CN114716427A - Compound serving as RIP inhibitor and preparation method and application thereof - Google Patents

Compound serving as RIP inhibitor and preparation method and application thereof Download PDF

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CN114716427A
CN114716427A CN202210015083.6A CN202210015083A CN114716427A CN 114716427 A CN114716427 A CN 114716427A CN 202210015083 A CN202210015083 A CN 202210015083A CN 114716427 A CN114716427 A CN 114716427A
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compound
salt
stereoisomer
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CN114716427B (en
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广兵
阳泰
董韧涵
谢建
彭坚
黄胜
刘进
覃传军
占伟
赖永新
彭向阳
许庆
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Chengdu Benuo Kecheng Biotechnology Co ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
    • C07D413/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings linked by a chain containing hetero atoms as chain links
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
    • Y02P20/00—Technologies relating to chemical industry
    • Y02P20/50—Improvements relating to the production of bulk chemicals
    • Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Abstract

The invention provides a compound serving as a RIP inhibitor, and a preparation method and application thereof, and belongs to the field of chemical medicines. The compound is a compound represented by the formula (I), or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof. The compound of the invention has good inhibitory activity to human RIP1 kinase, has better effect than the compound of the prior art, and can be used for treating diabetesFor preparing RIP1 inhibitor; meanwhile, the compound has a remarkably better effect on inhibiting apoptosis of pancreatic cancer cells, colon cancer cells, lymph cancer cells and immortalized keratinocytes induced by TNF than the existing compound, and can effectively inhibit apoptosis of cells. Furthermore, the compound can be used for preparing medicines for treating diseases related to RIP1 and apoptosis necrosis, such as various inflammations, psoriasis, tumors and the like, and has wide application prospect.

Description

Compound serving as RIP inhibitor and preparation method and application thereof
Technical Field
The invention belongs to the field of chemical medicine, and particularly relates to a compound serving as a RIP inhibitor, and a preparation method and application thereof.
Background
Programmed necrosis (also known as necroptosis) is a novel regulated mode of cell death characterized by necrotic morphology. Programmed necrosis plays a key role in embryonic development and the dynamic balance of the adult body, and is also widely involved in the pathophysiological processes of various diseases. Receptor-interacting protein 1 (RIP 1) is located at the key position of programmed necrosis pathway, is a kinase which is located upstream and plays a role in regulation, and is used as an upstream regulatory molecule of a signal pathway, the abnormal activation of which can cause a series of reactions, and RIP1 has become a central controller in the death receptor signal pathway for determining cell fate.
Numerous studies have shown that cell death and inflammatory responses mediated by activation of RIP1 and the programmed necrosis pathway are implicated in a variety of human diseases including stroke, myocardial infarction, retinal damage, lethal systemic inflammatory response syndrome and chronic enteritis and malignancies, among others. RIP1 kinase is a potential target in the programmed necrosis pathway, and inhibition of its kinase activity can inhibit the progression of the disease. The RIP1 inhibitor has important application value in treating inflammation, cardiovascular and cerebrovascular diseases, neurodegenerative diseases, tumor and other diseases. RIP inhibitors, and in particular RIP1 inhibitors, have received considerable attention from researchers in medicinal chemistry.
Philip a. harris et al reported a class of RIP inhibitors with the following representative structure and found their application in the treatment of inflammation or tumors. However, the therapeutic effect of RIP inhibitors still needs to be further improved, and RIP inhibitors having a better therapeutic effect are still in urgent clinical need.
Figure BDA0003459534320000011
Disclosure of Invention
The invention aims to provide a compound serving as a RIP inhibitor and a preparation method and application thereof.
The present invention provides a compound represented by formula (I), or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof:
Figure BDA0003459534320000021
wherein, the first and the second end of the pipe are connected with each other,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
x is selected from O, CH2；
R4Selected from the S configuration-CH3Or R configuration-CH3；
The A ring is selected from five-membered heterocyclic rings.
