide (2 mL) was added N,N-diisopropylethylamine (433.14 mg). After stirring for 5 minutes, 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (95.57 mg) was added. The resulting residue was stirred for 1 hour. The reaction solution was directly purified by HPLC (XBridge BEH C18 OBD preparative column, 30 mm × 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 25% B to 51% B) to obtain the title compound 141 (27.11 mg).

[0329] LC-MS (ESI): m / z = 496.10 [M+H] +

[0330] 1 H NMR(400MHz,DMSO-d6)δ8.21(d,1H),7.84–7.79(m,1H),6.86–6.74(m,1H),6.72(d,1H),6.45–6.35(m,1H),4.16–4.00(m,2H),3.84(d,3H),3.7 8(d,3H),3.72–3.51(m,3H),3.49(d,0.5H),3.24(d,0.5H),2.56(d,3H) ,2.40(d,3H),1.36–1.23(m,3H),0.89–0.77(m,1H),0.73–0.45(m,3H).

[0331] Example 18 (2R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl]amino}-5-azaspiro[2.4]heptan-5-yl]propan-1-one 142

[0332] Step 1: (S)-tert-Butyl 7-((6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 142a

[0333] Under nitrogen, 136b (250 mg) and tert-butyl (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylate (254 mg) were added to a reaction flask. Tris(dibenzylideneacetone)dipalladium (109 mg), sodium tert-butoxide (230 mg), and 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (98 mg) were then added. Ultra-dry toluene (4 mL) was added, and the atmosphere was replaced with nitrogen. The reaction was incubated at 100°C for 2 h. The reaction mixture was cooled to room temperature, diluted with water (30 mL), and extracted with ethyl acetate (3 × 30 mL). The organic phases were combined, backwashed with saturated sodium chloride (1 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using methanol / dichloromethane (0-50%) as the eluent to afford the title compound 142a (200 mg).

[0334] LC-MS: (ESI, m / z) = 385.10 [M+H] +

[0335] Step 2 (S)-N-(6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 142b

[0336] To the reaction flask, 142a (40 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (1 mL) were added and allowed to react at room temperature for 30 minutes. The reaction solution was concentrated under reduced pressure to afford the hydrochloride salt of the title compound 142b (35 mg, crude product). The crude product was used in the next step without further purification.

[0337] LC-MS: (ESI, m / z) = 285.10 [M+H] +

[0338] Step 3 (2R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-[(7S)-7-{[6-methyl-5-(1-methyl-1H-1,2,4-triazol-3-yl)pyridin-2-yl]amino}-5-azaspiro[2.4]heptan-5-yl]propan-1-one 142

[0339] At room temperature, the hydrochloride salt of 142b (35 mg) and 083-1f (26 mg) were added to a reaction flask and dissolved in N,N-dimethylacetamide (2 mL). N,N-diisopropylethylamine (94 mg) was then added and stirred for 5 minutes. Finally, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (70 mg) was added. The mixture was stirred for 2 hours. The crude product was purified by HPLC (XBridge BEH C18 column, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 16% B to 51% B over 10 minutes) to afford the title compound 142 (17.09 mg).

[0340] LC-MS: (ESI, m / z) = 481.90 [M+H] +

[0341] 1 H NMR(400MHz,DMSO-d6)δ8.44(d,1H),8.22(d,1H),7.88–7.82(m,1H),6.91–6.78(m,1H),6.72(d,1H),6.47–6.37(m,1H),4.16–4.01(m,2H),3.8 8(s,3H),3.84(d,3H),3.72–3.51(m,3H),3.49(d,0.5H),3.24(d,0.5H) ,2.57(d,3H),1.35–1.25(m,3H),0.89–0.78(m,1H),0.74–0.45(m,3H).

[0342] Example 19 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((7S)-7-((6-methyl-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one 143

[0343] Step 1 6-chloro-2-methyl-3-(1-methyl-1H-pyrazol-4-yl)pyridine 143a

[0344] To a solution of compound 083-1a (1 g) and 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (1.01 g) in 1,4-dioxane (20 mL) and water (5 mL) were added cesium carbonate (4.73 g) and (2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl)(2'-amino-1,1'-biphenyl-2-yl)methanesulfonate palladium(II) (188.95 mg). Under nitrogen protection, the resulting residue was stirred and reacted at 100 degrees Celsius for 2 hours. The reaction mixture was diluted with water (30 mL) and extracted with ethyl acetate (50 mL*3). The organic phases were combined, backwashed with saturated brine (30 mL*1), and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using petroleum ether / ethyl acetate (1:1) to afford the title compound 143a (480 mg).

[0345] LC-MS: (ESI, m / z) = 208.10 [M+H] +

[0346] Step 2 (S)-tert-Butyl 7-((6-methyl-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 143b

[0347] To a toluene (3 mL) solution of 143a (250 mg) and tert-butyl (7S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylate (255.58 mg) were added tris(dibenzylideneacetone)dipalladium (55.12 mg), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (49.42 mg), and sodium tert-butoxide (231.40 mg). Under nitrogen, the mixture was stirred at 100°C for 2 hours. The reaction mixture was then cooled to room temperature. The reaction mixture was diluted with water (20 mL) and extracted with ethyl acetate (20 mL x 3). The organic phases were combined, backwashed with saturated brine (20 mL x 1), and dried over sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (1:1) to obtain the title compound 143b (320 mg).

[0348] LC-MS: (ESI, m / z) = 384.10 [M+H] +

[0349] Step 3 (S)-N-(6-methyl-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)-5-azaspiro[2.4]heptane-7-amine 143c

[0350] To the reaction flask were added 143b (70 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (2 mL). The mixture was stirred at room temperature for 1 hour to afford the hydrochloride salt of the title compound 143c (50 mg). The crude product was used in the next step without further purification.

[0351] LC-MS: (ESI, m / z) = 284.45 [M+H] +

[0352] Step 4 (2R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((7S)-7-((6-methyl-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-yl)propan-1-one 143

[0353] To a solution of the hydrochloride salt of 143c (25 mg) and compound 1f (17.57 mg) in N,N-dimethylacetamide (1 mL) was added N,N-diisopropylethylamine (228.05 mg). After stirring for 5 minutes, 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (50.32 mg) was added. The resulting mixture was stirred for 1 hour. The reaction solution was purified by HPLC (column specifications: YMC Triart C18 ExRs preparative column, 30 mm × 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 35% B to 57% B) to obtain the title compound 143 (10.96 mg).

[0354] LC-MS (ESI): m / z = 465.00 [M+H] +

[0355] 1 H NMR(400MHz,DMSO-d6)δ8.13–8.09(m,1H),7.79–7.75(m,1H),7.54–7.50(m,1H),7 .36–7.30(m,1H),6.72–6.68(m,1H),6.61–6.47(m,1H),6.43–6.32(m,1H),4.15–4 .01(m,2H),3.85(d,3H),3.82(d,3H),3.75–3.48(m,3H),3.48–3.41(m,0.5H),3.2 4(d,0.5H),2.33(d,3H),1.35–1.27(m,3H),0.87–0.76(m,1H),0.70–0.42(m,3H).

[0356] Example 20 (2R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((7S)-7-[6-methyl-5-(1-methyl-1H-pyrazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptan-5-yl]propan-1-one 144

[0357] To a solution of the hydrochloride salt of compound 143c (25 mg) and compound 083-1f (19.02 mg) in N,N-dimethylacetamide (1 mL) was added N,N-diisopropylethylamine (228.05 mg). After stirring for 5 minutes, 2-(7-azobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (50.32 mg) was added. The resulting residue was stirred for 1 hour. The reaction solution was purified by HPLC (column specifications: YMC Triart C18 ExRs preparative column, 30 mm × 150 mm, 5 μm, mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 40% B to 60% B) to obtain the title compound 144 (12.19 mg).

