Synthesis and application of small molecule compounds with 2-pyridine-substituted carboxamide structures

Abstract

The present invention relates to a class of small molecule inhibitors of ASK1 kinase, and specifically to the synthesis and application of a small molecule compound having a 2-pyridine-substituted carboxamide structure, as well as a pharmaceutical composition thereof and its use in preparing ASK1 small molecule inhibitors, or in drugs for preventing and / or treating ASK1-related diseases, especially liver diseases, lung diseases, tumors, cardiovascular diseases, kidney diseases and metabolic diseases.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and specifically relates to the synthesis and application of a small molecule compound having a 2-pyridine-substituted carboxamide structure, and a pharmaceutical composition thereof and its use in preventing or treating diseases or conditions mediated by apoptosis signal-regulating kinase 1 (ASK1, also known as MAP3K5, MAPKKK5, MEKK5), especially its use in drugs for liver diseases, lung diseases, cardiovascular diseases, kidney diseases and metabolic diseases. Background Art

[0002] ASK1 kinase, also known as Apoptosis Signal Regulating Kinase 1 (ASK1), is a member of the MAP3K family and is located upstream of Jun N-terminal kinase (JNK) and p38. Numerous experiments have demonstrated that ASK1 plays a crucial role in mammalian cell stress responses and apoptosis-induced diseases.

[0003] ASK1 kinase inhibitors hold potential therapeutic promise for the treatment of common liver diseases, such as nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, chronic liver disease, metabolic liver disease, liver fibrosis, primary sclerosing cholangitis, hepatic ischemia-reperfusion injury, primary biliary cirrhosis, and diabetes-related liver disease. Biological and medical research has also demonstrated that ASK1 kinase inhibitors are effective in treating lung diseases (such as pulmonary hypertension and pulmonary fibrosis). Furthermore, ASK1 kinase inhibition has potential for treating kidney diseases (glomerulonephritis, diabetic nephropathy, and hypertensive nephropathy) and tumors. Inhibiting ASK1 kinase activity has shown significant therapeutic effects in diabetic mouse models; the small molecule ASK1 inhibitor GS-444217 has been shown to reduce diabetic parameters. Therefore, ASK1 is a potential target for the treatment of these diseases, and the development of novel ASK1 inhibitors is crucial for alleviating or even curing these conditions.

[0004] Currently, some ASK1 inhibitors have been reported in the literature to have potential issues, including low molecular-level activity, poor target selectivity, unclear dose-response relationships in animals, and poor drug-like properties. Furthermore, some compounds have potential issues with chemical stability and metabolic stability in vivo. Some small-molecule ASK1 kinase inhibitors are highly water-soluble, resulting in low oral exposure in animals and suboptimal pharmacokinetic properties in animals, leading to a need for improved in vivo efficacy.

[0005] Therefore, there is still a need in the art to develop a more effective small molecule inhibitor of ASK1 kinase. Summary of the Invention

[0006] The purpose of the present invention is to provide a class of small molecule inhibitors of ASK1 kinase with novel structure.

[0007] The present invention also aims to provide a synthesis method and use of the above-mentioned compound.

[0008] The first aspect of the present invention provides a small molecule compound having a 2-pyridine-substituted carboxamide structure as described in formula (I), or its enantiomer, racemate or mixture, or its pharmaceutically acceptable salt, hydrate or solvate.

[0009]

[0010] in,

[0011] R1 represents a halogen, a deuterium atom, a cyano group, a hydroxyl group, an amino group, a nitro group, an oxy group, an alkyl group, a substituted or unsubstituted 5-12 membered heteroaromatic ring containing N, O or S atoms, a substituted or unsubstituted 5-13 membered saturated heterocyclic ring containing N, O or S atoms, a substituted or unsubstituted C1-C4 alkyl group, a substituted or unsubstituted C3-C6 cycloalkyl group, a substituted or unsubstituted C1-C4 alkoxy group, a substituted or unsubstituted C2-C4 alkenyl group, a substituted or unsubstituted C2-C4 alkynyl group, a substituted or unsubstituted C 3-C6 cycloalkyl, substituted or unsubstituted C1-C4 alkylethynyl, substituted or unsubstituted C1-C4 alkylamino, substituted or unsubstituted C1-C4 alkylcarbonylamino, substituted or unsubstituted C1-C4 alkoxycarbonylamino, substituted or unsubstituted C1-C4 sulfonyl, substituted or unsubstituted C1-C4 alkyl-S-, substituted or unsubstituted C2-C10 acyl, substituted or unsubstituted C1-C4 alkylcarbonyl, substituted or unsubstituted C1-C4 alkylaminocarbonyl, or a combination thereof;

[0012] represents a quinoline ring with different nitrogen atom positions or a 6-membered heterocyclic ring containing two nitrogen atoms;

[0013] X represents N, CH or C=O;

[0014] a, b represent the positions of amide substitution on the N-containing heterocycle.

[0015] R2 represents H or methyl;

[0016] R3 represents a substituted or unsubstituted 5-6 membered heteroaromatic ring containing 1-4 heteroatoms of N, O or S, and one or more hydrogen atoms on the substituent group are substituted by a group selected from the group consisting of halogen, deuterium atom, cyano, hydroxyl, amino, nitro, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C3-C6 cycloalkyl, substituted or unsubstituted C1-C6 alkoxy; wherein one or more hydrogen atoms on the corresponding substituent group in the substituent group are substituted by a group selected from the group consisting of hydroxyl, amino and carboxyl.

[0017] Another aspect of the present invention provides a compound of the present invention or a pharmaceutically acceptable salt, stereoisomer, polymorph, solvate, N-oxide, isotope-labeled compound, metabolite or prodrug thereof. The compound is specifically selected from one of the following structural formulas:

[0018]

[0019]

[0020] The present invention also provides an ASK1 inhibitor, which includes the compound, or its enantiomers, diastereomers, racemates and mixtures thereof, or pharmaceutically acceptable salts, crystalline hydrates or solvates thereof.

[0021] The present invention also provides a pharmaceutical composition comprising: (A) a therapeutically effective amount of the compound, and one or more of its enantiomers, diastereomers, racemates and mixtures thereof, and pharmaceutically acceptable salts, crystalline hydrates and solvates; and (B) a pharmaceutically acceptable carrier.

[0022] Preferably, the pharmaceutical composition further comprises a pharmaceutically acceptable excipient, which is selected from one of adhesives, fillers, diluents, disintegrants, suspending agents, suspending aids, sustained-release agents or control agents, lyoprotectants, coatings, enteric materials, lubricants, glidants, anti-adherents, sweeteners, flavors, plasticizers, opacifiers, solubilizers, humectants, solvents, osmotic pressure regulators, colorants, pigments, surfactants, emulsifiers, water-soluble matrices, fat-soluble matrices, oily matrices, pore-forming agents, gelling agents, preservatives, buffers, chelating agents and antioxidants, or a combination thereof.

[0023] Provided is a use of the pharmaceutical composition of the present invention in preparing a medicament for preventing or treating a disease or condition mediated by apoptosis signal-regulating kinase 1 (ASK1).

[0024] The present invention also provides a use of the compound in preparing a drug for treating diseases related to the activity or expression level of ASK1 kinase.

[0025] Preferably, the disease associated with the activity or expression of ASK1 kinase is selected from one of liver disease, lung disease, cardiovascular disease, kidney disease, metabolic disease and tumor disease.

[0026] Preferably, the liver disease is selected from the group consisting of non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, chronic liver disease, metabolic liver disease, liver fibrosis, primary sclerosing cholangitis, liver ischemia-reperfusion injury, primary biliary cirrhosis and diabetes-related liver disease;

[0027] The lung disease is pulmonary hypertension or pulmonary fibrosis;

[0028] The cardiovascular disease is heart failure;

[0029] The kidney disease is selected from one of glomerulonephritis, diabetic nephropathy and hypertensive nephropathy;

[0030] The metabolic disease is type 2 diabetes or type 1 diabetes;

[0031] The tumor disease is selected from one of liver cancer, lung cancer, breast cancer, gastric cancer, colorectal cancer, esophageal cancer, cervical cancer, ovarian cancer, bladder cancer and pancreatic cancer.