Further, the air conditioner is characterized in that,
a ring is selected from
Figure BDA0003459534320000022
Figure BDA0003459534320000023
Further, the compound is represented by formula (II):
Figure BDA0003459534320000024
wherein, the first and the second end of the pipe are connected with each other,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
R4selected from the S configuration-CH3Or R configuration-CH3；
The A ring is selected from five-membered heterocycle;
preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure BDA0003459534320000031
Figure BDA0003459534320000032
Further, the compound is represented by formula (III):
Figure BDA0003459534320000033
wherein the content of the first and second substances,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
the A ring is selected from five-membered heterocycle;
Preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure BDA0003459534320000034
Figure BDA0003459534320000035
Further, the compound is represented by formula (IV):
Figure BDA0003459534320000036
wherein the content of the first and second substances,
R1selected from hydrogen, methyl, deuterated methyl;
the A ring is selected from five-membered heterocycle;
preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure BDA0003459534320000041
Figure BDA0003459534320000042
Further, the salt is a mesylate, a hydrochloride, a sulfate, an organic acid salt.
Further, the compound is one of the following compounds:
Figure BDA0003459534320000043
Figure BDA0003459534320000051
the invention also provides the use of the compound or the salt thereof, or the stereoisomer thereof, or the solvate thereof, or the prodrug thereof in the preparation of a RIP inhibitor;
preferably, the RIP inhibitor is a RIP1 inhibitor;
more preferably, the RIP inhibitor is a drug that inhibits apoptosis.
The invention also provides the application of the compound or the salt thereof, or the stereoisomer thereof, or the solvate thereof, or the prodrug thereof in preparing a medicament for treating diseases related to programmed cell necrosis;
preferably, the medicament is an anti-inflammatory, tumor-preventing and/or treating medicament, psoriasis-preventing and/or treating medicament;
more preferably, the medicament is a medicament for preventing and/or treating pancreatic cancer, colon cancer, lung cancer, liver cancer, breast cancer, lymph cancer;
and/or, the medicament is a medicament for preventing and/or treating rheumatoid arthritis and ulcerative colitis.
The invention also provides a medicament which is prepared by taking the compound, or the salt, the stereoisomer, the solvate or the prodrug thereof as an active ingredient and adding pharmaceutically acceptable auxiliary materials or auxiliary ingredients.
As used herein, "deuterated" refers to a compound or group in which one or more hydrogens are replaced by deuterium (D). If deuterated methyl is-CH2D、-CHD2or-CD3。
In the present invention, halogen is fluorine, chlorine, bromine or iodine.
In the present invention, the room temperature is 25. + -. 5 ℃ and the overnight time is 12. + -.2 h.
The compound has good inhibitory activity to human RIP1 kinase, has better effect than the compound in the prior art, and can be used for preparing RIP1 inhibitor; meanwhile, the inhibition effect of the compound on TNF-induced apoptosis necrosis of pancreatic cancer cells, colon cancer cells, lymph cancer cells and immortalized keratinocytes is obviously superior to that of the existing compound, and the compound can effectively inhibit apoptosis necrosis of cells. Furthermore, the compound can be used for preparing medicines for treating diseases related to RIP1 and apoptosis necrosis, such as various inflammations, psoriasis, tumors and the like, and has wide application prospect.
Obviously, many modifications, substitutions, and variations are possible in light of the above teachings of the invention, without departing from the basic technical spirit of the invention, as defined by the following claims.
The present invention will be described in further detail with reference to the following examples. This should not be understood as limiting the scope of the above-described subject matter of the present invention to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
Detailed Description
The raw materials and equipment used in the embodiment of the present invention are known products and commercially available products.
Synthesis of intermediates in the A Ring
1. Synthesis of intermediate A-1
Figure BDA0003459534320000061
Step 1: cooling 20 ml of ethanol to-5 ℃, dropwise adding 10 g of acetyl chloride (0.13mol), and reacting at low temperature (0-5 ℃) for half an hour to obtain the freshly prepared ethanolic acid gas. Adding a mixed solution of 12.5 g of ethyl cyanoformate (0.13mol) and 30 ml of dichloromethane dropwise at low temperature (0-5 ℃), stirring at 0-5 ℃ for reaction overnight, filtering a solid generated by the reaction to obtain imine intermediate hydrochloride, suspending the hydrochloride in 50 ml of methyl tert-butyl ether, adding 20 g of triethylamine, stirring at room temperature for 5 hours, filtering to remove triethylamine hydrochloride, and concentrating the methyl tert-butyl ether under reduced pressure to obtain 12.3 g of intermediate AM 1-1.
And 2, step: 9.6 g of intermediate AM1-1(66.1mmol) prepared in step 1 was dissolved in a mixed solution of 40 ml of ethanol and 120 ml of methyl tert-butyl ether, 10 g of phenylacethydrazide (66.6mmol) was added, the mixture was stirred at room temperature overnight, filtered, the solid was washed twice with ethanol and dried to obtain 8.7 g of intermediate AM1-2, which was used in the next reaction.
