Jnk-nlrp3 dual-target inhibitor and preparation method and application thereof

Abstract

This invention discloses compounds of Formula 1, or their tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof. These compounds exhibit excellent JNK inhibitory activity and NLRP3 inflammasome inhibitory activity. Therefore, the compounds of this invention lay a new material foundation for the development of therapeutic drugs for inflammation-related diseases.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry. Specifically, this invention relates to compounds having dual inhibitory activity against JNK-NLRP3, pharmaceutical compositions comprising said compounds, and their use in the preparation of remedies for JNK-NLRP3-related diseases. Background Technology

[0002] JNK (c-Jun N-terminal kinase) is a member of the MAPK (mitogen-activated protein kinase) family and plays a crucial role in cellular responses to various abiotic and biotic stresses. JNK regulates important physiological processes, including neuronal function, immune function, and embryonic development, by influencing gene expression, cytoskeletal protein dynamics, and cell death / survival pathways. Dysregulation of the JNK signaling pathway is associated with the development and progression of several human diseases, including diabetes, inflammatory diseases, and neurodegenerative diseases.

[0003] NLRP3 (NOD-like receptor protein 3) is a member of the NLR (NOD-like receptor) family. It forms the inflammasome complex in the cytoplasm and participates in the regulation of the innate immune system and inflammatory responses. The NLRP3 inflammasome can recognize pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs), activate caspase-1, and promote the maturation and secretion of IL-1β and IL-18, thereby mediating the inflammatory response. Aberrant activation of the NLRP3 inflammasome is associated with a variety of diseases, including metabolic diseases, autoinflammatory diseases, and tumors.

[0004] In some studies, it has been found that JNK can promote the activation of the NLRP3 inflammasome by directly phosphorylating NLRP3. This phosphorylation is one of the key steps in the activation of the NLRP3 inflammasome. For example, in diabetic nephropathy, the Syk / JNK / NLRP3 signaling pathway is involved in the pathogenesis of kidney disease, and the activation of JNK is associated with the expression level of the NLRP3 inflammasome.

[0005] Furthermore, JNK activation can also affect the NLRP3 inflammasome in other ways, such as by influencing NLRP3 ubiquitination and self-polymerization. In certain infectious diseases, such as Streptococcus pneumoniae infection, the ALK / JNK signaling pathway activates the NLRP3 inflammasome, thereby affecting pyroptosis and inflammatory responses.

[0006] In summary, the relationship between JNK and NLRP3 plays a crucial role in the development and progression of inflammatory diseases. JNK activation can promote the assembly and activation of NLRP3 inflammasomes, thereby influencing inflammatory responses and cell death processes. These findings offer potential therapeutic approaches for inflammatory diseases or other JNK & NLRP3-related conditions.

[0007] Aberrant activation of JNK and NLRP3 plays a crucial role in various diseases, and the two may have synergistic effects. Dual-target compounds offer multiple advantages, simplifying treatment regimens, improving patient compliance, reducing drug interactions, and enhancing therapeutic efficacy compared to simple drug combinations. Therefore, the development of dual-target drugs targeting JNK and NLRP3 has significant clinical implications and promising application prospects for the treatment of inflammatory diseases, cancer, and other related conditions. Summary of the Invention

[0008] The purpose of this invention is to provide a drug that targets JNK and NLRP3, thereby effectively treating inflammatory diseases, cancer and other related diseases.

[0009] Another object of the present invention is to provide a pharmaceutical composition comprising the said compound.

[0010] Another object of the present invention is to provide the use of the said compound or pharmaceutical composition in the preparation of a therapeutic remedy for JNK & NLRP3-related diseases and a method of treating JNK & NLRP3-related diseases using the said compound or pharmaceutical composition.

[0011] In a first aspect, the present invention provides compounds of Formula 1, or tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof.

[0012]

[0013] In the formula,

[0014] R1 is selected from: H, halogen, cyano, or optionally substituted C. 1-10 Alkyl, optionally substituted C 1-10 Alkoxy, -C(O)R 12 ;

[0015] R2 is H or C with optional substitution. 1-10 Alkyl groups, or R2 and R1 together with the carbon atoms attached to them, form optional substituted 3- to 10-membered heteroaryl or heterocyclic groups containing 1, 2 or 3 heteroatoms selected from N, O or S;

[0016] R3 is H or C with optional substitution. 1-10 Alkyl groups; or, R3 together with R1 or R4, along with the atoms attached to them, forms an optionally substituted 3- to 10-membered heteroaryl or heterocyclic group containing one, two, or three independent heteroatoms selected from N, O, or S.

[0017] Rings A and B are independently selected from: C 5-10 Aryl, 5-10 heteroaryl, C 3-10Cycloalkyl groups, 3-10 membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S, C 3-10 Cycloalkenyl, 3-10 membered heterocyclic alkenyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 8-10 membered bicyclic aryl or 8-10 membered bicyclic heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 8-10 membered bicyclic carbocyclic or 8-10 membered bicyclic heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S;

[0018] Where valence is permissible, x can be 0, 1, 2, 3, 4 or 5, and y can be 0, 1, 2, 3, 4 or 5;

[0019] R4 is selected from: H, hydroxyl group, or optionally substituted C. 1-10 Alkyl, optionally substituted C 5-10 Aryl, halogen, cyano, nitro, -C 0-6 -C(O)R7、-C 0-6 -OR8、-C 0-6 -N(R9)2, optionally substituted sulfonyl group, optionally substituted phosphonoyl group, optionally substituted 5-7 membered heteroaryl or heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S;

[0020] Alternatively, two adjacent R4s together with the atoms attached to them form an optionally substituted 3-10 membered heteroaryl or heterocyclic group containing one, two or three independent heteroatoms selected from N, O or S.

[0021] R5 is selected from: H, hydroxyl, halogen, cyano, nitro, -C 0-6 -C(O)R 13 -C 0-6 -OR 14 -C 0-6 -N(R 15 2. Optionally substituted sulfonyl group, Optionally substituted C 1-10 Alkyl, optionally substituted C 2-10 alkenyl, optionally substituted C 2-10 alkynyl group, optionally substituted C 1-10 Alkoxy, optional substituted C 5-10 aryl or optionally substituted C-type compounds containing one, two, or three heteroatoms independently selected from N, O, or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S;

[0022] J is selected from: -C(R) 16 )2-、-C(R 17 )=C(R 18)-、-N(R 19 )-, or J does not exist;

[0023] Where valence allows, W can be selected from: =O, =NR 10 -NH2; Q1 is selected from: =N-, -NR 20 -; Q2 is selected from: -NR 21 -、-C(R 11 )2-;

[0024] R6 is selected from C with optional substitution. 5-10 aryl, or optionally substituted C-type compounds containing one, two, or three heteroatoms independently selected from N, O, or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S;

[0025] Each R7, R 12 R 13 R 16 R 17 and R 18 Each is independently selected from: H, optionally substituted hydroxyl group, optionally substituted amino group, optionally substituted C group. 1-10 Alkyl, optionally substituted C 1-10 Alkoxy, optional substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S;

[0026] Each R8 and R 14 Each is independently selected from: H, and optionally substituted C. 1-3 Acyl group, optionally substituted C 1-10 Alkyl, optionally substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S;

[0027] Each R9, R 15 and R 19 Independently selected from: H, optional substituted C 1-3 Acyl group, optionally substituted C 1-10 Alkyl, optionally substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S.5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S;

[0028] R 10 It is H or cyano;

[0029] R 11 Selected from H, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NH-C 1-6 Alkyl, or -N-(C 1-6 Alkyl)2;

[0030] Each R 20 and R 21 Independently selected from: H, optional substituted C 1-10 alkyl.

[0031] In a preferred embodiment, R6 has 1-5 substituents; the substituents are selected from halogens or C. 1-10 Alkyl group; or, two substituents and the atoms attached to them form an optionally substituted C-shape. 5-10 aryl, or optionally substituted C-type compounds containing one, two, or three heteroatoms independently selected from N, O, or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S.

