Synthesis method of compound containing oxo-pyridino-alicyclic ring or alicyclic ring structure and novel intermediate

Abstract

The invention belongs to the technical field of medicines, and provides a novel synthetic route of a compound containing an oxo-pyridino-alicyclic ring or alicyclic ring structure and a novel intermediate compound in the oxo-pyridino-alicyclic ring or alicyclic ring structure.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and specifically relates to a new synthesis method of a compound containing an oxopyridine heterocyclic or aliphatic ring structure and a novel intermediate compound involved therein. Background Art

[0002] Migraine is a common neurovascular disease characterized by recurrent, mostly unilateral, moderate to severe throbbing headaches that usually last for 4 to 72 hours and are accompanied by symptoms such as nausea, vomiting, and sensitivity to sound and light (phonophobia and photophobia) [1]. A period of sensory disturbance often occurs before a migraine attack. This type of migraine is called migraine with aura. One in seven migraine patients in my country have aura symptoms [2]. The main types of aura include visual disturbances such as dark spots, bright spots, and flashes, as well as symptoms such as parosmia, dizziness, tinnitus, and difficulty in speaking. Currently, more than 1 billion people worldwide are affected by migraine [3]. The rate of medical consultation for migraine patients in my country is only 52.9%, and the correct diagnosis rate by physicians is only 13.8%. In addition, there is a widespread lack of preventive treatment and excessive use of analgesics [2]. Migraine attacks can last for hours or even days and may seriously affect normal daily activities. This has created a significant demand for acute therapies that can quickly relieve headaches. In recent years, with the progress of migraine research at home and abroad, new therapeutic targets such as calcitonin gene-related peptide (CGRP) have emerged, enriching the treatment options for migraine.

[0003] Calcitonin gene-related peptide (CGRP) is a 37-amino acid polypeptide, whose N-terminal disulfide bond and amino C-terminal play an important role in activating receptors [4]. CGRP is mainly located in the trigeminal ganglion, c-fibers and Aδ-fibers in the dorsal root ganglion and the central nervous system [5]. There are two forms of CGRP in the human body, namely α-CGRP and β-CGRP, which differ by only 1-3 amino acids in different species. Among them, α-CGRP is highly expressed in sensory neurons, while β-CGRP mainly plays a role in the enteric nervous system [6]. CGRP exerts its biological function by binding to membrane receptors. CGRP receptors are mainly composed of calcitonin receptor-like receptors (CLR) and receptor activity modifying protein 1 (RAMP1), and receptor component protein (RCP) is also required to exert signal transduction function [7]. CGRP plays a major role in vasodilation in the body. Studies have shown that CGRP plays an important role in the occurrence and development of migraine. Clinical evidence shows that during migraine attacks, the level of CGRP in the jugular vein of patients increases. Intravenous injection of CGRP can cause moderate to severe headaches, while the use of CGRP antagonists can relieve the pain and related symptoms of migraine[4].

[0004] Currently, specific treatment modalities for acute phase in the migraine diagnosis and treatment guidelines include triptans, ergotamines, ditans, and gempans [2]. Triptans are 5-HT1B / 1D receptor agonists and have been widely used for acute treatment of migraine in the past two decades. Triptans have a high level of evidence and recommendation in the guidelines, but they also have significant cardiovascular risks, and many patients have low response rates. Ditans are 5-HT1F receptor agonists and do not have the adverse effect of vasoconstriction of triptans, but they have central nervous system depressant effects [8]. Gepans, antagonists developed for CGRP receptors, are a new treatment for migraine without cardiovascular risks and central nervous system depressant effects [7]. This provides a new option for patients who do not respond to triptans and ditans. Currently, the marketed gepant drugs developed for CGRP receptors include AbbVie's ubrogepant, atogepant, and Pfizer's rimegepant. All three drugs are oral tablets. Migraine patients often experience symptoms such as nausea and vomiting. At this time, oral medications are not feasible, and migraine attacks last for a long time, which seriously affects daily life. Pfizer's new drug Zavegepant (vazegepant) was launched in the United States in March 2023. The marketing dosage form is nasal spray. The approved indication is acute treatment of adult migraine with or without aura, but it cannot be used for the prevention of migraine, and it has the most common adverse reactions (at least 2% of ZAVZPRET-treated patients are greater than placebo) - taste disorders, nausea, nasal discomfort and vomiting.

[0005] BMS disclosed and protected the compound structure of Zavegepant in CN102834388B applied in 2011. Before CN102834388B, BMS also applied for related patents such as CN100558728C, CN100558428C, CN1914193A, and CN1972929A from 2003 to 2005, which also disclosed a series of heterocyclic CGRP antagonists related to the Zavegepant structure for the treatment of migraine.

[0006] Boehringer Ingelheim also applied for CN101146799A and WO2005084672A in 2005-2006, which also disclosed a CGRP antagonist, hoping to be used in the treatment of pain and other related diseases.

[0007] The above-mentioned previously marketed drugs or compounds disclosed in the literature related to the action of CGRP are different from those in the present invention. Considering the current status of the treatment of migraine and neuropathic pain, in order to meet and enrich clinical demand options, there is still a demand for compounds with new structures that specifically treat migraine and neuropathic headache.

[0008] The prior related application CN202310479996.8 of the present invention provides a new compound containing an oxo-substituted pyridine heterocyclic or aliphatic ring structure, and the exemplary structure of the new compound containing an oxo-substituted pyridine heterocyclic or aliphatic ring structure is as follows:

[0009] Where: X 1 , X 2 Each independently is CH or N, X 2 Preferably N; X 3 , X 4 are each independently C, CH or N; alternatively, X 3 , X 4 Optional not N; X 5 is C, CH, CH2, NH or N; Y 1 is CH2, O or NH, Y 1 Preferably O or NH; Y 2 O or S, Y 2 Preferably O; R 2 is independently at each occurrence -H, -(C1-C6)alkyl or -(C3-C8)cycloalkyl, n is 0, 1, 2, 3, 4, 5 or 6, or, -(R 2 ) n The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 2 ) n The attached ring forms a spiro ring structure, a cyclic structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S; R 3 is independently -H, -(C1-C6)alkyl or -(C3-C8)cycloalkyl at each occurrence, o is 0, 1, 2, 3, 4, 5 or 6, or, -(R 3 ) o The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 3 ) o The attached ring forms a spiro ring structure, a cyclic structure or a bridged ring structure, the heteroatom in the aliphatic heterocycle is O, N or S; A is a structure shown in the following formula: The G chain group and the two carbon atoms to which it is attached together form a 5- to 7-membered aliphatic ring or aliphatic heterocyclic ring, wherein the heteroatoms constituting the ring are -O-, -S- or R 1 is independently at each occurrence -H, -(C1-C6)alkyl or -(C3-C8)cycloalkyl, m is 0, 1, 2, 3, 4, 5 or 6, or -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

[0010] In some embodiments, the selection of the G chain group in A is: if the G chain group and the two carbon atoms to which it is attached together form a 5- or 6-membered aliphatic ring, the G chain group may optionally have 0 or 1 double bonds, and the rest are single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 7-membered aliphatic ring, the G chain group may optionally have 0, 1 or 2 double bonds, and the rest are single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 5-membered aliphatic heterocycle, the G chain group may have single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 6- or 7-membered aliphatic heterocycle, the G chain group may optionally have 0 or 1 double bonds, and the rest are single bonds.

[0011] In some embodiments, R 1 is independently -H, -(C1-C3)alkyl or -(C3-C6)cycloalkyl at each occurrence, m is 0, 1, 2 or 3, or, -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

[0012] In some embodiments, R 1 is independently -H, methyl, ethyl or cyclopropane at each occurrence, m is 0, 1 or 2, or, -(R 1 ) m The atoms to which it is attached together form cyclopropane, cyclobutane or cyclopentane, and the cyclopropane, cyclobutane or cyclopentane and -(R 1 ) m The attached rings constitute a spiro ring structure, a fused ring structure or a bridged ring structure.

[0013] In some embodiments, the A group is preferably a structure represented by the following formula IV-1, IV-2, IV-3, IV-4 or IV-5: In formula IV-1, IV-2, IV-3, IV-4 or IV-5, X 6 -O-, -S- or In formula IV-2, two "---"s both refer to single bonds, or any one of them is a double bond and the other is a single bond; in formula IV-3, three "---"s both refer to single bonds, or any one of them is a double bond and the other two are single bonds; in formula IV-4, "---"s both refer to single bonds or double bonds; in formula IV-5, two "---"s both refer to single bonds, or any one of them is a double bond and the other is a single bond.

[0014] In some embodiments, the G chain group and the two carbon atoms to which it is attached together form a 5-7 membered aliphatic ring, and m is 0.

[0015] In some embodiments, more than one H in the compound is replaced by D, and optionally, a G chain group or R 1 One or more H in is replaced by D.

[0016] Representative examples of the novel compounds containing oxopyridoaliphatic heterocyclic structures provided are

[0017] Studies have shown that the above-mentioned compounds containing oxopyridoaliphatic heterocyclic or aliphatic ring structures have excellent medicinal properties and can effectively inhibit the production of cAMP stimulated by CGRP, thereby preventing, alleviating or treating CGRP-related diseases, especially migraine or neuropathic pain, as CGRP antagonists. Specifically, the novel compounds provided by the present invention have a low IC 50 values, especially some specific structural compounds with common characteristics have significantly lower IC 50 Other experimental data show that the compounds of the present invention also have good liver mitochondrial stability, are expected to have potentially lower side effects, and have better PK properties.

[0018] A method for synthesizing such oxo-pyridine-containing heterocyclic or aliphatic ring structure compounds has been provided in the previous related application CN202310479996.8. The purpose of the present invention is to provide a new synthetic route and method for the synthesis of such oxo-pyridine-containing heterocyclic or aliphatic ring structure compounds. The provided new method has advantages in shortening the reaction route, improving the yield or being more suitable for industrial applications.

[0019] References:

[0020] [1]Ashina M, Terwindt GM, Al-Karagholi MA, et al. Migraine: disease characterisation, biomarkers, and precision medicine. Lancet.2021;397(10283):1496-1504.doi:10.1016 / S0140-6736(20)32162-0

[0021] [2] Chinese Medical Association Neurology Branch, Chinese Society of Research Hospitals Headache and Sensory Disorders Professional Committee. Chinese Guidelines for the Diagnosis and Treatment of Migraine (2022 Edition)[J]. Chinese Journal of Pain Medicine, 2022, 28(12): 881-898.

[0022] [3]Ashina M,Katsarava Z,Do TP,et al.Migraine:epidemiology and systems of care.Lancet.2021;397(10283):1485-1495.doi:10.1016 / S0140-6736(20)32160-7

[0023] [4]Russo AF.Calcitonin gene-related peptide(CGRP):a new target formigraine.Annu Rev Pharmacol Toxicol.2015;55:533-552.doi:10.1146 / annurev-pharmtox-010814-124701

[0024] [5]Iyengar S, Ossipov MH, Johnson KW. The role of calcitonin gene-related peptide in peripheral and central pain mechanisms including migraine. Pain. 2017; 158(4): 543-559. doi: 10.1097 / j.pain.0000000000000831

[0025] [6]Hargreaves R,Olesen J.Calcitonin Gene-Related Peptide Modulators-The History and Renaissance of a New Migraine Drug Class.Headache.2019;59(6):951-970.doi:10.1111 / head.13510

[0026] [7] Russell FA, King R, Smillie SJ, Kodji

[0027] [8]Clemow DB,Johnson KW,Hochstetler HM,Ossipov MH,Hake AM,BlumenfeldAM.Lasmiditan mechanism of action-review of a selective 5-HT1F agonist.JHeadache Pain.2020;21(1):71.Published 2020Jun 10.doi:10.1186 / s10194-020-01132-3. Summary of the invention

[0028] The object of the present invention is to provide a new synthetic route and method for the synthesis of such oxopyridine-containing heterocyclic or aliphatic ring structure compounds, and the provided new method has advantages in shortening the reaction route, improving the yield or being more suitable for industrial application.

[0029] Based on the new route synthesis method, the present invention first provides the following novel important intermediates, which are the key parts for synthesizing the target active products.