[0032] The present invention also provides a method for synthesizing the compound, and the synthetic route is as follows:

[0033]

[0034] Preferably, I-1 in the synthetic route is prepared by a one-step reaction, wherein step 1 is specifically as follows: amine S1 and the corresponding acid chloride S2 are mixed in a suitable solvent, such as tetrahydrofuran, 2-methyltetrahydrofuran, dichloromethane, or N,N-dimethylformamide, etc., a base (triethylamine, diisopropylethylamine, or N-methylmorpholine, etc.) is added, and the reaction is carried out at room temperature, a suitable low temperature condition (-10°C to 0°C), or a suitable elevated temperature condition (e.g., 40-50°C) to obtain the final product I-1; step 2 is specifically as follows: I-1 is dissolved in DMF, NaH and iodomethane are added, and the reaction is stirred at room temperature, a low temperature condition, or an elevated temperature condition to prepare the final product I-2. Implementation Method

[0035] definition

[0036] Unless otherwise defined below, all technical and scientific terms used herein are intended to have the same meanings as those commonly understood by those skilled in the art. References to technology used herein are intended to refer to technologies commonly understood in the art, including variations or equivalents that would be apparent to those skilled in the art. While detailed terminology is well understood by those skilled in the art, the following definitions are provided to better explain the present invention.

[0037] As used herein, the terms "comprises," "comprising," "having," "containing," or "involving," and variations thereof herein, are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[0038] As used herein, the term "hydrogen" refers to protium (H), deuterium (D), tritium (T).

[0039] As used herein, the term "alkyl" is defined as an offline or branched saturated aliphatic hydrocarbon. 1-12 Alkyl refers to a group having 1 to 12, for example 1 to 6, carbon atoms (C 1-6 alkyl) or 1 to 4 carbon atoms (C 1-4 For example, as used herein, the term "C 1-6 "Alkyl" refers to an offline or branched group of 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl or n-hexyl), which is optionally substituted with one or more (e.g., 1 to 3) suitable substituents such as halogen (in which case the group is referred to as "haloalkyl") (e.g., CH2F, CHF2, CF3, CCl3, C2F5, C2Cl5, CH2CF3, CH2Cl or -CH2CH2CF3, etc.). The term "C 1-4 "Alkyl" refers to a linear or branched aliphatic hydrocarbon chain of 1 to 4 carbon atoms (ie, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl).

[0040] As used herein, the term "alkenyl" means an linear or branched monovalent hydrocarbon radical containing one or more double bonds and having 2 to 6 carbon atoms ("C 2-6 The alkenyl group is, for example, vinyl, 1-propenyl, 2-butenyl, 3-butenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 2-methyl-2-propenyl and 4-methyl-3-pentenyl. When the compound of the present invention contains an alkenyl group, the compound may be present in the pure E (entgegen) form, the pure Z (zusammen) form or a mixture thereof in any ratio.

[0041] As used herein, the term "alkynyl" refers to a monovalent hydrocarbon group containing one or more triple bonds, for example having 2, 3, 4, 5 or 6 carbon atoms, such as ethynyl or propynyl.

[0042] As used herein, the term "cycloalkyl" refers to a saturated monocyclic or polycyclic (such as bicyclic) hydrocarbon ring (e.g., a monocyclic ring such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, or a bicyclic ring, including spirocyclic, fused or bridged systems (such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl or bicyclo[5.2.0]nonyl, decahydronaphthyl, etc.), which is optionally substituted with one or more (such as one to three) suitable substituents. The cycloalkyl group has 3 to 15 carbon atoms. For example, the term "C 3-6 "Cycloalkyl" refers to a saturated monocyclic or polycyclic (such as bicyclic) hydrocarbon ring of 3 to 6 ring carbon atoms (for example cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl), which is optionally substituted by 1 or more (such as 1 to 3) suitable substituents, for example methyl substituted cyclopropyl.

[0043] As used herein, the term "heterocyclyl" refers to a saturated or partially unsaturated monovalent monocyclic or bicyclic group having 2, 3, 4, 5, 6, 7, 8 or 9 carbon atoms or one or more (e.g., one, two, three or four) heteroatom-containing groups selected from C(=O), O, S, S(=O), S(=O)2 and NRa, wherein Ra represents a hydrogen atom.

[0044] As used herein, the term "5-12 membered heteroaromatic ring" refers to a monocyclic aromatic group having 5-12 ring members, wherein at least one (e.g., 1, 2, 3 or 4) heteroatoms selected from N, O, and S are saturated in the ring members, for example, "5-6 membered heteroaryl", 5 membered heteroaryl, 6 membered heteroaryl, etc. Specific examples include, but are not limited to, furyl, thienyl, pyrrolyl, thiazolyl, isothiazolyl, thiadiazolyl, oxazolyl, isoxazolyl, imidazolyl, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,3-oxadiazole, 1,2,4-oxadiazole, 1,2,5-oxadiazole, 1,3,4-oxadiazole, pyridyl, 2-pyridonyl, 4-pyridonyl, pyrimidinyl, 2H-1,2-oxazinyl, 4H-1,2-oxazinyl, 6H-1,2-oxazinyl, 4H-1,3-oxazinyl, 6H-1,3-oxazinyl, 4H-1,4-oxazinyl, pyridazinyl, pyrazinyl, 1,2,3-triazinyl, 1,3,5-triazinyl, 1,2,4,5-tetrazinyl, and the like.

[0045] As used herein, the term "halo" or "halogen" group is defined to include F, Cl, Br, or I.

[0046] As used herein, the term "substituted" refers to the replacement of one or more (e.g., one, two, three, or four) hydrogen atoms on the designated atom with a selected group selected from the indicated group, provided that the designated atom's normal valence in the present context is not exceeded and the substitution results in a stable compound. Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0047] Unless otherwise indicated, as used herein, the point of attachment of a substituent can be from any suitable position of the substituent.

[0048] The present invention also includes all pharmaceutically acceptable isotopically labeled compounds, which are identical to the compounds of the present invention, except that one or more atoms are replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number prevalent in nature. Isotopes suitable for inclusion in the compounds of the present invention, such as deuterium ( 2 H) Tritium ( 3 H)): Carbon isotopes (e.g. 11 C. 13 C and 14 C); isotopes of chlorine (e.g. 36 Cl); isotopes of fluorine (e.g. 18 F); isotopes of iodine (such as 123 I and 125 I); isotopes of nitrogen (e.g. 13 N and 15 N); oxygen isotopes (e.g. 15 O. 17 O and 18 O); isotopes of phosphorus (such as 32 P); sulfur isotopes (e.g. 35 S). Certain isotopically labeled compounds of the invention (e.g., incorporating a radioactive isotope) are useful in drug and / or substrate tissue distribution studies (e.g., assays). The radioactive isotope tritium ( 3 H) and carbon-14 ( 14 C) is particularly useful for this purpose because it is incorporated and easily detected. 11 C. 18 F. 15 O and 13N) substitution can be used to examine substrate receptor occupancy in positron emission tomography (PET) studies. Isotopically labeled compounds of the present invention can be prepared by methods analogous to those described in the accompanying schemes and / or examples and preparations by using appropriate isotopically labeled reagents instead of the non-labeled reagents previously employed. Pharmaceutically acceptable solvates of the present invention include those in which the crystallization solvent is isotopically substituted, for example, D2O, acetone-d6 or DMSO-d6.

[0049] Pharmaceutically acceptable salts of the compounds of the present invention include acid addition salts and base addition salts thereof.

[0050] Suitable acid addition salts are formed from acids that form pharmaceutically acceptable salts, examples of which include aspartate, gluconate, lactate, palmitate, hydrochloride and other similar salts.

[0051] Suitable base addition salts are formed from bases which form pharmaceutically acceptable salts. Examples include aluminum, sodium, calcium, potassium, choline and other similar salts.

[0052] The compounds of the present invention may exist in the form of solvates (preferably hydrates), wherein the compounds of the present invention contain a polar solvent as a structural element of the crystal lattice of the compound, in particular water, methanol or ethanol. The amount of polar solvent, in particular water, may be present in a stoichiometric or non-compound stoichiometric ratio.

[0053] The term "about" means within ±10%, preferably within ±5%, and more preferably within ±2% of the stated numerical value.

[0054] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present invention only and are not intended to limit the scope of the invention. The experimental methods in the following examples, for which specific conditions are not specified, are generally based on conventional conditions or the conditions recommended by the manufacturer. Unless otherwise stated, percentages and parts are calculated by mass.