[0032] In a specific implementation method

[0033] R1 is selected from: H, halogen, or optionally substituted C. 1-6 alkyl;

[0034] R2 is H;

[0035] R3 is H;

[0036] Ring A is selected from: phenyl, 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 3-6 membered cycloalkyl or 5-6 membered heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S;

[0037] Ring B is a phenyl group;

[0038] x is 0, 1, 2, or 3;

[0039] y is 0, 1, or 2;

[0040] R4 is selected from: H, hydroxyl, halogen, cyano, nitro, or optionally substituted C. 1-6 Alkyl, optionally substituted C 1-6Alkoxy, -C 0-6 -C(O)R7、-C 0-6 -OR8、-C 0-6 -N(R9)2;

[0041] R5 is selected from: H, hydroxyl, halogen, cyano, nitro, or optionally substituted C. 1-6 Alkyl, optionally substituted C 1-6 Alkoxy, -C 0-6 -C(O)R 13 -C 0-6 -OR 14 -C 0-6 -N(R 15 )2;

[0042] J is -C(R) 16 )2- or does not exist;

[0043] W = O; Q1 = -NH-; Q2 = -NH-;

[0044] R6 is selected from: 2,6-dialkylphenyl, 2,6-dialkyl-4-halophenyl, or

[0045] Each R7, R 13 and R 16 Each is independently selected from: H, hydroxyl, optionally substituted amino, optionally substituted C. 1-6 Alkoxy;

[0046] Each R8 and R 14 Each is independently selected from: H, and C with optional substitution. 1-6 alkyl;

[0047] Each R9 and R 15 Independently selected from: H, optional substituted C 1-6 alkyl.

[0048] R 22 Selected from: hydrogen, halogen, cyano, hydroxyl, C 1-6 Alkyl, C 1-6 Haloalkyl, C 3-5 cycloalkyl and C 1-6 alkoxy groups, all of which may be halogenated, cyanoated, or C-shaped. 1-6 Alkoxy groups may be optionally substituted.

[0049] In a specific implementation, R1 is a halogen; preferably bromine.

[0050] In a specific implementation, R4 is -C(O)R7 or -OR8;

[0051] R7 is selected from: H, hydroxyl group, optionally substituted amino group, optionally substituted C group.1-6 Alkoxy;

[0052] R8 is selected from: H, C which is optionally substituted by one or more groups independently selected from the following 1-6 Alkyl groups: halogen, hydroxyl, carboxyl, C 1-3 Acyloxy group, -N(R) 23 )2;

[0053] Each R 23 H independently, or C with optional substitution 1-6 Alkyl; or, two Rs 23 Together with the nitrogen atom it is bonded to, it forms a 3-7 membered heterocycle with 1, 2 or 3 independently selected heteroatoms chosen from N, O or S, which may be substituted.

[0054] x is 1, 2 or 3, and at least one of them R4 is -C(O)R7 and is substituted at the -NH- or adjacent position.

[0055] In a specific implementation, R1 is selected from bromine;

[0056] Ring A is a phenyl group;

[0057] x is 1, 2, or 3;

[0058] y is 0;

[0059] R4 is selected from -C 0-6 -C(O)R7;

[0060] J does not exist;

[0061] R6 is selected as

[0062] R7 is selected from C 1-6 Alkyl (preferably C) 1-3 Alkyl-substituted amino groups;

[0063] R 22 Selected from: hydrogen, C 1-6 alkyl.

[0064] In a specific embodiment, the compound is selected from the group consisting of:

[0065]

[0066] Preferred

[0067] In a second aspect, the present invention provides a pharmaceutical composition comprising the compound described in the first aspect or its tautomers, mesosomes, racemates, enantiomers, diastereomers, mixtures thereof or pharmaceutically acceptable salts thereof, and optionally a pharmaceutically acceptable transporter.

[0068] In a third aspect, the present invention provides the use of the compound described in the first aspect or its tautomers, mesosomes, racemates, enantiomers, diastereomers, mixtures thereof or pharmaceutically acceptable salts thereof in the preparation of JNK inhibitors, NLRP3 inhibitors or JNK&NLRP3 dual-target inhibitors.

[0069] In a specific implementation, the JNK inhibitor, NLRP3 inhibitor, or JNK&NLRP3 dual-target inhibitor is a drug for treating and / or preventing JNK / NLRP3-related diseases.

[0070] In specific implementations, the JNK / NLRP3-related diseases include fibrosis, neurodegenerative diseases, diabetes, inflammatory diseases, tumors, central nervous system diseases, gout, etc.

[0071] In specific implementations, the fibrosis includes, but is not limited to, pulmonary fibrosis;

[0072] The neurodegenerative diseases mentioned include, but are not limited to, Alzheimer's disease;

[0073] The inflammatory diseases include, but are not limited to, arthritis and heart inflammation;

[0074] The tumors include, but are not limited to, lung cancer and liver cancer;

[0075] The central nervous system diseases mentioned include, but are not limited to, cerebral ischemia-reperfusion.

[0076] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Detailed Implementation

[0077] Through extensive and in-depth research, the inventors unexpectedly discovered a series of novel compounds with JNK & NLRP3 inhibitory activity, which enable the preparation of therapeutic drugs for JNK & NLRP3-related diseases, thereby treating these diseases. This invention was completed based on this discovery.

[0078] Terminology Definition

[0079] The terms used herein to refer to groups, substituents, or structures of compounds have the same meaning as understood by those skilled in the art. For clarity, the terms used in this specification are defined as follows.

[0080] In this invention, "a", "a type" or "a class" means at least one / a type or more than one / a type.

[0081] In this paper, expressions such as "1-n" refer to groups having 1 to n carbon atoms. For example, "1-10" indicates that a group has 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms; similarly, "C6~C10" indicates that a group has 6, 7, 8, 9, or 10 carbon atoms. Furthermore, the descriptions of the range of carbon atom counts in this paper also include sub-ranges. For instance, when referring to 1-10 carbon atoms, this paper also includes cases with 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, and 1-3 carbon atoms.

[0082] As used herein, the term "alkyl" has the same meaning as commonly understood by those skilled in the art, referring to various saturated or unsaturated straight-chain, side-chain, or cyclic hydrocarbon groups. For example, the alkyl group referred to herein refers to a lower alkyl group with 1-10 carbon atoms; preferably a lower alkyl group with 1-8 carbon atoms; more preferably a lower alkyl group with 1-6 carbon atoms. In specific embodiments, the alkyl group referred to herein includes, but is not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, heptyl, etc. Similarly, the terms "alkenyl" or "alkynyl" as used herein refer to various unsaturated straight-chain, side-chain, or cyclic hydrocarbon groups containing carbon-carbon double or triple bonds.

[0083] As used herein, the terms "aryl" or "aromatic ring" have the same meaning as commonly understood by those skilled in the art, referring to a cyclic conjugated aromatic system; for example, the term "C6-C10 aryl" refers to an aromatic cyclic group with 6 to 10 carbon atoms that does not contain heteroatoms in the ring, such as phenyl or naphthyl. The term "heteroaryl," as used herein, refers to a cyclic conjugated aromatic system containing one or more heteroatoms such as N, O, or S in the ring; for example, pyridyl or pyrazinyl.

[0084] As used herein, the term "halogen" has the meaning commonly understood by one of ordinary skill in the art. In specific embodiments, halogen refers to fluorine, chlorine, bromine, or iodine.

[0085] As used herein, the term "substitution" refers to the replacement of one or more hydrogen atoms on a specific group by a specific substituent. A specific substituent may be a substituent described above or a specific substituent appearing in the various embodiments. Therefore, in this invention, the substituents in Formula 1 can each independently be the corresponding group in the specific compounds of the embodiments; that is, this invention includes combinations of the substituents in Formula 1 above, as well as combinations of some of the substituents shown in Formula 1 with other specific substituents appearing in the embodiments.