[0358] LC-MS (ESI): m / z = 480.90 [M+H] +

[0359] 1 H NMR(400MHz,DMSO-d6)δ8.21(d,1H),7.79–7.76(m,1H),7.53–7.51(m,1H) ),7.36–7.30(m,1H),6.72(d,1H),6.60–6.48(m,1H),6.43–6.33(m,1H), 4.13–3.98(m,2H),3.88–3.81(m,6H),3.70–3.47(m,3.5H),3.25(d,0.5H ),2.33(d,3H),1.35–1.25(m,3H),0.88–0.76(m,1H),0.72–0.44(m,3H).

[0360] Example 21 (R)-1-((S)-7-((5-(1,2-dimethyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 149

[0361] Step 1 6-chloro-3-(1,2-dimethyl-1H-imidazol-4-yl)-2-methylpyridine 149b

[0362] Compound 149a (200 mg) was dissolved in a solution of 1,4-dioxane (2 mL) and water (0.2 mL), and 083-1b (144.85 mg), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (41.81 mg), and potassium carbonate (157.92 mg) were added sequentially. The atmosphere was replaced with nitrogen, and the reaction was allowed to proceed overnight at 60°C. The mixture was then cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (3 x 10 mL). The organic phases were combined and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-100%) to obtain the title compound 149b (130 mg).

[0363] LC-MS: (ES, m / z) = 221.90 [M+H] +

[0364] Step 2 (S)-tert-Butyl 7-((5-(1,2-dimethyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 149c

[0365] Compound 149b (100 mg), (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (95.76 mg), tris(dibenzylideneacetone)dipalladium (41.31 mg), 2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl (37.04 mg), and sodium tert-butoxide (86.70 mg) were added sequentially to a sample vial. Toluene (1 mL) was added to displace the nitrogen atmosphere. The reaction was then continued at 100°C for 2 hours. The mixture was then cooled to room temperature, diluted with water (5 mL), and extracted with ethyl acetate (3 x 5 mL). The organic phases were combined and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by reverse-phase column chromatography using the following conditions (C18 column, mobile phase: water and methanol, gradient from 0% to 100% over 20 minutes). This afforded the title compound 149c (30 mg).

[0366] LC-MS: (ES, m / z) = 398.25 [M+H] +

[0367] Step 3 (S)-N-(5-(1,2-dimethyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 149d

[0368] Compound 149c (30 mg) was added to a solution of hydrogen chloride in 1,4-dioxane (4 M, 1 mL) and stirred at room temperature for 30 minutes. The reaction solution was then concentrated under reduced pressure to obtain the hydrochloride salt of the title compound 149d (20 mg). The crude product was used directly in the next step without further purification.

[0369] LC-MS: (ES, m / z) = 298.45 [M+H] +

[0370] Step 4 (R)-1-((S)-7-((5-(1,2-dimethyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 149

[0371] To a solution of the hydrochloride salt of compound 149d (20 mg) and compound 1f (20 mg) in N,N-dimethylformamide (1 mL) at room temperature was added dropwise N,N-diisopropylethylamine (64.89 mg). After the addition was complete, the mixture was stirred for 5 minutes, followed by the addition of propylphosphonic anhydride (79.87 mg). The reaction system was stirred at room temperature for 1 hour. The crude product was purified by HPLC (YMC Triart C18 ExRs 5 μm, 30 mm x 150 mm; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 30% B to 52% B over 10 minutes) to afford the title compound 149 (1 mg).

[0372] LC-MS: (ES, m / z) = 479.25 [M+H] +

[0373] 1 H NMR(400MHz,DMSO-d6)δ8.12–8.08(m,1H),7.72–7.66(m,1H),7.09(s,1H), 6.72–6.67(m,1H),6.56–6.43(m,1H),6.42-6.31(m,1H),4.16–4.02(m,2H), 3.82(d,3H),3.74–3.67(m,0.5H),3.65–3.41(m,6H),3.24(d,0.5H),2.39(d ,3H),2.30(s,3H),1.36–1.26(m,3H),0.88–0.75(m,1H),0.69–0.42(m,3H).

[0374] Example 22 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methyl-5-(1-methyl-1H-imidazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 150

[0375] Step 1 6-chloro-2-methyl-3-(1-methyl-1H-imidazol-4-yl)pyridine 150b

[0376] To a solution of compound 150a (200 mg) and compound 083-1b (314.94 mg) in 1,4-dioxane (5 mL) was added 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloromethane complex (90.89 mg) and a 2M aqueous sodium carbonate solution (1 mL). Under nitrogen, the reaction mixture was stirred at 100°C for 2 hours. The reaction mixture was diluted with water (20 mL) and then extracted with ethyl acetate (20 mL x 3). The combined organic phases were backwashed with saturated brine (20 mL) and dried over anhydrous sodium sulfate. The resulting mixture was filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (1:1) to obtain the title compound 150b (45 mg).

[0377] LC-MS: (ESI, m / z) = 208.05 [M+H] +

[0378] Step 2 (S)-tert-Butyl 7-((6-methyl-5-(1-methyl-1H-imidazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 150c

[0379] To a toluene (2 mL) solution of 150b (45 mg) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (46.0 mg) were added tris(dibenzylideneacetone)dipalladium (19.84 mg), 2-dicyclohexylphosphine-2',6'-dimethoxybiphenyl (17.79 mg) and sodium tert-butoxide (41.65 mg). Under nitrogen protection, the reaction was stirred at 100°C for 2 hours. The reaction mixture was then cooled to room temperature. The reaction mixture was diluted with water (5 mL) and then extracted with ethyl acetate (5 mL*3). The organic phases were combined, backwashed with saturated brine (5 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered and concentrated under reduced pressure. Purification by silica gel column chromatography with petroleum ether / ethyl acetate (1:1) gave the title compound 150c (70 mg).

[0380] LC-MS: (ESI, m / z) = 383.95 [M+H] +

[0381] Step 3 (S)-N-(6-methyl-5-(1-methyl-1H-imidazol-4-yl)pyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 150d

[0382] Compound 150c (70 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (2 mL) were added to the reaction flask. The resulting mixture was stirred for 1 hour. The residue was concentrated under reduced pressure to afford the hydrochloride salt of the title compound 150d (70 mg). The crude product was used in the next step without further purification.

[0383] LC-MS: (ESI, m / z) = 284.15 [M+H] +

[0384] Step 4 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((6-methyl-5-(1-methyl-1H-imidazol-4-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 150

[0385] To a solution of the hydrochloride salt of 150d (50 mg) in N,N-dimethylacetamide (2 mL) were added compound 1f (31.14 mg) and N,N-diisopropylethylamine (101.02 mg). After stirring for 5 minutes, 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (89.16 mg) was added. The resulting residue was stirred for 1 hour. The reaction solution was directly purified by HPLC (XBridge BEH C18 OBD Prep Column, 30 mm × 150 mm, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 23% B to 47% B) to obtain the title compound 150 (10.3 mg).

[0386] LC-MS (ES+): m / z = 465.20 [M+H] +

[0387] 1H NMR(400MHz,DMSO-d6,ppm)δ8.12–8.08(m,1H),7.74–7.68(m,1H),7.60–7.57(m ,1H),7.18–7.15(m,1H),6.72–6.68(m,1H),6.57-6.43(m,1H),6.43-6.33(m,1H ),4.17-4.03(m,2H),3.82(d,3H),3.75–3.57(m,5H),3.56-3.42(m,1.5H),3.24 (d,0.5H),2.41(d,3H),1.35–1.28(m,3H),0.89–0.77(m,1H),0.70–0.42(m,3H).