[0030] The first type of novel important intermediate compound is shown in the following formula I:

[0031]

[0032] In Formula I:

[0033] R ais -H, -(C1-C6)alkyl, and may also be -(C1-C3)alkyl, preferably methyl or ethyl;

[0034] R b is -H or an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, and the amino protecting group is preferably -Boc;

[0035] The G chain group and the two carbon atoms to which it is attached together form a 5- to 7-membered aliphatic ring or aliphatic heterocyclic ring, wherein the heteroatoms constituting the ring in the aliphatic heterocyclic ring are -O- or -S-;

[0036] R 1 is independently at each occurrence -H, -(C1-C6)alkyl or -(C3-C8)cycloalkyl, m is 0, 1, 2, 3, 4, 5 or 6,

[0037] Or, -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

[0038] According to a specific embodiment of the present invention, in some schemes, if the G chain group and the two carbon atoms to which it is attached together form a 5- or 6-membered aliphatic ring, the G chain group may optionally have 0 or 1 double bonds, and the rest are single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 7-membered aliphatic ring, the G chain group may optionally have 0, 1 or 2 double bonds, and the rest are single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 5-membered aliphatic heterocyclic ring, the G chain group may have single bonds; if the G chain group and the two carbon atoms to which it is attached together form a 6- or 7-membered aliphatic heterocyclic ring, the G chain group may optionally have 0 or 1 double bonds, and the rest are single bonds; R 1 is independently -H, -(C1-C3)alkyl or -(C3-C6)cycloalkyl at each occurrence, m is 0, 1, 2 or 3, or, -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

[0039] According to a specific embodiment of the present invention, in some schemes, R 1is independently -H, methyl, ethyl or cyclopropane at each occurrence, m is 0, 1 or 2, or, -(R 1 ) m The atoms to which it is attached together form cyclopropane, cyclobutane or cyclopentane, and the cyclopropane, cyclobutane or cyclopentane and -(R 1 ) m The attached rings constitute a spiro ring structure, a fused ring structure or a bridged ring structure.

[0040] According to a specific embodiment of the present invention, in some schemes, the intermediate compound is represented by the following formula I-1, I-2, I-3, I-4 or I-5:

[0041]

[0042] In formula I-1, I-2, I-3, I-4 or I-5, X 6 is -O- or -S-;

[0043] In formula Ⅰ-2, two "---"s both represent single bonds, or any one of them is a double bond and the other is a single bond;

[0044] In formula Ⅰ-3, three "---" all represent single bonds, or any one of them is a double bond and the other two are single bonds;

[0045] In formula Ⅰ-4, "---" refers to a single bond or a double bond;

[0046] In formula Ⅰ-5, two "---"s both represent single bonds, or any one of them is a double bond and the other is a single bond.

[0047] According to a specific embodiment of the present invention, in some schemes, the G chain group and the two carbon atoms to which it is attached together form a 5-7 membered aliphatic ring, and m is 0.

[0048] According to a specific embodiment of the present invention, in some embodiments, more than one H in the intermediate compound is replaced by D, and optionally, the G chain group or R 1 One or more H in is replaced by D.

[0049] Important intermediate compounds of this first type may be selected from:

[0050]

[0051]

[0052] The present invention also provides the second and third important intermediate compounds, as shown in the following formula II or III:

[0053]

[0054] R a , G chain group, R 1 and m are independently defined at each occurrence as in any place in the intermediate compound of formula I above; in formula II, when R a When is ethyl, the intermediate compound shown in formula II is not In formula III, when R a When is methyl, the intermediate compound shown in formula III is not

[0055] The important intermediate compounds of the second and third categories can be selected from:

[0056]

[0057]

[0058] In addition to the above first, second and third types of intermediate compounds, the present invention also provides the following important intermediate compounds: R b is -H or an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, and the amino protecting group is preferably -Boc.

[0059] According to the specific use and storage requirements, the important intermediate compound provided by the present invention may exist in the form of a suitable salt, which is also within the protection scope of the present invention, such as hydrochloride or trifluoroacetate.

[0060] In the implementation of the specific preparation examples and embodiments of the present invention, the obtained intermediate compounds were subjected to MS and 1 To avoid redundant description, the following only provides specific examples of some intermediate compounds. 1 H NMR characterization information:

[0061]

[0062]

[0063] In addition to the above corresponding intermediate compounds 1 The intermediate compound of the present invention can also be directly characterized by the MS provided and the final active compound product synthesized by the intermediate compound. 1 HNMR was combined to confirm.

[0064] On the basis of the various novel important intermediate compounds provided above, the present invention also provides a novel synthesis method for synthesizing compounds containing oxopyridoaliphatic heterocyclic or aliphatic ring structures or the important intermediate compounds required therefor, which comprises at least one of the following synthesis routes (1) to (5):

[0065] (1)

[0066] (2)

[0067] (3)

[0068] (4)

[0069] (5)

[0070] in:

[0071] R a Each occurrence is independently -(C1-C6)alkyl, and may optionally be -(C1-C3)alkyl, preferably methyl or ethyl;

[0072] R b Each occurrence is independently an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, the amino protecting group is preferably -Boc;

[0073] G chain group, R 1 and m are independently defined at each occurrence as above in any of the intermediate compounds provided by the present invention.

[0074] According to a specific embodiment of the present invention, in some schemes, the new synthesis preparation method provided by the present invention may further include the following synthesis route:

[0075]

[0076] in:

[0077] R b Each occurrence is independently an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, the amino protecting group is preferably -Boc;

[0078] G chain group, R 1 and m are independently defined at each occurrence as above in any of the intermediate compounds provided by the present invention.

[0079] According to a specific embodiment of the present invention, the present invention also provides a method for synthesizing representative exemplary compounds, including the following synthetic routes:

[0080]

[0081]

[0082] According to a specific embodiment of the present invention, the synthesis method of the representative exemplary compound may further include the following synthesis route:

[0083] DETAILED DESCRIPTION

[0084] Unless stated to the contrary or otherwise defined, the following terms used in the specification and claims have the following meanings.

[0085] "Alkyl" refers to an aliphatic hydrocarbon group, which refers to a saturated hydrocarbon group. The alkyl group may be a straight chain alkyl group or a branched chain alkyl group. For example, -(C1-C6)alkyl or -(C1-C3)alkyl. -(C1-C6)alkyl refers to an alkyl group having 1 to 6 carbon atoms, such as an alkyl group having 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms. Non-limiting examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, neopentyl, n-hexyl, and the like.

[0086] "Ring" refers to any covalently closed structure, including, for example, carbocyclic rings (e.g., aryl, cycloalkyl), heterocyclic groups (e.g., heteroaryl, heterocycloalkyl), aromatic groups (e.g., aryl, heteroaryl), non-aromatic groups (e.g., cycloalkyl, heterocycloalkyl, etc.). The ring can be a monocyclic ring or a polycyclic ring. Typical polycyclic rings generally include bicyclic rings and tricyclic rings. The ring of the present invention usually has 3-20 ring atoms, for example, 3 ring atoms, 4 ring atoms, 5 ring atoms, 6 ring atoms, 7 ring atoms, 8 ring atoms, 9 ring atoms, 10 ring atoms, 11 ring atoms, 12 ring atoms, 13 ring atoms, 14 ring atoms, 15 ring atoms, 16 ring atoms, 17 ring atoms, 18 ring atoms, 19 ring atoms or 20 ring atoms.

[0087] "Aliphatic ring" is a saturated or unsaturated cyclic hydrocarbon without aromatic properties, and the skeleton atoms constituting the ring are all carbon atoms. In the present invention, the aliphatic ring can usually have 3 to 8, 3 to 6 or 5 to 7 ring carbon atoms, and can also be called a 3 to 8-membered aliphatic ring, a 3-6-membered aliphatic ring or a 5-7-membered aliphatic ring. Typical examples of aliphatic rings include cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane, cyclooctane, cyclopropene, cyclobutene, cyclopentene, cyclohexene, cycloheptene, etc.

[0088] "Cycloalkyl" refers to a saturated cyclic hydrocarbon substituent, preferably cyclopropane, cyclobutane, cyclopentane, cyclohexane, cycloheptane or cyclooctane. The cycloalkyl group may be substituted or unsubstituted.

[0089] “Aliphatic heterocycle” is also referred to as “aliphatic heterocycle” in the present invention. It is a heterocycle without aromatic properties formed by replacing one or more of the carbon atoms in the skeleton of the “aliphatic ring” with heteroatoms. The replaced heteroatoms can usually be selected from O, S, N, etc.

[0090] "Member" refers to the number of skeleton atoms constituting the ring. Typical 5-membered rings include, for example, cyclopentyl, pyrrole, imidazole, thiazole, furan, and thiophene; typical 6-membered rings include, for example, cyclohexane, pyridine, pyran, pyrazine, thiopyran, pyridazine, pyrimidine, benzene, etc. Among them, a ring containing heteroatoms in the skeleton atoms is a heterocycle.

[0091] "Spirocyclic ring" or "spirocyclic structure" refers to a structure formed by two rings sharing a backbone ring atom, such as wait.

[0092] "Bridged ring" or "bridged ring structure" refers to a structure formed by two rings sharing two backbone ring atoms, and the two shared backbone ring atoms are not adjacent, for example wait.

[0093] "Both rings" or "both ring structures" refer to structures formed by two rings sharing two adjacent skeleton ring atoms, for example wait.

[0094] "Oxo" refers to the replacement of a hydrogen on a carbon with =0.

[0095] "Substituted" means that one or more hydrogen atoms, preferably up to 5 (e.g. 1, 2, 3, 4, 5), more preferably 1 to 3 hydrogen atoms in the group can be replaced independently of each other by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art can determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom with an unsaturated bond.

[0096] "Antagonist" refers to a drug that has no intrinsic activity but can block the effects mediated by receptor agonists after binding to the receptor.

[0097] "Optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes that the event or circumstance occurs or does not occur.

[0098] The term "substituted or unsubstituted" herein refers to any group being monosubstituted or polysubstituted by a specified substituent to the extent that such monosubstituted or polysubstituted (including multiple substitutions in the same moiety) is chemically permitted, and each substituent can be located at any available position on the group and can be connected through any available atom on the substituent. "Any available position" refers to any position on the group that is chemically accessible by methods known in the art or methods taught herein and does not produce excessively unstable molecules. When there are two or more substituents on any group, each substituent is defined independently of any other substituent and can therefore be the same or different.

[0099] In the chemical structure of this article It means that the structure here has a chirality, but the chirality has not been split in the current compound, which means that the current compound contains different chirality structures.

[0100] In the chemical structure of this article It means that the double bond structure here contains cis and trans configurations, but the cis and trans separation is not done in the current compound, that is, the meaning of the current compound contains different cis and trans configurations.

[0101] It should be understood that when selecting the various groups in the compound structure of the present invention, the groups that are connected, coordinated or influenced by each other should be selected accordingly under the premise of complying with the chemical valence bond rules.

[0102] According to the contents of the present invention, in accordance with common technical knowledge and means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0103] In order to further illustrate the present invention, the active compound provided by the present invention and its preparation method and application are described in detail below in conjunction with examples.

[0104] The following abbreviations or terms have the following meanings:

[0105] K2CO3 represents potassium carbonate; H2O represents water;

[0106] DMF stands for N,N-dimethylformamide; KF stands for potassium fluoride;

[0107] DIPEA or DIEA represents N,N-diisopropylethylamine; n-BuLi represents n-butyllithium;

[0108] THF stands for tetrahydrofuran; TFA stands for trifluoroacetic acid;

[0109] HCl / Dioxane means hydrogen chloride dioxane solution; HCl means hydrogen chloride solution;

[0110] DMSO means dimethyl sulfoxide; CH3I or MeI means iodomethane;

[0111] H2 represents hydrogen; Pd2(dba)3 represents tris(dibenzylideneacetone)dipalladium;

[0112] t-BuOK represents potassium tert-butoxide; NMP represents N-methylpyrrolidone;

[0113] MeOH means methanol; EtOH means ethanol;

[0114] Pd(PPh3)2Cl2 represents bistriphenylphosphine palladium dichloride; DIBAL-H represents diisobutylaluminum hydride;

[0115] TEA represents triethylamine; DCM represents dichloromethane;

[0116] DCE means 1,2-dichloroethane; dioxane means 1,4-dioxane;

[0117] NBS represents N-bromosuccinimide; NaH represents sodium hydride;

[0118] LiHMDS represents lithium bis(trimethylsilyl)amide; NaBH4 represents sodium borohydride;

[0119] Pd(dppf)Cl2 represents [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride; Zn represents zinc;

[0120] Ruphos represents 2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1'-biphenyl; I2 represents iodine;

[0121] NaSH represents sodium hydrosulfide; CuI represents cuprous iodide;

[0122] Pd / C represents palladium on carbon; AcSH represents thioacetic acid;

[0123] DIAD represents diisopropyl azodicarboxylate; PPh3 represents triphenylphosphine;

[0124] CH3CN or MeCN represents acetonitrile; PtO2 represents platinum dioxide;

[0125] NaOMe represents sodium methoxide; KHMDS represents potassium bis(trimethylsilyl)amide;

[0126] DEAD stands for diethyl azodicarboxylate; MWI stands for microwave-induced reaction;

[0127] AcCl represents acetyl chloride; NaOH represents sodium hydroxide;

[0128] TBTU stands for 2-(1H-benzotriazol L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate;

[0129] DMAP represents 4-dimethylaminopyridine; Ms2O represents methanesulfonic anhydride;

[0130] LiOH represents lithium hydroxide; B(OCH3)3 represents trimethyl borate;

[0131] Cs2CO3 represents cesium carbonate; TCDI represents N,N'-thiocarbonyldiimidazole;

[0132] TosCl stands for p-toluenesulfonyl chloride; DSC stands for N,N'-disuccinimidyl carbonate.