And 3, step 3: 5 g of intermediate AM1-2(20.0mmol) were suspended in 100 ml of xylene, reacted at 170 ℃ under reflux for 24 hours, cooled to room temperature and filtered to obtain 4.3 g of intermediate AM 1-3.
And 4, step 4: suspending 4 g of intermediate AM1-3(17.3mmol) in 40 ml of water, adding 1.2 g of lithium hydroxide (50.1mmol), stirring at room temperature for 3 hours, adjusting the pH value to 2 with 2M dilute hydrochloric acid, separating out a solid, filtering, washing the solid twice with water, and drying to obtain 3.3 g of intermediate A-1.
2. Synthesis of intermediate A-2
Figure BDA0003459534320000071
Step 1: 5 g of commercially available methyl 1, 2, 4-triazole-3-carboxylate (39.3mmol) were dissolved in 100 ml of acetone, 8.1 g of potassium carbonate (58.6mmol) and 6.7 g of benzyl bromide (39.1mmol) were added, the mixture was stirred at room temperature overnight, and the starting material was detected by TLC to be completely reacted. 100 ml of water was added, extraction was performed 3 times with 150 ml of ethyl acetate, and organic layers were combined, washed with saturated brine 1 time and dried over 50 g of anhydrous sodium sulfate. The sodium sulfate is removed by filtration, the mixture is concentrated to dryness under reduced pressure, and 50 ml of petroleum ether is added for crystallization to obtain a wet product of 6.6 g. The 6.6 g wet product was recrystallized from 30 ml ethyl acetate petroleum ether mixed solution (EA: PE ═ 1:1), filtered and dried to give 4.1 g total of intermediate AM 2-1.
And 2, step: dissolving 2 g of intermediate AM2-1(9.2mmol) in 20 ml of methanol, dissolving 0.7 g of lithium hydroxide (29.2mmol) in 20 ml of water, adding into a methanol system, stirring at room temperature for 5 hours, detecting by TLC that the raw material is completely reacted, adjusting the pH value to 2-3 by using 2M dilute hydrochloric acid, separating out a solid, filtering, washing the solid to be neutral by using water, drying to obtain 1.7 g of intermediate A-2,
3. Synthesis of intermediate A-10
Figure BDA0003459534320000072
1-H-imidazole-4-methyl formate is used as a raw material, and the synthesis method is similar to that of the intermediate A-2 to obtain an intermediate A-10.
The remaining intermediates A-3, A-4, A-5, A-6, A-7, A-8, A-9 are all commercially available and have the following structures:
Figure BDA0003459534320000073
Figure BDA0003459534320000081
example 1 Synthesis of Compound I-1
Figure BDA0003459534320000082
The synthetic route is as follows:
Figure BDA0003459534320000083
i：NaH/DMF；ii：H2，Pd/C，MeOH；iii：TBTU，DIPEA，DMSO；iv：MeI，Cs2CO3，DMF；v：HCl，MTBE；vi：TBTU，DIPEA，DCM
the specific operation is as follows:
1. synthesis of intermediate M1-1:
dissolving 5 g of BOC-L-threonine (cas:2592-18-9,22.8mmol) in 50 ml of DMF, cooling the ice salt bath to 0 ℃, suspending 1.4 g of NaH (57.0mmol) in 10 ml of DMF, dropwise adding the solution, removing the ice salt bath, stirring at room temperature until bubbles disappear, dropwise adding 3.2 g of 1-fluoro-2-nitrobenzene (22.7mmol), stirring at room temperature for 4 hours, adding ice water, and quenching the reaction. Adding 2M dilute hydrochloric acid into the reaction system to the pH value of 2-3, extracting with 50 ml ethyl acetate for 3 times respectively, combining organic layers, washing and drying the organic layers, concentrating under reduced pressure, and purifying residues by column chromatography to obtain 6.1 g yellow semisolid, namely an intermediate M1-1 (molecular weight 340.13).
