Heterocyclic JAK inhibitors

Abstract

The present invention relates to a heterocyclic compound or a pharma- ceutically acceptable salt thereof as an inhibitor of Janus kinase (JAK). Specifically, the present invention relates to a compound represented by general formula (I) or a pharma- ceutically acceptable salt thereof. The present invention further relates to a method for preparing said compound or a pharma- ceutically acceptable salt thereof. The compound of the present invention can be applied to the treatment and / or prevention of diseases mediated by JAK, particularly inflammatory diseases, autoimmune diseases and cancer. However, the definition of each group in general formula (I) is as described in the instructions. TIFF2024522210000058.tif27150

Description

[Technical Field]

[0001] This invention relates to heterocyclic compounds or pharmaceutically acceptable salts thereof that modulate or inhibit Janus kinase (JAK) activity. The invention also relates to methods for producing such compounds or pharmaceutically acceptable salts thereof. Furthermore, the invention relates to the use and methods of use of such compounds or pharmaceutically acceptable salts in the treatment and / or prevention of inflammatory diseases, autoimmune diseases, and cancer. [Background technology]

[0002] JAKs are non-receptor tyrosine kinases consisting of four families: JAK1, JAK2, JAK3, and TYK2. JAKs have seven homology domains (JAK Homology Domains, JH), of which the JH1 domain is the kinase domain, the JH2 domain is the pseudokinase domain (which regulates the kinase activity of JH1), and JH6 and JH7 are receptor-binding domains. When a cytokine binds to the cell surface region of a cytokine receptor, the intracellular region to which JAK is bound is phosphorylated, creating a binding site for signal transduction and transcriptional activation protein (STAT). STAT proteins are further phosphorylated by activated JAK to form dimers, which enter the nucleus, regulating the expression and transcription of related genes and enabling cell membrane-to-nuclear signaling (Lionard et. al, Ann. Rev. Immunol. 1998, 16, 293-322). Therefore, JAKs play a crucial role in many cellular functions, including cytokine-mediated signaling via the JAK-STAT pathway, and cytokine-dependent regulation of cell proliferation, differentiation, apoptosis, and immune responses, making them common targets for the treatment of inflammatory diseases, autoimmune diseases, and cancer (Alicea-Velazquez et. al, Curr. Drug Targets 2011, 12, 546-55). JAK-modulating drugs have been approved for sale, including the JAK1 / JAK2 inhibitor ruxolitinib and the JAK2 inhibitor fedratinib used to treat myelofibrosis, and the pan-JAK inhibitor tofacitinib, the JAK1 / JAK2 inhibitor baricitinib, the pan-JAK inhibitor peficitinib, and the JAK1 inhibitor upadacitinib used to treat rheumatoid arthritis.

[0003] JAKs and STATs act with highly specificity in regulating different immune responses. One type of JAK kinase can be involved in the signaling processes of multiple cytokines, and one type of cytokine signaling pathway can activate multiple JAK kinases. However, cytokines have selectivity for activated STAT proteins; for example, interleukin (IL)-4 activates STAT1 / 3 / 5 / 6, and IL-12 specifically activates STAT4. JAK1, JAK2, and TYK2 are widely present in various tissues and cells, and JAK1 is closely related to the activation of inflammatory factors such as IL-6 and interferon (IFN). Therefore, JAK1 selective inhibitors are thought to have promising therapeutic effects in the treatment of autoimmune diseases such as rheumatoid arthritis (RA) and psoriasis. JAK2 can independently mediate the signaling of cytokines such as erythropoietin (EPO) and thrombopoietin (TPO) (Won et al., BMC Bioinformatics 2009, 10, S53) and is closely correlated with the proliferation and differentiation of blood cells. TYK2 is involved in the signaling of inflammatory cytokines such as interferons (IFNs), IL-12, and IL-23, playing a crucial role in innate and adaptive immunity. Therefore, it is attracting considerable attention as a target for autoimmune diseases, such as psoriasis, systemic lupus erythematosus (SLE), and inflammatory bowel disease (IBD), and TYK2 inhibitors are being developed for their treatment. JAK3 is present only in the myeloid and lymphoid systems and mediates the signaling of IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21. These cytokines play important roles in inducing T cell proliferation and differentiation, activating B cell-produced antibodies, activating macrophages, enhancing natural killer (NK) cell activity, and inducing other cytokines such as IFN. Therefore, JAK3 selective inhibitors are expected to play an important role in the treatment of organ transplantation, autoimmune diseases, inflammatory pneumonia, and other conditions.

[0004] Inflammatory bowel disease (IBD) is a common chronic inflammatory bowel disease that includes Crohn's disease (CD), ulcerative colitis (UC), and non-definitive IBD (IBDU). IBD affects 5 million people worldwide, and its prevalence is increasing year by year. Clinical symptoms include diarrhea, bloody stools, abdominal pain, fatigue, and high fever. While treatments such as 5-aminosalicylic acid (5-ASAs), glucocorticoids, immunosuppressants (e.g., azathioprine), and biological agents (e.g., anti-TNF, IL-12 / IL-23 monoclonal antibodies) are commonly used, many patients do not achieve remission, and 80% of Crohn's disease patients and 30% of UC patients eventually require surgery. This field also presents a huge medical need for more effective and safer medications.

[0005] During this disease development process, the expression of pro-inflammatory factors such as IL-13 and IL-17 increases, inducing inflammation via the JAK-STAT pathway. JAK inhibitors have potential uses as novel oral small molecule drugs for the treatment of CD and UC. Tofatinib, for example, is approved for the treatment of UC in many countries, but it non-selectively inhibits JAK1 / 2 / 3, causing serious side effects such as severe infections and the development of malignancies. However, due to the high sequence similarity of the catalytic active sites of members of the JAK kinase family, designing an oral selective JAK inhibitor with the systemic exposure required for treatment is quite difficult. Therefore, new JAK inhibitors are being developed to enrich the local exposure required for treatment only at the site of action, such as the colon or skin, while keeping the solubility in the blood extremely low to avoid systemic side effects, exert efficacy, and enhance safety. Developed by Theravance, TD-1473 is a pan-JAK inhibitor with local intestinal absorption and has progressed to Phase 3 clinical trials, where it demonstrated good tolerance. [Overview of the project]

[0006] definition Unless otherwise specified, the following terms used in this specification shall have the following meanings:

[0007] "C x-y " refers to the range of carbon atoms, where x and y are both integers, for example, C 3-8 Cycloalkyl refers to cycloalkyls having 3 to 8 carbon atoms, i.e., cycloalkyls having 3, 4, 5, 6, 7, or 8 carbon atoms. Also, "C 3-8 " is any subrange within it, for example, C 3-7 , C 3-6 , C 4-7 , C 4-6 , C 5-6 It should be understood that this also includes such things.

[0008] "Alkyl" refers to saturated, linear or branched hydrocarbyl substituents containing 1 to 20 carbon atoms, for example, 1 to 8 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbon atoms. Non-limiting examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, and 2-ethylbutyl.

[0009] "Cycloalkyl" refers to a saturated cyclic hydrocarbyl substituent containing 3 to 14 carbocyclic atoms. Cycloalkyls may also be monocyclic carbocyclics typically containing 3 to 8, 3 to 7, or 3 to 6 carbocyclic atoms. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl. Cycloalkyls may also be bicyclic or tricyclic, condensed, cross-linked, or spirocyclic, such as decahydronaphthalenyl, bicyclo[2.2.2]octane, and spiro[3.3]heptane.

[0010] "Heterocyclyl or heterocyclic" means a saturated or partially unsaturated monocyclic or polycyclic cyclic group containing 3 to 20 ring atoms, for example, 3 to 14, 3 to 12, 3 to 10, 3 to 8, 3 to 6, or 5 to 6 ring atoms, where one or more of these are selected from nitrogen, oxygen, or S(O)m (where m is an integer from 0 to 2) (except for -OO-, -OS-, or -SS- ring portions), and the remaining ring atoms are carbon. Preferably, it contains 3 to 12 ring atoms, more preferably 3 to 10 ring atoms, more preferably 4 to 7 ring atoms, more preferably 4 to 6 ring atoms, most preferably 5 or 6 ring atoms, of which 1 to 4 are heteroatoms, more preferably 1 to 3 are heteroatoms, and most preferably 1 to 2 are heteroatoms. Non-exclusive examples of monocyclic heterocyclils include pyrrolidinyl, oxetanil, piperidinyl, piperazinyl, tetrahydrofuranil, tetrahydropyranil, tetrahydrothiopyranil, morpholinil, thiomorpholinil, homopiperazinyl, and azetidinil. Non-exclusive examples of polycyclic heterocyclils include polycyclic heterocyclic groups formed by condensation, cross-linking, or spiro, such as octahydrocyclopenta[c]pyrrole, octahydropyrrolo[1,2-a]pyrazine, 3,8-diazabicyclo[3.2.1]octane, 5-azaspiro[2.4]heptane, and 2-oxa-7-azaspiro[3.5]nonane.

[0011] "Aryl or aromatic ring" means an aromatic monocyclic or fused polycyclic group containing 6 to 14 carbon atoms, preferably 6 to 10 members, such as phenyl and naphthyl, most preferably phenyl. The aryl ring may be fused to a heteroaryl, heterocyclyl, or cycloalkyl ring, provided that the ring bonded to the parent structure is an aryl ring, and non-limiting examples include [ka] These are some examples.

[0012] "Heteroaryl or heteroaromatic ring" means a heteroaromatic system containing 5 to 14 ring atoms, of which 1 to 4 ring atoms are selected from heteroatoms containing oxygen, sulfur, and nitrogen. Heteroaryls are preferably 5 to 10-membered, more preferably 5 or 6-membered, and include, for example, furyl, thienyl, pyridinyl, pyrrolyl, pyrimidinyl, pyrazinyl, pyrazolyl, imidazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, quinolinyl, isoquinolyl, indolyl, isoindolyl, etc. The heteroaryl ring may be condensed with an aryl, heterocyclyl, or cycloalkyl ring, provided that the ring bonded to the parent structure is a heteroaryl ring, and non-limiting examples include: [ka] These are some examples.

[0013] "Halogen" refers to fluorine, chlorine, bromine, or iodine.