[0133] LDA represents lithium diisopropylamide; XantPhos represents 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene; LAH stands for lithium aluminum tetrahydride; DPPA stands for diphenylphosphoryl azide;

[0134] DMC stands for dimethyl carbonate; TMSCN stands for trimethylsilyl cyanide;

[0135] s-BuLi represents sec-butyllithium; PyBOP represents 1H-benzotriazol-1-yloxytripyrrolidino hexafluorophosphate;

[0136] DBU represents 1,8-diazabicyclo[5.4.0]undec-7-ene; DMSu represents dimethyl succinate;

[0137] Pd(dppf)Cl2·DCM represents [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride dichloromethane complex.

[0138] Intermediate Preparation Example 1:

[0139]

[0140] Step 1: Preparation of (S)-(2-((tert-butoxycarbonyl)amino)-3-methoxy-3-oxypropyl)zinc(II) iodide

[0141]

[0142] In a nitrogen atmosphere, add the activated zinc powder (2.37 g) to a 100 mL three-necked flask, dissolve iodine (926.37 mg) in anhydrous N,N-dimethylformamide, dropwise add to the three-necked flask, and then place the system at 60°C for 15 minutes. Dissolve (S)-2-((tert-butoxycarbonyl)amino)-3-iodopropionic acid methyl ester (6.00 g) in anhydrous N,N-dimethylformamide (35 mL), slowly dropwise add to the above reaction system, and react at 70°C for 2.5 hours. After the reaction is completed, move the reaction system to room temperature, and the resulting reaction solution is directly used for the next step.

[0143] Step 2: Preparation of methyl (R)-2-((tert-butoxycarbonyl)amino)-3-(7-methyl-1H-indazol-5-yl)propanoate

[0144]

[0145] In a nitrogen atmosphere, 5-bromo-7-methyl-1H-indazole (2.26 g), tris(dibenzylideneacetone)dipalladium (489.52 mg) and 2-dicyclohexylphosphine-2',6'-diisopropoxy-1,1'-biphenyl (499.29 mg) were dissolved in anhydrous N,N-dimethylformamide (5 mL), and the solution obtained in the first step was added to the reaction system, and heated at 70°C for 3 hours. After the reaction was completed, the reaction system was filtered through diatomaceous earth, the filtrate was collected, 30 mL of water was added, and ethyl acetate was extracted. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated and purified by silica gel column chromatography to obtain 2.1 g of the title compound.

[0146] MS (ESI) m / z (M+H) + =334.1.

[0147] Step 3: Preparation of (R)-2-amino-3-(7-methyl-1H-indazol-5-yl)propionic acid methyl ester trifluoroacetate

[0148]

[0149] Methyl (R)-2-((tert-butoxycarbonyl)amino)-3-(7-methyl-1H-indazol-5-yl)propanoate (1 g) was dissolved in dichloromethane (10 mL) and trifluoroacetic acid (10 mL) and reacted at room temperature for 1 hour. After the reaction, the reaction system was concentrated to obtain a crude product, which was directly used in the next step without purification.

[0150] MS (ESI) m / z (M+H) + =234.1.

[0151] Step 4: Preparation of methyl (R)-3-(7-methyl-1H-indazol-5-yl)-2-((phenoxycarbonyl)amino)propanoate

[0152]

[0153] In an ice-water bath, (R)-2-amino-3-(7-methyl-1H-indazol-5-yl)propionic acid methyl ester trifluoroacetate (990 mg) was dissolved in dichloromethane (20 mL), and N,N-diisopropylethylamine (1.16 g) was added dropwise. Then, phenyl chloroformate (469.71 mg) was dissolved in dichloromethane (10 mL), and added dropwise to the above reaction system to react for 1 hour. After the reaction, the reaction solution was poured into water, extracted with ethyl acetate, the organic phases were combined, washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was separated and purified by silica gel column chromatography to obtain 1 g of the title compound. MS (ESI) m / z (M+H) + =354.1.

[0154] Intermediate Preparation Example 2:

[0155]

[0156] In a nitrogen atmosphere, 1-Boc-4-ethynylpiperidine (5.0 g) and bis(cyclopentadienyl)zirconium chloride hydride (307.32 mg) were dissolved in toluene (50 mL), 4,4,5,5-tetramethyl-1,3,2-dioxaborane (3.67 g) was added dropwise, and the mixture was heated at 60°C for overnight reaction. After the reaction was completed, the reaction solution was concentrated, and the crude product was purified by column chromatography to remove another configuration, to obtain 5.3 g of the title compound.

[0157] Intermediate Preparation Example 3:

[0158]

[0159] In a nitrogen atmosphere, 1-Boc-4-ethynylpiperidine (3.0 g) and catechol borane (1.72 g) were dissolved in tetrahydrofuran (30 mL) and heated at 60° C. for 24 h. After the reaction was completed, the reaction solution was concentrated and the crude product was purified by column chromatography to obtain 0.7 g of the title compound.

[0160] Intermediate Preparation Example 4: Preparation of 3-(piperidin-4-yl)-5,8-dihydro-1H-thiopyrano[3,4-b]pyridin-2(6H)-one hydrochloride (can be used in Example 3)

[0161]

[0162] Step 1: Preparation of methyl 4-oxotetrahydro-2H-thiopyran-3-carboxylate

[0163]

[0164] In a nitrogen atmosphere, under ice-water bath conditions, slowly dropwise add a solution of 3,3'-dimethylthiodipropionate (5.0 g) in anhydrous tetrahydrofuran (30 mL) to a suspension of sodium hydride (1.45 g) in anhydrous tetrahydrofuran (15 mL), and move to room temperature to react overnight. LCMS shows that the raw material has reacted completely, and an appropriate amount of saturated ammonium chloride solution is added to quench the reaction, and ethyl acetate is extracted 3 times, the organic phases are combined, dried over anhydrous sodium sulfate, filtered, and concentrated, and the resulting crude product is purified by column chromatography to obtain 3.65 g of the title compound.

[0165] MS (ESI) m / z (M+H) + =175.0.

[0166] Step 2: Preparation of methyl 4-(((trifluoromethyl)sulfonyl)oxy)-5,6-dihydro-2H-thiopyran-3-carboxylate

[0167]

[0168] In a nitrogen atmosphere, under ice-water bath conditions, slowly drop a solution of 4-oxotetrahydro-2H-thiopyran-3-carboxylic acid methyl ester (3.65g) in anhydrous tetrahydrofuran (30mL) to a suspension of sodium hydride (1.09g) in anhydrous tetrahydrofuran (10mL), and move to room temperature to react for 1 hour. Add a solution of 1,1,1-trifluoro-N-phenyl-N-(trifluoromethyl)sulfonyl)methanesulfonamide (8.23g) in anhydrous tetrahydrofuran (20mL), and react at room temperature overnight. LCMS shows that the raw material reacts completely, and an appropriate amount of saturated ammonium chloride solution is added to quench the reaction, and the mixture is extracted with ethyl acetate 3 times. The organic phases are combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting crude product is purified by column chromatography to obtain 5.2g of the title compound.

[0169] MS (ESI) m / z (M+H) + =306.9.

[0170] Step 3: Preparation of tert-butyl (E)-4-(2-(5-(methoxycarbonyl)-3,6-dihydro-2H-thiopyran-4-yl)vinyl)piperidine-1-carboxylate

[0171]

[0172] In a nitrogen atmosphere, tert-butyl 4-ethynylpiperidine-1-carboxylate (1.0 g) was dissolved in anhydrous tetrahydrofuran (4 mL), and a solution of catechol borane (0.69 g) in anhydrous tetrahydrofuran (4 mL) was slowly added, and the mixture was reacted at 70°C for 4 hours, and then cooled to room temperature. Methyl 4-(((trifluoromethyl)sulfonyl)oxy)-5,6-dihydro-2H-thiopyran-3-carboxylate (1.5 g), 1,1'-bis(diphenylphosphino)ferrocenepalladium dichloride (II) (183 mg) and potassium carbonate (2.03 g) were dissolved in dioxane (10 mL) and water (1 mL), and slowly added dropwise to the above reaction solution, and the temperature was raised to 70°C and the reaction was continued for 1 hour. LCMS showed that the raw material was completely reacted. The reaction solution was filtered, the filtrate was collected, and concentrated, and the crude product was purified by column chromatography to obtain 1.3 g of the title compound.

[0173] MS (ESI) m / z (M+H-100) + =267.9.

[0174] Step 4: Preparation of (E)-4-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)-5,6-dihydro-2H-thiopyran-3-carboxylic acid

[0175]

[0176] Dissolve (E)-tert-butyl 4-(2-(5-(methoxycarbonyl)-3,6-dihydro-2H-thiopyran-4-yl)vinyl)piperidine-1-carboxylate (230 mg) in methanol (2 mL) and water (0.5 mL), add lithium hydroxide monohydrate (131.4 mg), and react overnight. LCMS shows that the reaction is complete. The reaction solution is neutralized with 2M hydrochloric acid, extracted with ethyl acetate three times, the organic phases are combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting crude product is purified by column chromatography to obtain 200 mg of the title compound.

[0177] MS (ESI) m / z (M+H-100) + =253.9.

[0178] Step 5: Preparation of tert-butyl 4-(2-oxo-2,5,6,8-tetrahydro-1H-thiopyrano[3,4-b]pyridin-3-yl)piperidine-1-carboxylate

[0179]

[0180] In a nitrogen atmosphere, (E)-4-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)-5,6-dihydro-2H-thiopyran-3-carboxylic acid (200 mg) and diphenylphosphoryl azide (155.7 mg) were dissolved in anhydrous toluene (3 mL), triethylamine (85.9 mg) was added, and the mixture was reacted at room temperature for 2 hours. The temperature was raised to 80°C and the reaction was continued for 1 hour. LCMS showed that the raw materials were completely reacted. The reaction solution was concentrated, and the crude product was purified by column chromatography to obtain 100 mg of the title compound.

[0181] MS (ESI) m / z (M+H-100) + =250.9.

[0182] Step 6: Preparation of 3-(piperidin-4-yl)-5,8-dihydro-1H-thiopyrano[3,4-b]pyridin-2(6H)-one hydrochloride

[0183]

[0184] Dissolve tert-butyl 4-(2-oxo-2,5,6,8-tetrahydro-1H-thiopyrano[3,4-b]pyridin-3-yl)piperidine-1-carboxylate (100 mg) in dichloromethane (2 mL), add hydrochloric acid / 1,4-dioxane solution (4M, 1 mL), and react at room temperature for 1 hour. LCMS showed that the raw material was completely reacted. The reaction solution was concentrated to obtain 80 mg of crude product, which was directly used in the next step.

[0185] MS (ESI) m / z (M+H) + =250.9.

[0186] Intermediate Preparation Example 5: Preparation of 3-(piperidin-4-yl)-5,6,7,8-tetrahydro-5,8-methylquinolin-2(1H)-one hydrochloride (can be used in Example 5)

[0187]

[0188] Step 1: Preparation of methyl 3-oxobicyclo[2.2.1]heptane-2-carboxylate

[0189]

[0190] In a nitrogen atmosphere, sodium hydride (726 mg) was dissolved in dimethyl carbonate (9 mL), and then bicyclo[2.2.1]heptane-2-one (1 g) was dissolved in dimethyl carbonate (6 mL), and then slowly added dropwise to the sodium hydride system, and reacted at 90°C for 2 hours. After the reaction, the reaction solution was quenched with water, extracted with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was separated and purified by column chromatography to obtain 1.10 g of the title compound.

[0191] MS (ESI) m / z (M+H) + =169.1.

[0192] Step 2: Preparation of methyl 3-((trifluoromethyl)sulfonyl)oxy)bicyclo[2.2.1]hept-2-ene-2-carboxylate

[0193]

[0194] Under ice-water bath conditions, sodium hydride (385 mg) was dissolved in anhydrous tetrahydrofuran (10 mL), and then 3-oxobicyclo[2.2.1]heptane-2-carboxylic acid methyl ester (1.08 g) was dissolved in anhydrous tetrahydrofuran (5 mL), and slowly added dropwise to the sodium hydride system, and stirred for 30 minutes. N-phenylbis(trifluoromethanesulfonyl)imide (2.98 g) was then dissolved in anhydrous tetrahydrofuran (10 mL), and slowly added dropwise to the above system, and reacted at room temperature overnight. After the reaction was completed, water was added to quench the reaction, and ethyl acetate was extracted, the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was separated and purified by column chromatography to obtain 1.70 g of the title compound.

[0195] MS (ESI) m / z (M+H) + =301.1.