[0086] Unless otherwise specified, a substituted group may have a particular substituent at any substituted site on that group, and the substituents may be the same or different at each position. Cyclic substituents, such as heterocyclic groups, may be attached to another ring, such as a cycloalkyl group, thereby forming a spirobicyclic system, for example, where the two rings share a common carbon atom.

[0087] In particular, the various substituents defined above also include groups formed by further substitution of these substituents, which may also contain other groups. For example, the hydrogen atoms on alkyl and aryl groups are replaced by amino, halogen, or other groups to become groups belonging to the above definitions.

[0088] As used herein, the term "optionally substituted" means that a group may be substituted or not substituted. In specific embodiments, "optionally substituted" means optionally substituted by one or more substituents selected from the following: cyano, halogen, hydroxyl, optionally substituted amino, nitro, carboxyl, ester, oxo, deuterated, optionally substituted C 1-3 Alkyl, optionally substituted C 1-3 Alkoxy, optional substituted C 1-3 Acyloxy group, optionally substituted C 5-10 Aryl or heteroaryl, optionally substituted 3-7 membered cycloalkyl or heterocyclic, optionally substituted sulfonyl, optionally substituted acyl.

[0089] The compounds of the present invention

[0090] The compound of this invention is a dual-target compound; that is, a single-component compound that can act on two or more molecular targets simultaneously. This drug design strategy is of great significance for the treatment of complex diseases, such as cancer, metabolic diseases, autoimmune diseases, and neurodegenerative diseases. Compared with single-target drugs, dual-target compounds have the following advantages: (1) Improved therapeutic effect: By acting on multiple key aspects of the disease simultaneously, dual-target compounds may produce a synergistic effect, thereby improving the therapeutic effect; (2) Reduced drug resistance: Since the drug acts on multiple targets, disease cells need to generate drug resistance mutations on multiple targets simultaneously to evade the drug's effect, thus making it more difficult to develop drug resistance; (3) Reduced side effects: The synergistic effect of dual targets can achieve therapeutic effects at lower doses, and side effects can be reduced by lowering the dose of single-target drugs; (4) Simplified treatment regimen: Compared with multi-drug combination therapy, dual-target compounds can simplify the treatment regimen, make it easier for patients to take, and improve compliance.

[0091] Given the important role of the relationship between JNK and NLRP3 in the occurrence and development of inflammatory diseases, the inventors have provided pyrimidine derivatives that target and inhibit JNK and NLRP3, the structural formula of which is shown in Formula 1:

[0092]

[0093] The substituents in the formula are as described above.

[0094] Based on the compounds of the present invention, the present invention also provides a pharmaceutical composition comprising the above-described compounds or their tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof or pharmaceutically acceptable salts thereof, and optionally a pharmaceutically acceptable carrier.

[0095] Based on the teachings of this invention, those skilled in the art will understand that the compounds of this invention can be used to prepare JNK & NLRP3 inhibitors. In specific embodiments, the JNK & NLRP3 inhibitors of this invention are drugs for treating and / or preventing JNK & NLRP3-related diseases. In specific embodiments, JNK & NLRP3-related diseases include, but are not limited to, neurodegenerative diseases, diabetes, inflammatory diseases, central nervous system diseases, fibrosis, gout, etc.

[0096] Based on the teachings of this invention, those skilled in the art can prepare the compounds of this invention into various dosage forms and determine the dosage, administration method, and administration time according to the actual situation of the patient to be treated, including but not limited to age, gender, disease severity, and previous treatment history. Furthermore, based on the teachings of this invention, those skilled in the art can understand that the compounds of this invention are compounds with therapeutic activity; in other words, the compounds of this invention have the potential to be used as drugs. Therefore, those skilled in the art can qualitatively and quantitatively detect various characteristics of the compounds of this invention as drugs using conventional techniques, including but not limited to therapeutic activity, toxicity, bioavailability, and drug-likeness. Performing these tests is obvious to those skilled in the art and requires no inventive effort.

[0097] Advantages of this invention:

[0098] 1. The compounds of the present invention exhibit excellent JNK inhibitory activity;

[0099] 2. The compounds of the present invention exhibit excellent NLRP3 inhibitory activity;

[0100] 3. The compounds of this invention lay a new material foundation for the development of therapeutic drugs for JNK & NLRP3-related diseases.

[0101] The technical solution of the present invention is further described below with reference to specific implementation examples. However, the following implementation examples do not constitute a limitation on the present invention. All application methods based on the principles and technical means of the present invention are within the scope of the present invention. Experimental methods in the following embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are by weight.

[0102] There are no particular restrictions on the source of any raw materials used in this invention; they can be purchased from the market or prepared using conventional methods known to those skilled in the art.

[0103] Example

[0104] Synthesis of intermediates

[0105] Intermediate 1

[0106]

[0107] Step 1:

[0108]

[0109] 2,4-Dichloro-5-bromopyrimidine (1 eq), 2-amino-N-methylbenzamide (1.2 eq), and N,N-diisopropylethylamine (3 eq) were dissolved in isopropanol and reacted with the solution at 90 °C for 16 h. After the reaction was completed, the solution was cooled to room temperature, filtered, and the residue was washed three times with isopropanol to obtain a white to pale yellow solid.

[0110] 1 H NMR (400MHz, DMSO-d6) δ12.02(s,1H),8.98(d,J=4.8Hz,1H),8.56(s,1H),8.48(dd,J=8.5,1.2Hz,1H),7.84( dd,J=7.9,1.6Hz,1H),7.60(ddd,J=8.6,7.4,1.5Hz,1H),7.23(td,J=7.6,1.2Hz,1H),2.81(d,J=4.5Hz,3H).

[0111] Step 2:

[0112]

[0113] The product from step 1 (1 eq), 4-aminobenzenesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with stirring at 110 °C for 16 h. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white to pale yellow solid.

[0114] 1H NMR (400MHz, DMSO-d6) δ11.56(s,1H),10.15(s,1H),8.91(q,J=4.5Hz,1H),8.58(d,J=8.4Hz,1H),8.41( s,1H),7.86–7.79(m,3H),7.71(d,J=8.9Hz,2H),7.58–7.52(m,1H),7.22(m,3H),2.81(d,J=4.4Hz,3H).

[0115] Intermediate 2

[0116]

[0117] Step 1: Same as step 1 for intermediate 1;

[0118] Step 2:

[0119]

[0120] The product from step 1 (1 eq), methyl 4-amino-2-aminosulfonylbenzoate (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with stirring at 110 °C for 16 h. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0121] 1 H NMR (400MHz, DMSO-d6) δ11.61(s,1H),10.18(s,1H),8.89(q,J=4.5Hz,1H),8.75(d,J=8.5Hz,1H),8.39(s,1H),8.30(d,J=2.2Hz,1H),8.21(dd,J= 8.6,2.3Hz,1H),7.80(dd,J=8.0,1.6Hz,1H),7.71(d,J=8.5Hz,1H),7.63 –7.51(m,1H),7.19(d,J=9.0Hz,3H),3.85(s,3H),2.81(d,J=4.5Hz,3H).

[0122] Intermediate 3

[0123]

[0124] Step 1: Same as step 1 for intermediate 1;

[0125] Step 2:

[0126]

[0127] The product from step 1 (1 eq), 3-aminobenzenesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with stirring at 110 °C for 16 h. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0128] 1 H NMR (400MHz, DMSO-d6) δ11.78(s,1H),10.23(s,1H),8.93(q,J=4.5Hz,1H),8.61(d,J=8.4Hz,1H),8.42(s,1H),8.02(s,1H),7.95(d q,J=6.3,4.1,3.5Hz,1H),7.81(dd,J=7.9,1.6Hz,1H),7.56–7.46(m,3H),7.40(s,2H),7.19(t,J=7.5Hz,1H),2.81(d,J=4.4Hz,3H).

[0129] Intermediate 4

[0130]

[0131] Step 1:

[0132]

[0133] 2-tert-butoxycarbonylamino-5-hydroxybenzoic acid (1 eq), methylamine hydrochloride (3.25 eq), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.5 eq), 1-hydroxybenzotriazole (1.65 eq), and N,N-diisopropylethylamine (6.5 eq) were dissolved in N,N-dimethylformamide and reacted at room temperature for 16 h. After the reaction was completed, the mixture was extracted three times with ethyl acetate / water and filtered through a silica gel column to obtain a brown solid.