[0014] "Cyano" means -CN.

[0015] "In some cases" means that the event or situation described later may occur but does not necessarily occur, and this expression means that it includes both the case where the event or situation occurs and the case where it does not occur. For example, "a heterocyclic group optionally substituted by an alkyl group" means that an alkyl group may or may not be present, and this expression includes the case where the heterocyclic group is substituted by an alkyl group and the case where the heterocyclic group is not substituted by an alkyl group.

[0016] "Substituted" means that one or more hydrogen atoms in the group, preferably 5, more preferably 1 to 3 hydrogen atoms, are independently substituted by the corresponding number of substituents. The substituents are only at their possible chemical positions, and it is needless to say that a person skilled in the art can determine possible or impossible substitutions (by experiment or theory) without undue effort. For example, an amino group or a hydroxyl group having free hydrogen may become unstable when bonded to a carbon atom having an unsaturated (e.g., ethylenic) bond. Examples of substituents include halogen, cyano, nitro, oxo, -SF5, C 1-4 alkyl, C 3-7 cycloalkyl, 4- to 7-membered heterocyclyl, phenyl, 5- to 6-membered heteroaryl, etc., but are not limited thereto.

[0017] "Isomer" means a compound having the same molecular formula but different in the nature or order of atomic bonds or the spatial arrangement of atoms. Isomers with different spatial arrangements of atoms are called "stereoisomers". Stereoisomers include optical isomers, geometric isomers, conformational isomers, etc.

[0018] The compounds of the present invention may exist in the form of optical isomers. Optical isomers include enantiomers and diastereoisomers. Enantiomers are two stereoisomers that are mirror images of each other but cannot be superimposed. A racemic mixture or racemate means a mixture of equal amounts of left-hand and right-hand enantiomers of a chiral molecule. Diastereoisomers mean two stereoisomers that are not mirror images of each other and cannot be superimposed. If an optical isomer is a single isomer and its absolute configuration is determined, it will have an absolute configuration of "R" or "S" according to the arrangement of substituents on the chiral carbon atom; if the absolute configuration of the optical isomer is not determined, it will be (+) or (-) according to the measured optical rotation. Methods for preparing and separating optical isomers are known in the art.

[0019] The compounds of the present invention may exist as geometric isomers. The present invention considers various geometric isomers and mixtures thereof arising from the distribution of substituents around carbon-carbon double bonds, carbon-nitrogen double bonds, cycloalkyl groups, or heterocyclic groups. Substituents around carbon-carbon double bonds or carbon-nitrogen bonds are assigned a Z or E configuration, and substituents around cycloalkyl or heterocyclic groups are assigned a cis or trans configuration.

[0020] Furthermore, the compounds of the present invention may exhibit tautomerism, such as keto-enol tautomerism.

[0021] It should be understood that the present invention includes any tautomer or stereoisomer and mixtures thereof, and is not limited to any one tautomer or stereoisomer used in the nomenclature or chemical structure of the compound.

[0022] "Isotope" refers to all isotopes of atoms present in the compound of the present invention. Isotopes include atoms that have the same atomic number but different mass numbers. Examples of isotopes suitable for incorporation into the compound of the present invention include hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, for example, 2 H(D), 3 H, 13 C,14 C, 15 N, 17 O, 18 O, 31 P, 32 P, 35 S, 18 F and 36 Cl is present, but is not limited to it. The isotope-labeled compounds of the present invention can generally be prepared by methods similar to those of the prior art known to those skilled in the art or by methods of the appended examples, using a suitable isotope-labeled reagent instead of a non-isotope-labeled reagent. Such compounds have a variety of potential applications, for example, as standards and reagents in assays of biological activity. In the case of stable isotopes, such compounds have the potential to favorably alter their biological, pharmacological, or pharmacokinetic properties. Deuterium (D) is a preferred isotope of the present invention, and for example, hydrogen in methyl, methylene, or methine may be replaced by deuterium.

[0023] The compounds of the present invention may be administered in the form of prodrugs. A “prodrug” means a derivative that is converted to the bioactive compound of the present invention by oxidation, reduction, hydrolysis, etc. (either using enzymes or without enzyme involvement, respectively) under physiological conditions in vivo. Examples of prodrugs include the following compounds: those in which the amino group of the present invention is acylated, alkylated, or phosphorylated, e.g., eicosanoylamino, alanylamino, pivaloyloxymethylamino; or those in which the hydroxyl group is converted to acylated, alkylated, phosphorylated, or borate, e.g., acetoxy, palmitoyloxy, pivaloyloxy, succinyloxy, fumaryloxy, alanyloxy; or those in which the carboxyl group is esterified or amidated; or those in which the sulfhydryl group forms a disulfide bridge with a carrier molecule such as a peptide that selectively delivers the drug to the target and / or cell cytosol. These compounds can be prepared from the compounds of the present invention according to known methods.

[0024] "Pharmacologically acceptable salt" means a salt prepared from a pharmaceutically acceptable base or acid, including inorganic bases or acids and organic bases or acids. If a compound of the present invention contains one or more acidic or basic groups, the present invention also includes the corresponding pharmaceutically acceptable salts thereof. Compounds of the present invention containing acidic groups can therefore exist in the form of salts and can be used according to the present invention, for example, as alkali metal salts, alkaline earth metal salts or ammonium salts. More specific examples of such salts are sodium salts, potassium salts, calcium salts, magnesium salts or salts with ammonia or organic amines, such as ethylamine, ethanolamine, triethanolamine or amino acids. Compounds of the present invention containing basic groups can exist in the form of salts and can be used according to the present invention in the form of addition salts with inorganic or organic acids. Examples of suitable acids include hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, nitric acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, oxalic acid, acetic acid, tartaric acid, lactic acid, salicylic acid, and benzoic acid. These include formic acid, propionic acid, pivalic acid, malonic acid, succinic acid, pimelic acid, fumaric acid, maleic acid, malic acid, sulfamic acid, phenylpropionic acid, gluconic acid, ascorbic acid, isonicotinic acid, citric acid, adipic acid, and other acids known to those skilled in the art. Where the compounds according to the present invention contain both an acidic and a basic group in the molecule, the present invention includes, in addition to the salt forms mentioned, internal salts or betaines. Individual salts can be obtained by conventional methods known to those skilled in the art, for example, by contacting them with an organic or inorganic acid or base in a solvent or dispersion medium, or by anion exchange or cation exchange with other salts.

[0025] "Pharmaceutical composition" means a composition comprising one or more compounds of the present invention or pharmaceutically acceptable salts thereof, stable isotopic derivatives, isomers, prodrugs or mixtures thereof, and other components such as pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to facilitate administration to a living organism, facilitate the absorption of the active ingredient, and consequently exert biological activity.

[0026] Therefore, when the terms “compound,” “compound of the present invention,” or “compound according to the present invention” are used in this application, all forms of the compound, such as pharmaceutically acceptable salts, stable isotopic derivatives, isomers, prodrugs, or mixtures thereof, are included.

[0027] In this specification, “cancer / tumor” includes, but is not limited to, gastrointestinal cancers, colon cancers, liver cancers, breast cancers, ovarian cancers, prostate cancers, head and neck cancers, skin cancers, lymphomas, leukemias (including acute myeloid leukemia and chronic myeloid leukemia), kidney cancers, lung cancers, muscle cancers, bone cancers, bladder cancers, brain tumors, melanomas, multiple myelomas, and angiodysplasia-related diseases / tumors.

[0028] In this specification, "inflammatory diseases or autoimmune diseases" include arthritis, Hashimoto's thyroiditis, autoimmune hemolytic anemia, autoimmune atrophic gastritis with pernicious anemia, autoimmune encephalomyelitis, autoimmune orchitis, Goodpasture disease, autoimmune thrombocytopenia, sympathetic ophthalmitis, myasthenia gravis, Graves' disease, primary biliary cirrhosis, hepatitis, primary sclerosing cholangitis, chronic invasive hepatitis, non-alcoholic fatty liver disease, non-alcoholic fatty liver disease, ulcerative colitis, membranous glomerulosis, systemic lupus erythematosus, rheumatoid arthritis, psoriatic arthritis, Sjögren's syndrome, Reiter's syndrome, polymyositis, dermatomyositis, type I interferon disease (Aicardi-Goutieres syndrome), and overexpression type I interferon Other systemic sclerosis caused by feron, Mendelian disease, polyarteritis nodosa, multiple sclerosis, relapsing-remitting multiple sclerosis, primary progressive multiple sclerosis, secondary progressive multiple sclerosis, bullous pemphigoid, Crohn's disease, ulcerative colitis, inflammatory bowel disease, celiac disease, proctitis, eosinophilic gastroenteritis, mastocytosis, atopic dermatitis, eczema, psoriasis, scleroderma, prurigo or other pruritus, vitiligo, alopecia areata, Cogan's syndrome, ankylosing spondylitis, Wegener's granulomatosis, autoimmune alopecia, type 1 or juvenile diabetes mellitus, O-cell (humoral) or T-cell based autoimmune diseases including thyroiditis, allergic dermatitis, summer eczema, horseshoe papules. Horseshoes), convulsions, inflammatory airway diseases, recurrent airway obstruction, airway hypersensitivity and allergic diseases of chronic obstructive pulmonary disease, chronic or severe asthma, delayed-onset asthma, bronchitis, bronchial asthma, allergic asthma, endogenous asthma, exogenous asthma, asthma including dust mite-induced asthma, and other obstructive airway diseases.

[0029] In this specification, “therapeutic dose” means an amount of the compound of the present invention that is effective in treating or preventing a disease.

[0030] In this specification, “patient” means a mammal, in particular a human.