[0055] Experimental Example 1: Synthesis of Compounds

[0056] Synthetic intermediate 01: 6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-amine (12-4-41)

[0057]

[0058] Step 1: Dissolve methyl 6-aminopicolinate (500 mg, 3.289 mmol) in 25 mL of methanol. Under argon, slowly add hydrazine hydrate (318.75 μL, 6.578 mmol) dropwise. Raise the temperature to 75°C and reflux for 3 h. After the reaction is complete, cool to room temperature, dry over anhydrous sodium sulfate, and concentrate by rotary evaporation. This affords 475 mg of the crude target compound (12-3-41) as a white solid in a 95% yield.

[0059] Step 2: Dissolve 6-amino-picolinohydrazide (475 mg, 3.125 mmol) in 30 mL of toluene, add N,N-dimethylformamide dimethyl acetal (104.74 μL, 9.375 mmol) and isopropylamine (157.5 μL, 21.875 mmol). Cool to 0°C, and add acetic acid (45.1 μL, 9.375 mmol) dropwise. Under argon, raise the temperature to 95°C and reflux for 10 h. Then cool to room temperature, concentrate on a rotary evaporator, dissolve in 10 mL of purified water, and continue the reaction at 85°C for 10 h. After completion of the reaction, wash with 20 mL of ether and 20 mL of dichloromethane, dry over anhydrous sodium sulfate, concentrate by rotary evaporation, and purify by column chromatography (petroleum ether / ethyl acetate = 20 / 1). 243.485 mg of the target compound (12-4-41) as a white solid was obtained in a 51.26% yield. 1 H NMR (400MHz, DMSO-d6): δ8.78(s,1H),7.52(dd,J=8.1,7.6Hz,1H),7.23–7.13(m ,1H),6.58–6.47(m,1H),δ5.53(dt,J=13.5,6.7Hz,1H).,1.43(d,J=6.7Hz,6H).

[0060] Final product 1: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)isoquinoline-4-carboxamide (1,12-168-41)

[0061]

[0062] Intermediate 01 (12-4-41, 70.2 mg, 0.346 mmol) and isoquinoline-4-carboxylic acid (50 mg, 0.289 mmol) were dissolved in 5 mL of dichloromethane and cooled to 0°C. Triethylamine (280 μL, 2.02 mmol) and 1-propylphosphonic anhydride (688 μL, 2.310 mmol) were added. Under an argon atmosphere, the temperature was raised to 40°C and refluxed for 7 hours. After the reaction, the mixture was washed with 20 mL of saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by column chromatography (dichloromethane / methanol = 80 / 1) to obtain 10 mg of the target compound (1,12-168-41) as a white solid in a 25.8% yield. 1 H NMR (400MHz, CD3OD): δ9.48–9.32(m,1H),8.79(dd,J=15.6,3.6Hz,1H),8.44(t,J=8.9Hz,1H),8.23(tt,J=17.0,8.6Hz,1H),7.94(dt,J=15.3,7.7Hz ,1H),7.88–7.75(m,1H),7.60–7.55(m,1H),7.21(dd,J=12.9,6.8Hz,1H), 6.66(d,J=8.4Hz,1H), 5.51(dt,J=13.5,6.7Hz,1H), 1.52(t,J=4.8Hz,6H).

[0063] Final product 2: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-4-carboxamide (2,12-161-41)

[0064]

[0065] Intermediate 01 (12-4-41, 70.2 mg, 0.346 mmol) and quinoline-4-carboxylic acid (50 mg, 0.289 mmol) were dissolved in 5 mL of dichloromethane and cooled to 0°C. Triethylamine (280 μL, 2.02 mmol) and 1-propylphosphonic anhydride (688 μL, 2.310 mmol) were added. Under an argon atmosphere, the temperature was raised to 40°C and refluxed for 7 hours. After the reaction, the product was washed with 20 mL of saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, concentrated to dryness, and purified by column chromatography (dichloromethane / methanol = 80 / 1) to obtain 54.1 mg of the target compound (2,12-161-41) as a white solid in a 52.3% yield. 1H NMR (400MHz, CD3OD): δ9.01 (d, J = 4.4Hz, 1H), 8.80 (s, 1H), 8.45 (d, J = 8.3Hz, 1H), 8.26 (d, J = 8.4Hz, 1H), 8.15 (d, J = 8.5Hz, 1H), 8.09 (t, J = 8.0Hz, 1H),7.94(d,J=7.7Hz,1H),7.87(t,J=7.7Hz,1H),7.78–7.73(m,1H),7. 71(d,J=7.3Hz,1H), 5.76(dt,J=13.5,6.8Hz,1H), 1.48(d,J=6.7Hz,6H).

[0066] Final product 3: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)isoquinoline-1-carboxamide (3,12-68-41)

[0067]

[0068] Isoquinoline-1-carboxylic acid (50 mg, 0.289 mmol) was dissolved in 5 mL of dichloromethane, and carbodiimide hydrochloride (55.4 mg, 0.289 mmol) and N-hydroxysuccinimide (34.1 mg, 0.289 mmol) were added. Under argon, intermediate 01 (12-4-41, 59 mg, 0.289 mmol) was added, and the temperature was raised to 40°C and refluxed for 7 h. After the reaction, the mixture was washed with 20 mL of saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by column chromatography (dichloromethane / methanol = 80 / 1) to obtain 24.5 mg of the target compound (3,12-68-41) as a white solid in a 23.7% yield. 1 H NMR (400MHz, CDCl3): δ10.73 (s, 1H), 9.61 (dd, J=

[0069] 6.7,3.0Hz,1H),8.52(t,J=5.4Hz,2H),7.99(s,1H),7.93–7.81(m,3H),7.75–

[0070] 7.67(m,2H),5.73(s,1H),1.56(s,6H).

[0071] Final product 4: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoxaline-2-carboxamide (4,12-13-42)

[0072]

[0073] Quinoxaline-2-carboxylic acid (81.78 mg, 0.47 mmol) was dissolved in 6 mL of dichloromethane, and 0.06 mL of N,N-dimethylformamide dimethyl acetal was added. The mixture was cooled to 0°C and, under argon, oxalyl chloride (80 μL, 0.93 mmol) was added dropwise. The mixture was stirred at room temperature for 2 h. After the acid chloride was prepared, it was spin-dried and vacuum-dried to remove the reaction solvent and excess oxalyl chloride. A solution of intermediate 01 (12-4-41, 62.93 mg, 0.31 mmol) in dichloromethane (6 mL) was added, followed by the dropwise addition of triethylamine (85.8 μL, 0.62 mmol). The mixture was stirred at room temperature for 1 h. After the reaction, 20 mL of saturated sodium bicarbonate aqueous solution was added for washing, and the mixture was dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by column chromatography (dichloromethane / methanol = 80 / 1) to obtain 21.42 mg of the target compound (4,12-13-42) as a white solid in a yield of 42.84%. 1 H NMR (400MHz, CDCl3): δ9.72(s,1H),8.45(d,J=8.2Hz,1H),8.39(s,1H),8.18(d,J=8.1Hz,1H),8.13(d,J=7.7Hz,1H), 8.00(d,J=7.6Hz,1H),7.88(dd,J=16.2,8.9Hz,3H),7.20(s,1H),5.53(dt,J=13.3,6.7Hz,1H),1.58(d,J=6.7Hz,6H).

[0074] Final product 5: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)-4-oxo-1,4-dihydroquinazoline-2-carboxamide (5,12-163-41)

[0075]

[0076] 4-Oxo-1,4-dihydroquinazoline-2-carboxylic acid (40 mg, 0.210 mmol) and intermediate 01 (12-4-41, 51.2 mg, 0.252 mmol) were dissolved in 5 mL of dichloromethane and cooled to 0°C. Triethylamine (206 μL, 1.472 mmol) and 1-propylphosphonic anhydride (500 μL, 1.682 mmol) were added. Under an argon atmosphere, the temperature was raised to 40°C and refluxed for 7 hours. After the reaction, the mixture was washed with 20 mL of saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, concentrated by rotary evaporation, and purified by column chromatography (dichloromethane / methanol = 80 / 1) to obtain 32 mg of the target compound (5,12-163-41) as a white solid in a 40.5% yield. 1H NMR (400MHz, CD3OD) δ8.86 (s, 1H), 8.43 (d, J = 8.3Hz, 1H), 8.31 (d, J = 7.9Hz, 1H), 8.08 (t, J = 8.0Hz, 1H),7.98–7.89(m,3H),7.66(dd,J=9.8,4.2Hz,1H),5.69–5.62(m,1H),1.63(s,3H),1.62(s,3H).

