WO2022073424A1 - 一种作为jak激酶抑制剂的小分子化合物及其用途 - Google Patents

一种作为jak激酶抑制剂的小分子化合物及其用途 Download PDF

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WO2022073424A1
WO2022073424A1 PCT/CN2021/120119 CN2021120119W WO2022073424A1 WO 2022073424 A1 WO2022073424 A1 WO 2022073424A1 CN 2021120119 W CN2021120119 W CN 2021120119W WO 2022073424 A1 WO2022073424 A1 WO 2022073424A1
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compound
cycloalkyl
small molecule
molecule compound
alkyl
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French (fr)
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方文奎
李冠群
蔡雨婷
潘翔
朱文浩
汪杨
王增全
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嘉兴特科罗生物科技有限公司
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Priority to EP21876942.0A priority Critical patent/EP4227298A4/en
Priority to KR1020237015632A priority patent/KR20240004211A/ko
Priority to JP2023521673A priority patent/JP2023544431A/ja
Priority to US18/248,362 priority patent/US20230373933A1/en
Publication of WO2022073424A1 publication Critical patent/WO2022073424A1/zh

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    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
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    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
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    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
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    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
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    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
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    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings

Definitions

  • the present invention belongs to the field of small molecule compounds, and in particular, relates to an autoimmune disease that can be used to prevent or treat and alleviate autoimmune diseases such as rheumatoid arthritis, ulcerative colitis, systemic lupus erythematosus, etc., or related inflammatory skin diseases such as Small molecule compounds for psoriasis, eczema, vitiligo and alopecia areata.
  • autoimmune diseases such as rheumatoid arthritis, ulcerative colitis, systemic lupus erythematosus, etc.
  • related inflammatory skin diseases such as Small molecule compounds for psoriasis, eczema, vitiligo and alopecia areata.
  • JAK Janus Kinase, Janus Kinase
  • JAK-STAT Signal Transducer and Activator of Transcription proteins
  • JAK-STAT signaling pathway plays an indispensable driving role in the pathogenesis of many diseases, especially autoimmune diseases such as rheumatoid arthritis, lupus erythematosus, inflammatory bowel disease, multiple sclerosis, Sjögren's syndrome, psoriasis, alopecia areata and vitiligo, etc.; allergic diseases such as asthma, allergic rhinitis, allergic conjunctivitis, atopic dermatitis and eczema, etc.
  • autoimmune diseases such as rheumatoid arthritis, lupus erythematosus, inflammatory bowel disease, multiple sclerosis, Sjögren's syndrome, psoriasis, alopecia areata and vitiligo, etc.
  • allergic diseases such as asthma, allergic rhinitis, allergic conjunctivitis, atopic dermatitis and eczema, etc.
  • JAK kinase activity especially JAK1 and TYK2 kinase activity
  • JAK1 and TYK2 kinase activity can block cytokine-mediated signaling pathways involved in inflammatory responses, thereby controlling inflammation and effectively treating autoimmune diseases and/or allergies inflammatory disease.
  • T cells will differentiate in different directions according to different inflammation-inducing factors such as virus or bacterial infection, forming Th1, Th2, Th17 and other T cell subsets, and these T cells produce different cytokines accordingly.
  • these cytokines bind to cell surface receptors, transmit inflammatory signals through intracellular JAK, and drive the pathological process of the disease.
  • JAK1 inhibitors can specifically inhibit Th2 allergic inflammation
  • JAK1/Tyk2 dual inhibitors and Tyk2 inhibitors are clinically effective
  • JAK1/Tyk2 dual inhibitors and Tyk2 inhibitors are clinically effective
  • a potent inhibitor can bring good curative effect while avoiding the side effects caused by systemic medication, but this will require a strong inhibitory activity to achieve at the same time.
  • JAK1/Tyk2 dual inhibitors may be suitable for SLE, vitiligo , IBD and eczema and other diseases
  • Tyk2-specific inhibitors may be more suitable for the treatment of rheumatoid arthritis, psoriasis and alopecia areata, etc., while overcoming hematopoietic inhibition and coagulation abnormalities caused by JAK2 inhibition.
  • selecting suitable external administration routes such as external use of JAK family member inhibitors with different properties associated with disease etiology and symptom manifestations for inflammatory skin diseases, to intervene and control symptoms, can achieve good therapeutic effects.
  • the present invention provides a small molecule compound
  • the small molecule compound is a compound represented by the following formula I, or its stereoisomer, geometric isomer, interconversion Isomers, hydrates, solvates, and pharmaceutically acceptable salts or prodrugs:
  • R 1 to R 4 are each independently selected from C or N;
  • R is selected from cycloalkyl, substituted cycloalkyl, heterocycloalkyl, substituted heterocycloalkyl, aryl, substituted aryl, heteroaryl or substituted heteroaryl.
  • At most two of R 1 to R 4 are N.
  • R has the structure represented by Formula II shown below:
  • R 5 is selected from C or N;
  • R is selected from hydrogen, halogen, alkyl, amino, amido, cycloalkyl, heterocycloalkyl, aryl or heteroaryl ;
  • R7 is selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl.
  • R6 and R7 are each independently selected from hydrogen, alkyl or cycloalkyl.
  • R has the structure represented by Formula III shown below:
  • R 8 is selected from C or N;
  • R is selected from hydrogen, halogen, alkyl, amino, amido, cycloalkyl, heterocycloalkyl, aryl or heteroaryl;
  • R 10 is selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl.
  • R 9 and R 10 are each independently selected from hydrogen, alkyl or cycloalkyl.
  • the alkyl group is methyl, ethyl, propyl or isopropyl and the cycloalkyl group is cyclopropyl, cyclobutyl or cyclopropylmethyl.
  • the present invention also provides the use of the above-mentioned small molecule compound in inhibiting JAK kinase.
  • the present invention also provides the use of the above-mentioned small molecule compound in the preparation of a medicament for preventing or treating JAK-related autoimmune diseases and immune-related inflammatory skin diseases.
  • the autoimmune disease is selected from the group consisting of rheumatoid arthritis, ankylosing spondylitis, ulcerative colitis, Crohn's disease, systemic lupus erythematosus, dermatomyositis, multiple sclerosis, type I At least one of diabetes, Sjögren's syndrome, and vasculitis.
  • the immune-related inflammatory skin disease is selected from the group consisting of atopic dermatitis, eczema, alopecia areata, psoriasis, vitiligo, lichen planus, lichen lustrous, lichen atrophicus, panniculitis, acne and at least one of hidradenitis suppurativa.
  • the present invention carries out the purposeful and rational design of small molecular compounds.
  • the synthesized compound is first tested for the kinase biochemical activity of JAK, and SAR (structure-activity relationship) is established according to IC 50 . , and then perform cytological tests on potent inhibitors with IC 50 below 200 nM, and determine the selectivity of the compounds.
  • SAR structure-activity relationship
  • the present invention provides a small molecule compound, which is a compound represented by formula I shown below, or a stereoisomer, geometric isomer, tautomer, hydrated Compounds, solvates, and pharmaceutically acceptable salts or prodrugs:
  • R 1 to R 4 are each independently selected from C or N;
  • R is selected from cycloalkyl, substituted cycloalkyl, heterocycloalkyl, substituted heterocycloalkyl, aryl, substituted aryl, heteroaryl or substituted heteroaryl.
  • R 1 , R 2 , R 3 and R 4 may each be independently selected from C or N.
  • at most two (ie, 0, 1 or 2) of R 1 to R 4 may be N.
  • at most one (ie, 0 or 1) of R 1 to R 4 may be N.
  • they will constitute a benzene ring or a pyridine ring.
  • R may have the structure represented by formula II as shown below:
  • R 5 can be selected from C or N;
  • R is selected from hydrogen, halogen, alkyl, amino, amido, cycloalkyl, heterocycloalkyl, aryl or heteroaryl ;
  • R7 is selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl.
  • R 6 and R 7 can be independently selected from hydrogen, alkyl or cycloalkyl, further, the alkyl can be methyl, ethyl, propyl or isopropyl,
  • the cycloalkyl group is cyclopropyl, cyclobutyl or cyclopropylmethyl.