[0031] The present invention relates to a JAK inhibitor with general formula (I):

[0032] [ka] [In the formula, Bond a is either a single bond or a double bond. R 1 and R 2 These are H, D, and CN Sofa C 1-6 Alkyl or C 3-6 Selected from cycloalkyl groups, wherein one or more hydrogens of the alkyl group may be optionally substituted with D or fluoro. A is C 3-10 Cycloalkyl, 4-10 membered heterocyclyl, C 6-10 An aryl or 5-10 membered heteroaryl, wherein one or more hydrogens of the cycloalkyl, heterocyclyl, aryl, and heteroaryl are optionally D, halogen, cyano, -OR a , -NR a R b , -C(O)R a -C(O)NR a R b -S(O)2R a -P(O)(CH3)2, C 1-6 Alkyl, C 3-6 They may be substituted with substituents selected from cycloalkyl groups, 4- to 8-membered heterocyclines, and 5- to 6-membered heteroaryl groups. B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally D, halogen, cyano, OR a , -NR a R b ,-COOR a , -C(O)R a , -NR a C(O)R b -C(O)NR a R b -S(O)2R a -S(O)2NR a R b -S(O)(NR a )R b, -P(O)(CH3)2 and R 11 They may be substituted with substituents selected from the following: R 11 is C 1-6 Alkyl, C 3-6 A cycloalkyl, a 4- to 8-membered heterocyclyl, or a 5- to 6-membered heteroaryl, wherein one or more hydrogen atoms of the alkyl, cycloalkyl, heterocyclyl, and heteroaryl are optionally D, halogen, CN, -OH, -NH2, C 1-6 Alkyl, -OC 1-6 Alkyl, -COOR a , -C(O)R a and -C(O)NR a R b They may be substituted with substituents selected from the following: R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyl or 4- to 8-membered heterocyclines, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocycline are optionally D, halogen, or C 1-6 [May be substituted with alkyl groups] The present invention provides compounds represented by [formula], or pharmaceutically acceptable salts thereof, stable isotopic derivatives, isomers, and prodrugs.

[0033] In one preferred embodiment, R 1 and R 2 Both are H.

[0034] In one preferred embodiment, A is C 3-8 A cycloalkyl, a 4- to 8-membered heterocyclyl, a phenyl, or a 5- to 6-membered heteroaryl, wherein one or more hydrogens of the cycloalkyl, heterocyclyl, phenyl, and heteroaryl are optionally a halogen, -C(O)R a , C 1-6 Alkyl and C 3-6 It is substituted with substituents selected from cycloalkyl groups.

[0035] In a more preferred embodiment, A is phenyl, where one or more hydrogen atoms of phenyl are optionally substituted with halogens.

[0036] In one embodiment, B is a phenyl or a 5-6 membered heteroaryl group, wherein one or more hydrogen atoms of the phenyl and heteroaryl groups may be halogens, -COOR a , -C(O)R a -C(O)NR a R b -S(O)2R a , C 1-6 Alkyl, C 3-6 Substituting with substituents selected from cycloalkyl, N, S and / or O heteroatoms, and one or more hydrogens of the alkyl, cycloalkyl and heterocyclyl groups may be C 1-6 Alkyl, -C(O)R a and -C(O)NR a R b It is further substituted with substituents selected from the following.

[0037] In one embodiment, R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyls or 4-6 membered heterocyclies containing N, S and / or O heteroatoms, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocyclyl are optionally C 1-6 It is substituted with alkyl.

[0038] In some embodiments, the compound represented by formula (I) is represented by formula (II): [ka] [In the formula, A is C 3-8Cycloalkyl, 4- to 8-membered heterocyclyl, phenyl, or 5- to 6-membered heteroaryl, provided that one or more hydrogens of said cycloalkyl, heterocyclyl, phenyl, and heteroaryl may optionally be halogen, -C(O)R a , C 1-6 alkyl, and C 3-6 cycloalkyl, and may be substituted with a substituent selected from In a preferred embodiment, A is phenyl, provided that one or more hydrogens of said phenyl may optionally be substituted with halogen, B is phenyl or 5- to 6-membered heteroaryl, provided that one or more hydrogens of said phenyl and heteroaryl may optionally be halogen, -COOR a , -C(O)R a , -C(O)NR a R b , -S(O)2R a , C 1-6 alkyl, C 3-6 cycloalkyl, a 4- to 8-membered heterocyclyl containing a heteroatom of N, S and / or O, and may be substituted with a substituent selected from the group consisting of, and one or more hydrogens of said alkyl, cycloalkyl and heterocyclyl may optionally be C 1-6 alkyl, -C(O)R a [[ID=​​​​​​​​​​​​​​​​​​​​​​​ [ka] [In the formula, A is phenyl, wherein one or more hydrogen atoms of the phenyl may be substituted with halogens as applicable. B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally halogens, -COOR a , -C(O)R a -C(O)NR a R b -S(O)2R a , C 1-6 Alkyl, C 3-6 The substituents may be substituted with substituents selected from cycloalkyl, 4- to 8-membered heterocyclines containing N, S, and / or O heteroatoms, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocyclines may be C 1-6 Alkyl, -C(O)R a and -C(O)NR a R b They may be further substituted with substituents selected from, R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyls or 4-6 membered heterocyclines containing N, S and / or O heteroatoms, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocyclines may be C 1-6 [Alkyl substitution] It is a compound represented by [formula].

[0040] The present invention also relates to the following compounds 1 to 40, or pharmaceutically acceptable salts, stable isotopic derivatives, isomers, prodrugs, and mixtures thereof. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 1-9] [Table 1-10]

[0041] The compounds of the present invention can effectively inhibit the activity of JAK, preferably IC. 50 It is less than 100 nM, and more preferably IC 50 It is less than 10 nM.

[0042] The present invention also relates to a pharmaceutical composition comprising a compound represented by general formula (I), or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, an isomer, and a prodrug, and one or more pharmaceutically acceptable carriers or excipients.

[0043] In another embodiment, the present invention provides a method for treating or preventing a JAK-mediated disease, comprising administering to a patient in need a therapeutically effective amount of a compound represented by general formula (I), or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, an isomer, a prodrug, or a mixture thereof, or a pharmaceutical composition containing said compound. Examples of said diseases include, but are not limited to, inflammatory diseases, autoimmune diseases, and cancers, particularly inflammatory bowel disease, dermatitis, eczema, rheumatoid arthritis, systemic lupus erythematosus, psoriasis, and alopecia areata.

[0044] According to the present invention, the drug may be any pharmaceutical dosage form, including but not limited to tablets, capsules, solutions, lyophilized preparations, and injections.

[0045] The pharmaceutical formulation of the present invention may be administered in the form of dosage units containing a predetermined amount of the active ingredient per dose unit. Such units may contain, for example, 0.5 mg to 1 g, preferably 1 mg to 700 mg, and particularly preferably 5 mg to 500 mg of the compound of the present invention, depending on the condition being treated, the method of administration, and the patient's age, weight, and condition. Furthermore, this type of pharmaceutical formulation may be prepared using methods known in the field of pharmacy, such as mixing the active ingredient with one or more excipients and / or adjuvants.

[0046] The pharmaceutical formulations of the present invention can be adapted for administration by any desired and appropriate method, for example, by oral (including oral or sublingual), rectal, nasal, topical (including oral, sublingual, or transdermal), vaginal, or parenteral (including subcutaneous, intramuscular, intravenous, or intradermal) administration.

[0047] The present invention also provides a method for preparing the aforementioned compounds. The preparation of the compounds represented by the general formula (I) of the present invention can be achieved by the following exemplary methods and examples, but these methods and examples should not be considered to limit the scope of the present invention in any way. The compounds described in the present invention can also be synthesized by synthetic techniques known to those skilled in the art, or by using a combination of methods known to those skilled in the art and the methods of the present invention. Products obtained in each reaction step include, but are not limited to, extraction, filtration, distillation, crystallization, chromatographic separation, etc. They can be obtained by separation techniques known to those skilled in the art. The starting materials and chemical reagents required for synthesis can be synthesized conventionally according to the literature (available on SciFinder) or purchased.

[0048] Synthesis method The heterocyclic compounds represented by the general formula (I) of the present invention can be prepared by the following scheme: 1) condensation of carboxylic acid A1 and amine A2 to obtain A3; 2) dehydration cyclization of A3 under acidic conditions to obtain A4; 3) bromination of A4 with NBS to obtain A5; 4) reaction of A5 with sodium methoxide to obtain A6; 5) Bulkwald coupling reaction of A6 and amine B-NH2 to obtain A7; and 6) deprotection of A7 to obtain A8. Several functional groups FG1 and FG2 of A7 and A8 can be further derivatized and converted into other target compounds, for example, FG1 may contain protected amines, deprotected amines, amides or substituted amines produced by further amidation or reductive amination of amines, and the ester of FG2 may be hydrolyzed with a base (e.g., LiOH) to produce an acid, which can then be amidated to produce an amide.

[0049] [ka]

[0050] The heterocyclic compounds represented by the general formula (I) of the present invention can also be prepared by the following scheme: 1) They can be synthesized by hydrogenating A8 to obtain B1. Several functional groups FG1 and FG2 of B1 can be further derivatized and converted into other target compounds, for example, by hydrolyzing the ester of FG2 with a base (e.g., LiOH) to produce an acid, and then amidating the acid to produce an amide.

[0051] [ka] [Examples]

[0052] The starting materials of the present invention can be synthesized according to methods known in the art, or can be purchased from chemical companies, such as ABCR GmbH&Co. KG, Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc., and Beijing OUHE technology Co. Ltd.

[0053] The structures of the compounds of the present invention were identified by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR identification was performed using a Bruker ASCEND-400 nuclear magnetic analyzer, with solvents such as deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), or deuterated methanol (CD3OD), and tetramethylsilane (TMS) as an internal standard. -6 Chemical shifts were obtained in units of ppm. Mass spectrometry (MS) identification was performed using an Agilent SQD(ESI) mass spectrometer (Agilent 6120).

[0054] HPLC was performed using either an Agilent 1260 DAD high-pressure liquid chromatography (Poroshell 120 EC-C18, 50×3.0 mm, 2.7 μm chromatographic column) or a Waters Arc high-pressure liquid chromatography (Sunfirc C18, 150×4.6 mm, 5 μm chromatographic column).

[0055] Unless otherwise specified in the examples, the reaction temperature was room temperature (20°C to 30°C).

[0056] Unless otherwise specified in the examples, the reactions were carried out under an argon or nitrogen atmosphere. An argon or nitrogen atmosphere meant that the reaction vessel was connected to an argon or nitrogen balloon with a volume of approximately 1 L.

[0057] A hydrogen atmosphere means repeating the process of evacuating the reaction vessel and then filling it with hydrogen three times, and then connecting it to a hydrogen balloon with a volume of approximately 1 liter.