[0077] Final product 6: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (6,12-70-41)

[0078]

[0079] Quinoline-2-carboxylic acid (50 mg, 0.289 mmol) was dissolved in 5 mL of dichloromethane, and carbodiimide hydrochloride (55.4 mg, 0.289 mmol) and N-hydroxysuccinimide (34.1 mg, 0.289 mmol) were added. Under argon, intermediate 01 (12-4-41, 59 mg, 0.289 mmol) was added. The temperature was raised to 40°C and refluxed for 7 hours. After the reaction, the mixture was washed with 20 mL of saturated sodium bicarbonate solution, dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a reverse-phase silica gel column (dichloromethane / methanol = 80 / 1) to obtain 27.5 mg of the target compound (6,12-70-41) as a white solid in a 26.57% yield. 1 H NMR (400MHz, CDCl3) δ8.49(d,J=8.2Hz,2H),8.36(s,3H),8.11(d,J=8.4Hz,1 H), 7.90 (t, J = 10.2Hz, 3H), 7.63 (t, J = 7.5Hz, 1H), 5.23 (s, 1H), 1.63 (s, 6H).

[0080] Final product 7: 8-bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (7, 21-25-41)

[0081]

[0082] 8-Bromoquinoline-2-carboxylic acid (50 mg, 0.2 mmol) and Intermediate 01 (12-4-41, 40.3 mg, 0.2 mmol) were added to a 50 mL eggplant-shaped flask, followed by 5 mL of anhydrous dichloromethane. The mixture was cooled to 0°C, triethylamine (140.5 mg, 1.4 mmol) was added, and T3P (50% ethyl acetate, 504.9 mg, 0.8 mmol) was slowly added. The mixture was stirred at room temperature overnight. After completion of the reaction, the mixture was diluted with dichloromethane, washed with water and sodium bicarbonate solution, and the organic layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a normal-phase silica gel column. 35.7 mg of the target compound (7, 21-25-41) was obtained in a 40.9% yield. 1 H NMR (400MHz, CDCl3): δ10.86 (s, 1H), 8.45 (d, J = 8.2Hz, 1H), 8.37 (dt, J = 12.8, 6.4Hz, 3H), 8.11–8. 03(m,2H),7.88(d,J=24.2Hz,2H),7.48(t,J=7.8Hz,1H),5.81–5.67(m,1H),1.59(d,J=6.8Hz,6H).

[0083] Final product 8: 7-bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (8, 21-26-41)

[0084]

[0085] 7-Bromoquinoline-2-carboxylic acid (50 mg, 0.2 mmol) and Intermediate 01 (12-4-41, 40.3 mg, 0.2 mmol) were added to a 50 mL eggplant-shaped flask, followed by 5 mL of anhydrous dichloromethane. The mixture was cooled to 0°C, triethylamine (140.5 mg, 1.4 mmol) was added, and T3P (50% ethyl acetate, 504.9 mg, 0.8 mmol) was slowly added. The mixture was stirred at room temperature overnight. After completion of the reaction, the mixture was diluted with dichloromethane, washed with water and sodium bicarbonate solution, and the organic layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a normal phase silica gel column. 48.3 mg of the target compound (8, 21-26-41) was obtained in a 55.2% yield. 1H NMR (400MHz, CDCl3): δ10.51(s,1H),8.46(d,J=8.2Hz,1H),8.37(d,J=8.4Hz,2H),8.32(d,J=8.6Hz,1H),8.29(s,1H),8.00(d,J =7.6Hz,1H),7.89(t,J=8.0Hz,1H),7.76(d,J=8.7Hz,1H),7.70(dd,J=8.7,1.7Hz,1H),5.62–5.47(m,1H),1.60(d,J=6.7Hz,6H).

[0086] Final product 9: 6-bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (9, 21-18-41)

[0087]

[0088] 6-Bromoquinoline-2-carboxylic acid (50 mg, 0.2 mmol) and Intermediate 01 (12-4-41, 40.3 mg, 0.2 mmol) were added to a 50 mL eggplant-shaped flask, followed by 5 mL of anhydrous dichloromethane. The mixture was cooled to 0°C, triethylamine (140.5 mg, 1.4 mmol) was added, and T3P (50% ethyl acetate, 504.9 mg, 0.8 mmol) was slowly added. The mixture was stirred at room temperature overnight. After completion of the reaction, the mixture was diluted with dichloromethane, washed with water and sodium bicarbonate solution, and the organic layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a normal phase silica gel column. 48.5 mg of the target compound (9,21-18-41) was obtained in a 55.5% yield. 1 H NMR (400MHz, CDCl3): δ10.55(s,1H),8.46(d,J=8.2Hz,1H),8.37(d,J=8.5Hz,1H),8.26(d,J=8.5Hz,1H),8.05(d,J=1.5Hz,1H ),7.98(dd,J=8.0,5.7Hz,2H),7.89(d,J=15.9Hz,1H),7.85–7.81(m,1H),5.54(dt,J=13.4,6.7Hz,1H),1.59(d,J=6.7Hz,6H).

[0089] Final product 10: 5-bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (10, 21-19-41)

[0090]

[0091] 5-Bromoquinoline-2-carboxylic acid (50 mg, 0.2 mmol) and Intermediate 01 (12-4-41, 40.3 mg, 0.2 mmol) were added to a 50 mL eggplant-shaped flask, followed by 5 mL of anhydrous dichloromethane. The mixture was cooled to 0°C, triethylamine (140.5 mg, 1.4 mmol) was added, and T3P (50% ethyl acetate, 504.9 mg, 0.8 mmol) was slowly added. The mixture was stirred at room temperature overnight. After completion of the reaction, the mixture was diluted with dichloromethane, washed with water and sodium bicarbonate solution, and the organic layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a normal phase silica gel column. The target compound (56.2 mg, 10, 21-19-41) was obtained in a 64.3% yield. 1 H NMR (400MHz, CDCl3): δ10.57(s,1H),8.73(d,J=8.7Hz,1H),8.45(d,J=3.0Hz,1H),8.35(s,1H),8.08(d,J=8 .5Hz,1H),7.98(s,1H),7.91–7.87(m,1H),7.63(t,J=8.0Hz,1H),5.61–5.42(m,1H),1.59(d,J=6.7Hz,6H).

[0092] Final product 11: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)-8-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)quinoline-2-carboxamide (11, 21-59-36)

[0093]

[0094] 8-Bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (07,21-25-41, 40 mg, 0.09 mmol), 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,2,3,6-tetrahydropyridine (22.5 mg, 0.1 mmol), sodium carbonate (48.5 mg, 0.45 mmol), and tetrakistriphenylphosphine palladium (10.6 mg, 0.009 mmol) were added to a 50 mL eggplant flask. 1,2-dimethoxyethane and water (2 mL each) were added and the mixture was reacted at 80°C for 2 h. The mixture was concentrated on a rotary evaporator and purified on a normal phase silica gel column. 42 mg of the target compound (11,21-59-36) was obtained in a 100% yield. 1H NMR (400MHz, CDCl3): δ10.38(s,1H),8.42(d,J=8.2Hz,1H),8.33(d,J=6.1Hz,3H),7.90(t,J=7.9Hz,1H),7.80(dd,J=11.6,7.8Hz,2H),7.64 (d,J=6.5Hz,1H),7.56(t,J=7.6Hz,1H),5.92(s,1H),5.34–5.24(m,1H),3.30(s,2H),2.92–2.86(m,4H),2.45(s,3H),1.50(d,J=6.7Hz,6H).

