Tricyclic compounds as JAK inhibitors and PHD inhibitors

Abstract

The present invention discloses tricyclic compounds as JAK inhibitors and PHD inhibitors. Specifically, the present invention discloses a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, which can be used to treat inflammatory bowel disease.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and in particular relates to tricyclic compounds serving as JAK inhibitors and PHD inhibitors. Background Art

[0002] The JAK family of cellular protein tyrosine kinases (JAK1, JAK2, JAK3, and TYK2) play an important role in cytokine signal transduction. When cytokines bind to their receptors, the cytokines activate the JAKs, which then phosphorylate the cytokine receptors, thereby creating docking sites for signal transduction molecules, notably members of the signal transducer and activator of transcription (STAT) family that ultimately lead to gene expression. Many cytokines are known to activate the JAK family. These cytokines include interferon IFN family (IFN-α, IFN-β, IFN-ω, limitin, IFN-γ, IL-10, IL-19, IL-20, IL-22), gp130 family (IL-6, IL-11, OSM, LIF, CNTF, NNT-1 / BSF-3, G-CSF, CT-1, leptin, IL-12, IL-23), γC family (IL-2, IL-7, TSLP, IL-9, IL-15, IL-21, IL-4, IL-13), IL-3 family (IL-3, IL-5, GM-CSF), single chain family (EPO, GH, PRL, TPO), receptor tyrosine kinase (EGF, PDGF, CSF-1, HGF), and G-protein coupled receptor (AT1). JAK1 is expressed at different levels in all tissues. Animal studies have shown that JAK1 is essential for the development, function and homeostasis of the immune system. Modulating immune activity by inhibiting JAK1 kinase activity has been shown to be useful in treating a variety of immune disorders.

[0003] Inflammatory bowel disease (IBD) is a chronic disease in which inflammation and ulcers are caused in the intestinal mucosa due to excessive immune response. IBD includes, for example, ulcerative colitis and Crohn's disease. It is known that in the pathological conditions of IBD, the expression of genes related to the barrier function of the gastrointestinal epithelium is induced by hypoxia-inducible factor 1α (HIF-1α). HIF-1α is one of the subtypes of hypoxia-inducible factor α (HIF-α). HIF-α becomes stable in a hypoxic environment (hypoxia), and then it responds to hypoxia and activates the transcription of several genes. In contrast, the proline residues of HIF-α are hydroxylated by prolyl hydroxylases (PHDs) in an oxygen-rich environment (normoxia), and then the HIF-α is degraded by the proteasome pathway. It is known that PHD has 3 subtypes, namely PHD1, PHD2 and PHD3. Summary of the invention

[0004] The invention provides a compound with a novel structure and capable of simultaneously inhibiting JAK1 and PHD2.

[0005] In the first aspect of the present invention, the present invention provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,

[0006]

[0007] R 1 Selected from C 3-6 Cycloalkyl, the C 3-6 The cycloalkyl group is optionally substituted with 1 C 1-6 Alkyl, C 1-6 Alkyl cyano or -NHC(=O)OC 1-6 Alkyl substitution;

[0008] L is selected from -L 1 -L 2 -L 3 -L 4 -;

[0009] L 1 Selected from single bond, C 1-6 Alkyl, the C 1-6 The alkyl group is optionally substituted with 1 or 2 halogen or C 1-6 Alkyl substitution;

[0010] L 2 Selected from a single bond, O or -N(R 2 )-;

[0011] L 3 Selected from single bond, C 1-6 Alkyl, the C 1-6 The alkyl group is optionally substituted with 1 or 2 halogen or C 1-6 Alkyl substitution;

[0012] L 4 Selected from a single bond, O or -N(R 2 )-;

[0013] R 2 Select from H or C 1-6 alkyl.

[0014] In some embodiments of the present invention, L 1 Selected from C 1-3 Alkyl, the C 1-3 The alkyl group is optionally substituted with 1 or 2 halogen or C 1-3 Alkyl substitution.