[0058] A CEM Discover-SP type microwave reactor was used for the microwave reaction.

[0059] The progress of the reaction in the examples was monitored using an Agilent liquid chromatograph (1260 / 6120). Alternatively, thin-layer chromatography (TLC) with a silica gel plate thickness of 0.15–0.2 mm (Qingdao Haiyang GF254) could also be used.

[0060] The compounds were purified using column chromatography or thin-layer chromatography. For column chromatography, 200-300 mesh silica gel from Qingdao Haiyang was used, and for thin-layer chromatography, GF254 silica gel plates with a thickness of 0.4-0.5 mm from Qingdao Haiyang were used.

[0061] The solvent systems used for developing compounds in column chromatography or thin-layer chromatography were typically a) dichloromethane and methanol systems, b) petroleum ether and ethyl acetate systems, or those shown in the examples. The volume ratio of the solvents was adjusted according to the polarity of the compound and could be further adjusted by adding small amounts of triethylamine or other acidic or basic reagents.

[0062] Furthermore, the compounds were purified using Waters' mass spectrometer-guided automated preparation system (mass spectrometer detector: SQD2). Depending on the polarity of the compound, an appropriate acetonitrile / water (containing 0.1% trifluoroacetic acid or formic acid, or 0.05% aqueous ammonia) gradient was used to elute a reverse-phase high-pressure column (XBridge-C18, 19×150mm, 5μm) at a flow rate of 20 mL / min. In some of the examples, after purification using the automated preparation system, 1N dilute hydrochloric acid was added, and the solvent was removed under reduced pressure to produce the hydrochloride salt.

[0063] The abbreviation DMF stands for N,N-dimethylformamide.

[0064] The abbreviation TFA stands for trifluoroacetic acid.

[0065] The abbreviation DIPEA stands for N,N-diisopropylethylamine.

[0066] The abbreviation NBS stands for N-bromosuccinimide.

[0067] The abbreviation HATU stands for O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate.

[0068] The abbreviation XantPhos stands for 4,5-bis(diphenylphosphin)-9,9-dimethylxanthene.

[0069] The abbreviation Pd2(dba)3 stands for Tris(dibenzylideneacetone)dipalladium.

[0070] The abbreviation Brettphos stands for 2-(dicyclohexylphosphino)-3,6-dimethoxy-2',4',6'-triisopropyl-1,1'-biphenyl.

[0071] Example 1 1-Bromo-3-(2,6-difluorophenyl)-8-methoxyimidazo[1,5-a]pyrazine (intermediate 1f) [ka]

[0072] 1st step N-((3-chloropyrazine-2-yl)methyl)-2,6-difluorobenzamide(1c) 2,6-Difluorobenzoic acid 1a (5 g, 31.6 mmol) was dissolved in dichloromethane (100 mL), 3 drops of DMF were added, the mixture was cooled to 0°C, and oxalyl chloride (8 g, 63.3 mmol) was added dropwise. After stirring at room temperature for 1 hour, the mixture was concentrated to dryness. The residue was dissolved in dichloromethane (10 mL) to obtain solution A. In another 250 mL round-bottom flask, (3-chloropyrazine-2-yl)methaneamine hydrochloride 1b (5.7 g, 31.6 mmol) was added, dichloromethane (100 mL) and triethylamine (9.6 g, 94.8 mmol) were added, the mixture was cooled to 0°C, and solution A was added dropwise. After stirring the mixture at room temperature for 1 hour, it was quenched with water (50 mL). The separated organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (ethyl acetate / dichloromethane = 1 / 1) to obtain the target product 1c (6.8 g, 76%).

[0073] MS m / z (ESI): 284 [M+1] 1H NMR (400 MHz, CDCl3) δ 8.47 (d, J = 2.5 Hz, 1H), 8.39 - 8.30 (m, 1H), 7.44 - 7.37 (m, 2H), 7.03 - 6.90 (m, 2H), 4.93 (d, J = 4.5 Hz, 2H).

[0074] 2nd process 8-Chloro-3-(2,6-difluorophenyl)imidazo[1,5-a]pyrazine(1d) To a solution of 1c (6.8 g, 24 mmol) in acetonitrile (80 mL), phosphine oxychloride (18.4 g, 120 mmol) was added. The mixture was heated to 90°C and stirred for 16 hours. After cooling to room temperature, another 18.4 g, 120 mmol of phosphine oxychloride was added, and the mixture was heated again to 90°C and stirred for 28 hours. After cooling to room temperature, the mixture was concentrated to dryness, then saturated sodium bicarbonate solution (50 mL) was added, and the mixture was extracted with dichloromethane (2 × 100 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (ethyl acetate / dichloromethane = 1 / 2) to obtain the target product 1d (5.9 g, 92%).

[0075] MS m / z (ESI): 266 [M+1] 1 H NMR (400 MHz, CDCl3) δ 8.10 (s, 1H), 7.60 - 7.50 (m, 2H), 7.44 (d, J = 5.0 Hz, 1H), 7.14 (t, J = 8.1 Hz, 2H).

[0076] 3rd process 1-Bromo-8-chloro-3-(2,6-difluorophenyl)imidazo[1,5-a]pyrazine(1e) NBS (590 mg, 3.31 mmol) was added to a solution of 1d (880 mg, 3.31 mmol) in acetonitrile (40 mL). The mixture was stirred at room temperature for 18 hours and then concentrated to dryness. The residue was dissolved in ethyl acetate (100 mL), washed with water (2 × 50 mL), and then dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated to dryness under reduced pressure, and the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 2 / 1) to obtain the target product 1e (940 mg, 82%).

[0077] MS m / z (ESI): 344 [M+1] 1 H NMR (400 MHz, CDCl3) δ 7.57 (tt, J = 8.5, 6.3 Hz, 1H), 7.49 - 7.46 (m, 1H), 7.41 (d, J = 5.0 Hz, 1H), 7.13 (t, J = 8.1 Hz, 2H).

[0078] 4th step 1-Bromo-3-(2,6-difluorophenyl)-8-methoxyimidazo[1,5-a]pyrazine(1f) 1e (590 mg, 1.71 mmol) was dissolved in methanol (40 mL), cooled to 0°C, and sodium methoxide solution (463 mg, 2.57 mmol, 30% methanol solution) was added dropwise. The resulting mixture was stirred for 2 hours, then poured into saturated ammonium chloride solution (100 mL), added, and extracted with ethyl acetate (3 × 50 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 1) to obtain the target product 1f (570 mg, 98%).

[0079] MS m / z (ESI): 340 [M+1] 1H NMR (400 MHz, CDCl3) δ 7.56-7.49 (m, 1H), 7.21 (dt, J = 5.0, 1.9 Hz, 1H), 7.17 (d, J = 5.1 Hz, 1H), 7.13 - 7.08 (m, 2H), 4.18 (s, 3H) For all intermediates in the table below, the experimental procedure was carried out in accordance with Example 1, except that in the first step, 2,6-difluorobenzoic acid 1a was replaced with a different carboxylic acid. [Table 2]

[0080] The nuclear magnetic data for intermediates 3f and 19a were as follows: [Table 3]

[0081] Example 2 2-(4-aminophenyl)-2-methylpropanamide (intermediate 33c) [ka]

[0082] 1st step 2-Methyl-2-(4-nitrophenyl)propanamide (33b) To a solution of 2-methyl-2-(4-nitrophenyl)propionitrile 33a (2.00 g, 10.52 mmol) in ethanol / water (20 mL, 1 / 1 volume ratio), potassium carbonate (0.58 g, 4.21 mmol) and 30% aqueous hydrogen peroxide solution (35 mL) were added at 0°C. The mixture was stirred at 0°C for 0.5 hours, then raised to room temperature and stirred for 16 hours. After the reaction was complete, saturated sodium sulfite solution (50 mL) was added, and the mixture was stirred at room temperature for 0.5 hours. The mixture was then extracted with ethyl acetate (3 × 50 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 20 / 1) to obtain the target product 33b (0.76 g, 35%).

[0083] MS m / z (ESI): 209 [M+1] 2nd process 2-(4-aminophenyl)-2-methylpropanamide(33c) To a solution of 33b (0.76 g, 3.65 mmol) in methanol (10 mL), 20% palladium-carbon (0.15 g) was added. The mixture was stirred under a hydrogen atmosphere for 12 hours and then filtered. The filtrate was concentrated to dryness under reduced pressure to obtain the target product 33c (0.65 g, 100%).

[0084] MS m / z (ESI): 179 [M+1] Example 3 tert-butyl 4-(2-(4-aminophenyl)-2-methylpropanoyl)piperazine-1-carboxylate (intermediate 36d) [ka]

[0085] 1st step 2-Methyl-2-(4-nitrophenyl)propanoic acid (36a) A mixture of 33a (5.00 g, 26.29 mmol) and water (25 mL) was mixed with concentrated sulfuric acid (25 mL) at 0°C, then heated to 120°C and stirred for 12 hours. After cooling to room temperature, water (50 mL) was added, the pH was adjusted to 8 with saturated sodium bicarbonate solution, and then extracted with ethyl acetate (3 × 50 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure to obtain the target product 36a (5.00 g, 91%).

[0086] MS m / z (ESI): 210 [M+1]

[0087] 2nd process 2-Methyl-2-(4-nitrophenyl)propionyl chloride (36b) 36a (5.00 g, 23.90 mmol) was added to thionyl chloride at 0°C, then heated to 85°C and stirred for 12 hours. After cooling to room temperature, the mixture was concentrated to dryness to obtain the target product 36b (5.20 g, 96%).

[0088] 3rd process tert-butyl 4-(2-methyl-2-(4-nitrophenyl)propionyl)piperazine-1-carboxylate To a solution of tert-butyl-piperazine-1-carboxylate (1.84 g, 9.89 mmol) in dichloromethane (20 mL), triethylamine (2.00 g, 19.77 mmol) was added at room temperature and the mixture was stirred for 10 minutes. The mixture was then cooled to 0°C, and a solution of 36c (1.50 g, 6.59 mmol) in dichloromethane (10 mL) was added. After stirring at room temperature for 2 hours, water (20 mL) was added, and the mixture was extracted with dichloromethane (3 × 30 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 1) to obtain the target product 36c (1.50 g, 60%).