[0095] Final product 12: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)-7-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)quinoline-2-carboxamide (12, 21-36-36)

[0096]

[0097] 7-Bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (08,21-26-41, 30 mg, 0.07 mmol), 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,2,3,6-tetrahydropyridine (16.8 mg, 0.075 mmol), sodium carbonate (72.7 mg, 0.69 mmol), and tetrakistriphenylphosphine palladium (7.9 mg, 0.0073 mmol) were added to a 50 mL eggplant flask. 1,2-dimethoxyethane and water (2 mL each) were added and the mixture was reacted at 80°C for 2 h. The mixture was concentrated on a rotary evaporator and purified on a normal phase silica gel column. 40.1 mg of the target compound (12,21-36-36) was obtained in a 100% yield. 1 HNMR (400MHz, CDCl3): δ10.61(s,1H),8.45(dd,J=8.2,0.9Hz,1H),8.34(s,1H),8.29(s ,2H),8.01(d,J=1.4Hz,1H),7.95(d,J=7.0Hz,1H),7.88(t,J=7.9Hz,1H),7.80(d,J=8. 6Hz,1H),7.72(dd,J=8.6,1.8Hz,1H),6.29(t,J=3.5Hz,1H),5.59–5.47(m,1H),3.25(d ,J=2.6Hz,2H),2.79(s,2H),2.76(d,J=2.2Hz,2H),2.46(s,3H),1.58(d,J=6.7Hz,6H).

[0098] Final product 13: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)-6-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)quinoline-2-carboxamide (13, 21-45-36)

[0099]

[0100] 6-Bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (09,21-18-41, 40 mg, 0.09 mmol), 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,2,3,6-tetrahydropyridine (22.5 mg, 0.1 mmol), sodium carbonate (48.5 mg, 0.45 mmol), and tetrakistriphenylphosphine palladium (10.6 mg, 0.009 mmol) were added to a 50 mL eggplant flask. 1,2-dimethoxyethane and water (2 mL each) were added and the mixture was reacted at 80°C for 2 h. The mixture was concentrated on a rotary evaporator and purified on a normal phase silica gel column. 33.6 mg of the target compound (13,21-45-36) was obtained in an 82.3% yield. 1 H NMR (400MHz, CDCl3): δ10.62 (s, 1H), 8.45 (d, J = 7.5Hz, 1H), 8.33 (s, 1H), 8.3 0(d,J=4.7Hz,1H),8.01(d,J=8.9Hz,1H),7.96(d,J=6.9Hz,1H),7.89–7.85( m,2H),7.76(d,J=1.8Hz,1H),6.29(t,J=3.5Hz,1H),5.62–5.50(m,1H),3.17 (d, J = 3.0Hz, 2H), 2.71 (d, J = 3.8Hz, 4H), 2.40 (s, 3H), 1.58 (d, J = 6.7Hz, 6H).

[0101] Final product 14: N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)-5-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)quinoline-2-carboxamide (14, 21-46-36)

[0102]

[0103] 5-Bromo-N-(6-(4-isopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (10,21-19-41, 40 mg, 0.09 mmol), 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,2,3,6-tetrahydropyridine (22.5 mg, 0.1 mmol), sodium carbonate (48.5 mg, 0.45 mmol), and tetrakistriphenylphosphine palladium (10.6 mg, 0.009 mmol) were added to a 50 mL eggplant flask. 1,2-dimethoxyethane and water (2 mL each) were added, and the mixture was reacted at 80°C for 2 h. The mixture was concentrated on a rotary evaporator and purified on a normal phase silica gel column. 20.6 mg of the target compound (14,21-46-36) was obtained in a 50.5% yield. 1 H NMR (400MHz, CDCl3): δ10.65(s,1H),8.58(d,J=8.8Hz,1H),8.47–8.43(m,1H),8.33(s,1H) ),8.30(d,J=8.7Hz,1H),7.98(d,J=3.1Hz,1H),7.88(t,J=7.9Hz,1H),7.70(dd,J=8.5,7. 1Hz,1H),7.43(dd,J=7.1,1.0Hz,1H),5.76–5.72(m,1H),5.62–5.49(m,1H),3.21(d,J=2. 8Hz, 2H), 2.77 (t, J = 5.6Hz, 2H), 2.57 (d, J = 1.8Hz, 2H), 2.46 (s, 3H), 1.59 (d, J = 6.7Hz, 6H).

[0104] Final product 15: N-(6-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (15, 24-57-31)

[0105]

[0106] Step 1: Add methyl 6-aminopicolinate (250 mg, 1.6 mmol) to a 100 mL eggplant flask, add hydrazine hydrate (3 mL), and reflux at 120°C under argon for 1 h. After the reaction, concentrate on a rotary evaporator and dry the solid. A crude white solid (250.8 mg, 100% yield) is obtained. The crude product is added to a 100 mL eggplant flask and dissolved in toluene. 1,1-dimethoxy-N,N-dimethylmethanamine (573.7 mg, 4.8 mmol) and cyclopropylamine (635.4 mg, 11.2 mmol) are added. The mixture is cooled to 0°C, and glacial acetic acid (289.2 mg, 4.8 mmol) is added. Under argon, reflux at 95°C for 20 h. Concentrate on a rotary evaporator and dry to dryness. Add an appropriate amount of water and concentrate again on a rotary evaporator. Dissolve the residue in 10 times the amount of water and stir at 85°C overnight. After the reaction, the mixture was extracted twice with ether to obtain the aqueous layer, which was then extracted three times with dichloromethane. The dichloromethane layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a silica gel column. 163 mg of the target compound (22-55-33) was obtained as a white solid in a 50.7% yield. 1 H NMR (400MHz, CDCl3): δ8.14(s,1H),7.53(t,J=8.0Hz,1H),7.46(d,J=7.2Hz,1H ),6.58(d,J=8.0Hz,1H),3.86(m,1H),1.05(q,J=6.8Hz,2H),0.89–0.82(m,2H); 13 C NMR (101MHz, DMSO-d6): δ163.43,158.62,147.84,143.63,138.10,111.22,109.80,25.86,6.05; ESI-HRMS (m / z)[M+H] + , C 10 H 11 N5, calculated value: 202.1014, measured value: 202.1093.

[0107] Step 2: Add quinoline-2-carboxylic acid (30 mg, 0.17 mmol) and (6-(4-cyclopropyl-4H-1,2,4-triazol-3-yl)pyridin-2-amine (35 mg, 0.17 mmol) to a 50 mL eggplant flask, and add 3 mL of anhydrous pyridine. Under argon protection, phosphorus oxychloride (106.1 mg, 0.7 mmol) was slowly added and stirred at room temperature for 2 h. After the reaction was completed, the reaction was quenched with water and extracted three times with ethyl acetate. The ethyl acetate layer was dried over anhydrous sodium sulfate. The organic solvent was concentrated on a rotary evaporator and purified by silica gel column chromatography. 47 mg of the target compound (15,24-57-31) was obtained as a white powdery solid with a yield of 76.3%. 1H NMR (400MHz, CDCl3): δ10.75(s,1H),8.55(dd,J=8.0,0.8Hz,1H),8.41(s,2H),8.31(s,1H),8.17(d,J=8.4Hz,1H),7.99(dd,J=7.6 ,0.8Hz,1H),7.95(t,J=8.0Hz,2H),7.82(m,1H),7.68(m,1H),4.02–3.92(m,1H),1.24(m,2H),1.01(m,2H); ESI-HRMS(m / z)[M+Na] + , C 20 H 16 N6O, calculated value: 379.1386, measured value: 3791286.

[0108] Final product 16: N-(6-(1-cyclobutyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (16, 24-61-34)

[0109]

[0110] Step 1: Add methyl 6-aminopicolinate (250 mg, 1.6 mmol) to a 100 mL eggplant flask, add hydrazine hydrate (3 mL), and reflux at 120°C under argon for 1 h. After the reaction, concentrate on a rotary evaporator and dry the solid. A crude white solid (250.8 mg, 100% yield) is obtained. The crude product is added to a 100 mL eggplant flask and dissolved in toluene. 1,1-dimethoxy-N,N-dimethylmethanamine (573.7 mg, 4.8 mmol) and cyclobutylamine (800 mg, 11.2 mmol) are added. The mixture is cooled to 0°C, and glacial acetic acid (289.2 mg, 4.8 mmol) is added. Under argon, reflux at 95°C for 20 h. Concentrate on a rotary evaporator and dry to dryness. Add an appropriate amount of water and concentrate again on a rotary evaporator. Dissolve the residue in 10 times the amount of water and stir at 85°C overnight. After the reaction, the mixture was extracted twice with ether to obtain the aqueous layer, which was then extracted three times with dichloromethane. The dichloromethane layer was dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a silica gel column. 224 mg of the target compound (21-149-37) was obtained as a yellow solid in a 65.1% yield. 1H NMR (400MHz, CDCl3): δ8.36(s,1H),7.57–7.52(m,1H),7.49(dd,J=7.6,0.8Hz,1H),6.58(dd,J =7.6,0.8Hz,1H),5.53–5.41(m,1H),2.60–2.51(m,2H),2.32–2.28(m,2H),1.88–1.78(m,2H); 13 C NMR (101MHz, DMSO-d6): δ159.18,151.04,145.34,143.62,138.06,111.19,108.53,49.64,30.79,14.34; ESI-HRMS(m / z)[M+Na] + calcd for C 11 H 13 N5,238.1171,found 238.1061.