2. Synthesis of intermediate M2-1:
dissolving the intermediate M1-1 in 60 ml of methanol, carrying out catalytic hydrogenation reaction at room temperature by using palladium carbon as a catalyst, filtering to remove the palladium carbon after complete reaction, concentrating the filtrate under reduced pressure, stirring and crystallizing the residue with methyl tert-butyl ether, dissolving the solid obtained by drying in 25 ml of DMSO, adding 5.7 g of TBTU (17.9mmol) and 2.3 g of DIPEA (17.9mmol), stirring overnight at room temperature, adding 25 ml of ice water for quenching, extracting for 3 times with 30 ml of ethyl acetate respectively, combining organic layers, washing the organic layers with water, drying, concentrating under reduced pressure, crystallizing the residue with petroleum ether, filtering and drying to obtain 4.7 g of white solid, namely the intermediate M2-1 (molecular weight 292.14).
3. Synthesis of intermediate M3-1:
2.0 g of intermediate M2-1(6.8mmol) above was dissolved in 50 mL of DMF, and 3.3 g of cesium carbonate (10.2mmol), 1.4 g of methyl iodide (10.2mmol) were added and stirred at room temperature until the reaction was complete. After adding 50 ml of ice water for quenching, extracting with 80 ml of ethyl acetate for three times respectively, combining organic layers, washing and drying the organic layers with water, concentrating under reduced pressure, crystallizing with petroleum ether, and filtering to obtain a solid. 20 ml of methyl tert-butyl ether saturated with hydrochloric acid gas was added to the above solid, and after stirring at room temperature for 6 hours, the solvent was concentrated under reduced pressure, and the residue was crystallized from petroleum ether to give 1.7 g of a pale yellow solid, i.e., intermediate M3-1 (molecular weight 242.70).
4. Synthesis of target I-1:
after 1.7 g of intermediate M3-1(7.0mmol), 4.5 g of TBTU (14.0mmol) and 1.36 g of DIPEA (10.5mmol) were dissolved in 20 ml of DCM, 1.7 g of A-1 ring (5-benzyl-4H-1, 2, 4-triazole-3-carboxylic acid, 8.4mmol) was added, the mixture was stirred overnight at room temperature, ice water was added to quench the reaction, the organic layer was washed with water, dried and concentrated under reduced pressure, and the residue was isolated by column chromatography to give 1.5 g of compound I-1.
1HNMR(400MHz，DMSO-d6)δ：14.41(s,1H),7.90(s,1H),7.52-7.43(m,1H),7.38-7.21((m,8H),4.90-4.79(m,2H),4.16(s,2H),3.36(s,3H),1.28(d,J＝5.1Hz,3H).
ESI-MS m/z:390.11[M-1]-
Example 2: synthesis of Compound I-2:
Figure BDA0003459534320000091
the synthetic route is as follows:
Figure BDA0003459534320000092
iv：CD3I，Cs2CO3，DMF；v：HCl，MTBE；vi：TBTU，DIPEA，DCM
the specific operation is as follows:
1. synthesis of M3-2
M2-1 was used as a starting material, and the synthesis method was similar to the preparation of intermediate M3-1 in example 1, using deuterated iodomethane instead of iodomethane to obtain intermediate M3-2.
2. Synthesis of I-2
The same procedure for preparation of I-1 as in example 1 was repeated to give the objective compound I-2.
ESI-MS m/z:393.13[M-1]-
Example 3: synthesis of Compound I-3:
Figure BDA0003459534320000101
the synthetic route is as follows:
Figure BDA0003459534320000102
v：HCl，MTBE；vi：TBTU，DIPEA，DCM
the specific operation is as follows:
m2-1 was used as a starting material, and methyl t-butyl ether saturated with hydrochloric acid gas was added thereto, and after stirring at room temperature for 6 hours, the solvent was concentrated under reduced pressure, and the residue was crystallized from petroleum ether to give intermediate M3-3 (molecular weight 228.68).
The intermediate M3-3 and the intermediate A-1 are used as raw materials, and the synthesis method is similar to the step 4 in the example 1, so that the compound I-3 is obtained.
ESI-MS m/z:376.15[M-1]-
Example 4: synthesis of Compound I-4:
Figure BDA0003459534320000103
the synthetic route is as follows:
Figure BDA0003459534320000111
i：NaH/DMF；ii：H2，Pd/C，MeOH；iii：TBTU，DIPEA，DMSO；iv：MeI，Cs2CO3，DMF；v：HCl，MTBE；vi：TBTU，DIPEA，DCM
the specific operation is as follows:
the synthesis method was similar to example 1 using BOC-D-threonine as the starting material to give compound I-4.