  • R5 is C
  • R6 is hydrogen
  • R7 is ethyl
  • R can have a structure like this:
  • R5 is N
  • R6 is methyl
  • R7 is methyl
  • R can have a structure like this:
  • R5 is C
  • R6 is methyl
  • R7 is ethyl
  • R can have a structure like this:
  • R5 is N
  • R6 is methyl
  • R7 is hydrogen
  • R can have a structure like this:
  • R may have the structure represented by formula III as shown below:
  • R 8 can be selected from C or N;
  • R is selected from hydrogen, halogen, alkyl, amino, amido, cycloalkyl, heterocycloalkyl, aryl or heteroaryl;
  • R 10 is selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl or heteroaryl.
  • R 9 and R 10 can be independently selected from hydrogen, alkyl or cycloalkyl, further, the alkyl can be methyl, ethyl, propyl or isopropyl,
  • the cycloalkyl group is cyclopropyl, cyclobutyl or cyclopropylmethyl.
  • R 8 is N
  • R 9 is hydrogen
  • R 10 is methyl
  • R can also have the following structure:
  • the term "pharmaceutically acceptable” refers to a substance that does not affect the biological activity or properties of the compounds of the present invention, and is relatively non-toxic, ie, the substance can be administered to an individual without causing an adverse biological response or in an undesirable manner Interacts with any components contained in the composition.
  • “pharmaceutically acceptable salts” may include inorganic salts and organic salts, wherein the organic salts may include but are not limited to ammonium, lithium, sodium, potassium, cesium, calcium, magnesium, copper, aluminum, Zinc, barium, or quaternary ammonium salts, and the inorganic salts may include, but are not limited to, arginine, tert-butylamine, dimethylamine, diethanolamine, ethanolamine, ethylenediamine, imidazole, lysine, methylamine, pyridine, picolinyl ester, piperazine, triethylamine, triethanolamine, trimethylamine or urea salts.
  • the organic salts may include but are not limited to ammonium, lithium, sodium, potassium, cesium, calcium, magnesium, copper, aluminum, Zinc, barium, or quaternary ammonium salts
  • the inorganic salts may include, but are not limited to, arginine, tert-butylamine, dimethylamine
  • the present invention provides the use of the above-mentioned small molecule compounds in inhibiting JAK kinases, especially as JAK1/Tyk2 dual inhibitors and Tyk2 specific inhibitors.
  • the present invention also provides the use of the above-mentioned small molecule compounds in the preparation of medicaments for preventing or treating autoimmune diseases and immune-related inflammatory skin diseases. Studies have shown that the pathogenesis of these diseases is related to the dysregulation of JAK signaling.
  • treatment refers to any administration of a therapeutic agent according to a therapeutic regimen that achieves the desired effect, ie, partial or complete relief, amelioration, remission, inhibition, delay of onset, reduction in severity and/or reducing the incidence of one or more symptoms or characteristics of a particular disease, disorder and/or condition; in some embodiments, administration of a therapeutic agent according to a therapeutic regimen is associated with achievement of a desired effect.
  • Such treatment can be directed to subjects who do not exhibit the relevant disease, disorder and/or condition and/or to subjects exhibiting only early signs of the disease, disorder and/or condition. Alternatively or additionally, such treatment may be directed to subjects exhibiting one or more established signs of the relevant disease, disorder and/or condition.
  • treatment can be directed to a subject who has been diagnosed with the relevant disease, disorder, and/or condition. In some embodiments, treatment may be directed to a subject known to have one or more predisposing factors that are statistically associated with an increased risk of developing the relevant disease, disorder, and/or condition.
  • the medicament prepared in the above-mentioned use may comprise an effective amount of the small molecule compound of the present invention, and a pharmaceutically acceptable excipient, carrier or diluent.
  • the term "effective amount”, “therapeutically effective amount” or “pharmaceutically effective amount” refers to the amount of a therapeutic agent that confers a therapeutic effect to a treated subject at a reasonable benefit/risk ratio applicable to any drug treatment .
  • Such therapeutic effects may be objective (ie, measurable by some test or marker) or subjective (ie, the subject gives an indication or perceives the effect).
  • a "therapeutically effective amount” refers to an amount effective to treat, ameliorate, or prevent (eg, delay onset), such as by ameliorating symptoms associated with a disease, preventing or delaying the onset of a disease, and/or also reducing the severity or frequency of symptoms of a disease ) associated disease or disorder and/or an amount of a therapeutic agent or composition that exhibits a detectable therapeutic or prophylactic effect.
  • the therapeutically effective amount of the small molecule compound to be administered will vary depending on the nature and severity of the subject and disease, the physical condition of the subject, the treatment regimen ( For example, whether to use a second therapeutic agent), and the route of administration selected; appropriate dosages can be readily determined by those skilled in the art. Additionally, the optimal number and interval of individual doses of the drug will be determined by the nature and extent of the condition being treated, the form, route and location of administration, and the age and condition of the particular subject being treated, and the physician will ultimately An appropriate dose to be administered is determined. This dose can be repeated as many times as necessary. If side effects occur, the amount and/or frequency of the dose can be varied or reduced according to normal clinical practice.
  • pharmaceutically acceptable excipient, carrier or diluent includes, but is not limited to, any adjuvant, carrier, excipient, Glidants, sweeteners, diluents, preservatives, dyes/colorants, flavoring agents, surfactants, wetting agents, dispersing agents, suspending agents, stabilizers, isotonic agents, solvents or emulsifiers, etc. .
  • the medicament prepared in the above-mentioned use can also contain other substances that can be used to prevent or treat autoimmune diseases and immune-related inflammatory diseases.
  • a medicament for skin diseases as another active ingredient examples include, but are not limited to, vitamin D derivatives, vitamin A derivatives, glucocorticoids, calcineurin inhibitors, or non-steroidal anti-inflammatory drugs, and the like.
  • the medicament contains a plurality of active ingredients, the respective active ingredients may be administered simultaneously, sequentially or separately according to the judgment of the physician.
  • the small molecule compounds of the present invention can be administered to a patient by a variety of routes such as oral, transdermal, subcutaneous, intranasal, intravenous, intramuscular, intrathecal, regional or topical (eg mucosal).
  • routes such as oral, transdermal, subcutaneous, intranasal, intravenous, intramuscular, intrathecal, regional or topical (eg mucosal).
  • the most appropriate route of administration in any given situation will depend on the subject and the nature and severity of the disease, the subject's physical condition, and the like.
  • the small molecule compounds of the present invention can be administered intravenously.
  • the small molecule compounds of the present invention can be administered orally.
  • the medicament of the present invention can be prepared into different dosage forms.
  • the medicament can be prepared as tablets, capsules, pills, granules, aerosols, sprays, or injections.
  • the small molecule compound of the present invention or the medicine prepared therefrom can exert excellent effects in preventing or treating JAK-related autoimmune diseases and immune-related inflammatory skin diseases.
  • the autoimmune disease may include, but is not limited to, rheumatoid arthritis, ankylosing spondylitis, ulcerative colitis, Crohn's disease, systemic lupus erythematosus, dermatomyositis, multiple sclerosis, type I Diabetes, Sjögren's syndrome, vasculitis, etc.; and the immune-related inflammatory skin diseases may include, but are not limited to, atopic dermatitis, eczema, alopecia areata, psoriasis or vitiligo, lichen planus, lichen lustrous, sclerosing atrophic Lichen, panniculitis, acne and hidradenitis suppurativa.