[0388] Example 23 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-7-((5-(4-(difluoromethyl)pyrimidin-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 156

[0389] Step 1 4-(difluoromethyl)-2-(6-fluoro-2-methylpyridin-3-yl)pyrimidine 156b

[0390] Compound 156a (1.08 g) and 2-chloro-4-(difluoromethyl)pyrimidine (500 mg) were dissolved in a mixture of 1,4-dioxane (10 mL) and water (3 mL). [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex (249 mg), cuprous chloride (60 mg), and sodium carbonate (970 mg) were then added. The atmosphere was purged with nitrogen three times, and the temperature was raised to 100°C with stirring for 2 hours. The reaction mixture was then cooled to room temperature, diluted with water (50 mL), and extracted with ethyl acetate (3 x 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 x 50 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-21%) as the eluent to obtain the title compound 156b (550 mg).

[0391] LC-MS: (ESI, m / z) = 240.05 [M+H] +

[0392] Step 2 (S)-tert-Butyl 7-((5-(4-(difluoromethyl)pyrimidin-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 156c

[0393] Under nitrogen, 156b (202 mg) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid tert-butyl ester (150 mg) were added to a reaction flask. N,N-diisopropylethylamine (913 mg) was then added to displace the nitrogen atmosphere. Finally, methylpyrrolidone (3 mL) was added and the mixture was allowed to react overnight at 140°C. The reaction mixture was then diluted with water (30 mL) and extracted with ethyl acetate (3 x 30 mL). The combined organic phases were backwashed with saturated sodium chloride (1 x 30 mL) and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-50%) as the eluent to afford the title compound 156c (60 mg).

[0394] LC-MS: (ESI, m / z) = 432.45 [M+H] +

[0395] Step 3 (S)-N-(5-(4-(difluoromethyl)pyrimidin-2-yl)-6-methylpyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 156d

[0396] Compound 156c (60 mg) and a 4 M solution of hydrogen chloride in 1,4-dioxane (1 mL) were added to a reaction flask at room temperature and allowed to react for 30 minutes. The reaction solution was concentrated under reduced pressure to afford the hydrochloride salt of the title compound 156d (45 mg, crude product). The crude product was used in the next step without further purification.

[0397] LC-MS: (ESI, m / z) = 332.10 [M+H] +

[0398] Step 4 (R)-2-(5-chloro-2-methoxypyridin-4-yl)-1-((S)-7-((5-(4-(difluoromethyl)pyrimidin-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 156

[0399] Compound 156d (40 mg) and compound 083-1f (26 mg) were added to a reaction flask at room temperature and dissolved in N,N-dimethylformamide (2 mL). N,N-diisopropylethylamine (156 mg) was then added and stirred for five minutes. Finally, propylphosphonic anhydride (192 mg) was added and stirred at room temperature for 2 hours. The crude product was purified by HPLC (column specifications: XBridge BEH C18 column, 30x150, 5 μm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 45% B to 70% B over 8 minutes) to afford the title compound 156 (27.9 mg).

[0400] LC-MS: (ESI, m / z) = 528.80 [M+H] +

[0401] 1 H NMR(400MHz, DMSO-d6)δ9.06–9.02(m,1H),8.22(d,1H),8.10–8.04(m,1H),7.56–7.52(m,1H),7.24–6.83(m,2H),6.73(d,1H),6.56–6.45(m,1 H),4.22–4.05(m,2H),3.84(d,3H),3.75–3.47(m,3.5H),3.25(d,0.5H) ,2.63(d,3H),1.36–1.26(m,3H),0.90–0.78(m,1H),0.75–0.48(m,3H).

[0402] Example 24 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((5-(2-(2-hydroxypropan-2-yl)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 181

[0403] Step 1 2-(4-bromo-1-methyl-1H-imidazol-2-yl)propan-2-ol 181b

[0404] Under nitrogen protection, methylmagnesium chloride (13.5 mL, 1 M) was added dropwise to a solution of 181a (1 g) in tetrahydrofuran (15 mL) at 0°C. After the addition was complete, the system was stirred at 0°C for 2 hours. The reaction mixture was then quenched by the addition of saturated aqueous ammonium chloride (20 mL) at 0°C. The aqueous phase was extracted with ethyl acetate (3 × 30 mL). The organic phases were combined, backwashed with saturated brine (2 × 20 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain the title compound 181b (670 mg). The resulting mixture was not further purified and was directly used in the next step.

[0405] LC-MS: (ESI, m / z) = 218.90 [M+H] +

[0406] Step 2 (S)-7-((5-(2-(2-hydroxypropyl-2-yl)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 181d

[0407] To a solution of 181b (189.11 mg) and 181c (200 mg) in 1,4-dioxane (1 mL) / water (0.2 mL) at room temperature were added 1,1'-bis(di-tert-butylphosphino)ferrocenedichloropalladium (28.13 mg) and potassium phosphate (183.23 mg). After the addition was complete, the reaction system was stirred at 80°C for 4 hours. The reaction mixture was then diluted with 10 mL of water, and the resulting mixture was extracted with ethyl acetate (3 × 30 mL). The organic phases were combined, backwashed with saturated brine (2 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (2:3) to obtain the title compound 181d (175 mg).

[0408] LC-MS: (ESI, m / z) = 476.10 [M+H] +

[0409] Step 3 (S)-2-(4-(6-((5-azaspiro[2.4]hept-7-yl)amino)-2-methylpyridin-3-yl)-1-methyl-1H-imidazol-2-yl)propan-2-ol 181e

[0410] Compound 181d (60 mg) was dissolved in trifluoroacetic acid (1 mL) and stirred at 60°C for 2 h under nitrogen. The resulting residue was concentrated under reduced pressure to afford the hydrochloride salt of the title compound 181e (20 mg). The resulting mixture was used in the next step without further purification.

[0411] LC-MS: (ESI, m / z) = 342.00 [M+H] +

[0412] Step 4 (R)-2-(5-fluoro-2-methoxypyridin-4-yl)-1-((S)-7-((5-(2-(2-hydroxypropan-2-yl)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)propan-1-one 181

[0413] To a solution of the hydrochloride salt of 181e (20 mg) and compound 1f in N,N-dimethylacetamide (0.57 mL) were added dropwise N,N-diisopropylethylamine (37.85 mg) and N,N,N′,N′-tetramethyl-O-(7-azabenzotriazol-1-yl)uronium hexafluorophosphate (33.41 mg). The mixture was stirred at room temperature for 30 minutes. The reaction solution was purified by HPLC (column specifications: YMC Triart C18 ExRs 5μm, 30x150mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 28% B to 51% B over 10 minutes) to afford the title compound 181 (10 mg).

[0414] LC-MS: (ESI, m / z) = 522.90 [M+H] +

[0415] 1 H NMR(400MHz,DMSO-d6)δ8.12–8.08(m,1H),7.74–7.69(m,1H),7.07(d,1H),6.72–6.68 (m,1H),6.51(d,0.5H),6.44–6.33(m,1.5H),5.26(d,1H),4.16–4.03(m,2H),3.85–3.7 9(m,6H),3.75–3.68(m,0.5H),3.65–3.58(m,1.5H),3.55–3.41(m,1.5H),3.23(d,0.5 H),2.41(d,3H),1.53(s,6H),1.35–1.27(m,3H),0.89–0.77(m,1H),0.70–0.41(m,3H).

[0416] Synthesis of 181c

[0417] Step 1 (S)-7-((6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 181g

[0418] 2-Bromo-6-methylpyridine 181f (2.5 g) and (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester (3.6 g) were dissolved in 1,4-dioxane (30 mL). (2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1'-biphenyl)(2-amino-1,1'-biphenyl-2-yl)palladium(II) methanesulfonate (1.2 g) and cesium carbonate (9.5 g) were added. The mixture was heated to 90°C and stirred under a nitrogen atmosphere overnight. The reaction mixture was filtered, the filtrate was diluted with water (50 mL), and the mixture was extracted with ethyl acetate (3 × 100 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 100 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-25%) as eluent to give 181 g (2.5 g) of the title compound.