[0196] Step 3: Preparation of tert-butyl (E)-4-(2-(3-(methoxycarbonyl)bicyclo[2.2.1]hept-2-en-2-yl)vinyl)piperidine-1-carboxylate

[0197]

[0198] In a nitrogen atmosphere, methyl 3-((trifluoromethyl)sulfonyl)oxy)bicyclo[2.2.1]hept-2-ene-2-carboxylate (600 mg), tert-butyl (E)-4-(2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)vinyl)piperidine-1-carboxylate (742 mg), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (163 mg) and potassium carbonate (690 mg) were dissolved in 1,4-dioxane (10 mL) and water (2 mL) and reacted at 80°C for 2 hours. After the reaction was completed, water was added to quench the reaction, the filtrate was collected by suction, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was separated and purified by column chromatography to obtain 660.00 mg of the title compound. MS (ESI) m / z (M+H) + =362.2.

[0199] Step 4: Preparation of (E)-3-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)bicyclo[2.2.1]hept-2-ene-2-carboxylic acid

[0200]

[0201] Dissolve (E)-tert-butyl 4-(2-(3-(methoxycarbonyl)bicyclo[2.2.1]hept-2-en-2-yl)vinyl)piperidine-1-carboxylate (200 mg) in methanol (10 mL) and water (1 mL), add sodium hydroxide (155 mg), and react at 50° C. overnight. After the reaction, adjust the pH of the reaction system to 5-6 with 1N hydrochloric acid, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, filter, and concentrate to obtain 180 mg of a crude product.

[0202] MS (ESI) m / z (M+H) + =348.2.

[0203] Step 5: Preparation of tert-butyl 4-(2-oxo-1,2,5,6,7,8-hexahydro-5,8-methylquinolin-3-yl)piperidine-1-carboxylate

[0204]

[0205] In a nitrogen atmosphere, (E)-3-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)bicyclo[2.2.1]hept-2-ene-2-carboxylic acid (180 mg) was dissolved in toluene (2 mL), and diphenylphosphoryl azide (154 mg) and triethylamine (78 mg) were added, and the mixture was reacted at room temperature for 3 hours. After the reaction was complete, the system was pre-purified by preparative thin layer chromatography, dissolved in toluene (2 mL), and the reaction was continued at 90°C for 3 hours. After the reaction was completed, the mixture was concentrated, and the crude product was separated and purified by preparative thin layer chromatography to obtain 80.00 mg of the title compound.

[0206] MS (ESI) m / z (M+H) + =345.2.

[0207] Step 6: Preparation of 3-(piperidin-4-yl)-5,6,7,8-tetrahydro-5,8-methylquinolin-2(1H)-one hydrochloride

[0208]

[0209] Dissolve tert-butyl 4-(2-oxo-1,2,5,6,7,8-hexahydro-5,8-methylquinolin-3-yl)piperidine-1-carboxylate (80 mg) in hydrogen chloride / 1,4-dioxane solution (4M, 1 mL) and react at room temperature for 1 hour. After the reaction is complete, concentrate to obtain 78 mg of a crude product.

[0210] MS (ESI) m / z (M+H) + =245.1

[0211] Intermediate Preparation Example 6: Preparation of 6'-(piperidin-4-yl)-4',8'-dihydro-2'H,7'H-spiro[cyclopropane-1,3'-pyranylthio[2,3-b]pyridine]-7'-one hydrochloride (can be used in Example 6)

[0212]

[0213] Step 1: Preparation of methyl 2-(((1-(2-methoxy-2-oxoethyl)cyclopropyl)methyl)thio)acetate

[0214]

[0215] Under ice-water bath conditions, methyl 2-(1-(mercaptomethyl)cyclopropyl)acetate (600 mg) was dissolved in methanol (12 mL), sodium methoxide (750 mg) was added, and after stirring for 10 minutes, ethyl bromoacetate (750 mg) was added and reacted for 2 hours. LCMS showed that the reaction was complete. Hydrochloric acid solution (2N) was added to adjust the pH to 7-8, and ethyl acetate was extracted, the organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by column chromatography to obtain 650 mg of the title compound. MS (ESI) m / z (M+H) + = 233.1.

[0216] Step 2: Preparation of methyl 7-oxo-5-thiospiro[2.5]octane-6-carboxylate

[0217]

[0218] Under ice-water bath conditions, methyl 2-(((1-(2-methoxy-2-oxoethyl)cyclopropyl)methyl)thio)acetate (630 mg) was dissolved in tetrahydrofuran (10 mL), potassium tert-butoxide (608 mg) was added in batches, and the reaction was continued for 1 hour. LCMS showed that the reaction was complete. Hydrochloric acid solution (2N) was added to adjust the pH to 5-6, and the mixture was extracted with ethyl acetate. The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by column chromatography to obtain 380 mg of the target compound.

[0219] MS (ESI) m / z (M+H) + = 201.1.

[0220] The title compound was prepared by using the corresponding common commercially available reagents and subsequently using a preparation method similar to the above intermediate preparation example 5.

[0221] MS (ESI) m / z (M+H) + =277.1.

[0222] Intermediate Preparation Example 7: Preparation of 6-(piperidin-4-yl)-1a,2,4,7b-tetrahydrocyclopropane[4,5]thiopyrano[2,3-b]pyridine-5(1H)-one hydrochloride (can be used in Example 7)

[0223]

[0224] Step 1: Preparation of 2-(methoxycarbonyl)cyclopropane-1-carboxylic acid

[0225]

[0226] Dissolve triethylamine (901.8 mg) in methanol (15 mL), add 3-oxabicyclo[3.1.0]hexane-2,4-dione (1 g) in batches, and react at room temperature for 1 hour. After the reaction is completed, add water to quench, remove excess methanol, adjust the system pH to 3-4 with 1N hydrochloric acid, extract with ethyl acetate, combine the organic phases, dry over anhydrous sodium sulfate, filter, and concentrate to obtain 1.15 g of crude product.

[0227] Step 2: Preparation of methyl 2-(hydroxymethyl)cyclopropane-1-carboxylate.

[0228]

[0229] Under ice-water bath conditions, 2-(methoxycarbonyl)cyclopropane-1-carboxylic acid (1.15 g) was dissolved in anhydrous tetrahydrofuran (2 mL), and borane tetrahydrofuran solution (1 M, 16 mL) was added dropwise, and the mixture was moved to room temperature for 3 hours. After the reaction was completed, an appropriate amount of methanol was added to quench the reaction, and the crude product was concentrated to obtain 750 mg of the title compound.

[0230] MS (ESI) m / z (M+H) + =131.1.

[0231] Step 3: Preparation of methyl 2-(((methylsulfonyl)oxy)methyl)cyclopropane-1-carboxylate

[0232]

[0233] Under -10℃, methyl 2-(hydroxymethyl)cyclopropane-1-carboxylate (750mg) was dissolved in dichloromethane (10mL), and N,N-diisopropylethylamine (1.49g) was added. Methanesulfonyl chloride (856mg) was diluted with dichloromethane (3mL), added dropwise to the above reaction system, and returned to room temperature to react for 30 minutes. After the reaction was completed, water was added to quench, and ethyl acetate was used for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated and purified by column chromatography to obtain 1.1g of the title compound.

[0234] MS (ESI) m / z (M+H)+ =209.0.

[0235] Step 4: Preparation of methyl 2-(((2-ethoxy-2-oxoethyl)thio)methyl)cyclopropane-1-carboxylate

[0236]

[0237] Under ice-water bath conditions, in a nitrogen atmosphere, methyl 2-(((methylsulfonyl)oxy)methyl)cyclopropane-1-carboxylate (1.1 g) and ethyl 2-mercaptoacetate (936 mg) were dissolved in dry N,N-dimethylformamide (12 mL), sodium methoxide (421 mg) was added, and the mixture was returned to room temperature for 2 hours. After the reaction was completed, water was added to quench the mixture, and the mixture was extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated and purified by column chromatography to obtain 1.10 g of the title compound.

[0238] MS (ESI) m / z (M+H) + =233.0.

[0239] Step 5: Preparation of ethyl 5-oxo-3-thiabicyclo[4.1.0]heptane-4-carboxylate

[0240]

[0241] Under -10℃, in a nitrogen atmosphere, titanium tetrachloride (1.51g) was dissolved in dry dichloromethane (20mL), isopropanol (518mg) was added, and stirred for 30 minutes. Methyl 2-(((2-ethoxy-2-oxoethyl)thio)methyl)cyclopropane-1-carboxylate (1.54g) was dissolved in dry dichloromethane (5mL), and slowly added dropwise to the above reaction system, keeping the internal temperature at -10℃, and continued to stir for 30 minutes. Triethylamine (2.35g) was added, and the reaction was continued for 1.5 hours. After the reaction was completed, an appropriate amount of 1N hydrochloric acid was added to quench the reaction, and the dichloromethane was extracted. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated and purified by column chromatography to obtain 740mg of the title compound.

[0242] MS (ESI) m / z (M+H) + =201.0.

[0243] Step 6: Preparation of ethyl 5-(((trifluoromethyl)sulfonyl)oxy)-3-thiabicyclo[4.1.0]hept-4-ene-4-carboxylate

[0244]

[0245] Under -78°C, 5-oxo-3-thiabicyclo[4.1.0]heptane-4-carboxylic acid ethyl ester (570 mg) was dissolved in dry dichloromethane (15 mL), and N,N-diisopropylethylamine (1.1 g) was added. Trifluoromethanesulfonic anhydride (1.36 g) was diluted with dry dichloromethane (5 mL), added dropwise to the above system, and the reaction was continued for 30 minutes. After the reaction was completed, water was added to quench, and ethyl acetate was extracted. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated and purified by column chromatography to obtain 700 mg of the title compound.

[0246] MS (ESI) m / z (M+H) + =333.0.

[0247] The title compound was prepared by using the corresponding common commercially available reagents and subsequently using a preparation method similar to the above intermediate preparation example 5.

[0248] MS (ESI) m / z (M+H) + =263.1.

[0249] Intermediate Preparation Example 8: Preparation of 3-(piperidin-4-yl)-1,5,6,7-tetrahydro-2H-cyclopenta[b]pyridin-2-one hydrochloride (can be used in Example 9)

[0250]

[0251] The title compound was prepared by using the corresponding common commercially available reagents and subsequently using a preparation method similar to the above intermediate preparation example 5.

[0252] MS (ESI) m / z (M+H) + =219.1.

[0253] Preparation Example 9: Preparation of (R)-2-amino-3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)propan-1-one trifluoroacetate

[0254]

[0255] Step 1: Preparation of (R)-2-((tert-butoxycarbonyl)amino)-3-(7-methyl-1H-indazol-5-yl)propanoic acid

[0256]

[0257] Methyl (R)-2-((tert-butyloxycarbonyl)amino)-3-(7-methyl-1H-indazol-5-yl)propanoate (100 mg) was dissolved in tetrahydrofuran (3 mL) and water (1 mL), and sodium hydroxide (120 mg) was added to react at room temperature overnight. After the reaction, an appropriate amount of hydrochloric acid (0.5 N) was added to adjust the pH of the system to 5-6, and the mixture was extracted with ethyl acetate three times. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain 90.0 mg of a crude product.

[0258] MS (ESI) m / z (M+H) + =320.1.

[0259] Step 2: Preparation of tert-butyl (R)-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)carbamate

[0260]

[0261] (R)-2-((tert-butoxycarbonyl)amino)-3-(7-methyl-1H-indazol-5-yl)propionic acid (50 mg), 1-(1-methylpiperidin-4-yl)piperazine (57.18 mg), 2-(1H-benzotriazol L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (60.02 mg) and N,N-diisopropylethylamine (40.32 mg) were dissolved in acetonitrile (10 mL) and reacted at room temperature for 4 hours. After the reaction was completed, the crude product was concentrated and purified by column chromatography to obtain 30 mg of the title compound.

[0262] MS (ESI) m / z (M+H) + =485.2.

[0263] Step 3: Preparation of (R)-2-amino-3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)propan-1-one trifluoroacetate

[0264]

[0265] Tert-butyl (R)-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)carbamate (30 mg) was dissolved in dichloromethane (1 mL) and trifluoroacetic acid (0.5 mL) and reacted at room temperature for 1 h. After the reaction was completed, the crude product was concentrated and used in the next step without purification.

[0266] MS (ESI) m / z (M+H) + =385.2.

[0267] Example 1: Preparation of (R)-4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)piperidine-1-carboxamide

[0268] Step 1: Preparation of methyl 4-((2-methoxy-2-oxoethyl)thio)butyrate

[0269]

[0270] Under ice-water bath conditions, add sodium metal (1.16 g) to methanol (10 mL) and stir until the sodium crumbs disappear. Dissolve ethyl thioglycolate (6.04 g) in methanol (10 mL), slowly add dropwise to the above sodium methoxide solution, and stir for 0.5 h. Dissolve ethyl 4-bromobutyrate (9.8 g) in methanol (10 mL), add dropwise to the above solution, and move to room temperature to react overnight. After the reaction is completed, concentrate the reaction solution to remove the solvent, add water (100 mL) to dissolve the solid, adjust the pH to 6-7 with HCl (2N), extract with ethyl acetate (200 mL*3), combine the organic phases, wash three times with saturated NaCl aqueous solution, dry over anhydrous sodium sulfate, filter, and concentrate to obtain a crude product, which is purified by column chromatography to obtain 7.2 g of the title compound.