  • Step 2 Preparation of Compound 1e ((S)-N-(4-(2-chloropyrimidin-4-yl)phenyl)-2,2-difluorocyclopropane-1-carboxamide)
  • Step 3 Compound 1 ((S)-4-((4-(4-(4-(4-(2,2-difluorocyclopropane-1-carboxamido)phenyl)pyrimidin-2-yl)amino)- Preparation of N-ethylbenzamide)
  • Step 1 Compound 2 ((S)-5-((4-(4-(4-(4-(2,2-difluorocyclopropane-1-carboxamido)phenyl)pyrimidin-2-yl)amino)- Preparation of 3-picoline)
  • Step 2 Compound 3 ((S)-4-((4-(4-(4-(4-(2,2-difluorocyclopropane-1-carboxamido)phenyl)pyrimidin-2-yl)amino)- Preparation of N-ethyl-2-methylbenzamide)
  • Step 1 Compound 4 ((S)-5-((4-(4-(4-(4-(2,2-difluorocyclopropane-1-carboxamido)phenyl)pyrimidin-2-yl)amino)- Preparation of N,3-lutidine amide)
  • Step 5 Preparation of compound 5g (2-(5-aminopyridin-2-yl)propan-1-ol)
  • Step 6 Compound 5 ((1S)-2,2-difluoro-N-(4-(2-((6-(1-(1-hydroxypropan-2-yl)pyridin-3-yl)amino) Preparation of pyrimidin-4-yl)phenyl)cyclopropane-1-carboxamide)
  • Step 2 Preparation of compound 6e ((S)-N-(5-(2-chloropyrimidin-4-yl)pyridin-2-yl)-2,2-difluorocyclopropane-1-carboxamide)
  • Step 3 Compound 6 ((S)-4-((4-(6-(2,2-difluorocyclopropane-1-carboxamido)pyridin-3-yl)pyrimidin-2-yl)amino)- Preparation of N-ethylbenzamide)
  • Step 1 Compound 7 ((S)-5-((4-(6-(2,2-difluorocyclopropane-1-carboxamido)pyridin-3-yl)pyrimidin-2-yl)amino)- Preparation of N,3-lutidine amide)
  • Step 1 Compound 8 ((S)-5-((4-(6-(2,2-difluorocyclopropane-1-carboxamido)pyridin-3-yl)pyrimidin-2-yl)amino)- Preparation of 3-picoline amide)
  • Step 1 Compound 9 ((S)-4-((4-(6-(2,2-difluorocyclopropane-1-carboxamido)pyridin-3-yl)pyrimidin-2-yl)amino)- Preparation of N-ethyl-2-methylbenzamide)
  • Step 1 Compound 10 ((1S)-2,2-difluoro-N-(5-(2-((6-(1-(hydroxypropyl-2-yl)pyridin-3-yl)amino) Preparation of pyrimidinyl-4-yl)pyridyl-2-ylcyclopropane-1-carboxamide)
  • Kinase reaction buffer prepare the kinase reaction buffer with the following components: 50 mM HEPES, pH 7.5, 1 mM EGTA, 10 mM MgCl 2 , 2 mM DTT, 0.01% Tween 20.
  • 1X detection buffer prepare the detection buffer, deionized water 9: 1 Dilute 10X assay buffer to 1X.
  • 4X Kinase Solution Dilute JAK kinase to 4X final concentration in Kinase Reaction Buffer (JAK1: 80 nM, JAK2/JAK3/Tyk2: 4 nM).
  • 4X Substrate Solution Dilute ULight TM -JAK (Tyr1023) substrate to 200 nM (final concentration: 50 nM) in Kinase Reaction Buffer.
  • 4X ATP solution Dilute ATP to 4X final concentration in Kinase Reaction Buffer (JAK1: 160 ⁇ M, JAK2/JAK3/Tyk2: 40 ⁇ M).
  • 4X test compound solution DMSO dissolves the test compound to 10mM stock solution, 3-fold gradient dilution is configured to the desired concentration, each compound is set to 10 concentration points, and the final concentration range of the test compound is: 10 ⁇ M-0.5nM.
  • 4X Enzyme Reaction Stop Solution 1X Assay Buffer to dissolve EDTA to 40 mM (final EDTA concentration: 10 mM).
  • 4X detection antibody solution Dilute Eu-labeled detection antibody (anti-phosphotyrosine (PT66)) to 8 nM (final antibody concentration: 2 nM) in 1X detection buffer.
  • JAK1 was reacted at room temperature for 2 hours, and JAK2/JAK3/Tyk2 was reacted at room temperature for 1 hour.
  • the final concentrations of each component of the JAK1 reaction were: JAK1: 20 nM, substrate: 50 nM, ATP: 40 uM, and the final concentration of the test compounds ranged from 10 ⁇ M to 0.5 nM.
  • the final concentrations of each component in the JAK2/JAK3/Tyk2 reaction were: JAK2: 1 nM, substrate: 50 nM, ATP: 10 ⁇ M, and the final concentration of the test compounds ranged from 10 ⁇ M to 0.5 nM.
  • TDM-180972 Numbering Tyk2/ ⁇ M JAK1/ ⁇ M JAK2/ ⁇ M JAK3/ ⁇ M TDM-180972 0.005 0.004 0.003 0.117 TDM-180974 0.008 0.007 0.007 0.346 TDM-180977 0.004 0.003 0.004 0.057 TDM-180981 0.008 0.008 0.008 0.183 TDM-180989 0.014 0.011 0.013 0.177 TDM-180971 0.005 0.006 0.007 0.219 TDM-180979 0.017 0.025 0.033 0.931 TDM-180980 0.018 0.028 0.030 >5 TDM-180984 0.005 0.008 0.009 0.105 TDM-180987 0.017 0.025 0.022 0.310