[0419] LC-MS: (ESI, m / z) = 338.05 [M+H] +

[0420] Step 2: (S)-7-((5-bromo-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 181h: 181g (2.5g) was dissolved in dichloromethane (25mL), and a mixed solution of 1,3-dibromo-5,5-dimethylimidazolidin-4-one (1.05g) and dichloromethane (10mL) was added under ice-cooling, and stirring was continued for 1 hour. The reaction mixture was quenched with saturated aqueous sodium bicarbonate solution (50mL), and the mixture was then extracted with dichloromethane (3×50mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2×50mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-16%) as the eluent to give the title compound 181h (2.2g).

[0421] LC-MS: (ESI, m / z) = 416.00 [M+H] +

[0422] Step 3: (S)-Benzyl 7-((6-methyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylate 181c: 181h (1.07 g) and bis(boronic acid)pinacolato (1.31 g) were dissolved in 1,4-dioxane (10 mL). [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex (210 mg) and potassium acetate (560 mg) were then added. The atmosphere was purged with nitrogen three times, and the temperature was raised to 100°C with stirring for 2 hours. The reaction mixture was cooled to room temperature and diluted with water (30 mL). The mixture was then extracted with ethyl acetate (3 × 50 mL). The combined organic phases were backwashed with saturated sodium chloride solution (2 × 20 mL) and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-33%) as eluent to give the title compound 181c (780 mg).

[0423] LC-MS: (ESI, m / z) = 464.10 [M+H] +

[0424] Example 25 (R)-1-((S)-7-((5-(2-(difluoromethoxy)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 185

[0425] Step 1: 4-Bromo-2-methoxy-1-methyl-1H-imidazole 185b

[0426] Compound 185a (3 g) was dissolved in anhydrous methanol (30 mL). Sodium methoxide-methanol solution (30 wt%, 45 g) was slowly added dropwise at room temperature. After the addition was complete, the temperature was raised to 70°C and stirring was continued overnight. The reaction mixture was then cooled to room temperature, diluted with water (100 mL), and extracted with ethyl acetate (3 × 100 mL). The combined organic phases were backwashed with saturated sodium chloride solution (2 × 100 mL) and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure to yield the title compound 185b (1.1 g).

[0427] LC-MS: (ESI, m / z) = 191.25 [M+H] +

[0428] Step 2 6-chloro-3-(2-methoxy-1-methyl-1H-imidazol-4-yl)-2-methylpyridine 185d

[0429] Compounds 185b (600 mg) and 185c (976 mg) were dissolved in a mixture of 1,4-dioxane (6 mL) and water (1.5 mL). [1,1'-bis(di-tert-butylphosphino)ferrocene]palladium dichloride (206 mg) and potassium phosphate (1.34 g) were then added. The atmosphere was purged with nitrogen three times, and the temperature was raised to 80°C and stirred for 3 hours. The reaction mixture was cooled to room temperature and filtered. The filtrate was diluted with water (30 mL), and the mixture was extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-18%) as the eluent to afford the title compound 185d (350 mg).

[0430] LC-MS: (ESI, m / z) = 238.00 [M+H] +

[0431] Step 3 4-(6-chloro-2-methylpyridin-3-yl)-1-methyl-1H-imidazol-2-ol 185e

[0432] 185d (350 mg) was dissolved in tetrahydrofuran (4 mL), followed by the addition of dilute hydrochloric acid (1 M, 4 mL) and the mixture was heated to 60°C and stirred for 1 hour. The reaction system was cooled to room temperature, and the pH was adjusted to a weakly acidic state with saturated sodium bicarbonate solution. The mixture was then extracted with ethyl acetate (3 × 30 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure to afford the title compound 185e (300 mg).

[0433] LC-MS: (ESI, m / z) = 224.00 [M+H] +

[0434] Step 4 6-chloro-3-(2-(difluoromethoxy)-1-methyl-1H-imidazol-4-yl)-2-methylpyridine 185f

[0435] Compound 185e (300 mg) was dissolved in 1,4-dioxane (10 mL). Sodium hydroxide solution (20 wt%, 10 mL) was added and the temperature was raised to 65°C. Chlorodifluoromethane gas was then introduced and stirring continued at 65°C for 1 hour. The reaction system was cooled to room temperature and diluted with water (30 mL). The mixture was then extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-18%) as eluent to afford the title compound 185f (230 mg).

[0436] LC-MS: (ESI, m / z) = 273.95 [M+H] +7

[0437] Step 5 (S)-7-((5-(2-(difluoromethoxy)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 185h

[0438] 185f (200 mg) and 185g (198 mg) were dissolved in toluene (2 mL), followed by the addition of tris(dibenzylideneacetone)dipalladium (67 mg), 2-dicyclohexylphosphine-2′,6′-dimethoxy-biphenyl (60 mg), and sodium tert-butoxide (140 mg). The atmosphere was purged with nitrogen three times and the temperature was raised to 90°C and stirred for 1 hour. The reaction mixture was cooled to room temperature and filtered. The filtrate was diluted with water (30 mL), and the mixture was extracted with ethyl acetate (3×50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2×30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using ethyl acetate / petroleum ether (0-38%) as eluent to afford the title compound 185h (240 mg).

[0439] LC-MS: (ESI, m / z) = 484.10 [M+H] +

[0440] Step 6 (S)-N-(5-(2-(difluoromethoxy)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 185i

[0441] 185h (240 mg) was placed in a reaction flask, trifluoroacetic acid (5 mL) was added at room temperature, and the temperature was raised to 60°C and stirred for 1 hour. The reaction mixture was concentrated under reduced pressure to afford the trifluoroacetic acid salt of the title compound 185i (180 mg, crude product). The crude product was used in the next step without further purification.

[0442] LC-MS: (ESI, m / z) = 350.05 [M+H] +

[0443] Step 7 (R)-1-((S)-7-((5-(2-(difluoromethoxy)-1-methyl-1H-imidazol-4-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 185

[0444] The trifluoroacetic acid salt of 185i (90 mg, crude) and compound 1f (50 mg) were dissolved in N,N-dimethylacetamide (2 mL). N,N-diisopropylethylamine (322 mg) was added to make the mixture basic. 1-Propylphosphonic anhydride (50 wt% in ethyl acetate, 800 mg) was then added and stirred at room temperature for 1 hour. The resulting mixture was filtered, and the filter cake was washed with N,N-dimethylacetamide (2 × 1 mL). The crude product was purified by HPLC (column specifications: Kinetex 5μm EVO C18, 30 x 150 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 39% B to 54% B over 10 minutes) to afford the title compound 185 (46.67 mg).

[0445] LC-MS: (ESI, m / z) = 530.90 [M+H] +

[0446] 1 H NMR(400MHz, DMSO-d6)δ8.12–8.08(m,1H),7.71–7.31(m,2H),7.01(d,1H),6.73–6.68(m,1H),6.64–6.49(m,1H),6.45–6.33(m,1H),4.1 6–4.03(m,2H),3.82(d,3H),3.75–3.41(m,6.5H),3.23(d,0.5H),2.40(d,3H),1.36–1.27(m,3H),0.89–0.76(m,1H),0.70–0.42(m,3H).