[0015] In some embodiments of the present invention, L 1 is selected from -CH2-, -CH2CH2- or -CH2CH2CH2-, and the remaining variables are as defined herein.

[0016] In some embodiments of the present invention, L 2 Selected from a single bond, O or -N(R 2 )-, the R 2 Select from H or C 1-3 Alkyl, selected from H or C 1-6 The remaining variables are as defined herein.

[0017] In some embodiments of the present invention, L 2 is selected from a single bond, O or -NH-, and the remaining variables are as defined in the present invention.

[0018] In some embodiments of the present invention, L 3 Selected from single bond, C 1-3 Alkyl, the C 1-3 The alkyl group is optionally substituted with 1 or 2 halogen or C 1-3 Alkyl substitution.

[0019] In some embodiments of the present invention, L 3 is selected from a single bond, -CH2-, -CH2CH2- or -CH2CH2CH2-, and the remaining variables are as defined herein.

[0020] In some embodiments of the present invention, L 4 Selected from a single bond, O or -N(R 2 )-, the R 2 Select from H or C 1-3 Alkyl, selected from H or C 1-6 The remaining variables are as defined herein.

[0021] In some embodiments of the present invention, L 4 is selected from a single bond, O or -NH-, and the remaining variables are as defined in the present invention.

[0022] In some embodiments of the present invention, L is selected from The remaining variables are as defined in the present invention.

[0023] In some embodiments of the present invention, R 1 Selected from C 5-6 Cycloalkyl, the C 5-6 The cycloalkyl group is optionally substituted with 1 C 1-3 Alkyl, C 1-3 Alkyl cyano or -NHC(=O)OC 1-3 The remaining variables are as defined herein.

[0024] In some embodiments of the present invention, R 1 Selected from The remaining variables are as defined in the present invention.

[0025] In some embodiments of the present invention, the structure of the compound represented by formula (I) is shown as one of the following:

[0026]

[0027]

[0028] In another aspect of the present invention, the present invention provides a pharmaceutical composition, which comprises the aforementioned compound or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

[0029] In another aspect of the present invention, the present invention provides the use of the aforementioned compound or a pharmaceutically acceptable salt thereof or the aforementioned pharmaceutical composition in the preparation of a drug for treating diseases mediated by PHD2 and JAK1.

[0030] In some embodiments of the present invention, the PHD2 and JAK1 mediated disease is inflammatory bowel disease.

[0031] Unless otherwise specified, the term “C 1-6 "Alkyl" is used to refer to a straight or branched chain saturated hydrocarbon group consisting of 1 to 6 carbon atoms. 1-6 Alkyl groups include C 1-5 , C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-4 , C6 and C5 alkyl, etc.; it can be monovalent (such as CH3), divalent (-CH2-) or polyvalent (such as ).

[0032] Unless otherwise specified, the term “C 1-6 "Alkylcyano" represents a stable C 1-6 Alkyl. "Methyl cyano" represents -CH2CN, "ethyl cyano" represents -CH2CH2CN or -C(CN)HCH3 or -C(CN)HC(CN)H2, and the like.

[0033] Unless otherwise specified, “C 3-6 "Cycloalkyl" means a saturated cyclic hydrocarbon group consisting of 3 to 6 carbon atoms, which is a monocyclic or bicyclic system. 3-6 Cycloalkyl groups include C 3-5 , C 4-5 and C 5-6 Cycloalkyl, etc.; it may be monovalent, divalent or polyvalent. 3-6 Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