[0134] 1 H NMR (400MHz, DMSO-d6) δ10.10(s,1H),9.42(s,1H),8.59(d,J=5.3Hz,1H),7.91(d,J=9.0Hz, 1H), 7.02 (d, J = 2.8Hz, 1H), 6.88 (dd, J = 9.0, 2.8Hz, 1H), 2.74 (d, J = 4.5Hz, 3H), 1.44 (s, 9H).

[0135] Step 2:

[0136]

[0137] The product from step 1 (1 eq), 1,2-dichloroethane (2 eq), and potassium carbonate (3 eq) were dissolved in N,N-dimethylformamide and reacted at 80 °C for 16 h. After the reaction was completed, the mixture was extracted three times with ethyl acetate / water and filtered through a silica gel column to obtain a white solid.

[0138] 1 H NMR (400MHz, DMSO-d6) δ10.44(s,1H),8.72(d,J=4.9Hz,1H),8.09(d,J=9.1Hz,1H),7.27(d,J=2.9Hz,1H),7.11(d d,J=9.1,2.9Hz,1H),4.27(dd,J=6.1,4.2Hz,2H),3.96(dd,J=5.9,4.4Hz,2H),2.77(d,J=4.5Hz,3H),1.46(s,9H).

[0139] Step 3:

[0140]

[0141] Dissolve the product from step 2 in dichloromethane, and add trifluoroacetic acid (V) dropwise while stirring. 二氯甲烷 V 三氟乙酸 =3:1), stir the mixture at room temperature for 2 hours. After the reaction is complete, remove the liquid by rotary evaporation and use it directly in the next step.

[0142] Step 4:

[0143]

[0144] 2,4-Dichloro-5-bromopyrimidine (1 eq), the product of step 3 (1.2 eq), and N,N-diisopropylethylamine (3 eq) were dissolved in isopropanol and reacted with the solution at 90 °C for 16 h. After the reaction was completed, the solution was cooled to room temperature, filtered, and the residue was washed three times with isopropanol to obtain a white solid.

[0145] 1 H NMR (400MHz, DMSO-d6) δ11.64(s,1H),8.87(d,J=4.9Hz,1H),8.50(s,1H),8.35(d,J=9.1Hz,1H),7.37(d,J= 2.9Hz, 1H), 7.24 (dd, J = 9.2, 2.9Hz, 1H), 4.33 (t, J = 5.1Hz, 2H), 4.00 (t, J = 5.1Hz, 2H), 2.80 (d, J = 4.4Hz, 3H).

[0146] Step 5:

[0147]

[0148] The product from step 4 (1 eq), 4-aminobenzenesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with stirring at 110 °C for 16 h. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0149] 1 H NMR (400MHz, DMSO-d6) δ11.30(s,1H),10.10(s,1H),8.86(d,J=4.8Hz,1H),8.44(d,J=9.1Hz,1H),8.37(s,1H),7.80(d,J=8.9Hz,2H),7.75–7.69(m, 2H),7.38(d,J=3.0Hz,1H),7.25(s,2H),7.14(dd,J=9.2,2.9Hz,1H),4.35 (dd, J=6.0, 4.2Hz, 2H), 4.01 (dd, J=6.0, 4.2Hz, 2H), 2.81 (d, J=4.5Hz, 3H).

[0150] Step 6:

[0151]

[0152] The product from step 5 (1 eq), N-methylpiperazine (5 eq), potassium carbonate (5 eq), and potassium iodide (0.1 eq) were dissolved in N,N-dimethylformamide and reacted at 100°C for 16 h. After the reaction was completed, the reaction was quenched with water. After adding excess water, a solid precipitated out. The solid was filtered and slurried with ethyl acetate / methanol to obtain a white solid.

[0153] 1 H NMR (400MHz, DMSO-d6) δ11.05(s,1H),9.83(s,1H),8.76(d,J=4.8Hz,1H),8.45(d,J=9.1Hz,1H),8.32(s,1H),7.83(d,J=8.9Hz,2H),7.72–7.64 (m,2H),7.33(d,J=2.9Hz,1H),7.20(s,2H),7.11(dd,J=9.2,2.9Hz,1H),4.15(t,J=5.8Hz,2H),2.80(d,J=4.4Hz,3H),2.75–2.71(m,2H),2.20(m 11H).

[0154] Intermediate 5

[0155]

[0156] Steps 1-5: Same as steps 1-5 for intermediate 4;

[0157] Step 6:

[0158]

[0159] Take the product from step 5 (1 eq), dimethylamine aqueous solution (5 eq), potassium carbonate (5 eq), and potassium iodide (0.1 eq) and dissolve them in N,N-dimethylformamide. React at 100°C for 16 h. After the reaction is complete, quench the reaction with water. After adding excess water, a solid precipitates out. Filter the solid and slurry it with ethyl acetate / methanol to obtain a white solid.

[0160] 1 H NMR(400MHz,DMSO-d6)δ11.05(s,1H),9.83(s,1H),8.76(d,J=4.8Hz,1H),8 .45(d,J=9.1Hz,1H),8.32(s,1H),7.83(d,J=8.7Hz,2H),7.68(d,J=8.7Hz, 2H),7.33(d,J=2.9Hz,1H),7.20(s,2H),7.12(dd,J=9.1,2.9Hz,1H),4.12( t,J=5.8Hz,2H),2.81(d,J=4.5Hz,3H),2.66(t,J=5.8Hz,2H),2.24(s,6H).

[0161] Intermediate 6

[0162]

[0163] Steps 1-4: Same as steps 1-4 for intermediate 4;

[0164] Step 5:

[0165]

[0166] The product from step 4 (1 eq), 1-(4-aminophenyl)methanesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with the solution at 110 °C for 16 h. After the reaction was completed, the solution was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0167] 1H NMR (400MHz, DMSO-d6) δ11.49 (s, 1H), 10.01 (s, 1H), 8.88 (q, J = 4.5Hz, 1H), 8. 45(d,J=9.1Hz,1H),8.34(s,1H),7.59(d,J=8.2Hz,2H),7.37(d,J=3.0Hz,1H), 7.31(d,J=8.5Hz,2H),7.11(dd,J=9.2,2.9Hz,1H),6.86(s,2H),4.34(dd,J=6. 1,4.1Hz,2H),4.25(s,2H),4.00(dd,J=6.1,4.1Hz,2H),2.80(d,J=4.4Hz,3H).

[0168] Step 6:

[0169]

[0170] Take the product from step 5 (1 eq), dimethylamine aqueous solution (5 eq), potassium carbonate (5 eq), and potassium iodide (0.1 eq) and dissolve them in N,N-dimethylformamide. React at 100°C for 16 h. After the reaction is complete, quench the reaction with water. After adding excess water, a solid precipitates out. Filter the solid and slurry it with ethyl acetate / methanol to obtain a white solid.

[0171] 1 H NMR (400MHz, DMSO-d6) δ11.04(s,1H),9.50(s,1H),8.75(q,J=4.5Hz,1H),8.5 1(d,J=9.2Hz,1H),8.25(s,1H),7.65(d,J=8.3Hz,2H),7.30(d,J=2.9Hz,1H), 7.28–7.22(m,2H),7.10(dd,J=9.1,2.9Hz,1H),6.81(s,2H),4.20(s,2H),4.1 2(t,J=5.8Hz,2H),2.80(d,J=4.5Hz,3H),2.66(t,J=5.7Hz,2H),2.25(s,6H).