1HNMR(400MHz,CDCl3)δ：8.22(d,J＝8.9Hz,1H),7.33-7.19((m,7H),7.17-7.07(m,1H),4.84-4.74(m,1H),4.70-4.58(m,1H),4.26-4.13(m,2H),2.83(s,3H),1.38(d,J＝6.2Hz,3H).
ESI-MS m/z:390.08[M-1]-
Example 5: synthesis of Compound I-5:
Figure BDA0003459534320000112
the synthetic route is as follows:
Figure BDA0003459534320000113
iv：CD3I，Cs2CO3，DMF；v：HCl，MTBE；vi：TBTU，DIPEA，DCM
the specific operation is as follows:
compound I-5 was obtained in a manner analogous to example 2 starting from intermediate M4-2.
ESI-MS m/z:393.12[M-1]-
Example 6: synthesis of Compound I-6:
Figure BDA0003459534320000121
the compound I-6 is prepared by a similar method of the compound I-1 and by adopting an A-2 ring as a raw material.
1H NMR(400MHz,DMSO-d6)δ：8.85(s,1H),7.95(d,J＝5.8Hz,1H),7.50-7.43(m,1H),7.42-7.23(m,8H),5.55-5.40(m,2H),4.89-4.77(m,2H),3.35(s,3H),1.30-1.21(m,3H).
ESI-MS m/z:390.08[M-1]-
Example 7: synthesis of Compound I-7:
Figure BDA0003459534320000122
the compound I-7 is prepared by adopting an A-2 ring as a raw material and deuterated iodomethane as a raw material in a similar way to the compound I-2.
1H NMR(400MHz,DMSO-d6)δ:8.8(s,1H),8.00-7.91(m,1H),7.50-7.43(m,1H),7.43-7.22(m,8H),5.52-5.46(m,2H),4.89-4.80(m,2H),1.33-1.23(m,3H).
ESI-MS m/z:393.12[M-1]-
Example 8: synthesis of Compound I-8:
Figure BDA0003459534320000123
by analogy with the method for preparing the compound I-3, the compound I-8 is prepared by taking the A-2 ring as a raw material.
ESI-MS m/z:376.15[M-1]-
Example 9: synthesis of Compound I-9:
Figure BDA0003459534320000131
compound I-9 is prepared by a method similar to compound I-1, using the A-3 ring as the starting material.
ESI-MS m/z:390.07[M-1]-
Example 10: synthesis of Compound I-10:
Figure BDA0003459534320000132
the compound I-10 is prepared by a similar method to the compound I-1 and by taking the A-4 ring as a raw material.
ESI-MS m/z:390.07[M-1]-
Example 11: synthesis of Compound I-11:
Figure BDA0003459534320000133
the compound I-11 is prepared by a similar method to the compound I-1 and by taking the A-5 ring as a raw material.
ESI-MS m/z:389.03[M-1]-
Example 12: synthesis of Compound I-12:
Figure BDA0003459534320000134
the compound I-12 is prepared by a similar method to the compound I-1 and by taking the A-6 ring as a raw material.
ESI-MS m/z:391.09[M-1]-
Example 13: synthesis of Compound I-13:
Figure BDA0003459534320000141
compound I-13 is prepared by a method similar to that of compound I-1, using the A-7 ring as the starting material.
ESI-MS m/z:391.09[M-1]-
Example 14: synthesis of Compound I-14:
Figure BDA0003459534320000142
the compound I-14 is prepared by a similar method to the compound I-1 and by using the A-8 ring as a raw material.
ESI-MS m/z:390.08[M-1]-
Example 15: synthesis of Compound I-15:
Figure BDA0003459534320000143
the compound I-15 is prepared by a similar method to the compound I-1 and by taking the A-9 ring as a raw material.
ESI-MS m/z:391.12[M-1]-
Example 16: synthesis of Compound I-16:
Figure BDA0003459534320000144
the compound I-16 is prepared by a similar method to the compound I-1 and by using the A-10 ring as a raw material.
ESI-MS m/z:389.03[M-1]-
Example 17: synthesis of Compounds I-17:
Figure BDA0003459534320000151
the compound I-17 is prepared by a similar method of the compound I-2 by adopting an A-10 ring and deuterated iodomethane as raw materials.
ESI-MS m/z:392.15[M-1]-
Example 18: synthesis of Compound I-18:
Figure BDA0003459534320000152
compound I-18 is prepared analogously to compound I-3, using the A-10 ring.
