A class of tricyclic compounds that inhibit JAK and PHD

Abstract

The present invention discloses a class of tricyclic compounds that inhibit JAK and PHD. 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 specifically relates to a class of tricyclic compounds that inhibit JAK and PHD. 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] L is selected from -CH2-, -C(CH3)H-, -C(CH3)HCH2- or -CH2C(CH3)H-;

[0008] X is selected from -NH-, -N(CH3)- or -O-;

[0009] And when X is selected from O, L is not -CH2-.

[0010] In some embodiments of the present invention, -LX- is selected from -CH2NH-, -CH2N(CH3)-, -C(CH3)HNH-, -CH(CH3)CH2O- or -CH2CH(CH3)O-, and the remaining variables are as defined herein.

[0011] In some embodiments of the present invention, the structure of the compound represented by formula (I) is as follows:

[0012]

[0013] In some embodiments of the present invention, the structure of the compound represented by formula (I) is as follows:

[0014]

[0015] In some embodiments of the present invention, the structure of the compound represented by formula (I) is as follows:

[0016]

[0017] In some embodiments of the present invention, the structure of the compound represented by formula (I) is as follows:

[0018]

[0019] In some embodiments of the present invention, the structure of the compound represented by formula (I) is as follows:

[0020]

[0021] 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.

[0022] 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.

[0023] In some embodiments of the present invention, the PHD2 and JAK1 mediated disease is inflammatory bowel disease.

[0024] 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.

[0025] 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 described 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 described in the present invention can be synthesized according to the method described in the embodiment using appropriate starting materials according to their specific structure, or can be synthesized using a similar method described 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 described 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 described in the following embodiments, or reaction conditions known in the art can be used. DETAILED DESCRIPTION

[0026] 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.

[0027] 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.

[0028] The following abbreviations are used throughout this disclosure.

[0029] DCM represents dichloromethane, DIPEA represents N,N-diisopropylethylamine, DMA represents dimethylacetamide, DMSO represents dimethyl sulfoxide, EtOAc represents ethyl acetate, FA represents formic acid, i-PrOH represents isopropanol, Int represents 2-chloro-N-((6-cyanopyridin-3-yl)methyl)-5-hydroxy-1,7-naphthyridine-6-carboxamide (which can be prepared by the method of step 1 of Example 29 in WO2023072257), MeOH represents methanol, Na2S2 O4 represents sodium dithionite, Na2SO4 represents sodium sulfate, NaOH represents sodium hydroxide, NH4Cl represents ammonium chloride, NMP represents N-methyl-2-pyrrolidone, PE represents petroleum ether, THF represents tetrahydrofuran, TFA represents trifluoroacetic acid, TBS represents tert-butyldimethylsilyl, Ts represents p-toluenesulfonyl, Xantphos represents 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene, and Xphos represents 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl.

[0030] Example 1

[0031]

[0032] Step 1:

[0033] Compound 1-1 (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 1-2, which was used directly in the next step without purification. LCMS: 454.3 [M+H] + .

[0034] Step 2:

[0035] Compound 1-2 (1.00 g, 2.20 mmol) and N-tert-butyloxycarbonyl-2-aminoacetaldehyde (820 mg, 5.15 mmol) were dissolved in DMSO (3.00 mL), MeOH (10.0 mL) and H2O (5.00 mL), and Na2S2O4 (1.20 g, 6.89 mmol) was added. The mixture was stirred for 15 hours under N2 atmosphere at 100°C. After the reaction was cooled to room temperature, H2O (50.0 mL) was added to quench it, and DCM (50.0 mL×2) was used for extraction. The organic phase was washed with saturated brine (50.0 mL) and dried over Na2SO4. After filtration and concentration, the crude product was purified by column chromatography (SiO2, PE / EtOAc=5 / 1-1 / 3) to obtain compound 1-3. LCMS: 563.4 [M+H] + .

[0036] Step 3:

[0037] Compound 1-3 (900 mg, 1.60 mmol) was dissolved in THF (10.0 mL) and MeOH (10.0 mL), and NaOH (1.33 mL, 4.00 mmol) and tetrahydropyrrole (2.61 g, 6.40 mmol) were added. The mixture was stirred at room temperature for 4 hours. Then it was quenched with H2O (50.0 mL) and extracted with EtOAc (50.0 mL×2). The organic phase was washed with saturated brine (50.0 mL) and dried over Na2SO4. After filtration and concentration, the crude product 1-4 was obtained, which was used directly in the next step without purification. LCMS: 409.3 [M+H] + .

[0038] Step 4:

[0039] Compound 1-4 (300 mg, 0.734 mmol) was dissolved in EtOAc (1.0 mL), and then HCl in EtOAc (2.0 mL) was added. The mixture was stirred at room temperature for 30 minutes. After filtration, the crude product 1-5 was obtained and used directly in the next step without purification. LCMS: 309.0 [M+H] + .