[0447] Synthesis of intermediate 185c

[0448] Step 1 (6-chloro-2-methylpyridin-3-yl)boronic acid 185j

[0449] Under nitrogen, 083-1a (50 g) and tetrahydrofuran (500 mL) were added to a 2-liter three-necked flask and the temperature was lowered to -78°C. n-Butyllithium solution (2.5 mol / L, tetrahydrofuran / n-hexane solution, 117 mL) was added dropwise at -78°C. After complete addition, the reaction was stirred at -78°C for 40 minutes. Trimethyl borate (50.33 g) was then added dropwise at -78°C. The reaction was continued at -78°C for 60 minutes, and then quenched by the addition of saturated aqueous ammonium chloride (400 mL). The system was extracted with ethyl acetate (2 × 300 mL). The organic phases were combined, and the aqueous phase was adjusted to pH 6 with hydrochloric acid (2 M). The aqueous phase was extracted with methanol / dichloromethane (1 / 5) (4 × 500 mL). The combined organic phases were dried over anhydrous sodium sulfate and concentrated to yield the title compound 185j (42 g, crude product). The crude product was not further purified and was directly used in the next step.

[0450] LC-MS: (ES, m / z) = 172.05 [M+H] +

[0451] Step 2 6-Chloro-2-methyl-3-(4,4,5,5-tetraethyl-1,3,2-dioxaborolan-2-yl)pyridine 185c

[0452] Compound 185j (17 g) and 3,4-diethylhexane-3,4-diol (15.56 g) were dissolved in dichloromethane (170 mL) and reacted at room temperature for 12 hours. The reaction mixture was diluted with water (150 mL) and extracted with dichloromethane (4 × 150 mL). The organic phases were combined and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-15%) to afford the title compound 185c (20 g).

[0453] LC-MS: (ESI, m / z) = 310.50 [M+H] +

[0454] Synthesis of intermediate 185g

[0455] Step 1 (S)-7-((tert-Butoxycarbonyl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 185l

[0456] At 0°C, 185k (30 g) and triethylamine (42.90 g) dissolved in dichloromethane (300 mL) were added to a 1-liter three-necked flask. Benzyl chloroformate (36.16 g) was then added dropwise. After the addition was complete, the system was stirred at room temperature for 2 hours. The reaction mixture was quenched with ice water (100 mL) at room temperature. The reaction mixture was extracted with ethyl acetate (3 × 300 mL). The organic phases were combined, backwashed with saturated brine (1 × 100 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with petroleum ether / ethyl acetate (3:2) to obtain the title compound 185l (46.0 g).

[0457] LC-MS: (ES, m / z) = 347.10 [M+H] +

[0458] Step 2 (S)-7-amino-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 185g

[0459] At 0°C, add 185l (46g) dissolved in 1,4-dioxane (100mL) to a 1-liter three-necked flask. Add a 4M solution of hydrogen chloride in 1,4-dioxane (150mL) dropwise. After the addition is complete, continue stirring at room temperature for 1 hour. Quench the reaction mixture with saturated aqueous sodium bicarbonate at room temperature. Extract the reaction mixture with ethyl acetate (3 x 500mL). Combine the organic phases, backwash with saturated brine (1 x 300mL), and dry over anhydrous sodium sulfate. Filter the resulting mixture, and concentrate the filtrate under reduced pressure to obtain 185g (28.0g) of the title compound.

[0460] LC-MS: (ES, m / z) = 247.05 [M+H] +

[0461] Example 26 (R)-1-((S)-7-((5-(7,7-difluoro-6,7-dihydro-5H-pyrrolo[1,2-a]imidazol-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 189

[0462] Step 1 (S)-7-((5-(7,7-difluoro-6,7-dihydro-5H-pyrrolo[1,2-a]imidazol-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]heptane-5-carboxylic acid benzyl ester 189b

[0463] 189a (300 mg, prepared according to patent "WO2023280254A1") and 181c (626 mg) were dissolved in a mixture of 1,4-dioxane (4 mL) and water (1 mL). Subsequently, [1,1'-bis(di-tert-butylphosphino)ferrocene]palladium dichloride (91 mg) and potassium phosphate (572 mg) were added. The atmosphere was replaced with nitrogen three times and the temperature was raised to 80°C and stirred for 1 hour. The reaction mixture was cooled to room temperature and diluted with water (30 mL). The mixture was then extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, backwashed with saturated sodium chloride solution (2 × 30 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with ethyl acetate / petroleum ether (0-100%) as the eluent to obtain the title compound 189b (400 mg).

[0464] LC-MS: (ESI, m / z) = 480.10 [M+H] +

[0465] Step 2 (S)-N-(5-(7,7-difluoro-6,7-dihydro-5H-pyrrolo[1,2-a]imidazol-2-yl)-6-methylpyridin-2-yl)-5-azaspiro[2.4]hept-7-amine 189c

[0466] 189b (200 mg) was placed in a reaction flask, trifluoroacetic acid (5 mL) was added at room temperature, and the temperature was raised to 60°C and stirring was continued for 1 hour. The reaction mixture was concentrated under reduced pressure to obtain the title compound 189c (160 mg, crude product). The crude product was not further purified and was directly used in the next step. LC-MS: (ESI, m / z) = 346.00 [M+H] +

[0467] Step 3 (R)-1-((S)-7-((5-(7,7-difluoro-6,7-dihydro-5H-pyrrolo[1,2-a]imidazol-2-yl)-6-methylpyridin-2-yl)amino)-5-azaspiro[2.4]hept-5-yl)-2-(5-fluoro-2-methoxypyridin-4-yl)propan-1-one 189

[0468] Compound 189c (80 mg, crude) and compound 1f (42 mg) were dissolved in N,N-dimethylacetamide (2 mL). The mixture was made basic by the addition of N,N-diisopropylethylamine (271 mg). 1-Propylphosphonic anhydride (50 wt% in ethyl acetate, 334 mg) was then added and stirred at room temperature for 1 hour. The resulting mixture was filtered, and the filter cake was washed with N,N-dimethylacetamide (2 × 1 mL). The crude product was purified by HPLC (column specifications: XBridge BEH Shield RP18 5 μm, 30 × 150 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 32% B to 50% B over 8 minutes) to afford the title compound 189 (45.3 mg).

[0469] LC-MS: (ESI, m / z) = 526.90 [M+H] +

[0470] 1 H NMR(400MHz,DMSO-d6)δ8.13–8.08(m,1H),7.70–7.64(m,1H),7.43(d,1H) ),6.74–6.58(m,2H),6.46–6.35(m,1H),4.29–4.22(m,2H),4.17–4.04(m ,2H),3.82(d,3H),3.75–3.42(m,3.5H),3.24(d,0.5H),3.20–3.07(m,2H ),2.43(d,3H),1.36–1.27(m,3H),0.89–0.77(m,1H),0.71–0.42(m,3H).

[0471] The compounds listed in Table 1 below were prepared using methods similar to those in Examples 11, 14, 16, or 24-26. The structural characterization data of these compounds are also listed in Table 1.

[0472] Table 1

[0473] Test Example 1: In vitro cell activity test of MC4R antagonists

[0474] a. Experimental consumables:

[0475] b. Experimental steps

[0476] √ Cell line: flipin-293-MC4

[0477] √Cell culture medium: DMEM, 10% fetal bovine serum 1*PS, 200μg / ml hygromycin

[0478] √ Experimental buffer: HBSS, 20mM HEPES, 0.1% BSA, 500μM IBMX

[0479] √Positive compound: ML00253764

[0480] c. Antagonist detection

[0481] 1. After digestion, cells were suspended in assay buffer and then seeded into 384-well cell assay plates.

[0482] 2. Add the test compound to the cell plate and incubate at 37°C for 10 minutes.

[0483] 3. Add Melanotan I to the cell plate and incubate at 37°C for 30 minutes.

[0484] 4. Dissolve EU cAMP tracer and light anti-camp detection reagents and dilute with the lysis buffer in the kit.

[0485] 5. Add the diluted detection reagent to the cell plate and incubate at room temperature for 1 hour.

[0486] 6. Read the sample using Envision (excitation: 340 nm, emission: 615 nm and 665 nm)

[0487] d. Data analysis

[0488] 1. %Inhibition calculation:

[0489] %Inhibiiton=(Signalcmpd-SignalAve_VC) / (SignalAve_PC-SignalAve_VC)×100.