  • the small molecule compound of the present application is a kind of compound with strong targeting to JAK family and excellent enzymatic activity, and can be used as a dual inhibitor of JAK1/Tyk2 and a specific inhibitor of Tyk2.

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Abstract

本发明提供了一种小分子化合物，所述小分子化合物为由下式（I）表示的化合物，或其立体异构体、几何异构体、互变异构体、水合物、溶剂化物、以及药学上可接受的盐或前药，其中R 1至R 4各自独立地选自C或N；并且其中R选自环烷基、取代的环烷基、杂环烷基、取代的杂环烷基、芳基、取代的芳基、杂芳基或取代的杂芳基。本发明的小分子化合物能够抑制JAK激酶，更特别是作为JAK1/Tyk2双抑制剂和Tyk2特异性抑制剂。

Description

一种作为JAK激酶抑制剂的小分子化合物及其用途
本申请的交叉引用
本发明要求于2020年10月9日提交的申请号为CN202011072703.7、发明名称为“一种作为JAK激酶抑制剂的小分子化合物及其用途”的专利申请的优先权。
技术领域
本发明属于小分子化合物领域，具体地，涉及一种能够用于预防或治疗缓解自身免疫性疾病如风湿性关节炎、溃疡性结肠炎和系统性红斑狼疮等，或相关的炎症性皮肤病如银屑病、湿疹、白癜风和斑秃等的小分子化合物。
背景技术
JAK(Janus激酶，Janus Kinase)是细胞内非受体性酪氨酸蛋白激酶的一个家族，包括JAK1、JAK2、JAK3和Tyk2四个成员。JAK-STAT(转录蛋白的信号转导和激活剂，Signal Transducer and Activator of Transcription proteins)信号传导通路是炎症性细胞因子和受体相结合之后激发的信号在细胞内传导的主要通路。许多证据表明，JAK-STAT信号传导通路在很多疾病的发病机理中起到不可或缺的驱动作用，特别是自身免疫性疾病如风湿性关节炎、红斑狼疮、炎症性肠病、多发性硬化、干燥综合征、银屑病、斑秃和白癜风等；过敏性疾病如哮喘、过敏性鼻炎、过敏性结膜炎、特应性皮炎和湿疹等等。因此，利用高效的小分子对JAK激酶活性，特别是JAK1、TYK2激酶活性进行抑制可以阻断参与炎症反应的细胞因子介导的信号通路，从而控制炎症，有效治疗自身免疫性疾病和/或过敏性炎症性疾病。
不同炎症性疾病发病过程中T细胞会根据不同的炎症诱发因素如病毒或细菌感染而向不同的方向分化，形成Th1，Th2，Th17等T细胞亚群，这些T细胞相应地产生不同的细胞因子，如与病毒感染引起的急性炎症相关的Th1细胞 产生IFNγ，IL-2；与过敏有关的Th2细胞产生IL-4，IL-5，IL-13；与自身免疫有关的Th17细胞产生IL-17，IL-12，IL-21，IL-22，IL-23；这些细胞因子和细胞表面的受体结合后，通过细胞内的JAK传递炎症信号，驱动疾病的病理过程。更重要的是，很多病因不清楚的炎症性疾病发表机理复杂，在不同的阶段甚至同一阶段会涉及多种T细胞亚群，也就是会涉及多个JAK通路，这就对开发针对JAK的炎症性疾病治疗药物提出了新的要求。
虽然有研究报道JAK1抑制剂对Th2类过敏性炎症有特异性抑制，同时有效抑制JAK1和/或TYK2抑制剂的研究鲜有报道，尤其我们认为JAK1/Tyk2双抑制剂和Tyk2抑制剂在临床上有更广泛的前途，尤其发病机制涉及到自身免疫异常的炎症性疾病。此外，更多的炎症性疾病尤其是炎症性皮肤病的发病机理可能涉及多个JAK，因此开发强效JAK1和Tyk2的单抑制剂或双抑制剂具有重要意义，尤其用于皮肤病的外用治疗，强效抑制剂在带来很好的疗效的同时又能避免系统用药造成的副作用，但是这将同时需要很强的抑制活性才能实现。
发明内容
本发明的目的旨在获得高效的JAK1/Tyk2双抑制剂和Tyk2特异性抑制剂，为不同的炎症性疾病提供有针对性的靶向治疗，例如JAK1/Tyk2双抑制剂可能适合于SLE、白癜风、IBD和湿疹等疾病，Tyk2特异性抑制剂可能更适合于治疗类风湿性关节炎、银屑病和斑秃等，同时克服由于抑制JAK2带来的造血抑制和凝血异常。另外，选择适合用于外用给药途径如针对炎症性皮肤病外用与疾病病因和症状表现相关联的不同性质的JAK家族成员抑制剂对疾病进行干预和症状控制，能够获得好的治疗效应。
为了实现上述目的，在一方面，本发明提供了一种小分子化合物，所述小分子化合物为由如下所示的式I表示的化合物，或其立体异构体、几何异构体、互变异构体、水合物、溶剂化物、以及药学上可接受的盐或前药：
[式I]
Figure PCTCN2021120119-appb-000001
其中，R 1至R 4各自独立地选自C或N；并且
其中，R选自环烷基、取代的环烷基、杂环烷基、取代的杂环烷基、芳基、取代的芳基、杂芳基或取代的杂芳基。
在一个实施方式中，R 1至R 4中的至多两个为N。
在另一个实施方式中，R具有如下所示的式II表示的结构：
[式II]
Figure PCTCN2021120119-appb-000002
其中，R 5选自C或N；
其中，R 6选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
R 7选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
在另一个实施方式中，R 6和R 7各自独立地选自氢、烷基或环烷基。
在另一个实施方式中，R具有如下所示的式III表示的结构：
[式III]
Figure PCTCN2021120119-appb-000003
其中，R 8选自C或N；
其中，R 9选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
R 10选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
在另一个实施方式中，R 9和R 10各自独立地选自氢、烷基或环烷基。
在另一个实施方式中，所述烷基为甲基、乙基、丙基或异丙基，所述环烷基为环丙基、环丁基或环丙基甲基。
在另一方面，本发明还提供了如上所述的小分子化合物在抑制JAK激酶中的用途。
在另一方面，本发明还提供了如上所述的小分子化合物在制备用于预防或治疗JAK相关的自身免疫性疾病、以及与免疫有关的炎症性皮肤疾病的药物中的用途。
在一个实施方式中，所述自身免疫性疾病选自类风湿性关节炎、强直性脊柱炎、溃疡性结肠炎、克罗恩病、系统性红斑狼疮、皮肌炎、多发性硬化、I型糖尿病、干燥综合症和血管炎中的至少一种。
在另一个实施方式中，所述与免疫有关的炎症性皮肤疾病选自特应性皮炎、湿疹、斑秃、银屑病、白癜风、扁平苔藓、光泽苔藓、硬化萎缩性苔藓、脂膜炎、痤疮和化脓性汗腺炎中的至少一种。
本发明的作用和效果：
本发明根据JAK激酶的蛋白结构，特别是Tyk2的蛋白结构，进行了小分子化合物有目的的合理设计，合成的化合物首先进行JAK的激酶生化活性检测，根据IC 50建立SAR(structure-activity relationship)，对IC 50在200nM以下的强效抑制剂再进行细胞学的测试，并确定化合物的选择性。参见具体活性实验数据可以发现，本发明涉及的化合物具有良好的JAK激酶活性和细胞生物学活性的抑制能力。
具体实施方式
以下对本发明的具体实施方式进行详细说明。应当理解的是，此处所描述的具体实施方式仅用于说明和解释本发明，并不用于限制本发明。
在本文中所披露的范围的端点和任何值都不限于该精确的范围或值，这些 范围或值应当理解为包含接近这些范围或值的值。对于数值范围来说，各个范围的端点值之间、各个范围的端点值和单独的点值之间，以及单独的点值之间可以彼此组合而得到一个或多个新的数值范围，这些数值范围应被视为在本文中具体公开。
在详细描述本发明前，应了解，在此使用的术语只在于描述特定的实施方式，而不希望限制本发明的范围，本发明的范围仅由所附权利要求书限定。为了更完全地了解在此描述的本发明，采用以下术语，它们的定义如下所示。除非另外定义，在此使用的所有技术和科学术语具有与本发明所属领域的普通技术人员所理解的相同的含义。
在一方面，本发明提供了一种小分子化合物，所述小分子化合物为由如下所示的式I表示的化合物，或其立体异构体、几何异构体、互变异构体、水合物、溶剂化物、以及药学上可接受的盐或前药：
[式I]
Figure PCTCN2021120119-appb-000004
其中，R 1至R 4各自独立地选自C或N；并且