[0172] Intermediate 7

[0173]

[0174] Steps 1-4: Same as steps 1-4 for intermediate 4;

[0175] Step 5:

[0176]

[0177] The product from step 4 (1 eq), 1-(4-aminophenyl)methanesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with the solution at 110 °C for 16 h. After the reaction was completed, the solution was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0178] 1 H NMR (400MHz, DMSO-d6) δ11.40(s,1H),10.02(s,1H),8.85(d,J=4.9Hz,1H),8.48(d,J=9.2Hz,1H),8.35(s,1H),8.00(s,1H),7.92(dd,J=6.1,2.9H z,1H),7.50(d,J=6.6Hz,2H),7.40–7.34(m,3H),7.12(dd,J=9.2,2.9Hz,1H),4.34(t,J=5.1Hz,2H),3.99(d,J=5.3Hz,2H),2.80(d,J=4.4Hz,3H).

[0179] Step 6:

[0180]

[0181] Take the product from step 5 (1 eq), dimethylamine aqueous solution (5 eq), potassium carbonate (5 eq), and potassium iodide (0.1 eq) and dissolve them in N,N-dimethylformamide. React at 100°C for 16 h. After the reaction is complete, quench the reaction with water. After adding excess water, a solid precipitates out. Filter the solid and slurry it with ethyl acetate / methanol to obtain a white solid.

[0182] 1 H NMR(400MHz,DMSO-d6)δ11.15(s,1H),9.71(s,1H),8.75(d,J=4.7Hz,1H),8 .56(d,J=9.1Hz,1H),8.28(s,1H),8.06(s,1H),7.97(dt,J=7.9,1.8Hz,1H), 7.49–7.39(m,2H),7.31(d,J=7.9Hz,3H),7.11(dd,J=9.2,2.9Hz,1H),4.11 (t,J=5.8Hz,2H),2.80(d,J=4.5Hz,3H),2.66(t,J=5.8Hz,2H),2.24(s,6H).

[0183] Intermediate 8

[0184]

[0185] Dissolve 1,2,3,5,6,7-hexahydro-S-inden-4-amine (1 eq) in ultra-dry tetrahydrofuran. Add triethylamine (1.1 eq) dropwise with stirring at room temperature, followed by the dropwise addition of triphosgene (0.33 eq)-tetrahydrofuran solution. After the addition is complete, heat to reflux for 4 hours. After the reaction is complete, filter, remove the solvent by rotary evaporation, and use the residue directly in the next reaction step.

[0186] Intermediate 9

[0187]

[0188] The raw material was replaced with 8-chloro-1,2,3,5,6,7-hexahydro-S-indene-4-amine, and the rest was the same as in step 1 of intermediate 8.

[0189] Intermediate 10

[0190]

[0191] The raw material was replaced with 4-chloro-2,6-diisopropylaniline, and the rest was the same as in step 1 of intermediate 8.

[0192] Intermediate 11

[0193]

[0194] Step 1: Same as step 1 for intermediate 1;

[0195] Step 2:

[0196]

[0197] The product from step 1 (1 eq), 3-methyl-4-aminobenzenesulfonamide (2 eq), and trifluoroacetic acid (3 eq) were dissolved in sec-butanol and reacted with stirring at 110 °C for 16 h. After the reaction was completed, the mixture was cooled to room temperature, filtered, and the residue was washed three times with sec-butanol to obtain a white solid.

[0198] 1 H NMR (400MHz, DMSO-d6) δ11.90(s,1H),9.54(s,1H),8.83(q,J=4.6Hz,1H),8.35(d,J=13.7Hz,2H) ,7.77–7.64(m,4H),7.41–7.28(m,3H),7.14(t,J=7.5Hz,1H),2.80(d,J=4.5Hz,3H),2.30(s,3H).

[0199] Example 1: Synthesis of compound B12

[0200]

[0201] Intermediate 1 (1 eq) was dissolved in ultra-dry N,N-dimethylformamide. Under nitrogen protection, sodium hydride (1 eq) was added in an ice bath, and stirring was continued for 15 minutes. Intermediate 8 (1 eq) dissolved in ultra-dry N,N-dimethylformamide was added dropwise, and the reaction was carried out at room temperature for 6 hours. After the reaction was completed, ice water was added to quench the reaction, and the reaction solution was acidified to pH 5. The solution was filtered, and the residue was slurried with ethyl acetate / methanol to obtain a white to pale yellow solid.

[0202] 1 H NMR (400MHz, DMSO-d6) δ11.44(s,1H),10.97(s,1H),10.03(s,1H),8.85(p,J=4.5,4.0 Hz,1H),8.77(s,1H),8.61(d,J=8.5Hz,1H),8.39(s,1H),7.89(d,J=8.8Hz,2H),7.82–7 .75(m,3H),7.54(t,J=7.8Hz,1H),7.15(t,J=7.6Hz,1H),6.89(s,1H),2.81(d,J=4.4Hz ,3H),2.74(t,J=7.4Hz,4H),2.56(d,J=7.4Hz,4H),1.88(p,J=7.4Hz,4H).MS(ESI):m / z 678.13[M+H] + .

[0203] Example 2: Synthesis of compound B13

[0204]

[0205] The raw materials are intermediate 1 and intermediate 9, and the rest are the same as in Example 1.

[0206] 1 H NMR (400MHz, DMSO-d6) δ11.44(s,1H),10.73(s,1H),10.02(s,1H),8.78(d,J=5. 2Hz,1H),8.60(d,J=8.4Hz,1H),8.39(s,1H),8.28(s,1H),7.91–7.86(m,2H),7.8 0(d,J=8.8Hz,2H),7.77–7.73(m,1H),7.51(t,J=7.9Hz,1H),7.15(t,J=7.5Hz,1 H),2.87–2.76(m,7H),2.63(t,J=7.4Hz,4H),1.94(p,J=7.4Hz,4H).MS(ESI):m / z 712.09 [M+H] + .

[0207] Example 3: Synthesis of Compound B16

[0208]

[0209] The raw materials are intermediate 1 and intermediate 10, and the rest are the same as in Example 1.

[0210] 1 H NMR (400MHz, DMSO-d6) δ11.44(s,1H),11.17(s,1H),10.04(s,1H),8.85(q,J=4. 5Hz,1H),8.73(s,1H),8.60(d,J=8.4Hz,1H),8.39(s,1H),7.96(s,1H),7.90(d, J=8.7Hz,2H),7.77(d,J=8.8Hz,2H),7.58–7.52(m,1H),7.17(t,J=7.5Hz,1H),7 .07(s,2H),2.81(d,J=4.5Hz,3H),2.78(m,2H),1.06–0.79(m,12H).MS(ESI):m / z 716.12[M+H] + .

[0211] Example 4: Synthesis of compound B18

[0212]

[0213] The raw materials are intermediate 2 and intermediate 8, and the rest are the same as in Example 1.

[0214] 1 H NMR (400MHz, DMSO-d6) δ11.60(s,1H),10.58(s,1H),10.17(d,J=3.5Hz,1H),9.06(s,1H),8.86(d, J=4.7Hz,1H),8.77(d,J=8.2Hz,1H),8.41–8.37(m,2H),8.27(dd,J=8.6,2.2Hz,1H),7.79(dd,J=7 .9,1.6Hz,1H),7.66(d,J=8.6Hz,1H),7.57–7.52(m,1H),7.22–7.13(m,1H),6.89(s,1H),3.92(s, 3H),2.81(d,J=4.6Hz,3H),2.74(m,4H),2.58(t,J=7.4Hz,4H),1.88(p,J=7.4Hz,4H).MS(ESI):m / z 736.14[M+H] + .

[0215] Example 5: Synthesis of compound B19

[0216]

[0217] The raw materials are intermediate 3 and intermediate 8, and the rest are the same as in Example 1.

[0218] 1 H NMR(400MHz,DMSO-d6)δ11.54(s,1H),11.14(s,1H),9.87(s,1H),8.89–8.68(m,3 H),8.34(d,J=5.8Hz,1H),8.23(s,1H),8.09(d,J=6.8Hz,1H),7.77(d,J=7.9Hz,1H ),7.50(d,J=7.7Hz,3H),7.15(t,J=7.5Hz,1H),6.89(s,1H),2.81(d,J=4.3Hz,3H) ,2.74(t,J=7.7Hz,4H),2.55(t,J=7.4Hz,4H),1.88(p,J=7.5Hz,4H).MS(ESI):m / z 678.13[M+H] + .