[0490] Signalcmpd: compound signal value;

[0491] SignalAve_VC: signal value of negative control;

[0492] SignalAve_PC: signal value of positive control;

[0493] %inhibition: percentage of inhibition rate.

[0494] 2. Calculate compound IC using GraphPad nonlinear fitting formula 50 :

[0495] Y=Bottom+(Top-Bottom) / (1+10^((LogIC50-X)*HillSlope))

[0496] X: Logarithmic value of compound concentration; Y: Percentage of inhibition rate

[0497] The antagonistic activities of the test compounds against MC4R are shown in Table 2.

[0498] Table 2

[0499] Test Example 2 Pharmacokinetic Study of the Test Compound in SD Male Rats

[0500] 2.1 Experimental methods

[0501] On the day of administration, the test substance was prepared using a solvent formulation of 5% DMSO + 5% Solutol + 90% Saline, and the dosing solution was prepared and ready for use.

[0502] The IV group received a dose of 1 mg / kg at a concentration of 0.2 mg / mL, while the PO group received a dose of 2 mg / kg at a concentration of 0.2 mg / mL. Animals in the oral group fasted overnight, had free access to water, and were fed 4 hours after dosing. The IV group did not require fasting.

[0503] Animals were weighed before dosing, and the dose was calculated based on body weight. The animals were administered once intravenously and once PO on the day of dosing. Blood was collected venously at 0.083, 0.25, 0.5, 1.0, 2.0, 4.0, 8.0, and 24 hours after dosing for the IV group and at 0.25, 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, and 24 hours after dosing for the PO group. Approximately 0.20 mL of blood was collected in a K2-EDTA anticoagulant tube. Plasma was obtained by centrifugation within 1 hour of blood collection (centrifugation conditions: 6800 g, 6 minutes, 2-8°C). The samples were stored in a -80°C freezer until analysis.

[0504] The concentration of the test substance in the plasma of SD male rats was determined using a validated LC-MS / MS method.

[0505] 2.2 Experimental Results

[0506] The pharmacokinetics of the compounds of the present invention in SD male rats were determined by the above experiments. The results are shown in Table 3.

[0507] Table 3 Pharmacokinetic study results of the test compound in SD male rats (IV1mpk / PO2mpk)

[0508] 2.3 Experimental Conclusion

[0509] The AUC of compound 056-1 in SD male rats was higher than those of compounds 001-1 and 051-1, and its clearance was lower than those of compounds 001-1 and 051-1. The AUC and bioavailability of compound 114 in SD male rats were significantly higher than those of compounds 083-1 and 112, and its clearance was lower than those of compounds 083-1 and 112. The AUC and bioavailability of compound 185 in SD male rats were significantly higher than those of compound 197, and its clearance was lower than that of compound 197. In summary, structure-activity relationship studies revealed that in the structures of the MC4R antagonist compounds disclosed in the present invention, the specific structural modification of introducing a spiro three-membered ring into the tetrahydropyrrole ring can significantly improve the in vivo pharmacokinetic properties of such compounds and enhance their drugability.

[0510] Test Example 3 P450 enzyme inhibition experiment of test compound

[0511] 3.1 Experimental Purpose

[0512] The inhibitory effect of the test compounds on 3A4-M (with midazolam as substrate) in human liver microsomes was evaluated in vitro to assess the drug safety of the compounds.

[0513] 3.2 Experimental methods

[0514] This study used a human liver microsome system and a cytochrome P450 enzyme probe substrate recommended by the FDA's in vitro drug interaction study guidance to investigate the inhibitory effects of test compounds on the CYP3A4 enzyme in human liver microsomes. Human liver microsomes were incubated with varying concentrations of the test compound (0 to 30.0 μM) and the corresponding probe substrate. Changes in CYP enzyme activity were measured, and IC50 values ​​were calculated to evaluate the inhibitory potential of the test compounds against CYP3A4.

[0515] 3.2.1 Test concentration

[0516] The seven tested concentrations were 30.00, 10.00, 3.33, 1.11, 0.370, 0.123 and 0.0412 μM.

[0517] 3.2.2 Experimental and control groups

[0518] Test group (TG): human liver microsomes were incubated with different concentrations of test substances, NADPH (final concentration 2.00 mM) and probe substrates of different subenzymes (Table 4) for 5 minutes.

[0519] Positive control group (PC): human liver microsomes were incubated with inhibitors (Table 4), NADPH (final concentration 2.00 mM) and 3A4M probe substrate (Table 4) for 5 minutes.

[0520] Table 4 Probe substrates, inhibitors and incubation systems for each subenzyme

[0521] At the end of incubation, terminate the reaction with stop solution.

[0522] 3.2.3 Preparation of experimental solution

[0523] 3.2.3.1 Stop solution containing internal standard

[0524] Take 18.26 μL of 4.379 mg / mL tolbutamide stock solution and 13.02 μL of 3.071 mg / mL verapamil stock solution and add them to 4 L acetonitrile / methanol (volume 1:1) to prepare a stop solution with an internal standard concentration of 20.0 ng / mL tolbutamide and 10.0 ng / mL verapamil.

[0525] 3.2.3.2 Preparation of Human Liver Microsome (HLM) Working Solution

[0526] Dilute the 20.0 mg / mL HLM stock solution to a 0.2 mg / mL HLM working solution using phosphate buffer.

[0527] 3.2.3.3 Preparation of NADPH working solution

[0528] Weigh an appropriate amount of NADPH, prepare 8.00 mM NADPH working solution with phosphate buffer, and preheat to 37°C.

[0529] 3.2.3.4 Preparation of test sample stock solution

[0530] Weigh an appropriate amount of test sample and dissolve it in DMSO to make a 30 mM stock solution.

[0531] 3.2.3.5 Preparation of substrate working solution

[0532] The 80 mM testosterone stock solution was diluted to a 320 μM working solution concentration in phosphate buffered saline, and the 5 mM midazolam stock solution was diluted to a 20 μM working solution concentration in phosphate buffered saline.

[0533] 3.2.4 Experimental process

[0534] 3.2.4.1 Preheat phosphate buffer (pH 7.40 ± 0.05)

[0535] 3.2.4.2 Preparation of a series of test sample or positive control intermediate solutions

[0536] Transfer 8 μL of 30 mM test sample to 12 μL of DMSO to obtain a 12 mM test sample intermediate solution concentration, and continue to dilute with DMSO to obtain different concentrations of test sample working solutions (12 mM, 4 mM, 1.33 mM, 0.433 mM, 0.147 mM, 0.049 mM and 0.0164 mM).

[0537] 8 μL of 2.5 mM CYP3A4 inhibitor ketoconazole stock solution was added to 12 μL of DMSO to obtain a 1 mM positive control inhibitor working solution.

[0538] 3.2.4.3 Test Group (TG)

[0539] Mix 199 μL of 0.2 mg / mL HLM and 1 μL of the test article working solution (12 mM, 4 mM, 1.33 mM, 0.433 mM, 0.147 mM, 0.049 mM, and 0.0164 mM). Add 30 μL of this drug-containing liver microsome solution to a 96-well plate, followed by 10 μL of substrate working solution (320 μM testosterone or 20 μM midazolam) in duplicate. Preincubate at 37°C for 5 minutes. Initiate the reaction by adding 15 μL of 8 mM NADPH solution preheated at 37°C for 5-10 minutes. After incubating the 96-well plate at 37°C for 5 minutes, terminate the reaction by adding 180 μL of an acetonitrile:methanol (ACN:MeOH = 1:1) solution containing the internal standard (IS). After shaking the 96-well plate at 600 rpm for 5-10 minutes, centrifuge at 6000 rpm for 15 minutes. Take 80 μL of the supernatant, add it to 120 μL of ultrapure water, shake it evenly, and analyze it by LC-MS / MS.