[0271] MS (ESI) m / z (M+H) + =207.0.

[0272] Step 2: Preparation of methyl 3-oxotetrahydro-2H-thiopyran-2-carboxylate

[0273]

[0274] In an ice-water bath, methyl 4-((2-methoxy-2-oxoethyl)thio)butanoate (7.20 g) was dissolved in tetrahydrofuran (72 mL), and potassium tert-butoxide (7.83 g) was added in batches and reacted for 1 h. After the reaction was completed, the mixture was moved to room temperature, HCl (2N) was added dropwise to adjust the pH to 5-6, and ethyl acetate (150 mL*3) was used for extraction. The organic phases were combined, washed three times with saturated aqueous NaCl solution (150 mL*2), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified by column chromatography to obtain 5.4 g of the title compound.

[0275] MS (ESI) m / z (M+H) + =175.0.

[0276] Step 3: Preparation of methyl 5-(((trifluoromethyl)sulfonyl)oxy)-3,4-dihydro-2H-thiopyran-6-carboxylate

[0277]

[0278] In a nitrogen atmosphere, under ice-water bath conditions, 3-oxotetrahydro-2H-thiopyran-2-carboxylic acid methyl ester (5.4 g) was dissolved in tetrahydrofuran (54 mL), sodium hydride (1.36 g) was added in batches, and stirred for 0.5 h. 1,1,1-trifluoro-N-phenyl-N-[(trifluoromethyl)sulfonyl]methanesulfonamide (13.29 g dissolved in 54 mL tetrahydrofuran) solution was slowly added to the above system, moved to room temperature, and reacted for 4 h until the solution was clear. The reaction solution was poured into a saturated aqueous ammonium chloride solution (100 mL), extracted with ethyl acetate (150 mL*3), the organic phases were combined, washed twice with a saturated aqueous NaCl solution, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified by column chromatography to obtain 7.50 g of the title compound. MS (ESI) m / z (M+H) + =306.9.

[0279] Step 4: Preparation of tert-butyl (E)-4-(2-(6-(methoxycarbonyl)-3,4-dihydro-2H-thiopyran-5-yl)vinyl)piperidine-1-carboxylate

[0280]

[0281] In a nitrogen atmosphere, 5-(((trifluoromethyl)sulfonyl)oxy)-3,4-dihydro-2H-thiopyran-6-carboxylic acid methyl ester (3.50 g), (E)-4-(2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)vinyl)piperidine-1-carboxylic acid tert-butyl ester (3.85 g), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (836.23 mg) and potassium carbonate (4.74 g) were dissolved in tetrahydrofuran (35 mL) and water (12 mL), and heated at 70°C for 1 h. After the reaction, the reaction solution was poured into 30 mL of water, extracted with ethyl acetate (50 mL*3), the organic phases were combined, washed three times with a saturated NaCl aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product, which was purified by column chromatography to obtain 3.7 g of the title compound.

[0282] MS (ESI) m / z (M+H) + =368.1.

[0283] Step 5: Preparation of (E)-5-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)-3,4-dihydro-2H-thiopyran-6-carboxylic acid

[0284]

[0285] Dissolve (E)-4-(2-(6-(methoxycarbonyl)-3,4-dihydro-2H-thiopyran-5-yl)vinyl)piperidine-1-carboxylic acid tert-butyl ester (3.7 g) in methanol (37 mL), add sodium hydroxide (1.21 g dissolved in 9 mL water) solution, and heat at 50°C for 2 h. After the reaction, adjust the pH to 5-6 with HCl (2N) solution, extract with ethyl acetate (50 mL*3), combine the organic phases, wash three times with saturated NaCl aqueous solution, dry over anhydrous sodium sulfate, filter, and concentrate to obtain 3.1 g of crude product.

[0286] MS (ESI) m / z (M+H) + =354.1.

[0287] Step 6: Preparation of tert-butyl 4-(7-oxo-3,4,7,8-tetrahydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxylate

[0288]

[0289] In a nitrogen atmosphere, (E)-5-(2-(1-(tert-butoxycarbonyl)piperidin-4-yl)vinyl)-3,4-dihydro-2H-thiopyran-6-carboxylic acid (3.1 g) and triethylamine (1.33 g) were dissolved in toluene (31 mL), diphenylphosphoryl azide (2.41 g) was added, and the mixture was reacted at 80°C for 2 h. After the reaction was completed, the mixture was filtered, washed with toluene 1 to 2 times, and the filter cake was collected to obtain 2 g of the title compound.

[0290] MS (ESI) m / z (M+H) + =351.1.

[0291] Step 7: Preparation of 6-(piperidin-4-yl)-2,3,4,8-tetrahydro-7H-thiopyrano[2,3-b]pyridin-7-one hydrochloride

[0292]

[0293] Dissolve tert-butyl 4-(7-oxo-3,4,7,8-tetrahydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxylate (500 mg) in dichloromethane (10 mL), add hydrochloric acid / dioxane solution (4N, 5 mL), and react at room temperature for 1 hour. After the reaction is completed, concentrate to obtain 350 mg of crude product.

[0294] MS (ESI) m / z (M+H) + =251.1.

[0295] Step 8: Preparation of (R)-2-(4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)-3-(7-methyl-1H-indazol-5-yl)propanoic acid methyl ester

[0296]

[0297] Dissolve 6-(piperidin-4-yl)-3,4-dihydro-2H-thiopyrano[2,3-b]pyridine-7-ol hydrochloride (40 mg), triethylamine (48.5 mg) and (R)-3-(7-methyl-1H-indazol-5-yl)-2-((phenoxycarbonyl)amino)propionic acid methyl ester (40 mg) in acetonitrile (3.0 mL) and react at 85°C for 1 hour. LCMS showed that the raw material was completely reacted. The reaction solution was directly concentrated. The crude product was purified by column chromatography to obtain 40 mg of the title compound.

[0298] MS (ESI) m / z (M+H) + =510.1.

[0299] Step 9: Preparation of (R)-2-(4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)-3-(7-methyl-1H-indazol-5-yl)propanoic acid

[0300]

[0301] (R)-2-(4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)-3-(7-methyl-1H-indazol-5-yl)propanoic acid methyl ester (40 mg) and lithium hydroxide monohydrate (16.4 mg) were dissolved in methanol (2.0 mL) and water (0.3 mL) and reacted at room temperature for 1 hour. LCMS showed that the reaction was complete. The reaction solution was concentrated to obtain 40 mg of the title compound.

[0302] MS (ESI) m / z (M+H) + =495.9.

[0303] Step 10: Preparation of (R)-4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)piperidine-1-carboxamide

[0304]

[0305] (R)-2-(4-(7-hydroxy-3,4-dihydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)-3-(7-methyl-1H-indazol-5-yl)propanoic acid (40 mg), 1-(1-methylpiperidin-4-yl)piperazine (17.8 mg), O-benzotriazole-N,N,N',N'-tetramethyluronium tetrafluoroborate (39 mg) and N,N-diisopropylethylamine (31 mg) were dissolved in N,N-dimethylformamide (1.0 mL) and reacted at room temperature for 1 hour. LCMS showed that the reaction was complete. The reaction solution was concentrated and purified by preparative liquid phase to obtain 4.61 mg of the title compound.

[0306] MS (ESI) m / z (M+H) + =660.8.

[0307] 1 H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.31(s,1H),7.96(s,1H),7.36(s,1H),7.00(s,1H),6.86(s,1H),6.64(d,J=8. 0,1H),4.80-4.72(m,1H),4.14-4.02(m,2H),3.57-3.53(m,1H),3.27-3.18(m,2H),3.14-3.08(m,1H),3.04-3.00(m,2H ),2.98-2.93(m,1H),2.92-2.86(m,1H),2.74-2.60(m,6H),2.56-2.54(m,1H),2.47(s,3H),2.37-2.31(m,1H),2.25-2. 19(m,1H),2.09(s,3H),2.01-1.89(m,4H),1.74-1.61(m,4H),1.55-1.49(m,1H),1.40-1.34(m,2H),1.30-1.13(m,4H).

[0308] Example 2: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)-4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamide

[0309]

[0310] Step 1: Preparation of methyl 3-((2-ethoxy-2-oxoethyl)thio)propanoate

[0311]

[0312] Dissolve methyl 3-mercaptopropionate (10.0 g) in N,N-dimethylformamide (50 mL), add sodium methoxide (5.9 g), and react at room temperature for 10 minutes. Add ethyl bromoacetate (13.9 g), and react at room temperature for 1 hour. TLC shows that the reaction is complete. Add dilute hydrochloric acid aqueous solution to the reaction system under ice-water bath conditions, keep the pH neutral, extract with ethyl acetate three times, combine the organic phases, backwash once with saturated sodium chloride solution, dry over anhydrous sodium sulfate, and concentrate. The resulting crude product is purified by column chromatography to obtain 15.5 g of the title compound.

[0313] MS (ESI) m / z (M+H) + =207.1.

[0314] Step 2: Preparation of ethyl 3-oxotetrahydrothiophene-2-carboxylate

[0315]

[0316] Titanium tetrachloride (17.7 g) was dissolved in dry dichloromethane (120 mL), and the temperature was lowered to -10 ° C after replacing nitrogen. Isopropanol (5.35 g) was added dropwise, and the reaction was continued for 30 minutes. Methyl 3-((2-ethoxy-2-oxyethyl)thio) propionate (16.0 g) was slowly added, and the reaction was continued for 30 minutes. Triethylamine (27.5 g) was slowly added, and the reaction was continued for 30 minutes. Dilute hydrochloric acid (2M, 93 mL) was added, and the mixture was moved to room temperature and reacted for 1 hour. TLC showed that the reaction was complete, and dichloromethane was extracted three times, the organic phases were combined, backwashed once with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by column chromatography to obtain 12.1 g of the title compound.

[0317] MS (ESI) m / z (M+H) + =175.0.

[0318] Step 3: Preparation of ethyl 3-((trifluoromethyl)sulfonyl)oxy)-4,5-dihydrothiophene-2-carboxylate

[0319]

[0320] At 0°C, add a solution of ethyl 3-oxotetrahydrothiophene-2-carboxylate (2.0 g) in tetrahydrofuran (20 mL) to a suspension of sodium hydride (549.0 mg) in tetrahydrofuran (20 mL). After addition, react at 0°C for 30 minutes. Add N-phenylbis(trifluoromethanesulfonyl)imide (4.5 g) to the system, naturally warm to room temperature and react for 3 hours. TLC shows that the reaction is complete. Add the system to 50 mL of water at 0°C, extract three times with ethyl acetate, combine the organic phases, backwash once with saturated sodium chloride solution, dry over anhydrous sodium sulfate, filter, and concentrate. The resulting crude product is purified by column chromatography to obtain 3.3 g of the title compound.

[0321] MS (ESI) m / z (M+H) + =307.0.

[0322] Step 4: Preparation of tert-butyl (E)-4-(2-(2-(ethoxycarbonyl)-4,5-dihydrothiophen-3-yl)vinyl)piperidine-1-carboxylate

[0323]

[0324] Ethyl 3-((trifluoromethyl)sulfonyl)oxy)-4,5-dihydrothiophene-2-carboxylate (260.0 mg), tert-butyl (E)-4-(2-(benzo[d][1,3,2]dioxanol-2-yl)vinyl)piperidine-1-carboxylate (280.0 mg) and potassium carbonate (235.0 mg) were dissolved in 1,4-dioxane (3 mL) and water (0.6 mL). After replacing the nitrogen, 1,1-bis(diphenylphosphino)diferronichloridopalladium (62.0 mg) was added. After the nitrogen was replaced again, the temperature was raised to 70°C and reacted for 2 hours. TLC showed that the reaction was complete. 30 mL of water was added to the reaction system, and ethyl acetate was extracted three times. The organic phases were combined, backwashed once with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated. The obtained crude product was purified by column chromatography to obtain 620.0 mg of the title compound.

[0325] MS (ESI) m / z (M+H) + =368.2.

[0326] Step 5: Preparation of tert-butyl (E)-4-(2-(benzo[d][1,3,2]dioxin-2-yl)vinyl)piperidine-1-carboxylate

[0327]

[0328] Dissolve (E)-tert-butyl 4-(2-(2-(ethoxycarbonyl)-4,5-dihydrothiophen-3-yl)vinyl)piperidine-1-carboxylate (230.0 mg) in methanol (6 mL), add a solution of sodium hydroxide (75.0 mg) in water (1 mL), and heat to 50°C for 2 hours. TLC shows that the reaction is complete, and the system is adjusted to pH 6 with 1M dilute hydrochloric acid and concentrated. The crude product is purified by column chromatography to obtain 150.0 mg of the title compound.