[0089] MS m / z (ESI): 378 [M+1] 4th step tert-butyl4-(2-(4-aminophenyl)-2-methylpropanoyl)piperazine-1-carboxylate(36d) To a solution of 36c (1.50 g, 3.97 mmol) in methanol (250 mL), 20% palladium-carbon (0.3 g) was added, and the mixture was stirred under a hydrogen atmosphere for 12 hours. The mixture was filtered, and the filtrate was concentrated to dryness to obtain the target product 36d (1.38 g, 100%).

[0090] MS m / z (ESI): 348 [M+1]

[0091] Intermediate 35d was synthesized by referring to the experimental procedure of Example 3, but using N-methylpiperidine instead of tert-butyl-piperazine-1-carboxylate in the third step. [Table 4]

[0092] Example 4 3-(2,6-difluorophenyl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one (compound 1) [ka]

[0093] 1st step (4-((3-(2,6-difluorophenyl)-8-methoxyimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)(morpholino)methanone(1h) A mixture of 1f (200 mg, 0.59 mmol), (4-aminophenyl)(morpholino)methanone 1 g (121 mg, 0.59 mmol), and 1,4-dioxane (2 mL) was mixed with tert-butoxide sodium (144 mg, 1.5 mmol), followed by XantPhos (69 mg, 0.12 mmol) and Pd2(dba)3 (55 mg, 0.06 mmol). The mixture was heated to 100°C in a microwave reactor and stirred for 1 hour. After cooling to room temperature, the reaction mixture was diluted with water (20 mL) and then extracted with ethyl acetate (3 × 20 mL). The combined organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 20 / 1) to obtain the target product 1h (260 mg, 95%).

[0094] MS m / z (ESI): 466 [M+1] 2nd process 3-(2,6-difluorophenyl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one(1) To a solution of 260 mg (0.56 mmol) of acetonitrile in 10 mL, 10 mL of 6 N hydrochloric acid was added, and the mixture was heated to 70°C and stirred for 1 hour. After cooling to room temperature, the mixture was concentrated to dryness, and the residue was diluted with water (10 mL). The pH was then adjusted to 8 with saturated sodium bicarbonate solution, and the mixture was extracted with ethyl acetate (2 × 50 mL). The combined organic layers were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 20 / 1) to obtain target product 1 (solid, 150 mg, 59%).

[0095] MS m / z (ESI): 452 [M+1] 1H NMR (400 MHz, DMSO-d6) δ 10.56 (d, J = 5.3 Hz, 1H), 8.42 (s, 1H), 7.78 - 7.63 (m, 3H), 7.43 - 7.27 (m, 4H), 6.84 (d, J = 5.3 Hz, 1H), 6.59 (t, J = 5.7 Hz, 1H), 3.59 (d, J = 4.5 Hz, 4H), 3.50 (s, 4H). For all the compounds or intermediates listed in the table below, the experimental procedure in Example 4 was followed, except that in the first step, 1f and 1g were replaced with different compounds. [Table 5-1] [Table 5-2] [Table 5-3]

[0096] The nuclear magnetic data for compounds 3, 4, 6, 7, 9, 11, 14, 22, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, and 38 were as follows: [Table 6-1] [Table 6-2] [Table 6-3]

[0097] Example 5 3-(2,6-difluorophenyl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)-6,7-dihydroimidazo[1,5-a]pyrazine-8(5H)-one (compound 2) [ka] After mixing 1 (97 mg, 0.215 mmol), methanol (30 mL), and 10% palladium-carbon (97 mg), the mixture was heated to 30°C under a hydrogen atmosphere and stirred for 16 hours. After filtration, the filtrate was concentrated to dryness, and the residue was purified by reverse-phase preparative high-performance liquid chromatography to obtain target product 2 (solid, 25 mg, 26%).

[0098] MS m / z (ESI): 490 [M+1] 1 H NMR (400 MHz, CD3OD) δ 7.73 (d, J = 6.3 Hz, 1H), 7.66 (td, J = 6.2, 2.8 Hz, 3H), 7.58 (d, J = 8.3 Hz, 2H), 7.28 (ddd, J = 9.1, 7.6, 3.3 Hz, 1H), 7.19 (dd, J = 9.1, 4.2 Hz, 1H), 6.94 (d, J = 6.4 Hz, 1H), 4.73 (s, 2H), 4.52 (t, J = 11.5 Hz, 1H), 3.99 (s, 3H), 3.76 -3.70 (m, 1H), 2.21 -2.03 (m, 6H), 1.63 - 1.53 (m, 2H).

[0099] For all the compounds in the table below, the experimental procedure was carried out in accordance with Example 5, except that a different compound was used instead of compound 1 during the procedure. [Table 7]

[0100] The nuclear magnetic data for compounds 5, 8, 10, 12, 16, 18, and 23 were as follows: [Table 8]

[0101] Example 6 1-((4-(1-(azetidine-1-yl)-2-methyl-1-oxopropan-2-yl)phenyl)amino)-3-(2,6-difluorophenyl)-6,7-dihydroimidazo[1,5-a]pyrazine-8(5H)-one (compound 13) [ka]

[0102] 1st step tert-butyl 2-(4-((3-(2,6-difluorophenyl)-8-methoxyimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)-2-methylpropanoate(13b) To a solution of 1f (200 mg, 0.588 mmol) in 1,4-dioxane (5 mL), tert-butyl 2-(4-aminophenyl)-2-methylpropanoate 13a (280 mg, 1.17 mmol), Brettphos (62 mg, 0.118 mmol), Pd2(dba)3 (53 mg, 0.058 mmol), and cesium carbonate (600 mg, 1.76 mmol) were added under a nitrogen atmosphere. The mixture was then heated to 90°C in a microwave reactor and stirred for 1 hour. After cooling to room temperature, the mixture was concentrated to dryness under reduced pressure, and the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 0~7 / 3) to obtain the target product 13b (200 mg, 68%).

[0103] MS m / z (ESI): 495 [M+1]

[0104] 2nd process Ethyl 2-(4-((3-(2,6-difluorophenyl)-8-oxo-7,8-dihydroimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)-2-methylpropanoate(13c) To a solution of 13b (100 mg, 0.1 mmol) in ethanol (4 mL), a solution of HCl in ethanol (12N, 2 mL) was added. This solution was stirred at 70°C for 2 hours, then cooled to room temperature and concentrated to dryness to obtain the target product 13c (90 mg). The product was used directly in the next reaction without further purification.

[0105] MS m / z (ESI): 453 [M+1]

[0106] 3rd process Ethyl 2-(4-((3-(2,6-difluorophenyl)-8-oxo-5,6,7,8-tetrahydroimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)-2-methylpropanoate(13d) 13c (90 mg, 0.2 mmol), ethanol (5 mL), and 10% palladium carbon (100 mg) were mixed and stirred under a hydrogen atmosphere for 3 hours. The mixture was then filtered, and the filtrate was concentrated to dryness to obtain the target product 13d (80 mg, 88%).

[0107] MS m / z (ESI): 455 [M+1]

[0108] 4th step 2-(4-((3-(2,6-difluorophenyl)-8-oxo-5,6,7,8-tetrahydroimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)-2-methylpropanoic acid (13e) To a solution of 13d (90 mg, 0.2 mmol) in tetrahydrofuran (5 mL), lithium hydroxide monohydrate (17 mg, 0.4 mmol) was added, and the mixture was heated to 60°C and stirred for 18 hours. After cooling to room temperature, the reaction solution was concentrated to dryness, and the residue was purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 13e (containing 60 mg of TFA).

[0109] MS m / z (ESI): 427 [M+1]

[0110] 5th step 1-((4-(1-(azetidine-1-yl)-2-methyl-1-oxopropan-2-yl)phenyl)amino)-3-(2,6-difluorophenyl)-6,7-dihydroimidazo[1,5-a]pyrazine-8(5H)-one(13) Diisopropylethylamine (26 mg, 0.2 mmol) was added to a mixture of 13e (17 mg, 0.04 mmol), DMF (2 mL), HATU (20 mg, 0.052 mmol), and azetidine (10 mg, 0.12 mmol). The resulting solution was stirred at room temperature for 1 hour, and then directly purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 13 (solid, 2.69 mg, 14%).

[0111] MS m / z (ESI): 466 [M+1] 1 H NMR (400 MHz, DMSO-d6) δ 8.04 (s, 1H), 7.81 (s, 1H), 7.75 - 7.65 (m, 1H), 7.59 (d, J = 8.7 Hz, 2H), 7.42 - 7.29 (m, 2H), 7.10 (d, J = 8.7 Hz, 2H), 4.06 - 3.93 (m, 2H), 3.86 - 3.71 (m, 2H), 3.53 - 3.47 (m, 2H), 2.59 - 2.51 (m, 2H), 2.03 - 1.84 (m, 2H), 1.37 (s, 6H).

[0112] Example 7 3-(2-chloro-6-fluorophenyl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)-6,7-dihydroimidazo[1,5-a]pyrazine-8(5H)-one (compound 15) [ka] To a solution of 3 (140 mg, 0.3 mmol) in ethanol (10 mL), chlorobenzene (1 mL) and platinum dioxide (70 mg) were added. The mixture was stirred under a hydrogen atmosphere for 70 minutes and then filtered. The filtrate was concentrated to dryness, and the residue was purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 15 (solid, 62.7 mg, 44%).

[0113] MS m / z (ESI): 312 [M+1] 1 H NMR (400 MHz, DMSO-d6) δ 8.29 (s, 1H), 7.87 (s, 1H), 7.72 - 7.63 (m, 3H), 7.57 (d, J = 8.1 Hz, 1H), 7.47 (t, J = 8.7 Hz, 1H), 7.31 (d, J = 8.6 Hz, 2H), 3.95 - 3.90 (m, 2H), 3.61 - 3.55 (m, 4H), 3.54 - 3.45 (m, 6H).