[0540] 3.2.4.4 Positive Control Group (PC)

[0541] Mix 199 μL of 0.2 mg / mL HLM and 1 μL of 1 mM positive control inhibitor (ketoconazole) working solution. Add 30 μL of this drug-containing liver microsome solution to a 96-well plate, then add 10 μL of substrate working solution (20 μM midazolam) in duplicate. Pre-incubate at 37°C for 5 minutes. Initiate the reaction by adding 15 μL of 8 mM NADPH solution preheated at 37°C for 5-10 minutes. Incubate the 96-well plate at 37°C for 5 minutes, then terminate the reaction by adding 180 μL of a 1:1 solution of IS in ACN:MeOH. Shake the 96-well plate at 600 rpm for 5-10 minutes, then centrifuge at 6000 rpm for 15 minutes. Add 80 μL of the supernatant to 120 μL of ultrapure water, shake well, and analyze by LC-MS / MS.

[0542] 3.2.5 Data Analysis

[0543] 3.2.5.1 Calculate the relative activity of CYP450 subenzymes at different concentrations by dividing the peak area ratio of metabolites in the presence of the test article or positive control (analyte peak area / internal standard peak area) by the peak area ratio of metabolites in the solvent control sample without the test article or positive control. Calculate according to the following formula:

[0544] Relative activity (% of NC) = metabolite peak area ratio in the test group or positive control group / metabolite peak area ratio in the negative control group × 100

[0545] PC group enzyme activity inhibition rate % = 100% - relative enzyme activity % at the test concentration

[0546] 3.2.5.2 Half-maximal inhibitory dose (IC 50 ) was obtained by fitting the following model formula using Prism software:

[0547] Y=Bottom+(Top-Bottom) / (1+10^((Log IC 50 -X)×HillSlope))

[0548] X is the logarithm-converted test substance concentration (Log μM), Y is the relative activity (% of NC), Bottom and Top represent the percentage of enzyme activity at the bottom and top plateaus of the curve, respectively, and HillSlope represents the slope. To optimize the fit, the use of other (nonlinear) dose-response models was not considered a deviation from the protocol.

[0549] When the relative activity of the PC group (% of NC) is ≤50.0%, it indicates that the known inhibitor exhibits an inhibitory effect, indicating that the test system is qualified.

[0550] 3.3 Experimental Results

[0551] The experimental results of the P450 enzyme inhibition experiment of the test compounds are shown in Table 5.

[0552] Table 5

[0553] 3.4 Experimental Conclusion

[0554] The inhibition rate of compound 083-1 on CYP 3A4M is much stronger than that of compound 114 on CYP 3A4M. Similarly, the inhibition rate of compound 111 on CYP 3A4M is much stronger than that of compound 116 on CYP 3A4M. The inhibition rate of compound 200 on CYP 3A4M is much stronger than that of compound 201 on CYP 3A4M.

[0555] From this, it can be seen that in the molecular structure of this structural type of MC4R antagonist, when the specific structure of the spiro three-membered ring is introduced on the tetrahydropyrrole ring, the inhibition of the compound on CYP 3A4M can be significantly reduced, the risk of drug interactions can be reduced, and the drugability of the compound can be improved.

[0556] The above is an exemplary description of the implementation methods of the technical solution of the present invention. It should be understood that the scope of protection of the present invention is not limited to the above implementation methods. Any modifications, equivalent substitutions, improvements, etc. made by those skilled in the art within the spirit and principles of the present invention shall be included in the scope of protection of the claims of this application.

Claims

1. A compound of formula I and its racemate, stereoisomer, tautomer, isotope-labeled form, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug, or pharmaceutically acceptable salt: in, X is selected from NR', O or S; R' is selected from H, C 1-12 Alkyl or C 3-12 Cycloalkyl; Y is selected from CH or N; to exist or not to exist; R is selected from C 1-12 Alkyl, deuterated C 1-12 Alkyl, halogenated C 1-12 alkyl; A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl, 3-14 membered heterocyclic group, C 3-12 Cycloalkyl; each R a are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: C 1-12 Alkyl, C 1-12 Alkoxy; each R a1 the same or different, independently selected from deuterium, halogen, CN, C 1-12 Alkyl, C 1-12 alkoxy; E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted 3-14 membered N-containing heterocyclic group; each R e the same or different, independently selected from H, deuterium, halogen, CN, OH, oxo (=O), C 1-12 Alkyl, C 1-12 Alkoxy; or two R e The atoms connected to it form C 3-12 Cycloalkyl or 3-14 membered heterocyclic group; N in E is Connected, C in E is connected to X; G is selected from unsubstituted or optionally substituted with one, two or more R g Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl; each R g are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R g1 Substituted with the following groups: amino, C 1-12 Alkyl, C 1-12 Alkoxy, C 3-12 Cycloalkyl, C 6-14 Aryl, 5-14 membered heteroaryl; each R g1 the same or different, independently selected from deuterium, halogen, CN, C 1-12 Alkyl, C 1-12 alkoxy; M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted with the following groups: C 6-14 Aryl, 5-14 membered heteroaryl, 3-14 membered heterocyclic group, C 3-12 Cycloalkyl; each R m are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R m1 Substituted with the following groups: C 1-12 Alkyl, C 1-12 Alkoxy, C 3-12 Cycloalkyl; each R m1 the same or different, independently selected from deuterium, halogen, CN, OH, C 1-12 Alkyl, C 1-12 Alkoxy.