其中，R选自环烷基、取代的环烷基、杂环烷基、取代的杂环烷基、芳基、取代的芳基、杂芳基或取代的杂芳基。
也就是说，R 1、R 2、R 3和R 4可以各自独立地选自C或N。在一个优选的实施方式中，R 1至R 4中的至多两个(即0、1或2个)可以为N。在另一个优选的实施方式中，R 1至R 4中的至多一个(即0或1个)可以为N。当R 1至R 4中的至多一个(即0或1个)为N时，它们将构成苯环或吡啶环。
根据本发明，在一个优选的实施方式中，R可以具有如下所示的式II表示的结构：
[式II]
Figure PCTCN2021120119-appb-000005
其中，R 5可以选自C或N；
其中，R 6选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
R 7选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
在一个更优选的实施方式中，R 6和R 7可以各自独立地选自氢、烷基或环烷基，进一步，所述烷基可以为甲基、乙基、丙基或异丙基，所述环烷基为环丙基、环丁基或环丙基甲基。
例如，在一个具体的实施方式中，R 5为C，R 6为氢，且R 7为乙基。在这种情况下，R可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000006
例如，在一个具体的实施方式中，R 5为N，R 6为甲基，且R 7为甲基。在这种情况下，R可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000007
例如，在一个具体的实施方式中，R 5为C，R 6为甲基，且R 7为乙基。在这种情况下，R可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000008
例如，在一个具体的实施方式中，R 5为N，R 6为甲基，且R 7为氢。在这种情况下，R可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000009
根据本发明，在另一个优选的实施方式中，R可以具有如下所示的式III表示的结构：
[式III]
Figure PCTCN2021120119-appb-000010
其中，R 8可以选自C或N；
其中，R 9选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
R 10选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
在一个更优选的实施方式中，R 9和R 10可以各自独立地选自氢、烷基或环烷基，进一步，所述烷基可以为甲基、乙基、丙基或异丙基，所述环烷基为环丙基、环丁基或环丙基甲基。
例如，在一个具体的实施方式中，R 8为N，R 9为氢，且R 10为甲基。在这种情况下，R可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000011
根据本发明，在另一个优选的实施方式中，R还可以具有如下所示的结构：
Figure PCTCN2021120119-appb-000012
如本文所用，术语“药学上可接受的”是指不影响本发明化合物的生物活性或性质的物质，并且相对无毒，即该物质可施用于个体而不造成不良的生物反应或以不良方式与组合物中包含的任意组分相互作用。在本发明中，“药学上可接受的盐”可以包括无机盐和有机盐，其中，所述有机盐可以包括但不限于铵、锂、钠、钾、铯、钙、镁、铜、铝、锌、钡或季铵盐，并且所述无机盐可以包括但不限于精氨酸、叔丁胺、二甲胺、二乙醇胺、乙醇胺、乙二胺、咪唑、赖氨酸、甲胺、吡啶、吡啶甲酸酯、哌嗪、三乙胺、三乙醇胺、三甲胺或脲盐。
在另一方面，本发明提供了上述小分子化合物在抑制JAK激酶中的用途，特别是作为JAK1/Tyk2双抑制剂和Tyk2特异性抑制剂。
在另一方面，本发明还提供了上述小分子化合物在制备用于预防或治疗自身免疫性疾病、以及与免疫有关的炎症性皮肤疾病的药物中的用途。研究表明，这些疾病的发病机理均与JAK信号传导的失调相关。
如本文所用，术语“治疗”是指根据治疗性方案的治疗性试剂的任何施用，所述治疗性方案达到所需效果，即部分或完全减轻、改善、缓解、抑制、延迟发作、降低严重程度和/或降低特定疾病、障碍和/或病症的一种或多种症状或特征的发生率；在一些实施方式中，根据治疗性方案的治疗性试剂的施用与所需效果的实现相关。这种治疗可以针对没有表现出相关疾病、障碍和/或病症的受试者和/或针对仅表现出疾病、障碍和/或病症的早期迹象的受试者。替代地或另外地，这种治疗可以针对表现出相关疾病、障碍和/或病症的一种或多种所确定迹象的受试者。在一些实施方式中，治疗可以针对已被诊断患有相关疾病、障碍和/或病症的受试者。在一些实施方式中，治疗可以针对已知具有一种或多种易感因素的受试者，所述易感因素在统计学上与相关疾病、障碍和/或病症发展的风险增加相关。
根据本发明，上述用途中制得的药物可以包含有效量的本发明的小分子化合物，以及药学上可接受的赋形剂、载体或稀释剂。
如本文所用，术语“有效量”、“治疗有效量”或“药学有效量”是指对于治疗的受试者以适用于任何药物治疗的合理受益/风险比赋予治疗效果的治疗性试剂的量。这样的治疗效果可以是客观的(即可以通过某种测试或标记测量)或主观的(即受试者给出指示或感觉到效果)。在一些实施方式中，“治疗有效量”是指诸如通过改善与疾病有关的症状、预防或延迟疾病发作和/或还减轻疾病症状的严重性或频率来有效治疗、改善或预防(例如延迟发作)相关疾病或病症和/或表现出可检测的治疗或预防效果的治疗性试剂或组合物的量。
本领域的技术人员将认识到，待施用的所述小分子化合物的治疗有效量将根据以下各项而变化：受试者和疾病的性质和严重程度、受试者的身体状况、治疗方案(例如是否使用第二治疗剂)、以及所选择的施用途径；合适的剂量可以由本领域的技术人员容易地确定。另外，该药物的个体剂量的最佳数量和间 隔将通过所治疗的病状的性质和程度、施用的形式、途径和位置、以及所治疗的特定受试者的年龄和病状确定，并且医师将最终确定待施用的合适剂量。此剂量可以视需要重复多次。如果出现副作用，则可以根据正常临床实践改变或减少剂量的量和/或频率。
在本发明中，“药学上可接受的赋形剂、载体或稀释剂”包括但不限于任何被相关的政府管理部门许可为可接受供人类或家畜使用的佐剂、载体、赋形剂、助流剂、增甜剂、稀释剂、防腐剂、染料/着色剂、矫味剂、表面活性剂、润湿剂、分散剂、助悬剂、稳定剂、等渗剂、溶剂或乳化剂等。
根据本发明，进一步地，上述用途中制得的药物除了可以包含本发明的小分子化合物作为有效成分之外，还可以包含其他可用于预防或治疗自身免疫性疾病、以及与免疫有关的炎症性皮肤疾病的药剂作为另一种有效成分。所述药剂的实例包括但不限于维生素D衍生物、维生素A衍生物、糖皮质激素、钙调神经磷酸酶抑制剂或非甾体类抗炎药等。当该药物包含多种有效成分时，各有效成分可以根据医师的判断同时、依次或分开施用。
另外，本发明的小分子化合物可以通过多种途径施用于患者，这些途径诸如口服、透皮、皮下、鼻内、静脉内、肌内、鞘内、区域或局部(例如粘膜)。在任何给定情况下最适合的施用途径将取决于受试者和疾病的性质和严重程度、以及受试者的身体状况等。在一个实施方式中，本发明的小分子化合物可以经静脉内施用。在另一个实施方式中，本发明的小分子化合物可以口服施用。相应地，根据不同的施用方式，本发明的药物可以制备为不同的剂型。例如，在一个实施方式中，所述药物可以制备为片剂、胶囊剂、丸剂、颗粒剂、雾化剂、喷雾剂或注射剂。
经发明人研究发现，本发明的小分子化合物或其制得的药物在用于预防或治疗JAK相关的自身免疫性疾病、以及与免疫有关的炎症性皮肤疾病能够发挥优异的效果。具体地，所述自身免疫性疾病可以包括但不限于类风湿性关节炎、强直性脊柱炎、溃疡性结肠炎、克罗恩病、系统性红斑狼疮、皮肌炎、多发性硬化、I型糖尿病、干燥综合症和血管炎等；而所述与免疫有关的炎症性皮肤疾病可以包括但不限于特应性皮炎、湿疹、斑秃、银屑病或白癜风、扁平苔藓、光泽苔藓、硬化萎缩性苔藓、脂膜炎、痤疮和化脓性汗腺炎等。
以下，将通过实施例对本发明的特定小分子化合物的效果进行详细描述。
实施例
实施例1合成化合物1的一般方法(TDM-180972)
Figure PCTCN2021120119-appb-000013
步骤1：化合物1c(4-(2-氯嘧啶-4-基)苯胺)的制备
向三口烧瓶加入化合物1a(2g，9.129mmol)，化合物1b(1.36g，9.129mmol)，四三苯基膦钯(527g，0.45mmol)，碳酸钾(2.5g，18.258mmol)，二氧六环(20mL)和水(20mL)，用氮气置换数次，然后将混合物加热至80℃并搅拌45分钟。反应结束减压浓缩反应物，残余物通过硅胶色谱法进行纯化(石油醚/乙酸乙酯＝0-60％)，以得到淡黄色固体目标化合物(化合物1c，394mg，收率21％)。LCMS[M+1] +＝206。
步骤2：化合物1e((S)-N-(4-(2-氯嘧啶-4-基)苯基)-2,2-二氟环丙烷-1-羧酰胺)的制备