[0329] MS (ESI) m / z (M+H) + =340.2.

[0330] Step 6: Preparation of tert-butyl 4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxylate

[0331]

[0332] Dissolve (E)-4-(2-(Benzo[d][1,3,2]dioxanol-2-yl)vinyl)piperidine-1-carboxylic acid tert-butyl ester (50.0 mg) in toluene (1 mL), add triethylamine (45.0 mg) and diphenylphosphoryl azide (81.0 mg), and react at room temperature for 1 hour. After mass spectrometry detected that all the raw materials were converted into isocyanate, the system was heated to 60°C and reacted for 1 hour. TLC showed that the reaction was complete, 30 mL of water was added to the reaction system, and ethyl acetate was extracted three times. The organic phases were combined, backwashed once with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography to obtain 33.0 mg of the title compound.

[0333] MS (ESI) m / z (M+H) + =337.2.

[0334] Step 7: Preparation of 5-(piperidin-4-yl)-3,7-dihydrothieno[2,3-b]pyridin-6(2H)-one hydrochloride

[0335]

[0336] Dissolve tert-butyl 4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxylate (110 mg) in hydrogen chloride / 1,4-dioxane solution (4.0 M, 5 mL) and react at room temperature for 1 hour. Remove the solvent in vacuo to obtain 120 mg of crude product, which is directly used in the next step.

[0337] MS (ESI) m / z (M+H) + =237.1.

[0338] Step 8: Preparation of (R)-3-(7-methyl-1H-indazol-5-yl)-2-(4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamido)propanoic acid methyl ester

[0339]

[0340] In a nitrogen atmosphere, crude 5-(piperidin-4-yl)-3,7-dihydrothieno[2,3-b]pyridin-6(2H)-one hydrochloride (120 mg), (R)-methyl 3-(7-methyl-1H-indazol-5-yl)-2-((phenoxycarbonyl)amino)propanoate (110 mg) and triethylamine (51 mg) were dissolved in acetonitrile (5 mL) and reacted at 40° C. for 8 hours. After the reaction was completed, the system was concentrated under reduced pressure, and the crude product was purified by preparative TLC to obtain 90 mg of the title compound.

[0341] MS (ESI) m / z (M+H) + =496.1.

[0342] Step 9: Preparation of (R)-3-(7-methyl-1H-indazol-5-yl)-2-(4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamido)propanoic acid

[0343]

[0344] In a nitrogen atmosphere, (R)-3-(7-methyl-1H-indazol-5-yl)-2-(4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamido)propanoic acid methyl ester (60 mg) was dissolved in tetrahydrofuran (3 mL), and a solution of lithium hydroxide monohydrate (7.3 mg) in water (0.5 mL) was added, and the mixture was reacted at room temperature for 2 hours. After the reaction, the pH was adjusted to neutral with 2M dilute hydrochloric acid, and the system was concentrated under reduced pressure. The obtained crude product (70 mg) was used in the next step without purification.

[0345] MS (ESI) m / z (M+H) + =482.1.

[0346] Step 10: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropan-2-yl)-4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamide

[0347]

[0348] The crude product of (R)-3-(7-methyl-1H-indazol-5-yl)-2-(4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-carboxamido)propionic acid (70 mg), O-benzotriazole-N,N,N,N-tetramethyluronium tetrafluoroboron (57 mg) and 1-(1-methylpiperidin-4-yl)piperazine (27.5 mg) were dissolved in N,N-dimethylformamide (2 mL), and N,N-diisopropylethylamine (26 mg) was added and reacted at room temperature for 1 hour. After the reaction was completed, the system was purified by high pressure reverse preparation to obtain 39 mg of the title compound.

[0349] MS (ESI) m / z (M+H) + =647.1.

[0350] 1 H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.38(s,1H),7.96(s,1H),7.36(s,1H),7.09(s,1H),7.00(s,1H),6.65(d,J=8.0Hz,1H),4.77(q,J= 7.7Hz,1H),4.09(d,J=12.5Hz,2H),3.55(d,J=13.6Hz,1H),3.40(s,2H),3.24(s,1H),3.11(s,1H),3.06(t,J=8.1Hz,2H),2.96(dd,J=13.1, 7.8Hz,1H),2.88(dd,J=13.1,7.1Hz,1H),2.74–2.67(m,4H),2.63(d,J=11.1Hz,1H),2.47(s,3H),2.37–2.31(m,1H),2.23(d,J=10.1Hz,1H ),2.11(s,3H),1.99–1.90(m,2H),1.74(t,J=11.5Hz,2H),1.67–1.59(m,2H),1.57–1.51(m,1H),1.38(d,J=11.9Hz,2H),1.27–1.18(m,5H).

[0351] Example 3: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(2-oxo-2,5,6,8-tetrahydro-1H-pyranylthio[3,4-b]pyridin-3-yl)piperidine-1-carboxamide

[0352]

[0353] The title compound was prepared by using the corresponding common commercial reagents and the products in the above intermediate preparation examples and examples as raw materials and using a preparation method similar to that in Example 1.

[0354] MS (ESI) m / z (M+H) + =660.8.

[0355] 1 H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.36(s,1H),7.96(s,1H),7.36(s,1H),7.00(s,1H),6.89(s,1H),6.64(d,J=8.0 Hz,1H),4.82-4.72(m,1H),4.10(d,J=12.0Hz,2H),3.60-3.54(m,1H),3.50(s,2H),3.25-3.18(m,1H),3.15-3.08(m,1H ),2.98-2.92(m,1H),2.90-2.85(m,1H),2.81-2.78(m,3H),2.75-2.62(m,6H),2.47(s,3H),2.37-2.31(m,1H),2.25-2. 19(m,1H),2.09(s,3H),2.01-1.85(m,2H),1.75-1.64(m,4H),1.58-1.46(m,1H),1.43-1.32(m,2H),1.29-1.11(m,5H).

[0356] Example 4: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(6-oxo-2,3,6,7-tetrahydrothieno[2,3-b]pyridin-5-yl)piperidine-1-thiocarboxamide

[0357]

[0358] Dissolve (R)-2-amino-3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)propan-1-one (23.0 mg) in acetonitrile (1 mL), add N,N-diisopropylethylamine (54.0 mg) and N,N'-thiocarbonyldiimidazole (11.0 mg), and react at room temperature for 1 hour. Add 5-(piperidin-4-yl)-3,7-dihydrothieno[2,3-b]pyridine-6(2H)-one hydrochloride (15.0 mg), heat to 40°C and react for 1 hour. TLC shows that the reaction is complete, filter, collect the filtrate, purify by reverse phase preparation, and freeze-dry to obtain 2.0 mg of the title compound.

[0359] MS (ESI) m / z (M+H) + =663.3.

[0360] 1 H NMR (400MHz, DMSO-d6) δ13.04(s,1H),11.49(brs,1H),7.98(s,1H),7.67(d,J=7.5Hz,1H),7.40(s,1H),7.10(s,1 H),7.02(s,1H),5.54(q,J=7.5Hz,1H),4.80(t,J=12.5Hz,2H),3.50–3.45(m,1H),3.42–3.38(m,2H),3.25–3.18(m ,2H),3.09–2.96(m,4H),2.99–2.83(m,3H),2.72(d,J=10.9Hz,2H),2.48(s,3H),2.35–2.23(m,2H),2.10(s,3H), 2.08–1.99(m,1H),1.97–1.89(m,1H),1.77–1.67(m,4H),1.65–1.59(m,1H),1.44–1.31(m,4H),1.29–1.10(m,3H).

[0361] Example 5: Preparation of N-((R)-3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(2-oxo-1,2,5,6,7,8-hexahydro-5,8-methylquinolin-3-yl)piperidine-1-carboxamide

[0362]

[0363] The title compound was prepared by using the corresponding common commercial reagents and the products in the above intermediate preparation examples and examples as raw materials and using a preparation method similar to that in Example 1.

[0364] MS (ESI) m / z (M+H) + =655.4

[0365] 1H NMR(400MHz,DMSO-d6)δ13.01(s,1H),11.72(s,1H),7.96(s,1H),7.36(s,1H),7.06(s,1H),7.00(s,1H),6.6 3(d,J=8.0Hz,1H),4.77(q,J=7.7Hz,1H),4.08(d,J=12.7Hz,2H),3.55(d,J=13.6Hz,1H),3.38-3.33(m,1H), 3.26-3.20(m,2H),3.18–3.05(m,2H),3.00–2.84(m,2H),2.76-2.60(m,5H),2.47(s,3H),2.37–2.29(m,1H), 2.27–2.18(m,1H),2.09(s,3H),1.99–1.52(m,10H),1.43–1.33(m,3H),1.29-1.21(m,4H),1.06-0.96(m,2H).

[0366] Example 6: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(7'-oxo-7',8'-dihydro-2'H,4'H-spiro[cyclopropane-1,3'-pyranylthio[2,3-b]pyridine]-6'-yl)piperidine-1-carboxamide

[0367]

[0368] The title compound was prepared by using the corresponding common commercial reagents and the products in the above intermediate preparation examples and examples as raw materials and using a preparation method similar to that in Example 1.

[0369] MS (ESI) m / z (M+H)+ = 687.4.

[0370] 1H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.36(s,1H),7.95(s,1H),7.35(s,1H),7.00(s,1H),6.78(s,1H),6.64(d,J=8.0Hz,1H),4.76(q, J=7.6Hz,1H),4.08(d,J=12.8Hz,2H),3.55(d,J=13.2Hz,1H),3.23(s,1H),3.10(t,J=9.6Hz,1H),2.95(dd,J=13.2,7.6Hz,1H),2.88(d,J =16.0Hz,3H),2.67(dt,J=25.6,11.2Hz,5H),2.47(s,3H),2.41(s,2H),2.33(s,1H),2.22(s,1H),2.09(s,3H),1.92(dt,J=23.6,11.2Hz, 2H),1.72(t,J=11.6Hz,2H),1.64(d,J=12.6Hz,2H),1.54(t,J=9.2Hz,1H),1.38(d,J=12.0Hz,2H),1.32–1.09(m,5H),0.68–0.50(m,4H).

[0371] Example 7: Preparation of N-((R)-3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(5-oxo-1,1a,2,4,5,7b-hexahydrocyclopropyl[4,5]pyranthio[2,3-b]pyridin-6-yl)piperidine-1-carboxamide

[0372]

[0373] The title compound was prepared by using the corresponding common commercial reagents and the products in the above intermediate preparation examples and examples as raw materials and using a preparation method similar to that in Example 1.

[0374] MS (ESI) m / z (M+H) + =673.3

[0375] 1H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.31(s,1H),7.96(s,1H),7.36(s,1H),7.16(d,J=7.5Hz,1H),7.00(s,1H),6.65(d,J= 7.9Hz,1H),4.77(q,J=7.7Hz,1H),4.11(d,J=12.9Hz,2H),3.56(d,J=13.3Hz,1H),3.40-3.36(m,1H),3.24-3.19(m,1H),3.12- 3.06(m,3H),2.99-2.93(m,1H),2.90-2.84(m,1H),2.77–2.60(m,5H),2.47(s,3H),2.36-2.29(m,1H),2.26–2.17(m,1H),2.09 (s,3H),1.97–1.78(m,4H),1.76–1.61(m,4H),1.52(t,J=9.8Hz,1H),1.42–1.27(m,5H),1.24–1.10(m,2H),0.85-0.77(m,1H).

[0376] Example 8: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-oxo-1-(4-(piperidin-4-yl)piperazin-1-yl)propan-2-yl)-4-(7-oxo-3,4,7,8-tetrahydro-2H-pyranylthio[2,3-b]pyridin-6-yl)piperidine-1-carboxamide

[0377]

[0378] Step 1: Preparation of (R)-tert-butyl 4-(4-(3-(7-methyl-1H-indazol-5-yl)-2-(4-(7-oxo-3,4,7,8-tetrahydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)propanoyl)piperazin-1-yl)piperidine-1-carboxylate

[0379]

[0380] (R)-3-(7-methyl-1H-indazol-5-yl)-2-(4-(7-oxo-3,4,7,8-tetrahydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)propionic acid (66 mg), tert-butyl 4-(piperazine-1-yl)piperidine-1-carboxylate (43 mg) and N,N-diisopropylethylamine (43 mg) were dissolved in N,N-dimethylformamide (2 mL), and 2-(1H-benzotriazol L-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (43 mg) was added, and the mixture was reacted at room temperature for 4 hours. After the reaction was completed, the reaction solution was separated and purified by preparative HPLC to obtain 30 mg of the title compound.

[0381] MS (ESI) m / z (M+H) + =747.3.