2. The compound according to claim 1, characterized in that X is selected from NR', O or S; R' is selected from H, C 1-6 Alkyl or C 3-8 cycloalkyl, such as methyl, ethyl or cyclopropyl; Preferably, R is selected from C 1-6 Alkyl, deuterated C 1-6 Alkyl, halogenated C 1-6 Alkyl; for example, methyl; Preferably, A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl, 3-10 membered heterocyclic group, C 3-8 Cycloalkyl; each R a are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: C 1-6 Alkyl, C 1-6 Alkoxy; each R a1 the same or different, independently selected from deuterium, halogen, CN, C 1-6 Alkyl, C 1-6 alkoxy; Preferably, A is selected from unsubstituted or optionally substituted with one, two or more R a Substituted with the following groups: Each R a The same or different, independently selected from H, F, Cl, Br, I, CN, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl, halogenated C 1-6 alkoxy; Preferably, each R a are the same or different and are independently selected from H, F, Cl, Br, CN, methyl, methoxy, trifluoromethyl, difluoromethoxy; Preferably, A is selected from 3. The compound according to claim 1 or 2, characterized in that E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted 3-8 membered N-containing heterocyclic group; each R e the same or different, independently selected from H, deuterium, halogen, CN, OH, oxo (=O), C 1-6 Alkyl, C 1-6 Alkoxy; or two R e The atoms connected to it form C 3-8 Cycloalkyl; N in E and Connected, C in E is connected to X; Preferably, E is selected from unsubstituted or optionally substituted with one, two or more R e Substituted with the following groups: Each R e the same or different, independently selected from H, F, Cl, Br, I, CN, OH, oxo (=O), C 1-6 Alkyl; or two R e The atoms connected to it form C 3-8 Cycloalkyl; Preferably, each R e are the same or different and are independently selected from H, F, CN, OH, oxo (=O), methyl; or two R e The atom to which it is attached forms a cyclopropyl group; Preferably, E is selected from 4. The compound according to any one of claims 1 to 3, characterized in that G is selected from unsubstituted or optionally substituted with one, two or more R g Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl; each R g are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R g1 Substituted with the following groups: amino, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-8 Cycloalkyl, C 6-10 Aryl, 5-10 membered heteroaryl; each R g1 the same or different, independently selected from deuterium, C 1-6 Alkyl, C 1-6 alkoxy; Preferably, G is selected from unsubstituted or optionally substituted with one, two or more R g Substituted with the following groups: Each R g The same or different, independently selected from H, halogen, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-8 Cycloalkyl, amino, C 1-6 Alkylamino, (C 1-6 Alkyl)2amino, C 6-10 Aryl, C 1-6 Alkyl 5-8 membered heteroaryl; Preferably, each R g the same or different, independently selected from H, F, methyl, ethyl, cyclopropyl, cyclobutyl, methoxy, methylamino (CH3NH-), dimethylamino ((CH3)2N-), phenyl, Preferably, G is selected from 5. The compound according to any one of claims 1 to 4, characterized in that M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted with the following groups: C 6-10 Aryl, 5-10 membered heteroaryl, 3-8 membered heterocyclic group; each R m are the same or different and are independently selected from H, deuterium, halogen, CN, unsubstituted or optionally substituted by one, two or more R m1 Substituted with the following groups: C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl; each R m1 the same or different, independently selected from deuterium, halogen, CN, OH, C 1-6 Alkyl, C 1-6 alkoxy; Preferably, M is absent or selected from unsubstituted or optionally substituted with one, two or more R m Substituted groups: pyridyl, pyrimidinyl, phenyl, oxazolyl, pyrazolyl, triazolyl, 1,2,3-triazolyl, tetrazolyl, thiazolyl, tetrahydropyrrolyl, piperidinyl, morpholinyl, piperazinyl, naphthyl, quinolinyl, azepanyl, imidazolyl, Each R m The same or different, independently selected from H, deuterium, F, Cl, Br, I, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, halogenated C 1-6 Alkyl, halogenated C 1-6 Alkoxy, hydroxy-C 1-6 alkyl; Preferably, each R m are the same or different and are independently selected from H, F, CN, methyl, methoxy, difluoromethyl, difluoromethoxy, trifluoromethyl, cyclopropyl, Preferably, each R m are the same or different and are independently selected from H, F, CN, methyl, methoxy, difluoromethyl, difluoromethoxy, trifluoromethyl, and cyclopropyl; Preferably, R m Selected from CN; Preferably, M is absent or selected from Preferably, M is selected from one, two or more R m Substituted 5-6 membered heteroaryl, wherein at least one R m For CN, other R m is present or absent, and when present, is identical or different and is independently selected from halogen, CN, C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, halogenated C 1-6 Alkyl, halogenated C 1-6 alkoxy; Preferably, M is selected from 5-6 membered heteroaryl substituted by CN.

6. The compound according to any one of claims 1 to 5, characterized in that The compound represented by formula I is selected from the following structures: Among them, X, R, A, E, G, M, R a 、R e 、R g 、R m It has the definition of any one of claims 1 to 5; p1, p2, p3, and p4 are independently selected from 0, 1, 2, 3, 4, or 5; Preferably, the compound represented by formula I has a structure represented by formula II below: Among them, R, M, G, R a 、R e Having the definition of any one of claims 1-5; p1, p2 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by formula II-1, II-2 or II-3: Among them, R, M, G, R a Having the definition of any one of claims 1-5; p1 is selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by formula III below: Among them, M, R, R a 、R e 、R g Having the definition of any one of claims 1-5; p1, p2, p3 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I is selected from the following structures: Among them, M, R, R a 、R e 、R g Having the definition of any one of claims 1-5; p1, p2, p3 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by formula IV below: Among them, R, G, R a 、R e 、R m Having the definition of any one of claims 1-5; p1, p2, p4 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by the following formula V: Among them, R, E, R a 、R g 、R m Having the definition of any one of claims 1-5; p1, p3, p4 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by formula VI below: Among them, A, R, R e 、R g 、R m Having the definition of any one of claims 1-5; p2, p3, p4 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has a structure represented by formula VII below: Among them, R, R a 、R e 、R g 、R m Having the definition of any one of claims 1-5; p1, p2, p3, p4 are selected from 0, 1, 2, 3, 4 or 5; Preferably, the compound represented by formula I has the structure represented by the following formula VIII or VIII-A: Wherein, A and M have the definitions as described in any one of claims 1-5; Preferably, the compound represented by formula I has the structure represented by the following formula VIII-1 or VIII-2: Among them, R a , M has the definition of any one of claims 1-5; p1 is selected from 0, 1, 2, 3, 4 or 5; Y1 is selected from CH or N; Preferably, the compound represented by Formula I has a structure represented by Formula IX or Formula IX-1: Among them, R a Having the definition of any one of claims 1-5; p1 is selected from 0, 1, 2, 3, 4 or 5; W1 and W2 are the same or different, independently selected from CH or N.

7. The compound according to any one of claims 1 to 6, characterized in that The compound represented by formula I is selected from the following structures: And / or, the compound represented by formula I is selected from the following structures:

8. The method for preparing the compound according to any one of claims 1 to 7, characterized in that: The following steps are involved: Wherein, X, Y, R, A, E, G, and M have the definitions described in any one of claims 1 to 7; L is selected from a leaving group such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl, and methylsulfonyl; Preferably, the preparation method comprises the following steps: wherein X, Y, R, A, E, G, and M have the definitions according to any one of claims 1 to 7; L, L1, L2, and L3 are selected from leaving groups, such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl, and methylsulfonyl; and PG is selected from amino protecting groups, such as Cbz (benzyloxycarbonyl), Boc (tert-butyloxycarbonyl), Fmoc (methyloxycarbonyl), Bn (benzyl), and PMB (p-methoxybenzyl). Preferably, the preparation method comprises the following steps: Wherein, X, Y, R, A, E, G, and M have the definitions described in any one of claims 1 to 7; L, L1, L2, and L3 are selected from leaving groups such as OH, OTf, F, Cl, Br, I, SnBu3 (tributyltin), 4,4,5,5-tetramethyl-1,3,2-dioxaborol Heterocyclopentan-2-yl, methylsulfonyl; PG is selected from amino protecting groups such as Cbz (benzyloxycarbonyl), Boc (tert-butyloxycarbonyl), Fmoc (methoxycarbonyl), Bn (benzyl), PMB (p-methoxybenzyl); Preferably, the preparation method comprises the following steps: Among them, Y, R, A, R e 、R g 、R m , PG, L, L1, L2, L3 have the definitions as described in any one of claims 1-8.

9. A pharmaceutical composition comprising the compound of any one of claims 1 to 7 and its racemate, stereoisomer, tautomer, isotope-labeled substance, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt thereof.

10. Use of the compound according to any one of claims 1 to 7 and its racemate, stereoisomer, tautomer, isotope-labeled form, nitrogen oxide, solvate, polymorph, metabolite, ester, prodrug or pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 9 in the preparation of a medicament; Preferably, the drug is a drug for diagnosing, preventing and / or treating a disease or condition mediated by the MC4R receptor; Preferably, the drug is an MC4R antagonist; Preferably, the disease or condition is cachexia (cachexia associated with cancer, cachexia associated with acquired immune deficiency syndrome (AIDS), cachexia associated with congestive heart failure (CHF); cachexia associated with chronic kidney disease (CKD); cachexia associated with treatment of other chronic diseases); anorexia or anorexia nervosa (anorexia nervosa in the elderly, anorexia associated with chemotherapy and / or radiotherapy); nausea and vomiting; weight loss (involuntary weight loss); growth retardation; sarcopenia; muscle atrophy; muscle weakness; frailty; osteoporosis; bone disease (bone loss); pain (neuropathic pain); anxiety (post-traumatic stress disorder or PTSD); depression; hypertension; malnutrition obesity (such as sarcopenia caused by chronic obesity); sexual dysfunction; and inflammatory diseases (inflammatory diseases associated with anorexia or cachexia, sarcopenia or muscle atrophy).

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