[0114] Example 8 3-(2,6-difluorophenyl)-1-((4-(2-methyl-1-morpholino-1-oxopropan-2-yl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one (compound 17) [ka]

[0115] 1st step 2-(4-((3-(2,6-difluorophenyl)-8-oxo-7,8-dihydroimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)-2-methylpropanoic acid (17a) A solution of 13b (420 mg, 0.848 mmol) in trifluoroacetic acid (10 mL) was heated to 70°C and stirred for 1 hour. After cooling to room temperature, the reaction mixture was concentrated to dryness under reduced pressure to obtain the target product 17a (360 mg). This product was used directly in the next step without further purification.

[0116] MS m / z (ESI): 425 [M+1]

[0117] 2nd process 3-(2,6-difluorophenyl)-1-((4-(2-methyl-1-morpholino-1-oxopropan-2-yl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one hydrochloride (17) 17a (360 mg, 0.85 mmol), DMF (5 mL), HATU (420 mg, 1.1 mmol), morpholine (220 mg, 2.55 mmol), and DIPEA (442 mg, 3.2 mmol) were mixed. After stirring at room temperature for 1 hour, the reaction mixture was directly purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 17 (solid, 320 mg, 76%, hydrochloride).

[0118] MS m / z (ESI): 494 [M+1] 1 H NMR (400 MHz, DMSO-d6) δ 10.47 (d, J = 5.3 Hz, 1H), 8.12 (s, 1H), 7.78 - 7.67 (m, 1H), 7.64 (d, J = 8.7 Hz, 2H), 7.42 - 7.31 (m, 2H), 7.08 (d, J = 8.7 Hz, 2H), 6.81 (d, J = 5.5 Hz, 1H), 6.62 - 6.49 (m, 1H), 3.69 - 3.41 (m, 8H), 1.40 (s, 6H).

[0119] Example 9 3-(1-acetylpiperidine-4-yl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one (compound 19) [ka]

[0120] 1st step Benzyl 4-(8-methoxy-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-3-yl)piperidine-1-carboxylate(19b) A mixture of 19a (206 mg, 1 mmol), 1,4-dioxane (8 mL), and cesium carbonate (813 mg, 2.5 mmol) was mixed with XantPhos (116 mg, 0.2 mmol) and Pd2(dba)3 (92 mg, 0.1 mmol). The mixture was heated to 90°C in a microwave reactor and stirred for 1 hour. After cooling to room temperature, the mixture was filtered, the filtrate was concentrated to dryness, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 0~19 / 1) to obtain the target product 19b (490 mg, 86%).

[0121] MS M / z(ESI): 571 [M +1]

[0122] 2nd process (4-((8-methoxy-3-(piperidine-4-yl)imidazo[1,5-a]pyrazine-1-yl)amino)phenyl)(morpholino)methanone(19c) To a solution of 19b (230 mg, 0.403 mmol) in methanol (10 mL), 10% palladium-carbon (115 mg) was added. The mixture was stirred under a hydrogen atmosphere for 30 minutes and then filtered. The filtrate was concentrated to dryness to obtain the target product 19c (190 mg). The product was used directly in the next reaction without purification.

[0123] MS m / z (ESI): 437 [M+1]

[0124] 3rd process 1-(4-(8-methoxy-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-3-yl)piperidine-1-yl)ethane-1-one(19d) Triethylamine (85 mg, 0.837 mmol) was added to a solution of 19c (122 mg, 0.279 mmol) in dichloromethane (5 mL). After cooling to 0°C, acetyl chloride (22 mg, 0.279 mmol) was added dropwise. After stirring at room temperature for 30 minutes, the mixture was concentrated to dryness, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 0~19 / 1) to obtain the target product 19d (81 mg, 61%).

[0125] MS m / z (ESI): 479 [M+1]

[0126] 4th step 3-(1-acetylpiperidine-4-yl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one(19) To a solution of 19d (81 mg, 0.17 mmol) in acetonitrile (2 mL), a 33% ethanol solution of HCl (1 mL) was added, and the mixture was heated to 50°C and stirred for 30 minutes. After cooling to room temperature, the reaction mixture was concentrated to dryness to obtain a yellow solid (85 mg). 45 mg was taken out and purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 19 (solid, 31.1 mg, 69%).

[0127] MS m / z (ESI): 465 [M+1] 1 H NMR (400 MHz, CD3OD) δ 7.50 - 7.42 (m, 3H), 7.34 - 7.28 (m, 2H), 6.88 (d, J = 6.1 Hz, 1H), 4.72 (d, J = 13.5 Hz, 1H), 4.14 (d, J = 13.8 Hz, 1H), 3.85 - 3.57 (m, 9H), 3.38 (dd, J = 19.4, 7.5 Hz, 1H), 2.88 (t, J = 11.9 Hz, 1H), 2.19 (s, 3H), 2.17 - 2.06 (m, 2H), 1.97 - 1.75 (m, 2H).

[0128] Example 10 1 - ((4 - (Morpholine - 4 - carbonyl)phenyl)amino)-3 - (piperidin - 4 - yl)imidazo[1,5 - a]pyrazin - 8(7H)-one (Compound 20)

Chemical formula

[0129] MS m / z (ESI): 423 [M + 1] 1 H NMR (400 MHz, CD3OD) δ 7.66 - 7.61 (m, 2H), 7.44 - 7.39 (m, 2H), 7.28 (d, J = 6.1 Hz, 1H), 6.63 (d, J = 6.0 Hz, 1H), 3.72 (s, 8H), 3.62 (t, J = 3.5 Hz, 1H), 3.58 (t, J = 3.5 Hz, 1H), 3.56 - 3.49 (m, 1H), 3.30 - 3.20 (m, 2H), 2.29 - 2.17 (m, 4H).

[0130] Example 11 3 - (1 - Cyclopropylpiperidin - 4 - yl)-1 - ((4 - (morpholine - 4 - carbonyl)phenyl)amino)imidazo[1,5 - a]pyrazin - 8(7H)-one (Compound 21)

Chemical formula

[0131] The first step (4-((3-(1-cyclopropylpiperidine-4-yl)-8-methoxyimidazo[1,5-a]pyrazine-1-yl)amino)phenyl)(morpholino)methanone(21a) To a mixture of 19c (150 mg, 0.344 mmol), methanol (2 mL), and tetrahydrofuran (2 mL), (1-ethoxycyclopropoxy)trimethylsilane (120 mg, 0.687 mmol) was added, followed by the addition of acetic acid (103 mg, 1.72 mmol) and sodium borohydride cyanohydride (32 mg, 0.86 mmol). The mixture was heated to 60°C and stirred for 20 hours. After cooling to room temperature, saturated sodium bicarbonate solution (20 mL) was added, and then the mixture was extracted with ethyl acetate (3 × 20 mL). The combined organic phase was washed with saturated brine (20 mL) and dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated to dryness, and the residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 0~19 / 1) to obtain the target product 21a (46 mg, 28%).

[0132] MS m / z (ESI): 477 [M+1]

[0133] 2nd process 3-(1-cyclopropylpiperidine-4-yl)-1-((4-(morpholine-4-carbonyl)phenyl)amino)imidazo[1,5-a]pyrazine-8(7H)-one(21) To a solution of 21a (46 mg, 0.0965 mmol) in acetonitrile (2 mL), HCl solution (30% ethanol solution, 1 mL) was added, and the mixture was then heated to 50°C and stirred for 30 minutes. After cooling to room temperature, the mixture was concentrated to dryness, and the residue was purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 21 (solid, 16.9 mg, 38%).

[0134] MS m / z (ESI): 463 [M+1] 1H NMR (400 MHz, CD3OD) δ 7.77 (d, J = 8.6 Hz, 2H), 7.38 (d, J = 8.6 Hz, 2H), 7.15 (d, J = 6.0 Hz, 1H), 6.46 (d, J = 6.0 Hz, 1H), 3.71 (s, 4H), 3.68 (s, 4H), 3.28 (d, J = 11.9 Hz, 2H), 3.15-3.06 (m, 1H), 2.57 (t, J = 10.5 Hz, 2H), 2.13 - 1.94(m, 4H), 1.90 (s, 1H), 0.66 - 0.49 (m, 4H).

[0135] Example 12 3-(2,6-difluorophenyl)-1-((1-(2-morpholino-2-oxoethyl)-1H-pyrazole-4-yl)amino)-6,7-dihydroimidazo[1,5-a]pyrazine-8(5H)-one (compound 24) [ka]

[0136] 1st step tert-butyl 2-(4-nitro-1H-pyrazole-1-yl)acetate (24b) A mixture of 4-nitro-1H-pyrazole 24a (2 g, 17.7 mmol), potassium carbonate (4.8 g, 35 mmol), and DMF (10 mmol) was mixed with tert-butyl 2-bromoacetate (4 g, 17.7 mmol) at room temperature. The resulting mixture was stirred at room temperature for 12 hours, then diluted with water (200 mL), and extracted with ethyl acetate (2 × 150 mL). The combined organic phase was washed with saturated brine (2 × 150 mL) and then dried over anhydrous sodium sulfate. After filtration, the filtrate was concentrated to dryness under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 100 / 0~99 / 1) to obtain the target product 24b (5 g, 110%).

[0137] MS m / z (ESI): 228 [M+1]

[0138] 2nd process tert-butyl 2-(4-amino-1H-pyrazole-1-yl)acetate (24c) To a solution of 24b (5 g, 20 mmol) in ethanol (20 mL), 10% palladium carbon (700 mg) was added, and the mixture was stirred under a hydrogen atmosphere for 12 hours, after which it was filtered. The filtrate was concentrated to dryness under reduced pressure to obtain the target product 24c (3.45 g, 77%).

[0139] MS m / z (ESI): 198 [M+1]

[0140] 3rd process tert-butyl2-(4-((3-(2,6-difluorophenyl)-8-methoxyimidazo[1,5-a]pyrazine-1-yl)amino)-1H-pyrazole-1-yl)acetate(24d) A mixture of 1f (600 mg, 1.77 mmol), 1,4-dioxane (15 mL), 24c (720 mg, 3.53 mmol), Brettphos (192 mg, 0.353 mmol), Pd2(dba)3 (180 mg, 0.177 mmol), and cesium carbonate (1.7 g, 5.32 mmol) was heated to 90°C in a microwave reactor under a nitrogen atmosphere and stirred for 1 hour. After cooling to room temperature, the solvent was removed by distillation under reduced pressure, and the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 100 / 0~1 / 4) to obtain the target product 24d (450 mg, 47%).