[0034] Unless otherwise specified, the term "3-6 membered heterocyclyl" by itself or in combination with other terms refers to a saturated or partially unsaturated cyclic group consisting of 3 to 6 ring atoms, 1, 2, 3 or 4 of which are heteroatoms independently selected from O, S and N, and the rest are carbon atoms, wherein the nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms are optionally oxidized (i.e., NO and S(O) p , p is 1 or 2). It includes monocyclic and bicyclic systems, wherein bicyclic systems include spirocyclic, fused and bridged rings. In addition, with respect to the "3-6 membered heterocyclic group", a heteroatom may occupy the position where the heterocyclic group is connected to the rest of the molecule. The 3-6 membered heterocyclic group includes 4-6 membered, 5-6 membered, 4 membered, 5 membered and 6 membered heterocyclic groups, etc. Examples of 3-6 membered heterocyclic groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, 1,3-dioxolane, Pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothiophenyl (including tetrahydrothiophen-2-yl and tetrahydrothiophen-3-yl, etc.), tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2-piperidinyl and 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl and 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl and 4-morpholinyl, etc.), dioxanyl, dithianyl, isoxazolidinyl, isothiazolidinyl, 1,2-oxazinyl, 1,2-thiazinyl, hexahydropyridazinyl, homopiperazinyl or homopiperidinyl, etc.

[0035] The term "5-6 membered heteroaryl" means a 5- or 6-membered aromatic heterocyclic group having 1 to 4 heteroatoms selected from oxygen atoms, nitrogen atoms and sulfur atoms in the ring. For example, furyl, pyrrolyl, thienyl, imidazolyl, pyrazolyl, 1,2,4-triazolyl, isothiazolyl, isoxazolyl, oxazolyl, thiazolyl, 1,3,4-oxadiazolyl, 1,2,4-oxadiazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl and the like can be exemplified.

[0036] As used herein, the term "pharmaceutically acceptable salt" refers to a salt of a compound of the present invention that is pharmaceutically acceptable and has the pharmacological activity of the parent compound. Such salts include: acid addition salts formed with inorganic acids or organic acids, such as nitric acid, phosphoric acid, carbonic acid, etc.; organic acids such as propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, gluconic acid, stearic acid, muconic acid, etc.; or salts formed when the acidic protons present on the parent compound are replaced by metal ions, such as alkali metal ions or alkaline earth metal ions; or coordination compounds formed with organic bases, such as ethanolamine, diethanolamine, triethanolamine, N-methylglucamine, etc. The pharmaceutically acceptable salts of the present invention can be synthesized from parent compounds containing acid radicals or bases by conventional chemical methods. In general, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of an appropriate base or acid in water or an organic solvent or a mixture of the two. Generally, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred. In addition to the form of salts, compounds provided by the present invention also exist in prodrug forms. Prodrugs of compounds described herein easily undergo chemical changes under physiological conditions to be converted into compounds of the present invention. In addition, prodrugs can be converted to compounds of the present invention by chemical or biochemical methods in an in vivo environment.

[0037] As used herein, the term “C 1-6 The term "alkyl" refers to a straight or branched monovalent alkyl group having 1 to 6 carbon atoms. Specific examples include methyl, ethyl, n-propyl and isopropyl, and various branched isomers thereof.

[0038] As used herein, the term “C 1-3 The term "alkyl" refers to a straight or branched monovalent alkyl group having 1 to 3 carbon atoms. Specific examples include methyl, ethyl, n-propyl and isopropyl, and various branched isomers thereof.

[0039] As used herein, the term “C 1-6 The term "alkylene" refers to a straight or branched divalent alkyl group having 1 to 3 carbon atoms. Specific examples include methyl, ethyl, n-propyl and isopropyl, and various branched isomers thereof.

[0040] The compound shown in formula I of the present invention can be prepared using a synthetic method known in the art or using a method known in the art in combination with the method recorded in the present invention. The solvent, temperature and other reaction conditions given in the present invention are exemplary and can be changed according to methods well known in the art. The embodiment compounds recorded in the present invention can be synthesized according to the method recorded in the embodiment using appropriate starting materials according to their specific structure, or can be synthesized using a similar method recorded in the embodiment. The starting materials used to synthesize the embodiment compounds of the present invention can be prepared by known synthetic methods or similar methods recorded in the literature or obtained from commercial sources. The compound can be further separated by methods well known in the art, such as crystallization, chromatography, etc. to obtain its stereoisomers as needed, and the separation conditions are easily obtained by those skilled in the art through conventional means or limited experiments. As a further illustration, the compound of formula I of the present invention can be synthesized using the following method, wherein the solvent, temperature and other reaction conditions in each step can be the same or similar to those recorded in the following embodiments, or reaction conditions known in the art are used. DETAILED DESCRIPTION

[0041] The compounds of the present invention can be prepared by a variety of synthetic methods known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent substitutions known to those skilled in the art. Preferred embodiments include, but are not limited to, the embodiments of the present invention.