向三口烧瓶中加入化合物1c(300mg，1.459mmol)和化合物1d(187mg，1.531mmol)，将混合物用氮气置换几次，然后在0℃下加入吡啶(10mL)和三氯氧磷(335.6mg，2.189mmol)。将混合物在室温搅拌1h，然后减压浓缩，残余物用乙酸乙酯(30mL*3)萃取，合并有机层，并用水(50mL*3)和饱和盐水(50mL*2)洗涤，用硫酸钠干燥，将滤液减压浓缩，残余物通过硅胶色谱法进行纯化(石油醚/乙酸乙酯＝0-12％)，以得到黄色固体目标化合物(化合物1e，327.7mg，收率72.5％)。LCMS[M+H] +＝310。
步骤3：化合物1((S)-4-((4-(4-(4-(2,2-二氟环丙烷-1-羧酰胺基)苯基)嘧啶-2-基)氨基)-N-乙基苯甲酰胺)的制备
向化合物1e(80mg，0.258mmol)的正丁醇(8mL)溶液加入化合物1f(85mg，0.517mmol)和对甲苯磺酸一水合物(98mg，0.517mmol)。将所得混合物加热至110℃并搅拌3小时。反应结束将混合物在减压下浓缩，残余物通过制备型HPLC(甲酸)纯化，以得到黄色固体目标化合物TDM-180972(化合物1，19.7mg，收率17.5％)。LCMS[M+H] +＝438.2。
1H NMR(400MHz,DMSO)δ10.70(s,1H),9.91(s,1H),8.57(d,J＝5.3Hz,1H),8.28(t,J＝5.5Hz,1H),8.19(d,J＝8.8Hz,2H),7.92(d,J＝8.9Hz,2H),7.83(d,J＝8.9Hz,2H),7.78(d,J＝8.8Hz,2H),7.44(d,J＝5.3Hz,1H),3.28(dt,J＝12.7,6.4Hz,2H),2.86(ddd,J＝13.6,10.8,8.0Hz,1H),2.12–1.94(m,2H),1.13(t,J＝7.2Hz,3H)。
实施例2合成化合物2的一般方法(TDM-180974)
Figure PCTCN2021120119-appb-000014
步骤1：化合物2((S)-5-((4-(4-(4-(2,2-二氟环丙烷-1-羧酰胺基)苯基)嘧啶-2-基)氨基)-3-甲基吡啶啉)的制备
向化合物2a(80mg，0.258mmol)的正丁醇(8mL)溶液加入化合物2b(78mg，0.517mmol)和对甲苯磺酸一水合物(98mg，0.517mmol)，将混合物加热至115℃并搅拌过夜。反应结束将混合物在减压下浓缩，向残余物中加入甲醇，通过过滤收集固体，通过制备型HPLC(甲酸)纯化固体，以得到白色固体目标化合物TDM-180974(化合物2，18.9mg，收率10.6％)。LCMS[M+H] +＝425.2。
1H NMR(400MHz,DMSO)δ10.73(s,1H),10.08(s,1H),8.89(d,J＝2.2Hz,1H),8.60(d,J＝5.3Hz,1H),8.23(d,J＝2.0Hz,1H),8.18(d,J＝8.8Hz,2H),7.90(s,1H),7.79(d,J＝8.8Hz,2H),7.49(d,J＝5.3Hz,1H),7.26(s,1H),2.87(ddd,J＝13.6,10.8,8.1Hz,1H),2.61(s,3H),2.10–1.96(m,2H)。
实施例3合成化合物3的一般方法(TDM-180977)
Figure PCTCN2021120119-appb-000015
步骤1：化合物3b(4-氨基-N-乙基-2-甲基苯甲酰胺)的制备
向化合物3a(1.8g，11.91mmol)的N,N-二甲基甲酰胺(80mL)溶液中加入2-(7-氮杂苯并三氮唑)-N,N,N',N'-四甲基脲六氟磷酸酯(5.4g，14.289mmol)和N,N-二异丙基乙胺(3.8g，29.775mmol)，将混合物搅拌5分钟，然后加入乙胺的四氢呋喃(2M)(9mL，18mmol)溶液，将混合物在室温搅拌过夜。减压浓缩混合物除去一些溶剂，向残余物中加入水并用乙酸乙酯(100mL*3)萃取，合并有机相，用水(150mL*3)和饱和盐水(150mL)洗涤，用硫酸钠干燥，减压浓缩滤液，通过硅胶色谱法纯化(石油醚/乙酸乙酯＝0％-50％)，以得到黄色油状目标化合物(化合物3b，1.32g，收率62.2％)。LCMS[M+1] +＝179。
步骤2：化合物3((S)-4-((4-(4-(4-(2,2-二氟环丙烷-1-羧酰胺基)苯基)嘧啶-2-基)氨基)-N-乙基-2-甲基苯甲酰胺)的制备
向化合物3c(80mg，0.258mmol)的正丁醇(8mL)溶液加入化合物3b(92mg，0.517mmol)和对甲苯磺酸一水合物(98mg，0.517mmol)。将所得混合物加热至110℃并搅拌3小时。反应结束将混合物在减压下浓缩，残余物通过制备型HPLC(甲酸)纯化，以得到白色固体目标化合物TDM-180977(化合物3，19.4mg，收率13.3％。LCMS[M+H] +＝425.2。
1H NMR(400MHz,DMSO)δ10.70(s,1H),9.72(s,1H),8.54(d,J＝5.3Hz,1H),8.18(d,J＝8.8Hz,2H),8.08(d,J＝5.6Hz,1H),7.81–7.67(m,4H),7.40(d,J＝5.3Hz,1H),7.32(d,J＝8.4Hz,1H),3.28–3.17(m,2H),2.86(ddd,J＝13.6,10.8,8.1Hz,1H),2.37(s,3H),2.14–1.93(m,2H),1.11(t,J＝7.2Hz,3H)。
实施例4合成化合物4的一般方法(TDM-180981)
Figure PCTCN2021120119-appb-000016
步骤1：化合物4((S)-5-((4-(4-(4-(2,2-二氟环丙烷-1-羧酰胺基)苯基)嘧啶-2-基)氨基)-N,3-二甲基吡啶啉酰胺)的制备
以与实施例3相似的方法制备得到化合物4(白色固体，3.5mg，收率1.6％)。
1H NMR(400MHz,DMSO)δ10.74(s,1H),10.06(s,1H),8.92(d,J＝2.2Hz,1H),8.60(d,J＝5.3Hz,1H),8.48(d,J＝4.8Hz,1H),8.22–8.18(m,2H),8.17(s,1H),7.78(d,J＝8.8Hz,2H),7.49(d,J＝5.3Hz,1H),2.87(ddd,J＝13.6,10.8,8.0Hz,1H),2.78(d,J＝4.8Hz,3H),2.61(s,3H),2.03(ddd,J＝18.1,11.7,6.0Hz,2H)。LCMS[M+H] +＝439.2。
实施例5合成化合物5的一般方法(TDM-180989)
Figure PCTCN2021120119-appb-000017
步骤1：化合物5c(2-(5-硝基吡啶-2-基)丙二酸1-(叔丁基)3-甲基酯)的制备
在室温下向化合物5a(10g，63.076mmol)的N,N-二甲基甲酰胺(100mL)溶液中加入碳酸钾(17.435g，126.152mmol)和化合物5b(13.185g，75.691mmol)，混合物用氮气置换几次。将混合物加热至100℃并搅拌过夜。反应结束将混合物冷却至室温，然后添加饱和氯化铵溶液(150mL)，将混合物过滤，将滤液用乙酸乙酯(300mL*3)萃取，合并有机层，用饱和盐水(200mL*2)洗涤，用硫酸钠干燥，将滤液减压浓缩，残余物通过硅胶色谱法进行纯化(石油醚/乙酸乙酯 ＝0％-6％)，以得到黄色油状目标化合物(化合物5c，7.286g，收率39％)。LCMS[M-C 4H 9] +＝241.1。
步骤2：化合物5d(1-(叔丁基)3-甲基-2-甲基-2-(5-硝基吡啶-2-基)丙二酸酯)的制备
在0℃下，向化合物5c(7g，23.626mmol)的N,N-二甲基甲酰胺(150mL)溶液中加入碳酸铯(15.4g，47.252mmol)。然后将混合物温热至室温并搅拌10分钟，添加碘甲烷(13.4g，94.505mmol)，在室温搅拌3小时。反应结束在0℃下向混合物中加入水(150mL)，并用乙酸乙酯(150mL*3)萃取，合并有机层，并用饱和盐水(250mL*2)洗涤，用硫酸钠干燥，将滤液减压浓缩，残留物通过硅胶色谱纯化(石油醚/乙酸乙酯＝0％-3％)，得到黄色油状目标化合物(化合物5d，6.026g，收率82.2％)。LCMS[M-C 4H 9] +＝255.1。
步骤3：化合物5e(2-(5-硝基吡啶-2-基)丙酸丙酯)的制备
向化合物5d(6g，19.366mmol)的二氯甲烷(150mL)溶液中加入三氟乙酸(50mL)。将混合物在室温搅拌3小时。反应结束将混合物在减压下浓缩，残余物通过硅胶色谱法纯化(石油醚/乙酸乙酯＝0％-6％)，得到黄色油状目标化合物(化合物5e，3.671g，收率90.3％)。LCMS[M+1] +＝211.1。
步骤4：化合物5f(2-(5-氨基吡啶-2-基)丙酸甲酯)的制备