[0219] Example 6: Synthesis of compound B21

[0220]

[0221] The raw materials are intermediate 4 and intermediate 8, and the rest are the same as in Example 1.

[0222] 1 H NMR (400MHz, DMSO-d6) δ10.93 (s, 1H), 9.67 (s, 1H), 8.75 (d, J = 5.2Hz, 1H), 8. 41(d,J=9.0Hz,1H),8.28(s,1H),7.71–7.60(m,6H),7.30(d,J=2.7Hz,1H),7. 09(d,J=9.2Hz,1H),6.78(s,1H),4.11(t,J=5.9Hz,2H),2.79(d,J=4.4Hz,4H ),2.71(m,6H),2.59(t,J=7.4Hz,4H),2.21(s,3H),1.88(m,4H).MS(ESI):m / z 820.24[M+H] + .

[0223] Example 7: Synthesis of compound B22

[0224]

[0225] The raw materials are intermediate 5 and intermediate 8, and the rest are the same as in Example 1.

[0226] 1 H NMR (400MHz, Methanol-d4) δ8.27 (d, J = 9.1 Hz, 1H), 8.07 (s, 1H), 7.78 (d, J = 8. 7Hz,2H),7.68(d,J=8.8Hz,2H),7.31(d,J=2.9Hz,1H),7.07(dd,J=9.1,3.0Hz, 1H),6.83(s,1H),4.09(t,J=5.4Hz,2H),2.92(s,3H),2.78(t,J=7.4Hz,4H),2. 75–2.64(m,6H),2.33(s,6H),1.95(p,J=7.4Hz,4H).MS(ESI):m / z765.20[M+H] + .

[0227] Example 8: Synthesis of compound B23

[0228]

[0229] The raw materials are intermediate 6 and intermediate 8, and the rest are the same as in Example 1.

[0230] 1 H NMR (400MHz, Methanol-d4) δ8.30(t,J=9.5Hz,1H),8.20(s,1H),7.57–7.34(m,4H),7.22–7.11(m,2H),6.99(s,1H),4.71(s ,2H),4.43(m,2H),3.67–3.52(m,2H),3.02(s,3H),2.94(s,4H),2.90(m,4H),2.89–2.74(m,4H),2.09(m,4H).MS(ESI):m / z 779.22[M+H] + .

[0231] Example 9: Synthesis of compound B25

[0232]

[0233] The raw materials are intermediates 7 and 8, and the rest are the same as in Example 1.

[0234] 1H NMR (400MHz, Methanol-d4) δ8.42(d,J=9.1Hz,1H),8.10(t,J=2.0Hz,1H),8.04(s,1H),7. 73(dd,J=8.0,2.2Hz,1H),7.57(dt,J=7.8,1.3Hz,1H),7.32(t,J=8.0Hz,1H),7.18(d,J=2 .9Hz,1H),7.13(dd,J=9.1,2.9Hz,1H),6.80(s,1H),4.13(t,J=5.4Hz,2H),2.90(s,3H),2 .76(t,J=7.2Hz,6H),2.65(t,J=7.3Hz,4H),2.35(s,6H),1.95–1.88(m,4H).MS(ESI):m / z 765.20[M+H] + .

[0235] Example 10: Synthesis of compound B30

[0236]

[0237] The raw materials are intermediate 11 and intermediate 8, and the rest are the same as in Example 1.

[0238] 1 H NMR (400MHz, DMSO-d6) δ11.50(s,1H),8.86(s,1H),8.69(d,J=5.0Hz,1H),8.46( d,J=8.4Hz,1H),8.21(s,1H),7.65(d,J=9.4Hz,2H),7.61–7.53(m,2H),7.43(d, J=8.3Hz,1H),7.30(t,J=7.9Hz,1H),6.89(t,J=7.6Hz,1H),6.77(s,1H),2.82–2 .71(m,7H),2.67(t,J=7.3Hz,4H),2.21(s,3H),1.93–1.83(m,4H).MS(ESI):m / z 692.15 [M+H] + .

[0239] Determination of the inhibitory activity of the compounds of this invention against JNK kinase

[0240] (1) Reagents and materials: ADP-Glo TM Kinase Assay (Promega, V4071), DMSO (Aladdin, 67-68-5), 384 white well plate (Corning, 3570).

[0241] (2) Experimental instruments: microplate reader (Tecan Group Ltd., Swiss), microplate shaker (Hangzhou Allsheng Instruments CO., Ltd., MB100-2A).

[0242] (3) Experimental methods:

[0243] Dissolve the compound in DMSO to prepare a stock solution. Dilute the compound to eight final concentrations using Reaction Buffer according to the kit instructions, ensuring the final concentration of DMSO is 2%. Add 5 μL of the reaction mixture to a 384-well plate: 1 μL of the compound, 1 μL of p38 substrate (final concentration 0.2 μg / μL), 1 μL of ATP (final concentration 5 μM), and 2 μL of JNK enzyme (10 ng or 2 ng). Set up a positive control and a negative control. For the positive control, add 1 μL of 10% DMSO and 2 μL of Reaction Buffer instead of the compound and JNK enzyme. For the negative control, add 1 μL of 10% DMSO instead of the compound. Place the 384-well plate in a microplate shaker at 25°C for 1 h (JNK enzyme amount 10 ng) or 4 h (JNK enzyme amount 2 ng). Add 5 μL of LADP-Glo ​​reagent, place in a microplate shaker, and incubate at 25°C for 40 minutes. Terminate the reaction and remove any remaining ATP. Add 10 μL of Kinase Detection Reagent and incubate at 25°C for 30 minutes to convert ADP to ATP. The ATP then reacts with the newly synthesized ATP via a luciferase / luciferin reaction to produce chemiluminescence. Luminescence is detected using a microplate reader, and the RLU reading is recorded. The formula is: Compound Inhibition Rate (%) = [(RLU)] 阴 -RLU 样 ) / (RLU 阴 -RLU 阳 )]×100%, RLU 阴 For the negative control group, RLU readings. 阳 For the positive control group, RLU readings. 样 The data represents the experimental group readings. The inhibition rate data were imported into GraphPad Prism 8.0 software for fitting, and the experimental results are shown in the table below.

[0244] (4) Experimental Results

[0245] The bioactivity of the compounds described in this invention was determined through the above experiments. All compounds showed some inhibitory activity against JNK1, as shown in the table below. The compounds designated as "A" for JNK1 inhibitory activity provided IC50 values. 50 Value is IC 50≤0.1 μM; IC50 provided by compounds with activity specified as "B" 50 The value is 0.1 μM <IC 50 ≤1.0 μM; IC50 provided by compounds with activity specified as "C" 50 Value is IC 50 >1.0μM;

[0246]

[0247] Determination of the inhibitory activity of the compounds of this invention against NLRP3

[0248] I. Experimental Reagents, Materials, and Main Instruments

[0249] 1. Reagents and Materials: Dimethyl sulfoxide (Aladdin, 67-68-5), RPMI 1640 (VM-2101BM, Vistech), DMEM medium (Vistech, VM-1101BM), Opti-MEM medium (Gibco, 31985062), fetal bovine serum (FBS, Vistech, SE100-011), penicillin-streptomycin solution (double antibiotic) (PS, Gibco, 15140-122), macrophage colony-stimulating factor (M-CSFBio-techne, 416-ML), lipopolysaccharide (LPS, MedChemExpress, HY-D1056), nigericin (MedChemExpress, HY-127019), Mouse IL-1beta / IL-1F2 Quantikine HSELISA Kit (R&D Systems, MHSLB00), Mouse IL-1beta / IL-1F2 DuoSet ELISA (R&D Systems, DY401-05), ethanol (Shanghai Test, 10009159), RIPA lysis buffer (Beyotime, P0013B), chloroform (Shanghai Test, 10006818), methanol (Shanghai Test, 10014128), NLRP3 antibody (Cell Signaling Technologys, 15101), Caspase-1 antibody (Cell Signaling Technologys, 24232), IL-1β antibody (Cell Signaling Technologys, 12426), GAPDH antibody (Cell Signaling Technologys, 2118), HRP-Goat Anti-rabbit IgG (H+L) (Bioker, BK-R050), ECL exposure solution (NCM Biotech, P2300).