[0042] The present invention is described in detail below by way of examples, but it is not intended to limit the present invention in any adverse way. The present invention has been described in detail herein, and its specific embodiments are also disclosed. It will be apparent to those skilled in the art that various changes and modifications will be made to the specific embodiments of the present invention without departing from the spirit and scope of the present invention. If specific conditions are not specified in the examples, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer is not specified for the reagents or instruments used, they are all conventional products that can be obtained commercially.

[0043] The following abbreviations are used throughout this disclosure.

[0044] Fmoc represents 9-fluorenylmethoxycarbonyl, THF represents tetrahydrofuran, MeOH represents methanol, KOH represents potassium hydroxide, Cs2CO3 represents cesium carbonate, Na2SO4 represents sodium sulfate, DMA represents dimethylacetamide, Pd(OAc)2 represents palladium acetate, BINAP represents 1,1'-binaphthyl-2,2'-bisdiphenylphosphine, DCM represents dichloromethane, Na2S2O4 represents sodium dithionite, DMSO represents dimethyl sulfoxide, DIPEA represents N,N-diisopropylethylamine, i-PrOH represents isopropanol, EtOAc represents ethyl acetate, and NMP represents N-methyl-2-pyrrolidone.

[0045] Example 1

[0046]

[0047] Step 1:

[0048] Compound 1-1 (1.00 g, 2.84 mmol) and (1R, 3R)-3-aminocyclopentylcarbamic acid tert-butyl ester (0.540 g, 2.70 mmol) were dissolved in i-PrOH (2.00 mL), DIPEA (0.895 mL, 5.41 mmol) was added, and the mixture was stirred at 80°C for 12 hours. After concentration, the crude product 1-2 was obtained, which was used directly in the next step without purification. LCMS: 516.3 [M+H] + .

[0049] Step 2:

[0050] Compound 1-2 (500 mg, 0.970 mmol) was dissolved in DMSO (3.00 mL), and MeOH (10.0 mL), H2O (5.00 mL), N-fluorenylmethoxycarbonylglycinealdehyde (545 mg, 1.94 mmol) and Na2S2O4 (422 mg, 2.42 mmol) were added. The mixture was reacted at 100 °C for 12 hours. The reaction was quenched with H2O (100 mL) and extracted with EtOAc (50.0 mL×3). The organic phase was washed with saturated brine (100 mL×3), dried over Na2SO4, filtered and concentrated to obtain compound 1-3.

[0051] Step 3:

[0052] Compound 1-3 (600 mg, 0.928 mmol) was dissolved in MeOH (6.00 mL) and DCM (1.00 mL), (Boc)2O (1.06 mL, 4.63 mmol) was added, and the mixture was reacted at room temperature for 12 hours. The reaction was quenched with H2O (10.0 mL) and extracted with EtOAc (5.00 mL×3). The organic phase was washed with saturated brine (10.0 mL×3), dried over Na2SO4, filtered and concentrated. The crude product was purified by column chromatography to obtain compound 1-4.

[0053] Step 4:

[0054] Compound 1-4 (248 mg, 0.332 mmol) was dissolved in THF (2.00 mL), KOH (553.4 μL, 3.32 mmol, 6.00 M) was added, and the mixture was reacted at 80°C for 12 hours. The reaction was quenched with H2O (100 mL) and extracted with EtOAc (50.0 mL×3). The organic phase was washed with saturated brine (100 mL×3), dried over Na2SO4, filtered and concentrated. The crude product was purified by preparative TLC to obtain compound 1-5. LCMS: 371.2 [M+H] + .