[0111] Step 2: Add quinoline-2-carboxylic acid (30 mg, 0.17 mmol) and 6-(4-cyclobutyl-4H-1,2,4-triazol-3-yl)pyridin-2-amine (37 mg, 0.17 mmol) to a 50 mL eggplant-shaped flask, and add 3 mL of anhydrous pyridine. Under argon protection, slowly add phosphorus oxychloride (106.1 mg, 0.7 mmol) and stir at room temperature for 2 h. After the reaction is completed, quench with water and extract three times with ethyl acetate. Dry the ethyl acetate layer with anhydrous sodium sulfate. Concentrate the organic solvent on a rotary evaporator and purify by silica gel column chromatography. 47 mg of the target compound (16, 24-61-34) is obtained as a white solid in a yield of 74.9%. 1H NMR (400MHz, CDCl3): δ10.40(s,1H),9.80(s,1H),8.51(d,J=8.0Hz,2H),8.28–8.23(m,1H),8.21–8.16(m,1H),8.04(d,J=7.2 Hz,1H),7.99–7.89(m,3H),5.53(m,1H),2.76–2.64(m,2H),2.48–2.37(m,2H),2.05–1.98(m,2H); ESI-HRMS(m / z)[M+H]+calcd for C21H18N6O,371.1542,found371.1628.

[0112] Final product 17: N-(6-(1-cyclopentyl-4H-1,2,4-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (17, 24-58-31)

[0113]

[0114] Step 1: Add methyl 5-aminonicotinate (550 mg, 3.6 mmol) to a 100 mL eggplant-shaped flask and add 15 mL of anhydrous methanol. Add hydrazine hydrate (326.5 mg, 6.5 mmol) and stir under argon at reflux at 65°C overnight. After the reaction, concentrate on a rotary evaporator and dry the solid. A crude white solid (550.8 mg, 100% yield) is obtained. The crude product is added to a 100 mL eggplant-shaped flask and dissolved in toluene. Add 1,1-dimethoxy-N,N-dimethylmethanamine (1629.8 mg, 8.9 mmol) and cyclopentylamine (1627 mg, 19.1 mmol). Cool to 0°C, add glacial acetic acid (491.4 mg, 8.9 mmol), and reflux at 95°C under argon at reflux for 20 h. The residue was concentrated to dryness using a rotary evaporator. After drying, an appropriate amount of water was added and the mixture was concentrated again using a rotary evaporator. The residue was dissolved in 10 times the amount of water and stirred at 85°C overnight. After the reaction, the mixture was extracted twice with ether, leaving the aqueous layer. The aqueous layer was extracted three times with dichloromethane, dried over anhydrous sodium sulfate, concentrated using a rotary evaporator, and purified on a silica gel column. 110 mg of the target compound (23-143-36) was obtained as a white solid in a 17.6% yield. 1 H NMR (400MHz, CDCl3) δ8.28(s,1H),7.59–7.50(m,2H),6.58(dd,J=8.0,1.2Hz,1H),5.63–5.50(m,1H),2.30–2.18(m,2H),1.92–1.69(m,6H); 13 C NMR (101MHz, CDCl3): δ157.70,152.11,145.85,142.26,138.86,114.79,109.37,57.62,33.98,24.03.ESI-HRMS(m / z)[M+H] + calcd for C 12 H 15 N5,230.1327,found230.1408.

[0115] Step 2: Add quinoline-2-carboxylic acid (22.7 mg, 0.13 mmol) and 6-(4-cyclopentyl-4H-1,2,4-triazol-3-yl)pyridin-2-amine (30 mg, 0.13 mmol) to a 50 mL eggplant-shaped flask, and add 3 mL of anhydrous pyridine. Under argon protection, slowly add phosphorus oxychloride (80.4 mg, 0.5 mmol) and stir at room temperature for 2 h. After the reaction is completed, quench with water and extract three times with ethyl acetate. Dry the ethyl acetate layer with anhydrous sodium sulfate. Concentrate the organic solvent on a rotary evaporator and purify by silica gel column chromatography. 48 mg of the target compound (17, 24-58-31) is obtained as a white solid in a 96.4% yield. 1 H NMR (400MHz, CDCl3) δ10.75(s,1H),8.52(dd,J=8.4,0.8Hz,1H),8.43(s,1H),8.41(s,1H),8.13(d,J=8.4Hz,1H),8.04(dd,J=7.6, 0.8Hz,1H),7.99–7.92(m,2H),7.84(m,1H),7.69(m,1H),5.71–5.58(m,1H),2.41(m,2H),2.02–1.88(m,6H); ESI-HRMS(m / z)[M+Na] + calcd for C 22 H 20 N6O,407.1699,found407.1606.

[0116] Final product 18: (S)-8-bromo-N-(6-(4-(1-hydroxypropan-2-yl)-4H-1,24-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (18, 21-159-42)

[0117]

[0118] Step 1: Add 6-aminopicolinohydrazide (500 mg, 3.3 mmol) to a 100 mL eggplant-shaped flask, dissolve in acetonitrile, cool to 15°C, add 1,1-dimethoxy-N,N-dimethylmethanamine (1.57 g, 13.2 mmol), and stir at 90°C under argon for 4 h. After the reaction, cool to room temperature, add (R)-2-aminopropanol (1.24 g, 16.4 mmol), cool again to 15°C, add glacial acetic acid (394.7 mg, 6.6 mmol), and stir at 90°C under argon overnight. After the reaction, concentrate on a rotary evaporator and purify on a normal-phase silica gel column. 575.1 mg of the target compound (21-130-39) was obtained, with a yield of 79.5%. 1H NMR (400MHz, DMSO-d6): δ8.68(s,1H),7.51(t,J=8.0Hz,1H),7.17(d,J=7.2Hz,1H),6.51( d,J=8.0Hz,1H),6.17(s,2H),5.47–5.35(m,1H),3.65–3.59(m,2H),1.42(d,J=6.8Hz,3H); 13 CNMR(101MHz,DMSO-d6): δ159.17,151.33,145.84,143.32,138.17,111.68,108.56,64.37,45.29,17.71; ESI-HRMS(m / z)[M+H] + calcd for C 10 H 13 N5O,220.1120,found 220.1195.

[0119] Step 2: Add 8-bromoquinoline-2-carboxylic acid (62.1 mg, 0.25 mmol) to a 50 mL eggplant-shaped flask and add 4 mL of anhydrous toluene. Slowly add dichlorothionyl (39.1 mg, 0.33 mmol) dropwise and reflux at 80°C for 2 h. After completion of the reaction, concentrate to dryness on a rotary evaporator. Dissolve the residue in anhydrous toluene, add (S)-2-(3-(6-aminopyridin-2-yl)-4H-1,2,4-triazol-4-yl)propan-1-ol (30 mg, 0.14 mmol) and DIPEA (35.4 mg, 0.27 mmol), and reflux at 80°C for 2 h. After completion of the reaction, concentrate on a rotary evaporator, dilute the residue with dichloromethane, wash with saturated sodium bicarbonate solution, dry the organic layer over anhydrous sodium sulfate, concentrate on a rotary evaporator, and purify by silica gel column chromatography. 12 mg of the target compound (18,21-159-42) was obtained as a white to yellowish solid. The yield was 19.0%. 1 HNMR (400MHz, CD3OD): δ9.63(s,1H),8.65(t,J=8.4Hz,2H),8.44(d,J=8.4Hz,1H ),8.26(dd,J=7.6,1.2Hz,1H),8.20–8.13(m,1H),8.09(dd,J=8.4,1.2Hz,1H),8. 00(d,J=7.2Hz,1H),7.67–7.63(m,1H),5.88–5.77(m,1H),4.09(dd,J=12.0,3.6H z,1H),3.96(dd,J=12.0,5.6Hz,1H),1.74(d,J=7.2Hz,3H); ESI-HRMS(m / z)[M+H] + calcd for C20 H 17 BrN6O2,453.0596,found453.0659.