在室温下向化合5e(3.67g，17.460mmol)的甲醇(120mL)溶液中加入合适的钯碳，混合物用氢气置换几次，然后在室温下搅拌2小时。反应结束将混合物过滤并将滤液减压浓缩，残余物通过硅胶色谱法纯化(石油醚/乙酸乙酯＝0％-50％)，以得到黄色油状目标化合物(化合物5f，2.6025g，收率82.7％)。LCMS[M+1] +＝181。
步骤5：化合物5g(2-(5-氨基吡啶-2-基)丙-1-醇)的制备
在100mL三颈烧瓶中，用氮气置换瓶内空气，在冰浴下加入氢化铝锂(1.8g，47.26mmol)，然后逐滴加入四氢呋喃(180mL)，然后加入化合物5f(1.31g，7.27mmol)的四氢呋喃(20mL)溶液。将反应温热至室温并搅拌2小时。反应结束将反应液在冰浴中冷却，并依次滴加1.8mL水、1.8mL的15％氢氧化钠水溶液和5.4mL水。将混合物过滤以获得滤液，并将滤饼用乙酸乙酯洗涤。将滤 液用无水硫酸钠干燥，过滤并浓缩，以获得黄色油状目标化合物(化合物5g，1.2g，收率95％)。LCMS[M+1] +＝153。
步骤6：化合物5((1S)-2,2-二氟-N-(4-(2-((6-(1-(1-羟基丙烷-2-基)吡啶-3-基)氨基)嘧啶-4-基)苯基)环丙烷-1-羧酰胺)的制备
向化合物389h(100mg，0.323mmol)，化合物389g(98mg，0.646mmol)，醋酸钯(7.3mg，0.032mmol)，4,5-双(二苯基膦)-9,9-二甲基氧杂蒽(37.4mg，0.065mmol)和碳酸铯(316mg，0.969mmol)的混合物中加入二氧六环(25mL)，将混合物用氩气置换几次，将混合物加热至100℃并搅拌1小时。反应结束向反应混合物中加入水(30mL)，并用乙酸乙酯(30mL*3)萃取，合并有机层，然后用水(60mL)和饱和盐水(60mL)洗涤，用硫酸钠干燥，将滤液在减压下浓缩，残余物通过制备型HPLC(甲酸)进行纯化，以得到淡黄色固体目标化合物TDM-180989(化合物5，30.3mg，收率18.4％)。LCMS[M+H] +＝426.1。
1H NMR(400MHz,DMSO)δ10.70(s,1H),9.71(s,1H),8.86(d,J＝2.5Hz,1H),8.52(d,J＝5.3Hz,1H),8.20–8.10(m,3H),7.77(d,J＝8.8Hz,2H),7.39(d,J＝5.3Hz,1H),7.22(d,J＝8.5Hz,1H),4.60(s,1H),3.64(dd,J＝10.3,6.6Hz,1H),3.50(dd,J＝10.3,7.0Hz,1H),2.98–2.81(m,2H),2.13–1.94(m,2H),1.20(d,J＝7.0Hz,3H)。
实施例6合成化合物6的一般方法(TDM-180971)
Figure PCTCN2021120119-appb-000018
步骤1：化合物6c(5-(2-氯嘧啶-4-基)吡啶-2-胺)的制备
向250mL的三口烧瓶中加入化合物6a(1.5g,10mmol)，化合物6b(2.21g，1.01mmol)，四(三苯基膦)钯(1.16g,1mmol)，碳酸钾(2.76g，20mmol),1,4- 二氧六环(60mL)和水(60mL)，反应液用氮气置换数次，升温至80℃搅拌45分钟，LCMS[M+H] +＝227，检测反应完全。后处理：反应液浓缩拉干，得到的粗品过柱，[洗脱剂：(EA/PE)＝0-70％]得到黄色固体目标化合物(化合物6c，1.47g，收率71.36％)，LCMS[M+1] +＝207。
步骤2：化合物6e((S)-N-(5-(2-氯嘧啶-4-基)吡啶-2-基)-2,2-二氟环丙烷-1-羧酰胺)的制备
向化合物6c(1.47g，7.11mmol)的无水吡啶(50ml)溶液加入化合物6d(910mg，7.47mmol)和三氯氧磷(1.64g，10.67mmol)。反应液在室温下搅拌1小时，LCMS[M+H] +＝311，检测反应完全。后处理：反应液浓缩拉干，残余物加水和乙酸乙酯(3*100mL)萃取三次，合并有机相，并用饱和食盐水洗，无水硫酸钠干燥，过滤，浓缩拉干，得到的粗品过柱，[洗脱剂：(EA/PE)＝0-30％]，粗品再重结晶，以得到黄色固体目标化合物(化合物6e，1.73g，收率78.6％)，LCMS[M+H] +＝311。
步骤3：化合物6((S)-4-((4-(6-(2,2-二氟环丙烷-1-甲酰胺基)吡啶-3-基)嘧啶-2-基)氨基)-N-乙基苯甲酰胺)的制备
向化合物6e(100mg,0.32mmol)的正丁醇(10ml)溶液中加入化合物6f(105.7mg，0.64mmol)和一水合对甲苯磺酸(122.5mg，0.64mmol)。反应液升温至110℃搅拌2小时，LCMS[M+H] +＝439，检测反应完全。后处理：反应液浓缩拉干，制备得到黄色固体目标化合物(化合物6，12.3mg，收率8.7％)，LCMS[M+H] +＝439.2。
1H NMR(400MHz,DMSO)δ11.30(s,1H),9.99(s,1H),9.25–9.06(m,1H),8.62(d,J＝5.2Hz,1H),8.57(dd,J＝8.8,2.4Hz,1H),8.34–8.17(m,2H),7.90(d,J＝8.9Hz,2H),7.82(d,J＝8.9Hz,2H),7.53(d,J＝5.2Hz,1H),3.28(dt,J＝12.7,6.4Hz,2H),3.04(dd,J＝8.8,5.7Hz,1H),2.12–1.98(m,2H),1.13(t,J＝7.2Hz,3H)。
实施例7合成化合物7的一般方法(TDM-180979)
Figure PCTCN2021120119-appb-000019
步骤1：化合物7((S)-5-((4-(6-(2,2-二氟环丙烷-1-羧酰胺基)吡啶-3-基)嘧啶-2-基)氨基)-N,3-二甲基吡啶啉酰胺)的制备
向化合物7a(100mg，0.32mmol)的二氧六环(25ml)溶液中加入化合物7b(106.3mg，0.64mmol)，乙酸钯(3.6mg，0.02mmol)，4,5-双(二苯基膦)-9,9-二甲基氧杂蒽(18.6mg，0.03mmol)和碳酸铯(314.7mg，0.97mmol)。反应液用氩气置换数次，并升温至100℃搅拌2小时，LCMS[M+H] +＝440，检测反应完全。后处理：将反应液浓缩至干，残留物用甲醇打浆，粗品制备得到黄色固体目标化合物(化合物7，6.4mg，收率3.2％)，LCMS[M+1] +＝440.0。
1H NMR(400MHz,DMSO)δ11.34(s,1H),10.15(s,1H),9.17(d,J＝2.0Hz,1H),8.87(d,J＝2.2Hz,1H),8.65(d,J＝5.2Hz,1H),8.56(dd,J＝8.8,2.4Hz,1H),8.50(d,J＝4.8Hz,1H),8.30–8.18(m,2H),7.58(d,J＝5.3Hz,1H),3.04(dd,J＝8.9,3.5Hz,1H),2.77(d,J＝4.8Hz,3H),2.61(s,3H),2.05(dt,J＝10.7,5.2Hz,2H)。
实施例8合成化合物8的一般方法(TDM-180980)
Figure PCTCN2021120119-appb-000020
步骤1：化合物8((S)-5-((4-(6-(2,2-二氟环丙烷-1-羧酰胺基)吡啶-3-基)嘧啶-2-基)氨基)-3-甲基吡啶啉酰胺)的制备
以相似的方法制备得到化合物8(2.3mg，黄色固体，收率1.2％)。
1H NMR(400MHz,DMSO)δ11.35(s,1H),10.16(s,1H),9.18(d,J＝2.0Hz,1H),8.84(d,J＝2.2Hz,1H),8.66(d,J＝5.2Hz,1H),8.56(dd,J＝8.8,2.4Hz,1H),8.25(dd,J＝8.2,5.5Hz,2H),7.90(s,1H),7.58(d,J＝5.3Hz,1H),7.25(s,1H), 3.13–2.94(m,1H),2.61(s,3H),2.06(d,J＝9.6Hz,2H)。LCMS[M+H] +＝426.1。
实施例9合成化合物9的一般方法(TDM-180984)
Figure PCTCN2021120119-appb-000021
步骤1：化合物9((S)-4-((4-(6-(2,2-二氟环丙烷-1-羧酰胺基)吡啶-3-基)嘧啶-2-基)氨基)-N-乙基-2-甲基苯甲酰胺)的制备
以相似的方法制备得到化合物9(26.6mg，黄色固体，收率13.04％)。
1H NMR(400MHz,DMSO)δ11.33(s,1H),9.80(s,1H),9.17(d,J＝1.8Hz,1H),8.68–8.49(m,2H),8.22(d,J＝8.7Hz,1H),8.09(t,J＝5.6Hz,1H),7.80–7.61(m,2H),7.50(d,J＝5.2Hz,1H),7.32(d,J＝8.4Hz,1H),3.28–3.19(m,2H),3.04(dt,J＝13.5,9.8Hz,1H),2.37(s,3H),2.12–2.00(m,2H),1.11(t,J＝7.2Hz,3H)。LCMS[M+H] +＝453.2。
实施例10合成化合物10的一般方法(TDM-180987)