[0055] Step 5:

[0056] Compound 1-5 (60.0 mg, 0.162 mmol) was dissolved in NMP (2.00 mL), and 2-chloro-N-((6-cyanopyridin-3-yl)methyl)-5-hydroxy-1,7-naphthyridine-6-carboxamide (27.5 mg, 0.0810 mmol) and NMP (2.00 mL) were added. The mixture was reacted under microwave at 100°C for 4 hours. The crude product was purified by preparative HPLC (FA conditions) to obtain compound 1. LCMS: 674.5 [M+H] + . 1 HNMR: (400MHz, DMSO-d6)δ=11.89(br s,1H),8.76(s,1H),8.56(s,1H),8.48-8.29(m,2H),8.20(d,J=9.1Hz,1H),8.04-7.96(m,2H),7.49(br t,J=2.8Hz,1H),7.25-7.15(m,2H),6.57(br s,1H),5.36(br t,J=9.5Hz,1H),5.22-4.87(m,2H),4.63(br d,J=6.3Hz,2H),4.40-4.24(m,1H),2.41-2.17(m,5H),2.10-2.02(m,1H),1.78-1.62(m,1H),1.37(s,9H).

[0057] Example 2

[0058]

[0059] Step 1:

[0060] Compound 2-1 (3.28 g, 9.32 mmol) was dissolved in DMA (30.0 mL), and 2-[trans-4-aminocyclohexyl]acetonitrile hydrochloride (1.79 g, 10.257 mmol) and DIPEA (4.10 g, 31.7 mmol) were added under N2 atmosphere. The mixture was reacted at 80°C for 4 hours. After cooling to room temperature, the mixture was diluted with H2O (200 mL) and extracted with DCM (300 mL×2). The organic phase was washed with saturated brine (300 mL), dried over Na2SO4, filtered and concentrated to obtain crude 2-2, which was used directly in the next step without purification. LCMS: 454.3 [M+H] + .

[0061] Step 2:

[0062] Compound 2-2 (900 mg, 1.98 mmol) and 2-(2-((tert-butyldimethylsilyl chloride)-oxy)ethoxy)acetaldehyde (433 mg, 1.98 mmol) were dissolved in DMSO (3.0 mL), MeOH (10.0 mL) and H2O (5.0 mL), and N2S2O4 (1.07 g, 6.15 mmol) was added. The mixture was reacted at 100°C in N2 atmosphere for 15 hours. After cooling to room temperature, it was quenched with H2O (50.0 mL) and extracted with DCM (50.0 mL×2). The organic phase was washed with saturated brine (50.0 mL), dried over Na2SO4, filtered and concentrated. The crude product was purified by column chromatography to obtain compound 2-3. LCMS: 508.1 [M+H] + .

[0063] Step 3:

[0064] Compound 2-3 (500 mg, 0.985 mmol) was dissolved in THF (5.0 mL) and MeOH (5.0 mL), and NaOH (0.8 mL, 2.46 mmol) and tetrahydropyrrole (1.61 g, 3.94 mmol) were added. The mixture was stirred at room temperature for 12 hours, quenched with NH4Cl (50.0 mL), and extracted with EtOAc (50 mL×2). The organic phase was washed with saturated NH4Cl (50.0 mL), dried over Na2SO4, filtered and concentrated to obtain crude product 2-4, which was used directly in the next step without purification. LCMS: 354.1 [M+H] + .