[0040] Step 5:

[0041] Compound 1-5 (80.0 mg, 0.259 mmol) and Int (79.3 mg, 0.233 mmol) were dissolved in NMP (0.5 mL), and DIPEA (0.172 mL, 1.04 mmol) was added. The mixture was reacted under microwave at 100°C for 2 hours. The mixture was purified by preparative HPLC (0.1% FA). Compound 1 was obtained. LCMS: 612.4 [M+H] + . 1H NMR (400MHz, DMSO-d6) δ13.43-13.16(m,1H),11.91-11.84(m,1H),9.87-9.72(m,1H),8.76(s,1H),8.56(s,1H),8.48-8.43(m,1H),8.40(br d,J=0.9Hz,1H),8.21(d,J=9.1Hz,1H),8.00(s,2H),7.47(t,J=2.9Hz,1H),7.14(br d,J=8.6Hz,1H),6.71(br s,1H),5.11(br d,J=4.6Hz,2H),4.74-4.66(m,1H),4.63(br d,J=5.9Hz,2H),2.37(br s,2H),2.02-1.80(m,6H),1.31-1.20(m,1H),1.19-0.95(m,2H).

[0042] Example 2

[0043]

[0044] Step 1:

[0045] Compound N-tert-butyloxycarbonyl-(methylamino)acetaldehyde (381 mg, 2.20 mmol) and 1-2 (500 mg, 1.10 mmol) were added to DMSO (5 mL) and MeOH (5 mL), and then a solution of Na2S2O4 (479 mg, 2.75 mmol) in H2O (5 mL) was added. The mixture was reacted at 100°C in N2 atmosphere for 18 hours. The reaction was quenched with H2O (20 mL), extracted with EtOAc (20 mL×2), and the organic phase was dried with Na2SO4, filtered and concentrated to obtain compound 2-1. LCMS: 577.3 [M+H] + .

[0046] Step 2:

[0047] Compound 2-1 (900 mg, 1.56 mmol) was dissolved in THF (5 mL) and MeOH (5 mL), and NaOH solution (1.30 mL, 3.90 mmol) and tetrahydropyrrole (2.54 g, 6.24 mmol) were added, and the mixture was stirred at room temperature for 2 hours. The crude product was obtained after concentration. The crude product was diluted with H2O (20 mL), extracted with EtOAc (20 mL×2), and the organic phase was dried with Na2SO4, filtered and concentrated to obtain compound 2-2. LCMS: 423.3 [M+H] + .

[0048] Step 3:

[0049] Compound 2-2 (200 mg, 0.473 mmol) was dissolved in DCM (1 mL), TFA (0.2 mL, 0.473 mmol) was added, and the mixture was stirred at room temperature for 2 hours. After concentration, compound 2-3 was obtained. LCMS: 323.4 [M+H] + .

[0050] Step 4:

[0051] Compound 2-3 (75.0 mg, 0.233 mmol) and Int (79.0 mg, 0.233 mmol) were dissolved in NMP (2 mL), and DIPEA (0.077 mL, 0.465 mmol) was added, and the mixture was stirred at 100°C for 12 hours. After concentration, a crude product was obtained, and the crude product was subjected to preparative HPLC (FA conditions) to obtain compound 2. LCMS: 626.3 [M+H] + . 1 H NMR(400MHz, DMSO-d6)δ13.39(s,1H)11.87(s,1H)9.65-9.90(m,1H)8.76(s,1H)8.42-8.60(m,2H)8.26-8.39(m,1H)8.00(s,2H )7.37-7.54(m,2H)6.70(s,1H)5.44(s,2H)4.55-4.79(m,3H)3.22(s,3H)2.25-2.42(m,4H)1.78-1.98(m,5H)0.72-1.09(m,2H).

[0052] Example 3

[0053]

[0054] Step 1:

[0055] Compound 1-2 (5.00 g, 11.0 mmol), (R)-2-(tert-butoxycarbonylamino)propanal (3.82 g, 22.0 mmol) and Na2S2O4 (4.79 g, 27.5 mmol) were added to DMSO (50 mL), MeOH (50 mL) and H2O (25 mL), and the mixture was stirred at 100°C for 16 hours. The mixture was filtered and concentrated to obtain compound 3-1. LCMS: 577.4 [M+H] + .