[0382] Step 2: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-oxo-1-(4-(piperidin-4-yl)piperazin-1-yl)propan-2-yl)-4-(7-oxo-3,4,7,8-tetrahydro-2H-pyranothio[2,3-b]pyridin-6-yl)piperidine-1-carboxamide

[0383]

[0384] Dissolve (R)-4-(4-(3-(7-methyl-1H-indazol-5-yl)-2-(4-(7-oxo-3,4,7,8-tetrahydro-2H-thiopyrano[2,3-b]pyridin-6-yl)piperidine-1-carboxamido)propionyl)piperazin-1-yl)piperidine-1-carboxylic acid tert-butyl ester (30 mg) in acetic acid and water (2 mL, v / v = 1 / 5) and react at 100°C for 2 hours. After the reaction is completed, concentrate, dissolve the crude product in an appropriate amount of methanol, adjust the system pH to 8-9 with saturated potassium carbonate solution, filter, separate and purify the filtrate by preparative HPLC, and freeze-dry to obtain 10.00 mg of the title compound. MS (ESI) m / z (M+H) + =647.3.

[0385] 1H NMR(400MHz,DMSO-d6)δ13.02(s,1H),7.96(s,1H),7.36(s,1H),7.00(s,1H),6.86(s,1H),6.63 (d,J=8.0Hz,1H),4.77(q,J=7.7Hz,1H),4.08(d,J=12.6Hz,2H),3.60-3.47(m,1H),3.30-3.20( m,1H),3.15-3.06(m,2H),3.05–2.84(m,7H),2.75–2.59(m,3H),2.56-2.51(m,2H),2.47(s,3H) ,2.41–2.18(m,4H),2.06-1.91(m,4H),1.70–1.52(m,3H),1.41–1.17(m,4H),1.09–0.97(m,2H).

[0386] Example 9: Preparation of (R)-N-(3-(7-methyl-1H-indazol-5-yl)-1-(4-(1-methylpiperidin-4-yl)piperazin-1-yl)-1-oxopropane-2-yl)-4-(2-oxo-2,5,6,7-tetrahydro-1H-cyclopenta[b]pyridin-3-yl)piperidine-1-carboxamide

[0387]

[0388] The title compound was prepared by using the corresponding common commercial reagents and the products in the above intermediate preparation examples and examples as raw materials and using a preparation method similar to that in Example 1.

[0389] MS (ESI) m / z (M+H) + =629.3.

[0390] 1H NMR (400MHz, DMSO-d6) δ13.01(s,1H),11.66(s,1H),7.96(s,1H),7.36(s,1H),7.03(s,1H),7.00(d,J=1.2Hz,1H),6.6 4(d,J=8.1Hz,1H),4.77(q,J=7.7Hz,1H),4.16–4.03(m,2H),3.59–3.49(m,1H),3.38-3.36(m,1H),3.28–3.19(m,1H),3 .15-3.07(m,1H),2.99-2.92(m,1H),2.91-2.84(m,1H),2.79–2.56(m,9H),2.47(s,3H),2.37–2.28(m,1H),2.26–2.17( m,1H),2.09(s,3H),2.04–1.84(m,4H),1.78–1.60(m,4H),1.54(t,J=8.8Hz,1H),1.41-1.33(m,2H),1.29–1.09(m,4H).

[0391] With reference to the aforementioned synthesis method, the present invention further synthesized the following example compounds:

[0392]

[0393]

[0394]

[0395] Biological test data

[0396] Unless otherwise specified, the experimental materials, reagents, operations and methods used in the following activity test examples can be obtained from commercial channels or can be easily known or prepared based on existing technologies.

[0397] Test Example 1: Cellular Functional Antagonism Assay - cAMP Assay

[0398] 1. Experimental Principle

[0399] The CGRP receptor complex is coupled to Gs in the G protein. The binding of CGRP to the CGRP receptor complex leads to the activation of Gs and the production of cAMP (3',5'-cyclic adenosine monophosphate).

[0400] 2. Purpose of the experiment

[0401] The compounds of the invention were determined for their ability to inhibit CGRP-stimulated cAMP formation in SK-N-MC cells.

[0402] 3. Experimental Materials

[0403] 3.1 Experimental cell lines:

[0404] SK-N-MC (neuroepithelioma cells), source: National Biomedical Experimental Cell Resource Bank.

[0405] 3.2 Reagents and consumables

[0406]

[0407]

[0408] 4. Experimental Procedure

[0409] 4.1 Cell preparation:

[0410] Resuscitate and culture SK-N-MC cells in advance. Before use, the cell confluence should be 70%-80%.

[0411] 4.2 Prepare sample diluent, test compound and α-CGRP (human).

[0412] 4.3 Cell plating and drug effects:

[0413] Digest SK-N-MC cells, resuspend in sample diluent after centrifugation, and inoculate in 384-well plates. Add α-CGRP (human) and the test compound to each well, pipette and mix. Put the sealing film on the well plate and incubate it in an ELISA incubator at 25°C.

[0414] 4.4 cAMP determination: refer to the instructions of LANCE Ultra cAMP Kit. Prepare Eu-cAMP tracer and ULight-anti-cAMP working solution in a dark place. Add them to the well plate and mix them by pipetting. Put the well plate with a sealing film and place it in an ELISA incubator at 25℃.

[0415] 4.5 Use the chemiluminescence module of the fully automatic microplate reader to read the luminescence value.

[0416] 4.6 Data Analysis:

[0417] The IC50 value was calculated using the nonlinear formula with the compound concentration LOG value as the horizontal axis and the Inhibition as the vertical axis. The results are shown in the table below.

[0418] IC of compounds for inhibition of CGRP-stimulated cAMP production in SK-N-MC cells 50 value

[0419] Example No. <![CDATA[IC 50 (nM)]]> Example No. <![CDATA[IC 50 (nM)]]> 1 0.21 2 0.41 3 1.44 4 0.71 5 26.04 6 0.15 7 0.24 8 0.14 9 1.94 10 1.35 11 0.65 12 0.62 13 1.00 14 1.94 15 6.02 16 7.83 17 10.08 18 1.04

[0420] 5. Conclusion

[0421] The novel compounds provided by the present invention can significantly inhibit the production of cAMP stimulated by CGRP and have a low IC 50 The inhibitory activity of the compounds is very significant, and the IC50 value can reach the pM level. The different selection of variable groups has an important influence on the inhibitory activity of the compounds.

[0422] Test Example 2: In vitro 125 I] Competitive binding assay for CGRP

[0423] 1. Experimental purpose:

[0424] The compounds were tested for competition for the radiolabeled endogenous peptide human α-CGRP ([ 125 I]CGRP) capacity.

[0425] 2. Experimental principle:

[0426] SK-N-MC endogenously expresses a CGRP receptor that has the same sequence as the human CGRP receptor. Using SK-N-MC cell membrane homogenate as a receptor source, the endogenous peptide human αCGRP ([ 125 I]CGRP) and compounds compete with it for binding, and wash the unbound part [ 125 After PCR amplification, the binding of the compound to the receptor can be detected by radioactive signal.

[0427] 3. Experimental Materials:

[0428] 3.1 Experimental cell line: SK-N-MC (human neuroepithelioma cells), source: Peking Union Medical College Cell Resource Center 3.2 Detection equipment: MicroBeta2 (PerkinElmer), UNIFILTER-96 well cell collector (PerkinElmer, C961961)

[0429] 3.3 Experimental reagents and consumables

[0430] name brand Part Number Tris base Sigma T1503-1KG <![CDATA[MgCl2]]> Sigma M1028 PEI (Polyethyleneimine) Sigma P3143 Bovine Serum Albumin Sigma B2064-50G HEPES1M Gibco 15630080 Microscint 20cocktail PerkinElmer 6013329 Unifilter-96GF / B filter plates PerkinElmer 6055690 96well conical polypropylene plates Agilent 5042-1385 TopSeal-Asealing film PerkinElmer 6050185 <![CDATA[ 125 I-CGRP(Human)]]> NET354050UC αCGRP(Human) MCE HY-P1071

[0431] 3.4 Experimental procedures

[0432] 3.4.1 Membrane protein extraction and quantification: Collect fresh SK-N-MC cells. Resuspend the cell pellet with experimental buffer (Tris-HCl 50mM, pH7.4). Use a homogenizer to homogenize the cells 10 times, 1000g, 4℃, centrifuge for 10 minutes, remove nuclear / mitochondrial fragments, and collect the supernatant tissue homogenate. Centrifuge the supernatant at 50000g, 4℃ for 1 hour and remove the supernatant. Suspend the pellet with an appropriate buffer, determine the protein concentration, and store the suspension in aliquots at -80℃.

[0433] 3.4.2 Compound binding Ki (inhibition constant) determination: 1 μl of serially diluted test compound was transferred to the assay plate. 50 μl of membrane stock solution (15 μg) and 50 μl of 200 pM final concentration of 1% ... 125 I-CGRP (Human), shake and mix. Add 50μl 0.3% PEI to each well and soak for 1 hour. Filter the reaction mixture with a UNIFILTER-96 well cell collector and rinse the plate with pre-cooled buffer. After drying the filter plate, seal the bottom of the well plate with Unifilter-96 backing seal tape, add 50μl Microscint 20 cocktail to each well, and seal the top of the well plate with TopSeal-Asealing film. Count the cells captured on the filter using a MicroBeta2 Reader 3 H signal.

[0434] 3.4.3 Data Analysis

[0435] Data were analyzed using Prism 5. A “log(inhibitor) vs. response-Variable slope” model was used to fit the data, using the 125 I-CGRP (Human) Kd value is 240.6pM, calculated compound and 125 IC of I-CGRP(Human) competitively binds to SK-N-MC cell membrane 50 and Ki (Ki = IC50 / (1+[L] / Kd), [L]: Radioligand concentration (200 pM)).

[0436] 4. Experimental Results

[0437] The Ki values ​​of the compounds of the present invention for binding to CGRP receptors are shown in the following table:

[0438] Compound Ki(pM) Ki ratio Zavegepant 49.76 / Example 1 19.48 0.39 Example 2 11.58 0.23

[0439] Note: Ki ratio = Ki (compound) / Ki (Zavegepant).

[0440] From the in vitro data in the above table, it can be seen that the affinity of the compound of the present invention for the CGRP receptor reaches the pM level, and shows better in vitro target affinity properties compared with Zavegepant.

[0441] Test Example 3: Liver microsome stability test

[0442] 1. Purpose of the experiment

[0443] The stability of the compounds of the present invention in rat, monkey and human liver microsomes was determined. The compounds of the present invention tested were the compounds of Example 1 and Example 2, and the control drug was Zavegepant.

[0444] 2. Reagents and consumables

[0445] Reagent name supplier batch number Concentration (mg protein / mL) Rat liver microsomes RILD Wlq 20 Monkey liver microsomes RILD ZYM 20 Human liver microsomes BioreclamationIVT YB 20

[0446] 3. Experimental Procedure

[0447] 3.1 Experimental incubation system

[0448]

[0449] 3.2 Pipette an appropriate amount of liver microsome solution into a 1mL 96-deep-well plate, add the test solution (or probe substrate solution), and pre-incubate in a 37°C thermomixer for 5 minutes. Take out two mixed solutions from the system, add 1×PBS instead of NADPH, and take out the mixed solutions at 0 and 60 minutes respectively, and add methanol (including internal standard) to terminate the reaction. Add NADPH to each well of the remaining mixed system to start the reaction. Take out the mixed solution from the system at 0, 5, 15, 30, 45, and 60 minutes for the test group and 0, 30, and 60 minutes for the control group, and add methanol (including internal standard) to terminate the reaction. Mix all the samples that have terminated the reaction, centrifuge at 3800rpm for 15 minutes in a centrifuge, and take the supernatant for LC-MS / MS analysis.

[0450] 3.3 Data Analysis

[0451] The peak areas were determined from the extracted ion chromatograms. The slope value k was determined by linear regression of the residual percentage of the parent drug against the natural logarithm of the incubation time curve. The in vitro half-life (t 1 / 2 ), and calculated the in vitro intrinsic clearance (CL int , expressed in μL / min / mg protein). The calculation formula is as follows:

[0452] t 1 / 2 =ln2 / k=0.693 / k; CL int =0.693 / t 1 / 2 / liver microsomal protein concentration

[0453] The experimental results are shown in the following table:

[0454] Stability data of the compounds of the present invention in rat, monkey and human liver microsomes

[0455]

[0456] Test Example 4: Plasma protein binding rate test

[0457] 1. Purpose of the experiment

[0458] The protein binding rate of the compound of the present invention in the plasma of rats, monkeys and humans was determined by equilibrium dialysis. The tested compounds of the present invention were the compounds of Examples 1 and 2, and the control drug was Zavegepant.