[0065] Step 4:

[0066] Compound 2-4 (120 mg, 0.340 mmol) and 2-chloro-N-((6-cyanopyridin-3-yl)methyl)-5-hydroxy-1,7-naphthyridine-6-carboxamide (115 mg, 0.340 mmol) were dissolved in DMA (1.0 mL), and Pd(OAc)2 (7.62 mg, 0.034 mmol), BINAP (21.1 mg, 0.034 mmol) and Cs2CO3 (221 mg, 0.679 mmol) were added. The mixture was reacted at 130°C for 2 hours under N2 atmosphere. After cooling to room temperature, it was purified by preparative HPLC (0.1% FA) to obtain compound 2. LCMS: 657.5 [M+H] + . 1 HNMR: (DMSO-d6)δ=14.17-12.51(m,1H),10.21-9.76(m,1H),9.49(br d,J=8.6Hz,1H),8.82-8.70(m,4H),8.57(br d,J=3.6Hz,1H),8.05-7.97(m,2H),7.16(br d,J=3.8Hz,1H),4.88(br s,2H),4.67(br d,J=5.4Hz,4H),3.53(br dd,J=3.6,11.3Hz,4H),2.61(br d,J=5.9Hz,2H),2.31(br d,J=11.1Hz,2H),2.04(br d, J=9.6Hz, 5H), 1.46 (brd, J=11.1Hz, 2H).

[0067] Example 3

[0068]

[0069] Prepared in a similar manner to Example 2. LCMS: 613.5 [M+H] + . 1HNMR: (400MHz, DMSO-d6) δ13.18-13.98(m,1H),9.84-10.26(m,1H),9.48-9.68( m,1H),8.71-8.95(m,4H),8.63(d,J=3.63Hz,1H),7.96-8.08(m,2H),7.21(d,J=3 .00Hz,1H),5.80(t,J=5.38Hz,1H),4.81-4.89(m,2H),4.65-4.73(m,2H),2.63( d,J=6.00Hz,2H),2.32-2.39(m,3H),2.04(d,J=10.38Hz,4H),1.40-1.52(m,2H).

[0070] Example 4

[0071]

[0072] Prepared in a similar manner to Example 2. LCMS: 627.6 [M+H] + . 1 HNMR: (400MHz, DMSO-d6) δ9.60-9.31(m,1H),8.90-8.79(m,2H),8.78-8.66(m,2H ),8.60(d,J=3.1Hz,1H),8.22-8.17(m,1H),8.07-7.96(m,2H),7.21-7.13(m,1H), 5.03-4.81(m,1H),4.77-4.54(m,3H),3.95-3.84(m,2H),4.12-3.83(m,1H),2.62( d,J=6.0Hz,2H),2.37-2.27(m,2H),2.03(d,J=9.6Hz,7H),1.50(d,J=13.1Hz,2H).

[0073] Example 5

[0074]

[0075] Prepared in a similar manner to Example 2. LCMS: 641.4 [M+H] + . 1HNMR: (400MHz, DMSO-d6) δ9.60-9.31(m,1H),8.90-8.79(m,2H),8.78-8.66(m,2H ),8.60(d,J=3.1Hz,1H),8.22-8.17(m,1H),8.07-7.96(m,2H),7.21-7.13(m,1H), 5.03-4.81(m,1H),4.77-4.54(m,3H),3.95-3.84(m,2H),4.12-3.83(m,1H),2.62( d,J=6.0Hz,2H),2.37-2.27(m,2H),2.03(d,J=9.6Hz,7H),1.50(d,J=13.1Hz,2H).