[0056] Step 2:

[0057] Compound 3-1 (4.50 g, 7.80 mmol), NaOH solution (6.50 mL, 19.5 mmol) and tetrahydropyrrole (12.7 g, 31.2 mmol) were added to THF (40 mL) and MeOH (40 mL), and the mixture was reacted at room temperature for 3 hours. The reaction was quenched with saturated NH4Cl solution (50 mL) and extracted with EtOAc (100 mL). The organic phase was washed with saturated brine (100 mL), dried over Na2SO4, filtered and concentrated to obtain compound 3-2. LCMS: 423.5 [M+H] + .

[0058] Step 3:

[0059] Compound 3-2 (3.40 g, 8.04 mmol) was added to HCl / dioxane solution (50 mL), and the mixture was stirred at room temperature for 3 hours. After concentration, compound 3-3 was obtained. LCMS: 323.4 [M+H] + .

[0060] Step 4:

[0061] Compound 3-3 (2.80 g, 7.81 mmol), Int (1.86 g, 5.47 mmol), Xantphos (0.90 g, 1.56 mmol), Pd2(dba)3 (1.43 g, 1.56 mmol) and Cs2CO3 (7.64 g, 23.4 mmol) were added to dioxane (100 mL), and the mixture was reacted under N2 atmosphere at 100°C for 16 hours. After cooling to room temperature, the mixture was purified by preparative HPLC (0.1% FA) to obtain compound 3. LCMS: 626.4 [M+H] + . 1 H NMR(400MHz,DMSO-d6)δ10.47(br s,1H),9.48(br d,J=9.0Hz,1H),8.87-8.76(m,2H),8.71(s,1H),8.59(d,J=3.9Hz,1H),8.22(s,1H),8.02(s,2H),7.17(br d,J=3.3Hz,1H),4.89-4.77(m,1H),4.72-4.61(m,3H),2.63(br d,J=5.9Hz,2H),2.40-2.33(m,1H),2.28-1.92(m,6H),1.56(d,J=6.6Hz,3H),1.50(br d,J=12.1Hz,2H).

[0062] Example 4

[0063]

[0064] Step 1:

[0065] Compound 4-1 (5.00 g, 55.5 mmol) and TBSCl (6.70 g, 44.4 mmol) were added to THF (50 mL), and then imidazole (7.60 g, 110 mmol) was added, and the mixture was reacted at room temperature for 2 hours. After concentration, the crude product was purified by column chromatography (SiO2, PE / EtOAc=1 / 0-10 / 1) to obtain compound 4-2.

[0066] Step 2:

[0067] Compound 4-2 (6.40 g, 31.3 mmol) and Dess-Martin oxidant (19.9 g, 46.9 mmol) were dissolved in DCM (60 mL). The mixture was reacted at room temperature for 2 hours. The crude product was purified by PE to obtain compound 4-3.

[0068] Step 3:

[0069] Compound 1-2 (500 mg, 1.10 mmol) and 4-3 (446 mg, 2.21 mmol) were added to MeOH (5 mL), DMSO (5 mL) and H2O (2.5 mL), and then Na2S2O4 (384 mg, 2.21 mmol) was added. The mixture was reacted at 100°C for 16 hours. The reaction was quenched with a saturated solution of NH4Cl (20 mL) and extracted with EtOAc (20 mL). The organic phase was washed with saturated brine (50 mL), dried over Na2SO4, filtered and concentrated to obtain a crude product. The crude product was purified by column chromatography (SiO2, PE / EtOAc=10 / 1-1 / 1) to obtain compound 4-4. LCMS: 492.4 [M+H] + .

[0070] Step 4:

[0071] Compound 4-4 (165 mg, 0.336 mmol) and NaOH solution (0.280 mL, 0.839 mmol) were added to THF (3 mL) and MeOH (3 mL), and then tetrahydropyrrole (548 mg, 1.34 mmol) was added, and the mixture was stirred at room temperature for 4 hours. The reaction was quenched with H2O, extracted with EtOAc, and the organic phase was concentrated to obtain compound 4-5. LCMS: 338.1 [M+H] + .

[0072] Step 5:

[0073] Compound 4-5 (80.0 mg, 0.237 mmol) and Int (64.4 mg, 0.190 mmol) were added to DMA (5 mL), and then XPhos Pd G3 (5.00 mg, 0.006 mmol) and Cs2CO3 (154 mg, 0.474 mmol) were added. The mixture was reacted under microwave conditions at 110°C for 4 hours. The crude product was concentrated and purified by preparative HPLC to obtain compound 4. LCMS: 641.5 [M+H] + . 1 H NMR (400MHz, DMSO-d6) δ13.72-13.56(m,1H),10.10-9.93(m,1H),9.72-9.57(m,1H),8.94-8.86(m,2H),8.79(br d,J=2.8Hz,2H),8.62(d,J=3.9Hz,1H),8.12-7.96(m,2H),7.21(br s,1H),5.00-4.87(m,1H),4.69(br d,J=6.0Hz,3H),3.70(br s,2H),3.57-3.46(m,1H),3.29(s,2H),2.63(br d,J=6.0Hz,2H),2.06-1.97(m,4H),1.55-1.47(m,2H),1.36(d,J=6.6Hz,3H).