[0459] 2. Test Matrix

[0460] Matrix / species Anticoagulants SD rat plasma <![CDATA[EDTA-K2]]> Cynomolgus monkey plasma <![CDATA[EDTA-K2]]> Human plasma <![CDATA[EDTA-K2]]>

[0461] 3. Experimental Procedure

[0462] 3.1 Rinse the dry dialysis membrane with ultrapure water for 2-3 times, and soak it in phosphate buffer for 1 hour for use; soak the polytetrafluoroethylene module in 20% ethanol for 30 minutes, dry the surface moisture with dust-free paper and dry it for use. Assemble the pretreated dialysis membrane into the dialysis plate according to the product instructions, and add 100 μL of receiving solution (100 mM phosphate buffer solution plus 0.002% Tween 80) to one side of the permeable membrane in each dialysis well (receiving chamber).

[0463] 3.2 Place plasma in a centrifuge tube, add the working solution of the substance to be tested to the plasma to a final concentration of 1 μM, and mix by inverting (this step is performed on an ice bath); take 20 μL of drug-containing plasma into each 96-well sample plate, in parallel, as T0 samples, and store in a -20°C refrigerator.

[0464] 3.3 Take another 100 μL of the above drug-containing plasma to the other side of the membrane (sample chamber) of the dialysis device, and place two parallel copies at 37°C for constant temperature shaking incubation for 6 hours. After 6 hours of incubation, take 20 μL of each of the balanced receiving chamber and sample chamber to obtain samples B and A. Add the corresponding volume of blank plasma or phosphate buffer (containing 0.002% Tween 80) to sample B and sample A respectively, so that the volume ratio of plasma to buffer in each sample well is 1:1.

[0465] 3.4 Add 250 μL of internal standard methanol solution to all sample wells, mix thoroughly, and centrifuge at 3800 rpm for 10 minutes. Take 20 μL of supernatant and add 180 μL of methanol, vortex mix thoroughly, and then inject for LC-MS / MS analysis.

[0466] 3.5 Data Analysis

[0467] The plasma protein binding rate and recovery rate of the compound in plasma were calculated using the following formula:

[0468] Free percentage (%) = C B / C A ;

[0469] Plasma protein binding rate (f b%) = 1-free percentage (%);

[0470] Recovery rate (%) = (C B +C A ) / C T0

[0471] Among them C B is the concentration of the compound in the receiving fluid after equilibrium dialysis; C A is the concentration of the compound in the plasma after equilibrium dialysis; C T0 is the initial concentration of the compound in plasma.

[0472] The experimental results are shown in Table 3 below:

[0473]

[0474] Test Example 5: In vivo pharmacokinetic study of the test compound administered intravenously and intranasally to SD rats

[0475] 1. Experimental Animals

[0476] Species: SD rats, male, SPF grade. Source: purchased from Chengdu Dashuo Experimental Animal Co., Ltd., experimental animal production license number: SCXK (Chuan) 2020-030. Quantity: 2 rats were given intravenously and 4 rats were given nasal drops.

[0477] 2. Preparation of test samples

[0478] 2.1 Accurately weigh an appropriate amount of drug, add 50 mM succinate buffer (D5W dilution, pH = 5-6), and mix and dissolve thoroughly by ultrasonication and vortexing to obtain a 0.2 mg / mL dosing solution for intravenous (IV) administration.

[0479] 2.2 Accurately weigh an appropriate amount of drug, add 50 mM succinate buffer (diluted with purified water, pH = 5-6), and mix and dissolve thoroughly by ultrasonication and vortexing to obtain a 10 mg / mL dosing solution for intranasal (IN) administration.

[0480] 3. Experimental Design

[0481] Group quantity Dosage (mg / kg) Concentration (mg / mL) Dosing volume Dosage Collecting samples 1 2 1 0.2 5mL / kg IV plasma 2 4 2 10 50μL / pc IN plasma

[0482] The test drugs were Examples 1 and 2, and the control drug was Zavegepant.

[0483] 4. Blood collection time

[0484] 5min, 10min, 15min, 0.5h, 1h, 2h, 4h, 8h, and 24h after administration.

[0485] 5. Sample Collection and Disposal

[0486] Blood was collected from the jugular vein, and about 0.2 mL of blood was collected for each sample. EDTA-K2 was used for anticoagulation. The blood samples were placed on ice after collection and centrifuged to separate plasma within 2 hours (centrifugation conditions: 6000g, 5min, 2-8℃). The collected plasma samples were stored in a -80℃ refrigerator before analysis, and the remaining plasma samples were stored in a -80℃ refrigerator after analysis.

[0487] 6. Bioanalysis and Data Processing

[0488] The plasma concentration of the designated compound was determined by LC-MS / MS, and the main pharmacokinetic parameters were calculated by Winnolin 8.3 non-compartmental model. max The previous BLQ (including "No peak") is calculated as 0; C max BLQ (including "No peak") that appears later will not be included in the calculation.

[0489] In vivo pharmacokinetic study data of the test compound administered intravenously and intranasally to SD rats

[0490]

[0491] Test Example 6: In vivo pharmacodynamic study of the test compound on capsaicin-induced changes in forearm skin blood flow in cynomolgus monkeys 1. Experimental principle

[0492] Capsaicin induces the release of CGRP after acting on the skin. CGRP binds to receptors to dilate blood vessels and promote increased blood flow. The test compound inhibits the binding of CGRP to receptors, inhibits the increase in blood flow caused by the release of CGRP, and the blood flow is detected by a blood flowmeter to reflect the efficacy of the drug.

[0493] 2. Experimental Procedure

[0494] 2.1 Animal Information:

[0495] Species and strain: Cynomolgus monkey, male (5-8 kg), animal use license: SYXK (Su) 2019-0004, hair removal was performed 3 days before the experiment.

[0496] 2.2 Test sample configuration:

[0497] Accurately weigh an appropriate amount of the drug, dissolve it in sterile physiological saline, and mix and dissolve it thoroughly by ultrasonication and vortexing to obtain a 0.05 mg / ml dosing solution for intravenous (IV) administration.

[0498] 2.3 Preliminary stage: After the animals were anesthetized, a blood flow meter was used to scan the blood flow on the inner side of the forearm, and an O-shaped rubber ring was placed on the inner side of the arm, avoiding obvious blood vessels. After the position of the rubber ring was confirmed, the blood flow meter scanned the blood flow inside the ring as the baseline blood flow value, and then 2 mg of capsaicin was applied to the rubber ring. The blood flow meter recorded the blood flow at 10 minutes, 20 minutes and 30 minutes after capsaicin treatment.

[0499] 2.4 Drug administration phase: After the monkeys were anesthetized, a rubber ring was placed on the same part of the arm. After collecting baseline blood flow data, the test compound (dose: 0.05 mg / kg) was injected intravenously at a dose of 1 ml / kg. After administration, 2 mg of capsaicin was applied to the rubber ring, and the blood flow meter recorded the blood flow 10 min, 20 min and 30 min after capsaicin treatment.

[0500] 2.5 Data processing: The blood flow in the pre-test phase is recorded as P, and the blood flow in the rubber ring at time = t is recorded as P t The baseline blood flow is recorded as P0; the blood flow during the drug administration period is recorded as D, and the blood flow of the rubber ring at time = t is recorded as D t , the baseline blood flow is recorded as D0, then the blood flow inhibition rate I at time = t t The calculation formula is as follows:

[0501] I t =((P t –P0) / P0–(D t –D0) / D0)*100%

[0502] 3. Experimental results:

[0503] The blood flow inhibition rate at different time points Figure 1 As shown, the test compounds can inhibit the increase in blood flow induced by capsaicin. At 10 minutes, the blood flow inhibition rate of Example 1 (Mean=355.3%) is significantly higher than that of Zavegepant (Mean=89.7%). The blood flow inhibition effect of the test compounds lasts until 30 minutes.

[0504] The present invention is not limited to the above optional implementations, and anyone can derive other various forms of products under the enlightenment of the present invention. The above specific implementations should not be understood as limiting the scope of protection of the present invention. The scope of protection of the present invention should be based on the definition in the claims, and the description can be used to interpret the claims.

Claims

1. The intermediate compound described by the following formula or its salt: In Formula I: R a is -H, -(C1-C6)alkyl, and may also be -(C1-C3)alkyl, preferably methyl or ethyl; R b is -H or an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, and the amino protecting group is preferably -Boc; The G chain group and the two carbon atoms to which it is attached together form a 5- to 7-membered aliphatic ring or aliphatic heterocyclic ring, wherein the heteroatoms constituting the ring in the aliphatic heterocyclic ring are -O- or -S-; R 1 is independently at each occurrence -H, -(C1-C6)alkyl or -(C3-C8)cycloalkyl, m is 0, 1, 2, 3, 4, 5 or 6, or -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

2. The intermediate compound or its salt according to claim 1: For example, the G chain group and the two carbon atoms to which it is attached together form a 5- or 6-membered aliphatic ring, and the G chain group optionally has 0 or 1 double bonds, and the rest are single bonds; For example, the G chain group and the two carbon atoms to which it is attached together form a 7-membered aliphatic ring, and the G chain group optionally has 0, 1 or 2 double bonds, and the rest are single bonds; For example, the G chain group and the two carbon atoms to which it is attached together form a 5-membered aliphatic heterocyclic ring, and all the G chain groups have single bonds; For example, the G chain group and the two carbon atoms to which it is attached together form a 6- or 7-membered aliphatic heterocyclic ring, and the G chain group optionally has 0 or 1 double bonds, and the rest are single bonds; R 1 is independently at each occurrence -H, -(C1-C3)alkyl or -(C3-C6)cycloalkyl, m is 0, 1, 2 or 3, Or, -(R 1 ) m The atoms to which it is attached together form a 3-6 membered aliphatic ring or aliphatic heterocyclic ring, wherein the aliphatic ring or aliphatic heterocyclic ring is optionally connected with -(R 1 ) m The attached ring constitutes a spiro ring structure, a fused ring structure or a bridged ring structure, and the heteroatom in the aliphatic heterocycle is O, N or S.

3. The intermediate compound or its salt according to claim 1: R 1 is independently -H, methyl, ethyl or cyclopropane at each occurrence, m is 0, 1 or 2, Or, -(R 1 ) m The atoms to which it is attached together form cyclopropane, cyclobutane or cyclopentane, and the cyclopropane, cyclobutane or cyclopentane and -(R 1 ) m The attached rings constitute a spiro ring structure, a fused ring structure or a bridged ring structure.

4. The intermediate compound or a salt thereof according to claim 1: The intermediate compound is shown in the following formula Ⅰ-1, Ⅰ-2, Ⅰ-3, Ⅰ-4 or Ⅰ-5: In formula I-1, I-2, I-3, I-4 or I-5, X 6 is -O- or -S-; In formula Ⅰ-2, two "---" both represent single bonds, or any one of them is a double bond and the other is a single bond; In formula Ⅰ-3, three "---" all represent single bonds, or any one of them is a double bond and the other two are single bonds; In formula Ⅰ-4, "---" refers to a single bond or a double bond; In formula Ⅰ-5, two "---" both represent single bonds, or any one of them is a double bond and the other is a single bond.

5. The intermediate compound or a salt thereof according to claim 1: The G chain group and the two carbon atoms to which it is attached together form a 5- to 7-membered aliphatic ring, and m is 0.

6. The intermediate compound or a salt thereof according to claim 1: The salt is hydrochloride or trifluoroacetate.

7. The intermediate compound or a salt thereof according to claim 1: In the intermediate compound, more than one H is replaced by D, and optionally, a G chain group or R 1 One or more H in is replaced by D.

8. The intermediate compound or a salt thereof according to claim 1: The intermediate compound is selected from:

9. The intermediate compound described by the following formula or its salt: R a , G chain group, R 1 and m each occurrence is independently as defined in any intermediate compound of claims 1 to 5 and 7; In formula II, when R a When is ethyl, the intermediate compound shown in formula II is not In formula III, when R a When is methyl, the intermediate compound shown in formula III is not 10. The intermediate compound or a salt thereof according to claim 9: The intermediate compound is selected from:

11. The intermediate compound described by the following formula or its salt: R b is -H or an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, and the amino protecting group is preferably -Boc.

12. A method for preparing an intermediate compound, comprising at least one of the following synthetic routes (1) to (5): (1) (2) (3) (4) (5) in: R a Each occurrence is independently -(C1-C6)alkyl, and may optionally be -(C1-C3)alkyl, preferably methyl or ethyl; R b Each occurrence is independently an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, the amino protecting group is preferably -Boc; G chain group, R 1 and m are independently defined at each occurrence as in any one of the intermediate compounds of claims 1 to 5 and 7.

13. The method for preparing the intermediate compound according to claim 12, further comprising the following synthetic route: in: R b Each occurrence is independently an amino protecting group, the amino protecting group can be selected from -Cbz, -Boc, -Fmoc, -PMB, -Bn, -Trt, -Tos or -Alloc, the amino protecting group is preferably -Boc; G chain group, R 1 and m are independently defined at each occurrence as in any one of the intermediate compounds of claims 1 to 5 and 7.

14. A method for synthesizing a compound as a CGRP antagonist, comprising the following synthetic route:

15. The synthesis method according to claim 14, further comprising the following synthesis route:

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