[0076] Biological test cases

[0077] JAK

[0078] With Echo 655 (Beckman, 655SYSTEM) to transfer 40nL of compound to a 384 reaction plate (manufacturer: Greiner, catalog number: 784075). Prepare 2× kinase solution with 1× kinase reaction buffer (50mM Hepes, 10mM MgCl2, 0.01% Brij-35, 1mM EGTA, 2mM DTT), and transfer 2μL of JAK1 (25nM, manufacturer: Carna, catalog number: 08-144) solution to the 384 reaction plate. Centrifuge at 1000rpm for 1 minute and incubate at 25℃ for 10 minutes. Prepare a 2× mixture of substrate (IRS1: 0.05mg / mL) and ATP (20μM, manufacturer: Promega, catalog number: V915B) with kinase reaction buffer, start the reaction after adding 2μL of the mixture of substrate and ATP to the reaction plate, and centrifuge at 1000rpm for 1 minute. Incubate at 25°C for 60 minutes. Transfer 4 μL of ADP-Glo ​​to the 384 assay plate, centrifuge at 1000 rpm for 1 minute, and incubate at 25°C for 40 minutes. Transfer 8 μL of the assay solution to the 384 assay plate, centrifuge at 1000 rpm for 1 minute, incubate at 25°C for 40 minutes, and read the luminescence signal of each well in the ADP-Glo ​​mode on an ELISA reader (manufacturer: BMG, model: PHERAstar FSX). Set the reading of the negative control to 0% inhibition and the reading of the positive control to 100% inhibition, calculate the inhibition rate of each test solution, and then perform data analysis using GraphPad 8 software, and use the nonlinear fitting formula to obtain the IC of the compound. 50 (half inhibitory concentration), the results are shown in Table 1 below.

[0079] PHD2

[0080] Use DMSO to make gradient dilutions of the test compound. Use ECHO to add 40nL / well of the compound to the 384-well reaction plate. Seal the experimental plate and centrifuge at 1000rpm for 1 minute. Add 4μL / well 2× PHD2 enzyme working solution to the 384-well reaction plate with the above compounds. Seal the experimental plate, centrifuge at 1000rpm for 1 minute, and incubate at 25℃ for 30 minutes. Prepare 2× substrate working solution (CODD peptide and α-ketoglutarate) and add 4μL / well 2× substrate working solution to the 384-well reaction plate with the above compounds. Seal the experimental plate, centrifuge at 1000rpm for 1 minute, and incubate at 25℃ for 30 minutes. Add 4μL / well 4× prepared stop working solution to the 384-well reaction plate to stop the reaction. Seal the experimental plate and centrifuge at 1000rpm for 1 minute. Add 4 μL / well 4× detection working solution (Alphascreen streptavidin donor beads, AlphaScreen A protein acceptor beads and hydroxy-HIF-1α (Pro564) (D43B5) Rabbit mAb) into a 384 assay plate. The assay plate was sealed, centrifuged at 1000 rpm for 1 minute, and incubated at 25°C for 30 minutes. The fluorescence signal was detected on an Envision 2104 Reader. The results are shown in Table 1 below.

[0081] Table 1

[0082]

[0083] Although the specific embodiments of the present invention have been described in detail, according to all the teachings disclosed, those skilled in the art can make various modifications and substitutions to the details of the technical solution of the present invention, and these changes are within the protection scope of the present invention. The full scope of the present invention is given by the attached claims and any equivalents thereof.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof, R 1 Selected from C 3-6 Cycloalkyl, the C 3-6 The cycloalkyl group is optionally substituted with 1 C 1-6 Alkyl, -C 1-6 Alkyl cyano or -NHC(=O)OC 1-6 Alkyl substitution; L is selected from -L 1 -L 2 -L 3 -L 4 -; L 1 Selected from C 1-3 Alkylene; L 2 Selected from O or -NH-; L 3 is selected from a single bond or -CH2CH2-; L 4 is selected from a single bond or O.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein L 1 Independently selected from -CH2-, -CH2CH2- or -CH2CH2CH2-.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein L is selected from 4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 1 Selected from C 5-6 Cycloalkyl, the C 5-6 The cycloalkyl group is optionally substituted with 1 C 1-3 Alkyl, -C 1-3 Alkyl cyano or -NHC(=O)OC 1-3 Alkyl substitution.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 1 Selected from 6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The structure of the compound represented by formula (I) is shown as one of the following:

7. A pharmaceutical composition, wherein: The pharmaceutical composition comprises the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

8. Use of the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 7 in the preparation of a drug for treating diseases mediated by PHD2 and JAK1.

9. The use according to claim 8, wherein: The PHD2 and JAK1 mediated disease is inflammatory bowel disease.

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