[0120] Final product 19: (R)-8-bromo-N-(6-(4-(1-hydroxypropan-2-yl)-4H-1,24-triazol-3-yl)pyridin-2-yl)quinoline-2-carboxamide (19, 21-155-36)

[0121]

[0122] Step 1: Add 6-aminopicolinohydrazide (100 mg, 0.66 mmol) to a 100 mL eggplant-shaped flask, dissolve in acetonitrile, cool to 15°C, add 1,1-dimethoxy-N,N-dimethylmethanamine (313.2 mg, 2.6 mmol), and stir at 90°C under argon for 4 h. After the reaction, cool to room temperature, add (S)-2-aminopropanol (247.9 mg, 3.3 mmol), cool again to 15°C, add glacial acetic acid (78.9 mg, 1.3 mmol), and stir at 90°C under argon overnight. After the reaction, concentrate on a rotary evaporator and purify on a normal-phase silica gel column. The target compound (20-141-42) was obtained, yielding 38 mg (26.4%). 1 H NMR (400MHz, DMSO-d6): δ8.68(s,1H),7.55–7.48(m,1H),7.17(d,J=7.2Hz,1H),6.51(dd,J=8.4 ,0.8Hz,1H),6.16(d,J=5.2Hz,1H),5.46–5.36(m,1H),3.70–3.54(m,2H),1.42(d,J=6.8Hz,3H); 13 C NMR (101MHz, DMSO-d6): δ159.12,151.31,145.89,143.29,138.12,111.63,108.48,64.39,53.28,17.71; ESI-HRMS(m / z)[M+H] + calcd for C 10 H 13 N5O,220.1120,found220.1183.

[0123] Step 2: Add 8-bromoquinoline-2-carboxylic acid (30 mg, 0.12 mmol) and (R)-2-(3-(6-aminopyridin-2-yl)-4H-1,2,4-triazol-4-yl)propan-1-ol (26.1 mg, 0.12 mmol) to a 50 mL eggplant flask, and add 3 mL of anhydrous pyridine. Under argon protection, phosphorus oxychloride (73.0 mg, 0.5 mmol) was slowly added and stirred at room temperature for 2 h. After the reaction was completed, quenched with water and extracted three times with ethyl acetate. The ethyl acetate layer was dried over anhydrous sodium sulfate. The organic solvent was concentrated on a rotary evaporator and purified by silica gel column chromatography. 6.2 mg of the target compound (19, 21-155-36) was obtained as a white solid in an 11.4% yield. 1 H NMR (400MHz, DMSO-d6): δ10.38(s,1H),9.11(s,1H),8.58(d,J=8.8Hz,1H),8.25(dd,J=7.6,1.2Hz,1H),8.11(dd,J=8.0,1.2Hz,1H),8.02( dd,J=8.0,1.2Hz,1H),7.96(d,J=8.4Hz,1H),7.82–7.73(m,2H),7.67–7.61(m,1H),6.24–6.15(m,1H),4.68(m,2H),1.68(d,J=7.2Hz,3H).

[0124] Final product 20: N-(6-(1-isopropyl-1H-pyrazol-5-yl)pyridin-2-yl)quinoline-2-carboxamide (20, 21-127-36)

[0125]

[0126] Step 1: Add 6-bromopyridin-2-amine (100 mg, 0.58 mmol) to a 100 mL eggplant-shaped flask. Add 1-isopropyl-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborol-2-yl)-1H-pyrazole (273 mg, 1.16 mmol), Xphos-Pd-G2 (45.4 mg, 0.058 mmol), and sodium carbonate (612.68 mg, 5.78 mmol). Dissolve the mixture in 1,4-dioxane and water (5:1) and stir at 100°C for 2 h. After the reaction, dilute with water. Extract the aqueous layer three times with dichloromethane. The dichloromethane layer is collected, dried over anhydrous sodium sulfate, concentrated on a rotary evaporator, and purified on a normal-phase silica gel column. The target compound (14-163-41) is obtained in a yield of 51.3% (60 mg). 1H NMR (400MHz, CDCl3): δ7.53(d,J=1.6Hz,1H),7.50(dd,J=8.4,7.6Hz,1H),6.87(dd,J=7.2,0.8Hz,1H) ,6.48(dd,J=8.4,0.8Hz,1H),6.43(d,J=2.0Hz,1H),5.32(m,1H),4.59(s,2H),1.50(d,J=6.4Hz,6H); 13 C NMR (101MHz, CDCl3): δ157.99,148.66,141.03,138.48,138.18,113.91,107.56,106.26,50.79,22.89; ESI-HRMS (m / z)[M+H] + calcd for C 11 H 14 N4,203.1218,found 203.1297.

[0127] Step 2: Add quinoline-2-carboxylic acid (25.7 mg, 0.15 mmol) and 6-(1-isopropyl-1H-pyrazol-5-yl)pyridin-2-amine (30 mg, 0.15 mmol) to a 50 mL eggplant-shaped flask, followed by 3 mL of anhydrous pyridine. Under argon, slowly add phosphorus oxychloride (91.03 mg, 0.59 mmol) and stir at room temperature for 2 h. After the reaction, acidify with 2M hydrochloric acid solution until a solid precipitates. The precipitated solid is filtered through a Buchner funnel, washed with water, and finally dried. The target compound (54 mg, 20, 21-127-36) is obtained as a white to yellowish solid in a 75.6% yield. 1 H NMR (400MHz, CDCl3): δ10.80(s,1H),8.46(dd,J=8.4,0.8Hz,1H),8.41(s,2H),8.19(d,J=8.8Hz,1H),7.93(dd,J=8.4,0.8Hz,1H),7.90–7.81(m,2H), 7.72–7.65(m,1H),7.60(d,J=2.0Hz,1H),7.34(dd,J=7.6,0.8Hz,1H),6.54 (d,J=2.0Hz,1H),5.36(m,1H),1.61(d,J=6.8Hz,6H); ESI-HRMS(m / z)[M+H] + calcd for C 21 H 19 N5O,358.1590,found 358.1699.

[0128] Final product 21: (R)-N-(6-(1-(1-hydroxypropan-2-yl)-1H-pyrazol-5-yl)pyridin-2-yl)quinoline-2-carboxamide (21, 21-174-39)

[0129]

[0130] Step 1: Add 6-aminopicolinohydrazide (100 mg, 0.66 mmol) to a 100 mL eggplant-shaped flask, dissolve in acetonitrile, cool to 15°C, add 1,1-dimethoxy-N,N-dimethylmethanamine (313.2 mg, 2.6 mmol), and stir at 90°C under argon for 4 h. After the reaction, cool to room temperature, add (S)-2-aminopropanol (247.9 mg, 3.3 mmol), cool again to 15°C, add glacial acetic acid (78.9 mg, 1.3 mmol), and stir at 90°C under argon overnight. After the reaction, concentrate on a rotary evaporator and purify on a normal-phase silica gel column. The target compound (20-141-42) was obtained, yielding 38 mg (26.4%). 1 H NMR (400MHz, DMSO-d6): δ8.68(s,1H),7.55–7.48(m,1H),7.17(d,J=7.2Hz,1H),6.51(dd,J=8.4 ,0.8Hz,1H),6.16(d,J=5.2Hz,1H),5.46–5.36(m,1H),3.70–3.54(m,2H),1.42(d,J=6.8Hz,3H); 13 C NMR (101MHz, DMSO-d6): δ159.12,151.31,145.89,143.29,138.12,111.63,108.48,64.39,53.28,17.71; ESI-HRMS(m / z)[M+H] + calcd for C 10 H 13 N5O,220.1120,found220.1183.

[0131] Step 2: Dissolve quinoline-2-carboxylic acid (30 mg, 0.17 mmol) in 5 mL of dichloromethane, add carbodiimide hydrochloride (33.2 mg, 0.17 mmol) and N-hydroxysuccinimide (33.2 mg, 0.17 mmol). Under argon, add (R)-2-(3-(6-aminopyridin-2-yl)-4H-1,2,4-triazol-4-yl)propan-1-ol (38.0 mg, 0.17 mmol), raise the temperature to 40°C, and reflux overnight. After completion of the reaction, wash with 20 mL of saturated sodium bicarbonate solution, dry over anhydrous sodium sulfate, concentrate by rotary evaporation, and purify by column chromatography to obtain 19.9 mg of the target compound (21, 21-174-39) as a white solid in a 31.3% yield. 1 H NMR (400MHz, DMSO-d6): δ8.94(s,1H),8.47(d,J=8.4Hz,1H),8.08(d,J=8.4Hz,1H),8.04(d,J=7.6Hz,1H),7.87–7.80(m,2H),7.75–7.68(m,1H ),7.40–7.34(m,1H),7.10(d,J=6.8Hz,1H),6.42(d,J=7.6Hz,1H),6.06–5.96(m,1H),4.66(m,2H),1.60(d,J=7.2Hz,3H); ESI-HRMS(m / z)[M+H] + calcd for C 20 H 18 N6O2,375.1491,found 375.1565.