[0141] MS m / z (ESI): 457 [M+1]

[0142] 4th step 2-(4-((3-(2,6-difluorophenyl)-8-oxo-7,8-dihydroimidazo[1,5-a]pyrazine-1-yl)amino)-1H-pyrazole-1-yl)acetic acid (24e) A solution of 24d (450 mg, 0.98 mmol) in trifluoroacetic acid (10 mL) was heated to 80 °C and stirred for 12 h. After cooling to room temperature, the reaction solution was concentrated to dryness under reduced pressure to obtain the target product 24e (815 mg). The product was used directly in the next reaction without further purification.

[0143] MS m / z (ESI): 387 [M+1]

[0144] Step 5 2-(4-((3-(2,6-Difluorophenyl)-8-oxo-5,6,7,8-tetrahydroimidazo[1,5-a]pyrazin-1-yl)amino)-1H-pyrazol-1-yl)acetic acid (24f) To a solution of 24e (800 mg, 2.07 mmol) in ethanol (30 mL) was added 10% palladium on carbon (300 mg), and then the mixture was stirred under a hydrogen atmosphere for 12 h. After filtration, the filtrate was concentrated to dryness under reduced pressure to obtain the target product 24f (280 mg, 35%).

[0145] MS m / z (ESI): 389 [M+1]

[0146] Step 6 3-(2,6-Difluorophenyl)-1-((1-(2-morpholino-2-oxoethyl)-1H-pyrazol-4-yl)amino)-6,7-dihydroimidazo[1,5-a]pyrazin-8(5H)-one (24) To 24f (10 mg, 0.0257 mmol), HATU (13 mg, 0.0335 mmol), morpholine (0.025 mL) and DMF (0.5 mL) was added DIPEA (10 mg, 0.0771 mmol). After stirring at room temperature for 1 h, the reaction solution was directly purified by reverse-phase preparative high performance liquid chromatography to obtain the target product 24 (solid, 2.12 mg, 18%).

[0147] MS m / z (ESI): 458 [M+1] 1H NMR (400 MHz, DMSO-d6) δ 7.84 - 7.81 (m, 1H), 7.80 (s, 1H), 7.72 - 7.62 (m, 2H), 7.55 (s, 1H), 7.36 - 7.29 (m, 2H), 5.03 (s, 2H), 3.97 - 3.90 (m, 2H), 3.60 - 3.52 (m, 2H), 3.50 - 3.39 (m, 8H).

[0148] Compound 25 was prepared according to the experimental procedure of Example 12, except that in step 6, N-methylpiperazine was used instead of morpholine. [Table 9]

[0149] The nuclear magnetic data for compound 25 was as follows: [Table 10]

[0150] Example 13 4-((3-(2,6-difluorophenyl)-8-oxo-7,8-dihydroimidazo[1,5-a]pyrazine-1-yl)amino)benzoic acid (compound 39) [ka] To a mixed solution of 39a (150 mg, 0.37 mmol) of methanol, tetrahydrofuran, and water (3 / 3 / 1v / v / v, 7 mL), lithium hydroxide monohydrate (155 mg, 3.7 mmol) was added, and the mixture was heated to 50°C and stirred for 12 hours. After cooling to room temperature, the organic solvent was concentrated and removed under reduced pressure. The residue was adjusted to pH = 1 with 1N hydrochloric acid, stirred at room temperature for 5 hours, and then concentrated to dryness. The residue was purified by reverse-phase preparative high-performance liquid chromatography to obtain the target product 39 (solid, 25 mg, 18%).

[0151] MS m / z (ESI): 383 [M+1] 1 H NMR (400 MHz, DMSO-d6) δ 12.36 (s, 1H), 10.61 (d, J = 5.5 Hz, 1H), 8.62 (s, 1H), 7.81 (d, J = 8.8 Hz, 2H), 7.70 (d, J = 8.9 Hz, 3H), 7.39 (d, J = 8.2 Hz, 2H),6.87 (d, J = 5.6 Hz, 1H), 6.60 (s, 1H).

[0152] For compound 40, the procedure was carried out referring to the experimental procedure of Example 13, but 40a was used instead of 39a. [Table 11]

[0153] The nuclear magnetic data for compound 40 were as follows: [Table 12]

[0154] Biological experiments Example 14 JAK2 activity inhibition experiment The effect of the compound of the present invention on the enzyme activity of JAK2 was evaluated in vitro by kinase assay experiments.

[0155] The experimental method was outlined as follows: The enzyme activity of JAK2 was measured by detecting the level of substrate phosphorylation in the kinase reaction using a homogeneous time-resolved fluorescence (HTRF) kinase assay detection kit (Cisbio, 62TK0PEC). The reaction buffer contained the following components: enzyme reaction buffer (1×) included in the kit, 5 mM MgCl2, 1 mM DTT, and 0.01% 0.01% Brij35. Human recombinant JAK2 protein (Carna Biosciences, 08-045) was diluted to 0.15 ng / μL in the reaction buffer to form a kinase solution. The substrate reaction solution contained biotin-labeled tyrosine kinase substrate diluted to 0.25 μM in the reaction buffer and 2.5 μM ATP, and the detection buffer was Eu25 diluted to 0.1 ng / L in the reaction buffer. 3+ The test compound, containing labeled cage antibody (Cisbio, 61T66KLB) and 12.5 nM streptavidin-labeled XL665 (Cisbio, 610SAXLB), was dissolved in DMSO to a concentration of 10 μM, and then further diluted fourfold with DMSO to a minimum concentration of 0.061 nM. Each concentration of the sample was further diluted 40-fold with reaction buffer.

[0156] 4 μL of compound solution and 2 μL of kinase solution were added to a 384-well assay plate (Corning, 3674). After homogeneous mixing, the mixture was incubated at room temperature for 15 minutes, and then 4 μL of substrate reaction solution was added. After further incubation at room temperature for 30 minutes, 10 μL of detection buffer was added to the reaction mixture, homogeneous mixing was performed, and the mixture was left to stand at room temperature for 30 minutes. The progress of the reaction was measured at wavelengths of 620 nm and 665 nm using an Envision plate reader (Perkin Elmer). Signal values ​​(absorbance) were measured. 665 nm / absorbance 620 nm Since the IC50 index positively correlates with the degree of substrate phosphorylation, we detected the activity of JAK2 kinase. In this experiment, the group without JAK2 kinase protein was the 100% inhibition group, and the group with JAK2 kinase protein but without the test compound was the 0% inhibition group. The inhibition curves of the test compound were plotted using XLfit software, and the inhibition rate IC50 was calculated.50 The result was calculated and is shown in Table 1.

[0157] Example 15 TYK2 activity inhibition experiment The effect of the compound of the present invention on TYK2 activity was evaluated in vitro by kinase assay experiments.

[0158] The experimental method was outlined as follows: The enzymatic activity of TYK2 was measured by detecting the level of substrate phosphorylation in the kinase reaction using a homogeneous time-resolved fluorescence (HTRF) kinase assay detection kit (Cisbio, 62TK0PEC). The reaction buffer contained the following components: enzyme reaction buffer (1×) included in the kit, 5 mM MgCl2, 1 mM DTT, and 0.01% 0.01% Brij35. Human recombinant TYK2 protein (Carna Biosciences, 08-147) was diluted to 0.25 ng / μL in the reaction buffer to form a kinase solution. The substrate reaction solution contained biotin-labeled tyrosine kinase substrate diluted to 0.5 μM in the reaction buffer and 11.25 μM ATP, and the detection buffer was Eu25 diluted to 0.1 ng / L in the reaction buffer. 3+ The test compound, containing labeled cage antibody (Cisbio, 61T66KLB) and 25 nM streptavidin-labeled XL665 (Cisbio, 610SAXLB), was dissolved in DMSO to a concentration of 10 μM, and then further diluted fourfold with DMSO until a minimum concentration of 0.061 nM was obtained. Each concentration of the sample was further diluted 40-fold with reaction buffer.

[0159] 4 μL of compound solution and 2 μL of kinase solution were added to a 384-well assay plate (Corning, 3674). After homogeneous mixing, the mixture was incubated at room temperature for 15 minutes, and then 4 μL of substrate reaction solution was added. After further incubation at room temperature for 40 minutes, 10 μL of detection buffer was added to the reaction mixture, homogeneous mixing was performed, and the mixture was left at room temperature for 30 minutes. The progress of the reaction was measured at wavelengths of 620 nm and 665 nm using an Envision plate reader (Perkin Elmer). Signal values ​​(absorbance) were measured. 665 nm / absorbance 620 nm The activity of TYK2 kinase was detected because it positively correlates with the degree of substrate phosphorylation. In this experiment, the group without TYK2 kinase protein was the 100% inhibition group, and the group with TYK2 kinase protein but without the test compound was the 0% inhibition group. The inhibition curves of the test compound were plotted using XLfit software, and the inhibition rate IC was calculated. 50 The result was calculated and is shown in Table 1.

[0160] [Table 13]

[0161] The compounds of the present invention showed inhibitory activity against JAK2 kinase and TYK2 kinase. Preferably IC 50 It is less than 100 nM, and more preferably IC 50 It was less than 10 nM.

[0162] Example 16 Measurement of IL-12-induced IFN-γ secretion inhibition in NK92 cells The effect of the compound of the present invention on IFN-γ secretion in IL-12-induced NK92 cells was evaluated by enzyme-linked immunosorbent assay (ELISA).

[0163] The experimental principle was as follows: IL-12 was expressed in activated T cells, NK cells (NK92 is an NK cell line), DC cells, and B cells. Binding to IL-12 activated the JAK2 / TYK2 signaling pathway in NK cells and T lymphocytes, inducing IFN-γ secretion.

[0164] The experimental method was outlined as follows: The compound was dissolved in DMSO and diluted to 2.5 mM, then diluted fourfold in DMSO to a minimum concentration of 0.31 μM. Each solution at each concentration was further diluted 50-fold in FBS-free MEMα medium (Thermofisher, 12561-056).