[0250] 2. Experimental instruments: Microplate reader (Tecan Group Ltd., Swiss), water bath (Shanghai Boxun Medical Bio-Instrument Co., Ltd., DK-8D), high-speed centrifuge (Eppendorf AG, Centrifuge 5420), protein electrophoresis apparatus (Bio-Rad Laboratories Co., Ltd., Tetra Blotting Module), exposure apparatus (Ebiotrade, e-Blot).

[0251] 3. Experimental consumables: dissecting scissors, cell scraper, 12-well cell culture plate (Corning, 3737), 24-well cell culture plate (Corning, 3738), disposable bacterial culture dish (Bbi-life Sciences, F611004).

[0252] 4. Laboratory Animals: Male ICR mice (6-12 weeks old) were purchased from the Laboratory Animal Center of Hangzhou Normal University and housed in a temperature- and humidity-controlled environment. Husbandry and experimental use were carried out in accordance with the "Guidelines for the Husbandry, Management and Use of Laboratory Animals".

[0253] II. Experimental Methods:

[0254] 1. Isolation and Differentiation of BMDM Cells

[0255] 1) Mice leg bone harvesting: Mice were euthanized by dislocation, and their legs and backs were disinfected with sufficient 75% ethanol. The hind limb was then detached from the greater trochanter at the base of the thigh using scissors. After removing the muscle tissue, the limb was placed in cold PBS. (This step was performed in a laminar flow hood; subsequent steps were transferred to the cell culture room.)

[0256] 2) BMDM Cell Extraction and Induction: Rinse the leg bone with PBS to remove excess muscle, approximately 2-3 times. Cut the femur and tibia at both ends with scissors. Using a 1mL syringe, draw cold induction medium and blow bone marrow from the femur and tibia, repeating this process 3 times until no obvious red color is visible inside the leg bone. Use a pipette to repeatedly pipette the medium containing bone marrow cells to disperse cell clumps, transfer to a 15mL centrifuge tube, and centrifuge at 1500rpm for 5min. Discard the supernatant, add approximately 1-2mL of erythrocyte lysis buffer to resuspend the cells, and pipette to separate the cells into single cells. After incubating at room temperature for 5min, add 10mL of medium to terminate the reaction. Filter through a 0.45μm cell filter membrane and centrifuge at 1500rpm for 5min. Discard the supernatant, resuspend the cells, and count the cells. Resuspend the cells in 1640 medium (10% FBS, 2% PS) containing 20ng / mL M-CSF factor, and transfer the cell suspension to a no-treated disposable bacterial culture dish for culture.

[0257] 2. Activation of the classic NLRP3 inflammasome

[0258] Using well-differentiated BMDM cells (4-6 days old), discard the culture supernatant and wash the cells with an appropriate amount of PBS. Add 10 mL of DMEM medium and scrape the cells off using a cell scraper. Resuspend the cells in a centrifuge tube and centrifuge at 1500 rpm for 10 min. Remove the supernatant, resuspend the BMDM cells in DMEM medium, and divide them into 12-well plates, ensuring approximately 5 × 10⁶ cells per well. 5 / mL. The next day, 500 μL of Opti-MEM medium (containing 500 ng / mL LPS) was added to each well of BMDM cells for pretreatment for 4-5 hours. The compound was weighed using an analytical balance and placed in a sterile centrifuge tube, then dissolved in dimethyl sulfoxide (DMSO) to prepare the compound at the desired concentration. The compound was treated with the drug for 30 min, followed by stimulation with 10 μM Nigericin for 60 min. The cell supernatant (SN) and cell lysis buffer (Input) were collected. Based on the obtained cell supernatant (SN) and cell lysis buffer (Input), Western blotting and ELISA were performed.

[0259] 3. Protein extraction and sample preparation

[0260] 1) Extraction of protein from cell culture supernatant: Place the sample in a centrifuge and centrifuge at 13,000 rpm for 5 min to remove dead cells and impurities; transfer the supernatant to a new EP tube, add 500 μL methanol and 125 μL chloroform, vortex to mix well, and centrifuge at 13,000 rpm for 5 min; remove the upper liquid, at which point the middle layer is the protein solution, add 500 μL methanol, vortex to mix well, and centrifuge at 13,000 rpm for 5 min; the protein precipitates at the bottom of the EP tube, remove the supernatant, let stand for 10 min to evaporate the methanol completely, add 60 μL 1×SDS loading buffer, dissolve thoroughly, and denature at 100℃ for 10 min.

[0261] 2) Extraction of cell proteins: Add 120 μL of cell lysis buffer to the cells after removing the cell supernatant. After fully lysing the cells, add 5×SDS loading buffer and denature at 100℃ for 10 min.

[0262] 4. Western Blot Detection

[0263] Western blotting (WB) was performed using a 4-12% protein gel. The protein electrophoresis conditions were 70-80V for 20-30 minutes, and the separating gel conditions were 130-160V for 60-90 minutes. The PVDF membrane was pre-activated by soaking it in methanol for 5-10 minutes. The filter paper and sponge were pre-soaked in transfer buffer. After electrophoresis, the protein gel was soaked in transfer buffer for 5-10 minutes. The membrane was transferred using a sandwich structure of sponge-filter paper-gel-membrane-filter paper-sponge at 250mA for 90 minutes. The membrane was blocked with 5% skim milk for 1 hour. The primary antibody was diluted with 5% BSA according to the antibody dilution ratio, and incubated overnight at 4°C. The next day, the membrane was washed three times with 1×TBST, 5 minutes apart. The secondary antibody was diluted with 1×TBST according to the antibody dilution ratio, incubated for 1 hour, and washed three times with 1×TBST, 5 minutes apart. After washing, the membrane was exposed to ECL ultrasensitive exposure buffer.

[0264] The inhibitory effect of the compound on the NLRP3 inflammasome pathway was evaluated by measuring the amount of IL-1β secreted by BMDM cells stimulated with LPS+Nigericin after compound treatment. ImageJ was used to calculate the gray values ​​of the IL-1β band in the culture supernatant and the internal reference protein GAPDH band in the cell lysate, and their ratio was determined. The gray values ​​of the control group (DMSO treatment group) were normalized, and the IC50 was calculated using GraphPad Prism. 50 The experimental results are detailed in the table below.

[0265] III. Experimental Results

[0266] The bioactivity of the compounds described in this invention was determined through the above experiments. All compounds exhibited varying degrees of inhibition against NLRP3, as shown in the table below. The compounds designated as "A" for their NLRP3 inhibitory activity provided an IC50 value. 50 Value is IC 50 ≤0.2 μM; IC50 provided by compounds with activity specified as "B" 50 The value is 0.2 μM <IC 50 ≤1 μM; IC50 provided by compounds with activity specified as "C" 50 Value is IC 50 >1.0μM.

[0267] compound NLRP3 inhibitory activity compound NLRP3 inhibitory activity B12 A B13 B B16 C B18 C B19 A B21 B B22 B B23 B B25 B B30 A

[0268] All documents mentioned in this invention are incorporated herein by reference as if each document were individually incorporated by reference. Furthermore, it should be understood that after reading the foregoing teachings of this invention, those skilled in the art can make various alterations or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims.