[0132] Final product 22: (S)-N-(6-(1-(1-hydroxypropan-2-yl)-1H-pyrazol-5-yl)pyridin-2-yl)quinoline-2-carboxamide (22, 22-92-40)

[0133]

[0134] Step 1: Add 6-aminopicolinohydrazide (500 mg, 3.3 mmol) to a 100 mL eggplant-shaped flask, dissolve in acetonitrile, cool to 15°C, add 1,1-dimethoxy-N,N-dimethylmethanamine (1.57 g, 13.2 mmol), and stir at 90°C under argon for 4 h. After the reaction, cool to room temperature, add (R)-2-aminopropanol (1.24 g, 16.4 mmol), cool again to 15°C, add glacial acetic acid (394.7 mg, 6.6 mmol), and stir at 90°C under argon overnight. After the reaction, concentrate on a rotary evaporator and purify on a normal-phase silica gel column. 575.1 mg of the target compound (21-130-39) was obtained, with a yield of 79.5%.1 H NMR (400MHz, DMSO-d6): δ8.68(s,1H),7.51(t,J=8.0Hz,1H),7.17(d,J=7.2Hz,1H),6.51( d,J=8.0Hz,1H),6.17(s,2H),5.47–5.35(m,1H),3.65–3.59(m,2H),1.42(d,J=6.8Hz,3H); 13 CNMR(101MHz,DMSO-d6): δ159.17,151.33,145.84,143.32,138.17,111.68,108.56,64.37,45.29,17.71; ESI-HRMS(m / z)[M+H] + calcd for C 10 H 13 N5O,220.1120,found 220.1195.

[0135] Step 2: Add quinoline-2-carboxylic acid (70.9 mg, 0.41 mmol) to a 50 mL eggplant-shaped flask, followed by 4 mL of anhydrous toluene. Slowly add thionyl chloride (65 mg, 0.55 mmol) dropwise, and reflux at 80°C for 2 h. After completion of the reaction, concentrate the mixture to dryness on a rotary evaporator. Dissolve the residue in anhydrous toluene, add (S)-2-(3-(6-aminopyridin-2-yl)-4H-1,2,4-triazol-4-yl)propan-1-ol (50 mg, 0.23 mmol) and DIPEA (59 mg, 0.46 mmol), and reflux at 80°C for 2 h. After completion of the reaction, concentrate the mixture on a rotary evaporator, dilute the residue with dichloromethane, and wash with saturated sodium bicarbonate solution. Dry the organic layer over anhydrous sodium sulfate, concentrate the mixture on a rotary evaporator, and purify it by silica gel column chromatography. 23 mg of the target compound (22,22-92-40) was obtained as a white to yellowish solid in a 26.7% yield. 1 H NMR (400MHz, DMSO-d6): δ10.78(s,1H),8.80(s,1H),8.64(d,J=8.4Hz,1H), 8.31(d,J=8.4Hz,1H),8.25(d,J=8.8Hz,1H),8.18(d,J=8.4Hz,1H),8.10(d ,J=8.0Hz,1H),8.06(t,J=8.0Hz,1H),7.88(m,2H),7.74(t,J=8.0Hz,1H),5 .39(m,1H),3.74–3.65(m,2H),1.50(d,J=7.2Hz,3H); ESI-HRMS(m / z)[M+H] + calcd for C 20 H18 N6O2,375.1491,found 375.1553.

[0136] By selecting the corresponding raw materials, the following compounds can be synthesized:

[0137] Table 1

[0138]

[0139]

[0140]

[0141] Experimental Example 2: ADP-Glo ​​Kinase Assay Screening Method for ASK1: ADP-GLO luminescent kinase assay detects ADP formed in the kinase reaction. ADP is converted into ATP, which is then converted into light by Ultra-Glo luciferase.

[0142] Instruments: Multifunctional microplate reader (2104 Multilabel Reader, PerkinElmer, USA); biochemical incubator, purchased from Biochemical incubator Company; ultrasonic nanoliter liquid processing system, purchased from Echo Company.

[0143] Materials: ADP-Glo™ kit (Promega USA); ASK1 kinase, purchased from Eurofins. Principle: After the kinase reaction, the remaining ATP is completely consumed by the ADP-Glo ​​reagent. ADP is then converted to ATP, and the energy from ATP is transferred to luciferin, which is then converted into light by the Ultra-Glo™ luciferase. The luminescent signal is positively correlated with kinase activity. ADP-Glo™ can detect the activity of any enzyme that generates ADP, without the need for antibodies or radiolabeling.

[0144] Sample treatment: The sample was dissolved in DMSO and stored at low temperature. The concentration of DMSO in the final system was controlled within the range that did not affect the ADP-GloTM reagent.

[0145] ASK1 and substrates were diluted in HTRF kinase buffer (1X Kinase buffer, 25 mM MgCl2, 4 mM DTT, 20 mM HEPES, pH 7.5, 0.01% Triton X-100). 40 μL of compound was transferred from the source plate to a new 384-well plate used as an intermediate plate. 50 nL of compound was transferred to the assay plate via echo. ASK1 solution was prepared in 1X Kinase buffer at a concentration twice the final concentration of each reagent. 2.5 μL of kinase solution was added to each well of the assay plate. MBP substrate and ATP substrate solutions were prepared in 1X Kinase Reaction Buffer at a concentration four times the final concentration, corresponding to the concentration of each reagent required for the assay. The reaction was initiated by adding 2.5 μL of substrate solution to each well of the assay plate. The plate was shaken and incubated at 37°C for 60 minutes. 5 μL of ADP-Glo ​​reagent was added and incubated at 37°C for 180 minutes. Then, 10 μL of kinase assay reagent was added and allowed to equilibrate at room temperature for 30 minutes to convert ADP to ATP. Luciferase and luciferin were introduced to detect ATP. Data were collected on Envision. RLU values ​​were converted to percent inhibition values, and data were fitted in XLFit Excel add-in version 5.4.0.8 to obtain IC values. 50 The formula used is: Y = bottom + (top - bottom) / (1 + (IC 50 / X)^hillside).

[0146] Table 2

[0147]

[0148] (In the table, * indicates IC 50 >2001nM; ** indicates 501-2000nM; *** indicates IC 50 is 50-500 nM; **** indicates <50 nM, ***** indicates <20 nM).

Claims

1. A small molecule compound having a 2-pyridine-substituted carboxamide structure or a pharmaceutically acceptable salt thereof, characterized in that: The compound is specifically selected from one of the following structural formulas: 。 2. An ASK1 inhibitor, characterized in that: The inhibitor comprises the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

3. A pharmaceutical composition, characterized in that: include: (A) a therapeutically effective amount of the compound of claim 1 or a pharmaceutically acceptable salt thereof; and (B) a pharmaceutically acceptable carrier.

4. Use of the compound according to claim 1 in the preparation of a medicament for treating a disease associated with the activity or expression of ASK1 kinase.

5. The use according to claim 4, characterized in that: The disease associated with the activity or expression of ASK1 kinase is selected from one of liver disease, lung disease, cardiovascular disease, kidney disease, metabolic disease and tumor disease.

6. A method for synthesizing the compound according to claim 1, characterized in that: Its synthetic route is as follows: ； The definitions of the substituents in the general structure correspond to those of the compound of claim 1.

7. The method for synthesizing the compound according to claim 6, wherein: I-1 in the synthetic route is prepared by a one-step reaction. Step 1 is specifically: amine S2 is mixed with the corresponding acid S1 in a solvent, a base is added, and the reaction is carried out at room temperature, low temperature conditions or elevated temperature conditions to obtain the final product I-1; step 2 is specifically: I-1 is dissolved in DMF, NaH and iodomethane are added, and the reaction is stirred at room temperature, low temperature conditions or elevated temperature conditions to prepare the final product I-2.

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