[0074] Example 5

[0075]

[0076] Step 1:

[0077] Compound 5-1 (2.00 g, 16.9 mmol) and imidazole (2.30 g, 33.9 mmol) were dissolved in DCM (40 mL), and TBSCl (3.10 g, 20.3 mmol) was added at 0°C, and the mixture was stirred at room temperature for 3 hours. The reaction was quenched with H2O (10 mL), extracted with EtOAc (15 mL×2), and the organic phase was dried over Na2SO4, filtered and concentrated to obtain a crude product. The crude product was purified by column chromatography (SiO2, PE / EtOAc=20 / 1-1 / 1) to obtain compound 5-2.

[0078] Step 2:

[0079] Compound 5-2 (1.50 g, 6.45 mmol) was dissolved in DCM (10 mL), and DIBAL-H (7.10 mL, 7.10 mmol) was added under N2 atmosphere at -78°C and then reacted for 1 hour. The reaction was quenched with MeOH (10 mL), and then H2O (20 mL) was added to the mixture, extracted with DCM (20 mL×2), and the organic phase was dried over Na2SO4, filtered and concentrated to obtain compound 5-3.

[0080] Step 3:

[0081] Compound 1-2 (500 mg, 1.10 mmol) and 5-3 (446 mg, 2.21 mmol) were dissolved in MeOH (12 mL) and DMSO (12 mL), and then a solution of Na2S2O4 (480 mg, 2.75 mmol) in H2O (6 mL) was added. The mixture was reacted under N2 atmosphere at 100°C for 12 hours. The reaction was quenched with water (20 mL), extracted with EtOAc (20 mL×2), and the organic phase was dried over Na2SO4, filtered and concentrated to obtain compound 5-4. LCMS: 492.3 [M+H] + .

[0082] Step 4:

[0083] Compound 5-4 (110 mg, 0.220 mmol) was dissolved in THF (1 mL) and MeOH (1 mL), and NaOH solution (0.19 mL, 0.560 mmol) and tetrahydropyrrole (0.07 mL, 0.900 mmol) were added, and the mixture was stirred at room temperature for 12 hours. The mixture was quenched with water (10 mL), extracted with EtOAc (15 mL×2), and the organic phase was dried over Na2SO4, filtered and concentrated to obtain compound 5-5. LCMS: 338.2 [M+H] + .

[0084] Step 5:

[0085] Compound 5-5 (72.0 mg, 213 μmol), Int (36.3 mg, 106 μmol) and Cs2CO3 (139 mg, 426 μmol) were dissolved in DMA (2 mL), and then XPhos Pd G3 (18.0 mg, 21.3 μmol) was added, and the mixture was reacted under microwave N2 atmosphere at 110°C for 2 hours. The mixture was filtered and concentrated to obtain a crude product, which was purified by preparative HPLC (FA conditions) to obtain compound 5. LCMS: 641.5 [M+H] + . 1H NMR(400MHz, DMSO-d6)δ13.80(d,J=10.3Hz,1H),9.64-9.55(m,1H),8.90-8.72(m,4H),8.61(d,J=3 .9Hz,1H),8.07-7.99(m,2H),7.19(d,J=3.0Hz,1H),4.98(d,J=4.4Hz,1H),4.69(d,J=6.0Hz,3H),4. 24-4.11(m,1H),3.15-3.07(m,2H),2.67(td,J=1.8,3.7Hz,1H),2.62(d,J=6.0Hz,2H),2.36-2.30( m,2H),2.18-2.09(m,1H),2.01(s,2H),1.54-1.41(m,2H),1.25-1.25(m,1H),1.23(d,J=6.0Hz,2H).

[0086] Biological test cases

[0087] JAK

[0088] 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.

[0089] PHD2

[0090] 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.

[0091] Table 1

[0092]

[0093] 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, L is selected from -CH2-, -C(CH3)H-, -C(CH3)HCH2- or -CH2C(CH3)H-; X is selected from -NH-, -N(CH3)- or -O-; And when X is selected from O, L is not -CH2-.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein -LX- is selected from -CH2NH-, -CH2N(CH3)-, -C(CH3)HNH-, -CH(CH3)CH2O- or -CH2CH(CH3)O-.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The structure of the compound represented by formula (I) is shown below:

4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The structure of the compound represented by formula (I) is shown below:

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The structure of the compound represented by formula (I) is shown below:

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 below:

7. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The structure of the compound represented by formula (I) is shown below:

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

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

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

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