Claims

1. The compound represented by Formula 1, or its tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof, In the formula, R1 is selected from: H, halogen, cyano, or optionally substituted C. 1-10 Alkyl, optionally substituted C 1-10 Alkoxy, -C(O)R 12 ; R2 is H or C with optional substitution. 1-10 Alkyl groups, or R2 and R1 together with the carbon atoms attached to them, form optional substituted 3- to 10-membered heteroaryl or heterocyclic groups containing 1, 2 or 3 heteroatoms selected from N, O or S; R3 is H or C with optional substitution. 1-10 Alkyl groups; or, R3 together with R1 or R4, along with the atoms attached to them, forms an optionally substituted 3- to 10-membered heteroaryl or heterocyclic group containing one, two, or three independent heteroatoms selected from N, O, or S. Rings A and B are independently selected from: C 5-10 Aryl, 5-10 heteroaryl, C 3-10 Cycloalkyl groups, 3-10 membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S, C 3-10 Cycloalkenyl, 3-10 membered heterocyclic alkenyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 8-10 membered bicyclic aryl or 8-10 membered bicyclic heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 8-10 membered bicyclic carbocyclic or 8-10 membered bicyclic heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S; Where valence is permissible, x can be 0, 1, 2, 3, 4 or 5, and y can be 0, 1, 2, 3, 4 or 5; R4 is selected from: H, hydroxyl group, or optionally substituted C. 1-10 Alkyl, optionally substituted C 5-10 Aryl, halogen, cyano, nitro, -C 0-6 -C(O)R7、-C 0-6 -OR8、-C 0-6 -N(R9)2, optionally substituted sulfonyl group, optionally substituted phosphonoyl group, optionally substituted 5-7 membered heteroaryl or heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S; Alternatively, two adjacent R4s together with the atoms attached to them form an optionally substituted 3-10 membered heteroaryl or heterocyclic group containing one, two or three independent heteroatoms selected from N, O or S. R5 is selected from: H, hydroxyl, halogen, cyano, nitro, -C 0-6 -C(O)R 13 -C 0-6 -OR 14 -C 0-6 -N(R 15 2. Optionally substituted sulfonyl group, Optionally substituted C 1-10 Alkyl, optionally substituted C 2-10 alkenyl, optionally substituted C 2-10 alkynyl group, optionally substituted C 1-10 Alkoxy, optional substituted C 5-10 aryl or optionally substituted C-type compounds containing one, two, or three heteroatoms independently selected from N, O, or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S; J is selected from: -C(R) 16 )2-、-C(R 17 )=C(R 18 )-、-N(R 19 )-, or J does not exist; Where valence allows, W can be selected from: =O, =NR 10 -NH2; Q1 is selected from: =N-, -NR 20 -; Q2 is selected from: -NR 21 -、-C(R 11 )2-; R6 is selected from C with optional substitution. 5-10 aryl, or optionally substituted C-type compounds containing one, two, or three heteroatoms independently selected from N, O, or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S; Each R7, R 12 R 13 R 16 R 17 and R 18 Each is independently selected from: H, optionally substituted hydroxyl group, optionally substituted amino group, optionally substituted C group. 1-10 Alkyl, optionally substituted C 1-10 Alkoxy, optional substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S; Each R8 and R 14 Each is independently selected from: H, and optionally substituted C. 1-3 Acyl group, optionally substituted C 1-10 Alkyl, optionally substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S; Each R9, R 15 and R 19 Independently selected from: H, optional substituted C 1-3 Acyl group, optionally substituted C 1-10 Alkyl, optionally substituted C 5-10 aryl or C containing 1, 2 or 3 heteroatoms independently selected from N, O or S. 5-10 heteroaryl, optionally substituted C 3-10 Cycloalkyl groups, optionally substituted 3- to 10-membered heterocyclic groups containing 1, 2, or 3 heteroatoms independently selected from N, O, or S; R 10 It is H or cyano; R 11 Selected from H, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NH-C 1-6 Alkyl, or -N-(C 1-6 Alkyl)2; Each R 20 and R 21 Independently selected from: H, optional substituted C 1-10 alkyl.

2. The compound of claim 1, or its tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof, characterized in that, R1 is selected from: H, halogen, or optionally substituted C. 1-6 alkyl; R2 is H; R3 is H; Ring A is selected from: phenyl, 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, 3-6 membered cycloalkyl or 5-6 membered heterocyclic group containing 1, 2 or 3 heteroatoms independently selected from N, O or S; Ring B is a phenyl group; x is 0, 1, 2, or 3; y is 0, 1, or 2; R4 is selected from: H, hydroxyl, halogen, cyano, nitro, or optionally substituted C. 1-6 Alkyl, optionally substituted C 1-6 Alkoxy, -C 0-6 -C(O)R7、-C 0-6 -OR8、-C 0-6 -N(R9)2; R5 is selected from: H, hydroxyl, halogen, cyano, nitro, or optionally substituted C. 1-6 Alkyl, optionally substituted C 1-6 Alkoxy, -C 0-6 -C(O)R 13 -C 0-6 -OR 14 -C 0-6 -N(R 15 )2; J is -C(R) 16 )2- or does not exist; W = O; Q1 = -NH-; Q2 = -NH-; R6 is selected from: 2,6-dialkylphenyl, 2,6-dialkyl-4-halophenyl, or Each R7, R 13 and R 16 Each is independently selected from: H, hydroxyl, optionally substituted amino, optionally substituted C. 1-6 Alkoxy; Each R8 and R 14 Each is independently selected from: H, and C with optional substitution. 1-6 alkyl; Each R9 and R 15 Independently selected from: H, optional substituted C 1-6 alkyl. R 22 Selected from: hydrogen, halogen, cyano, hydroxyl, C 1-6 Alkyl, C 1-6 Haloalkyl, C 3-5 cycloalkyl and C 1-6 alkoxy groups, all of which may be halogenated, cyanoated, or C-shaped. 1-6 Alkoxy groups may be optionally substituted.

3. The compound of claim 2, or its tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof, characterized in that, R1 is a halogen; preferably bromine.

4. The compound of claim 2, or its tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof, characterized in that, R4 is either -C(O)R7 or -OR8; R7 is selected from: H, hydroxyl group, optionally substituted amino group, optionally substituted C group. 1-6 Alkoxy; R8 is selected from: H, C which is optionally substituted by one or more groups independently selected from the following 1-6 Alkyl groups: halogen, hydroxyl, carboxyl, C 1-3 Acyloxy group, -N(R) 23 )2; Each R 23 H independently, or C with optional substitution 1-6 Alkyl; or, two Rs 23 Together with the nitrogen atom it is bonded to, it forms a 3-7 membered heterocycle with 1, 2 or 3 independently selected heteroatoms chosen from N, O or S, which may be substituted. x is 1, 2 or 3, and at least one of them R4 is -C(O)R7 and is substituted at the -NH- or adjacent position.

5. The compound of claim 2, or its tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof, characterized in that, R1 is selected from bromine; Ring A is a phenyl group; x is 1, 2, or 3; y is 0; R4 is selected from -C 0-6 -C(O)R7; J does not exist; R6 is selected as R7 is selected from C 1-6 Alkyl (preferably C) 1-3 Alkyl-substituted amino groups; R 22 Selected from: hydrogen, C 1-6 alkyl.

6. Compounds selected from the group consisting of, or their tautomers, meso compounds, racemates, enantiomers, diastereomers, mixtures thereof, or pharmaceutically acceptable salts thereof: Preferred 7. A pharmaceutical composition comprising the compound of any one of claims 1-6 or its tautomers, mesosomes, racemates, enantiomers, diastereomers, mixtures thereof or pharmaceutically acceptable salts thereof, and optionally a pharmaceutically acceptable transporter.

8. Use of the compound of any one of claims 1 to 6, or its tautomer, meso compound, racemic compound, enantiomer, diastereomer, mixture thereof, or pharmaceutically acceptable salt thereof, in the preparation of a JNK inhibitor, an NLRP3 inhibitor, or a JNK & NLRP3 dual-target inhibitor.

9. The use as described in claim 8, characterized in that, The JNK inhibitor, NLRP3 inhibitor, or JNK&NLRP3 dual-target inhibitor is a drug for the treatment and / or prevention of JNK / NLRP3-related diseases.

10. The use as described in claim 9, characterized in that, The JNK / NLRP3-related diseases mentioned include fibrosis, neurodegenerative diseases, diabetes, inflammatory diseases, tumors, central nervous system diseases, gout, etc.

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