Figure PCTCN2021120119-appb-000022
步骤1：化合物10((1S)-2,2-二氟-N-(5-(2-((6-(1-(羟丙基-2-基)吡啶基-3-基)氨基)嘧啶基-4-基)吡啶基-2-基环丙烷-1-羧酰胺)的制备
以相似的方法制备得到化合物10(79mg，类白色固体，收率27.8％)。
1H NMR(400MHz,DMSO)δ11.30(s,1H),9.79(s,1H),9.13(d,J＝2.1Hz,1H),8.84(d,J＝2.5Hz,1H),8.65–8.46(m,2H),8.30–8.07(m,2H),7.48(d,J＝5.2Hz,1H),7.22(d,J＝8.5Hz,1H),4.59(t,J＝5.3Hz,1H),3.71–3.43(m,2H),3.04(dt,J＝13.5,9.9Hz,1H),2.91(dt,J＝14.0,7.0Hz,1H),2.13–1.95(m,2H),1.20(d,J＝6.9Hz,3H)。LCMS[M+H] +＝427.1。
测试例1 JAK激酶小分子抑制剂的酶活性抑制检测
实验方案
1、试剂准备
激酶反应缓冲液：配置激酶反应缓冲液，组分如下：50mM HEPES，pH 7.5，1mM EGTA，10mM MgCl 2，2mM DTT，0.01％Tween20。1X检测缓冲液：配置检测缓冲液，去离子水9:1稀释10X检测缓冲液至1X。4X激酶溶液：激酶反应缓冲液稀释JAK激酶至4X终浓度(JAK1：80nM，JAK2/JAK3/Tyk2：4nM)。4X底物溶液：激酶反应缓冲液稀释ULight TM-JAK(Tyr1023)底物至200nM(终浓度：50nM)。4XATP溶液：激酶反应缓冲液稀释ATP至4X终浓度(JAK1：160μM，JAK2/JAK3/Tyk2：40μM)。4X测试化合物溶液：DMSO溶解测试用化合物至10mM储存液，3倍梯度稀释配置成所需浓度，每个化合物设置10个浓度点，测试化合物终浓度范围为：10μM-0.5nM。4X酶反应终止液：1X检测缓冲液溶解EDTA至40mM(EDTA终浓度：10mM)。4X检测抗体溶液：1X检测缓冲液稀释Eu标记检测抗体(抗磷酸酪氨酸(PT66))至8nM(抗体终浓度：2nM)。
2、实验过程
向384微孔板中依次加入2.5μL的4X激酶溶液和2.5μL已经稀释好的不同浓度的4X测试化合物溶液，每个浓度设置2个复孔，同时设置酶溶液空白对照组和阴性对照组(DMSO组)。震荡384多孔板，混匀酶和化合物，1000转，离心1分钟，在室温下孵育60分钟。向384多孔板中加入2.5μL，4X底物溶液，1000转离心1分钟。向384多孔板中加入2.5μL，4XATP溶液，1000转离心1分钟，起始酶反应。JAK1室温反应2小时，JAK2/JAK3/Tyk2室温反应1小时。JAK1反应的各组分终浓度分别为：JAK1：20nM，底物：50nM，ATP：40uM，测试化合物终浓度范围为：10μM-0.5nM。JAK2/JAK3/Tyk2反应的各组分终浓度分别为：JAK2：1nM，底物：50nM，ATP：10μM，测试化合物终浓度范围为：10μM-0.5nM。酶反应结束后，向384多孔板每孔中加入5μL，4X酶反应终止液，1000转，离心1分钟，在室温下孵育5分钟。向384多孔板每孔中加入5μL，4X检测抗体溶液，(检测抗体终浓度为2nM)，1000转，离心1分钟，室温条件下孵育1小时。抗体孵育结束后，在Envision读板仪上测定各孔的信号值
3、数据分析
以酶溶液空白对照组为100％抑制率和阴性对照组(DMSO组)为0％抑制 率，计算检测各个浓度对应的百分比抑制率。在GraphPad Prism软件中对检测化合物的浓度对数值和相对应的百分比抑制率进行非线性回归分析，得到检测化合物的半数抑制浓度(IC 50)，针对实施例1-10的化合物所测得的实验结果列在下表1中。
表1
编号 Tyk2/μM JAK1/μM JAK2/μM JAK3/μM
TDM-180972 0.005 0.004 0.003 0.117
TDM-180974 0.008 0.007 0.007 0.346
TDM-180977 0.004 0.003 0.004 0.057
TDM-180981 0.008 0.008 0.008 0.183
TDM-180989 0.014 0.011 0.013 0.177
TDM-180971 0.005 0.006 0.007 0.219
TDM-180979 0.017 0.025 0.033 0.931
TDM-180980 0.018 0.028 0.030 >5
TDM-180984 0.005 0.008 0.009 0.105
TDM-180987 0.017 0.025 0.022 0.310
从上表1的结果可以看出，本申请的化合物的酶活性数据非常优异，以上具体化合物测得的半数抑制浓度较低，特别是针对Tyk2、JAK1和JAK2时基本上达到小于0.03μM，尤其是化合物TDM-180972、TDM-180974、TDM-180977、TDM-180981、TDM-180971和TDM-180984，其半数抑制浓度均小于0.01μM。因此，通过以上实验已经证明了本申请的小分子化合物是一类针对JAK家族靶向性强、酶活性优异的化合物，能够作为JAK1/Tyk2双抑制剂和Tyk2特异性抑制剂。
以上详细描述了本发明的优选实施方式，但是，本发明并不限于上述实施方式中的具体细节，在本发明的技术构思范围内，可以对本发明的技术方案进行多种简单变型，这些简单变型均属于本发明的保护范围。
另外需要说明的是，在上述具体实施方式中所描述的各个具体技术特征， 在不矛盾的情况下，可以通过任何合适的方式进行组合，为了避免不必要的重复，本发明对各种可能的组合方式不再另行说明。
此外，本发明的各种不同的实施方式之间也可以进行任意组合，只要其不违背本发明的思想，其同样应当视为本发明所公开的内容。

Claims (11)

  1. 一种小分子化合物，其特征在于，所述小分子化合物为由如下所示的式I表示的化合物，或其立体异构体、几何异构体、互变异构体、水合物、溶剂化物、以及药学上可接受的盐或前药：
    Figure PCTCN2021120119-appb-100001
    其中，R 1至R 4各自独立地选自C或N；并且
    其中，R选自环烷基、取代的环烷基、杂环烷基、取代的杂环烷基、芳基、取代的芳基、杂芳基或取代的杂芳基。
  2. 根据权利要求1所述的小分子化合物，其特征在于，R 1至R 4中的至多两个为N。
  3. 根据权利要求1所述的小分子化合物，其特征在于，R具有如下所示的式II表示的结构：
    Figure PCTCN2021120119-appb-100002
    其中，R 5选自C或N；
    其中，R 6选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
    其中，R 7选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
  4. 根据权利要求3所述的小分子化合物，其特征在于，R 6和R 7各自独立地选自氢、烷基或环烷基。
  5. 根据权利要求1所述的小分子化合物，其特征在于，R具有如下所示的式III表示的结构：
    Figure PCTCN2021120119-appb-100003
    其中，R 8选自C或N；
    其中，R 9选自氢、卤素、烷基、氨基、酰胺基、环烷基、杂环烷基、芳基或杂芳基；并且
    其中，R 10选自氢、烷基、环烷基、杂环烷基、芳基或杂芳基。
  6. 根据权利要求5所述的小分子化合物，其特征在于，R 9和R 10各自独立地选自氢、烷基或环烷基。
  7. 根据权利要求4或6所述的小分子化合物，其特征在于，所述烷基为甲基、乙基、丙基或异丙基，所述环烷基为环丙基、环丁基或环丙基甲基。
  8. 根据权利要求1-7中任一项所述的小分子化合物在抑制JAK激酶中的用途。
  9. 根据权利要求1-7中任一项所述的小分子化合物在制备用于预防或治疗自身免疫性疾病、以及与免疫有关的炎症性皮肤疾病的药物中的用途，其中，这些疾病的发病机理均与JAK信号传导的失调相关。
  10. 根据权利要求9所述的用途，其中，所述自身免疫性疾病选自类风湿性关节炎、强直性脊柱炎、溃疡性结肠炎、克罗恩病、系统性红斑狼疮、皮肌炎、多发性硬化、I型糖尿病、干燥综合症和血管炎中的至少一种。
  11. 根据权利要求9所述的用途，其中，所述与免疫有关的炎症性皮肤疾病选自特应性皮炎、湿疹、斑秃、银屑病、白癜风、扁平苔藓、光泽苔藓、硬化萎缩性苔藓、脂膜炎、痤疮和化脓性汗腺炎中的至少一种。
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EP4227298A1 (en) 2023-08-16
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