[0165] NK92 cells (Nanjing Cobioer, CBP60980) were incubated in complete MEMα medium containing 12.5% ​​FBS (Ausbian, VS500T), 12.5% ​​horse serum (Gibco, 16050-122), 0.02 mM folic acid (Sigma, F8758), 0.2 mM inositol (Sigma, 17850), 0.55 mM β-mercaptoethanol (Thermofish, 21985-023), 200 U / mL IL-2 (R&D Systems, 202-1L), and 100 U / mL penicillin (Thermofisher, 15140122). Once 80-90% of the culture vessel surface was covered, the cells were dispersed and seeded into a 96-well plate (ThermoFisher, 167425) with 100,000 cells per well (80 μL of complete MEMα medium without IL-2). The 96-well plate was then incubated overnight at 37°C / 5% CO2.

[0166] After overnight incubation, 10 μL of diluted test compound and 10 μL of 50 ng / mL IL-12 (R&D Systems, 219-1L) were added to each well and gently mixed. The 96-well plate was then incubated further in a 37°C / 5% CO2 incubator. After 24 hours, the plate was removed and centrifuged at room temperature at 800 rpm for 10 minutes. 50 μL of the supernatant was transferred to a 96-well plate coated with anti-IFN-γ antibody (Sigma, CLS3695). IFN-γ secretion was detected according to the instructions for the Human IFN-γ DuoSet ELISA kit (R&D Systems, DY285B). In the experiment, the group that added IL-12 and the test compound in MEMα medium without IL-12 was designated as the unstimulated control group (100% inhibition), and the group that added IL-12 and 0.2% DMSO was designated as the stimulated group (0% inhibition). The inhibition curves of the test compounds were plotted using XLfit software, and the inhibition rate IC was calculated. 50 The result was calculated and is shown in Table 2.

[0167] [Table 14]

[0168] Example 17 Mouse pharmacokinetic experiments The test compound was formulated in a 20% HP-β-CD vehicle to a dose of 5 mg / mL (suspension or solution).

[0169] Three female C57 mice were orally administered (PO) at a dose of 25 mg / kg of 5 mL / kg of the sample. Four hours after administration, blood was collected, and the mice were executed using CO2. The colon near the rectal end was collected, cut into sections approximately 4-6 cm in length, washed with cold saline solution, dried by suction with absorbent paper, and weighed.

[0170] The concentrations of the test compound in plasma and enterohomogenate samples were quantitatively analyzed by LC-MS / MS using an API-4500 mass spectrometer, and the limit of quantification (LOQ) in plasma was 1 ng / mL. Pharmacokinetic (PK) parameters were calculated using WinNonlin, and the results are summarized in Table 3.

[0171] [Table 15] Furthermore, the present invention includes the following embodiments. [Aspect 1] General formula (I): [ka] [In the formula, Bond a is either a single bond or a double bond. R 1 and R 2 These are H, D, and C, respectively, independently. 1-6 Alkyl or C 3-6 Selected from cycloalkyl groups, wherein one or more hydrogens of the alkyl group may be optionally substituted with D or fluoro. A is C 3-10 Cycloalkyl, 4-10 membered heterocyclyl, C 6-10 An aryl or 5-10 membered heteroaryl, wherein one or more hydrogens of the cycloalkyl, heterocyclyl, aryl, and heteroaryl are optionally D, halogen, cyano, -OR a , -NR a R b , -C(O)R a -C(O)NR a R b , -S(O) 2 R a ,-P(O)(CH 3 ) 2 、C 1-6 Alkyl, C 3-6 They may be substituted with substituents selected from cycloalkyl groups, 4- to 8-membered heterocyclines, and 5- to 6-membered heteroaryl groups. B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally D, halogen, cyano, OR a , -NR a R b ,-COORa , -C(O)R a , -NR a C(O)R b -C(O)NR a R b , -S(O) 2 R a , -S(O) 2 NR a R b -S(O)(NR a )R b ,-P(O)(CH 3 ) 2 and R 11 They may be substituted with substituents selected from the following: R 11 is C 1-6 Alkyl, C 3-6 The alkyl, cycloalkyl, 4-8 membered heterocyclyl, or 5-6 membered heteroaryl, wherein one or more hydrogen atoms of the alkyl, cycloalkyl, heterocyclyl, and heteroaryl are optionally D, halogen, CN, -OH, or -NH. 2 、C 1-6 Alkyl, -OC 1-6 Alkyl, -COOR a , -C(O)R a and -C(O)NR a R b They may be substituted with substituents selected from the following: R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyl or 4- to 8-membered heterocyclines, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocycline are optionally D, halogen, or C 1-6 [May be substituted with alkyl groups] Compounds represented by , or pharmaceutically acceptable salts thereof, stable isotopic derivatives, isomers, and prodrugs. [Aspect 2] R 1 and R 2 The compound according to Embodiment 1, wherein both atoms are H, or a pharmaceutically acceptable salt, stable isotopic derivative, isomer, and prodrug. [Aspect 3] General formula (II):

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Claims

1. General formula (I): 【Chemistry 1】 [In the formula, Bond a is either a single bond or a double bond. R 1 and R 2 These are H, D, CN, and C, respectively, independently. 1-6 Alkyl or C 3-6 Selected from cycloalkyl groups, wherein one or more hydrogens of the alkyl group may be optionally substituted with D or fluoro. A is C 3-10 Cycloalkyl, 4- to 10-membered heterocyclyl, C 6-10 Aryl or 5- to 10-membered heteroaryl, provided that one or more hydrogens of said cycloalkyl, heterocyclyl, aryl and heteroaryl may optionally be D, halogen, cyano, -OR a , -NR a R b , -C(O)R a , -C(O)NR a R b , -S(O) 2 R a , -P(O)(CH 3 ) 2 , C 1-6 Alkyl, C 3-6 Cycloalkyl, 4- to 8-membered heterocyclyl and 5- to 6-membered heteroaryl, and may be substituted with a substituent selected from B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally D, halogen, cyano, OR a , -NR a R b ,-COOR a , -C(O)R a , -NR a C(O)R b -C(O)NR a R b , -S(O) 2 R a , -S(O) 2 NR a R b -S(O)(NR a )R b ,-P(O)(CH 3 ) 2 and R 11 They may be substituted with substituents selected from the following: R 11 is C 1-6 Alkyl, C 3-6 The alkyl, cycloalkyl, 4- to 8-membered heterocyclyl, or 5- to 6-membered heteroaryl, wherein one or more hydrogen atoms of the alkyl, cycloalkyl, heterocyclyl, and heteroaryl are optionally D, halogen, CN, -OH, or -NH. 2 , C 1-6 Alkyl, -OC 1-6 Alkyl, -COOR a , -C(O)R a and -C(O)NR a R b They may be substituted with substituents selected from the following: R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyl or 4- to 8-membered heterocyclines, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocycline are optionally D, halogen, or C 1-6 [May be substituted with alkyl groups] A compound represented by , or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof.

2. R 1 and R 2 The compound according to claim 1, or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof, wherein both atoms are H.

3. General formula (II): 【Chemistry 2】 [In the formula, A is C 3-8 A cycloalkyl, a 4- to 8-membered heterocyclyl, a phenyl, or a 5- to 6-membered heteroaryl, wherein one or more hydrogens of the cycloalkyl, heterocyclyl, phenyl, and heteroaryl are optionally a halogen, -C(O)R a , C 1-6 Alkyl, and C 3-6 They may be substituted with substituents selected from cycloalkyl groups. B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally halogens, -COOR a , -C(O)R a -C(O)NR a R b , -S(O) 2 R a , C 1-6 Alkyl, C 3-6 The substituents may be substituted with substituents selected from cycloalkyl, 4- to 8-membered heterocyclines containing N, S and / or O heteroatoms, and one or more hydrogens of the alkyl, cycloalkyl and heterocyclines may be C 1-6 Alkyl, -C(O)R a and -C(O)NR a R b They may be further substituted with substituents selected from, R a and R b These are H and C, respectively, independently. 1-6 Alkyl, C 3-6 Selected from cycloalkyls or 4- to 6-membered heterocyclines containing N, S and / or O heteroatoms, wherein one or more hydrogens of the alkyl, cycloalkyl, and heterocyclines may be C 1-6 [May be substituted with alkyl] The compound according to claim 1 or 2, which is represented by , or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof.

4. A is phenyl, wherein one or more hydrogens of the phenyl may be optionally substituted with halogens, the compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof.

5. General formula (III): 【Transformation 3】 [In the formula, A is phenyl, wherein one or more hydrogen atoms of the phenyl may be substituted with halogens as applicable. B is a phenyl or a 5-6 member heteroaryl, wherein one or more hydrogens of the phenyl and heteroaryl are optionally halogens, -COOR a , -C(O)R a -C(O)NR a R b , -S(O) 2 R a , C 1-6 Alkyl, C 3-6 The substituents may be substituted with substituents selected from cycloalkyl, 4- to 8-membered heterocyclines containing N, S, and / or O heteroatoms, and one or more hydrogens of the alkyl, cycloalkyl, and heterocyclines may be C 1-6 Alkyl, -C(O)R a and -C(O)NR a R b They may be further substituted with substituents selected from, R a and R b each independently is selected from H, C 1-6 alkyl, C 3-6 cycloalkyl, or 4- to 6-membered heterocyclyl containing heteroatoms of N, S and / or O, provided that one or more hydrogens of said alkyl, cycloalkyl and heterocyclyl may optionally be substituted with C 1-6 alkyl] The compound according to claim 1, which is represented by , or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof.

6. The aforementioned compound is as follows: 【Chemistry 4-1】 【Chemistry 4-2】 【Chemistry 4-3】 A compound according to claim 1, selected from the compounds shown, or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer thereof.

7. The following compounds 【Chemistry 5-1】 【Chemistry 5-2】 【Chemistry 5-3】 A compound according to claim 6, or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer, selected from the above.

8. A pharmaceutical composition comprising a compound according to any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, a stable isotopic derivative, or a stereoisomer, and one or more pharmaceutically acceptable carriers or excipients.

9. A pharmaceutical composition according to claim 8 for treating or preventing a disease mediated by JAK, A pharmaceutical composition in which JAK is mediated by inflammatory diseases, autoimmune diseases, or cancer.

10. The pharmaceutical composition according to claim 9, wherein the disease is inflammatory bowel disease, dermatitis, eczema, rheumatoid arthritis, systemic lupus erythematosus, psoriasis, or alopecia areata.

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