ESI-MS m/z:375.11[M-1]-
Example 19: synthesis of Compounds I-19:
Figure BDA0003459534320000153
the compound I-19 is prepared by a method similar to the compound I-1 by using 2, 4-difluoronitrobenzene as a raw material.
ESI-MS m/z:408.12[M-1]-
Example 20: synthesis of Compound I-20:
Figure BDA0003459534320000154
the compound I-20 is prepared by a method similar to the compound I-3 by adopting 2, 4-difluoronitrobenzene as a raw material.
ESI-MS m/z:394.06[M-1]-
Example 21: synthesis of Compound I-21:
Figure BDA0003459534320000161
the compound I-21 is prepared by a similar method to the compound I-1 by adopting 2, 6-difluoronitrobenzene as a raw material.
ESI-MS m/z:408.12[M-1]-
Example 22: synthesis of Compound I-22:
Figure BDA0003459534320000162
the compound I-22 is prepared by a method similar to the compound I-3 by adopting 2, 6-difluoronitrobenzene as a raw material.
ESI-MS m/z:394.06[M-1]-
Example 23: synthesis of Compound I-23:
Figure BDA0003459534320000163
the compound I-23 is prepared by a method similar to the compound I-2 by adopting 2, 6-difluoronitrobenzene as a raw material.
ESI-MS m/z:411.13[M-1]-
Example 24: synthesis of Compound I-24:
Figure BDA0003459534320000164
the compound I-24 is prepared by a method similar to the compound I-1 by adopting 2,4, 6-trifluoronitrobenzene as a raw material.
ESI-MS m/z:412.09[M-1]-
Example 25: synthesis of Compound I-25:
Figure BDA0003459534320000171
the compound I-25 is prepared by a method similar to the compound I-8 by adopting 2,4, 6-trifluoronitrobenzene as a raw material.
ESI-MS m/z:412.09[M-1]-
Example 26: synthesis of Compounds I-26:
Figure BDA0003459534320000172
the compound I-26 is prepared by a method similar to the compound I-8 by adopting 2, 6-difluoronitrobenzene as a raw material.
ESI-MS m/z:394.06[M-1]-
Example 27: synthesis of Compound I-27:
Figure BDA0003459534320000173
the compound I-27 is prepared by a method similar to the compound I-11 by using 2,4, 6-trifluoronitrobenzene as a raw material.
ESI-MS m/z:411.09[M-1]-
The advantageous effects of the present invention are demonstrated by specific test examples below.
Test example 1 inhibitory Activity of the Compound of the present invention against RIP1 kinase
In assay buffer (50mM Hepes pH 7.5, 50mM NaCl, 30mM MgCl)21mM DTT, 0.02% CHAPS, 0.5mg/mL BSA) and a compound of the invention and a control compound were dissolved in 22-spot titration as described in 1: 1.5 serial dilutions (high concentration 2. mu.M) were added to 384 well plates. mu.L of each concentration of inhibitor and 3.5. mu.L of human RIP enzyme (25nM) dissolved in assay buffer were added to the plate. After pre-warming at 37 ℃ for 1 hour, 3.5 μ L of ATP (15 μ M to 1.5mM) in buffer was added to the plate to start the reaction. The reaction was carried out at room temperature for 5 hours. After completion of the reaction, 5 μ L of ADP-Glo reagent containing 0.02% CHAPS was added to each well and incubated at room temperature for 1 hour to terminate the kinase reaction and deplete all remaining ATP. Then 5 μ L of ADP-Glo assay solution containing 0.02% CHAPS was added to each well and incubated at room temperature for 30 minutes to measure the luminosity. For each ATP concentration, the luminescence data was subtracted from the control and expressed as activity data.
Figure BDA0003459534320000181
Determination of IC at Each ATP concentration by calculation50The results are given in table 1 below:
TABLE 1 inhibitory Activity IC of the Compounds of the invention against human RIP1 kinase50
Compound (I) IC50(nM)
I-1 10.19
I-2 8.68
I-4 8.97
I-6 7.36
I-7 8.18
I-16 14.40
Control Compound 1 20.35
Control Compound 2 18.29
Control Compound 3 22.64
The test result shows that: IC of the inventive compounds for inhibition of human RIP1 kinase50The value is obviously lower than that of the existing compound, which shows that the compound can be used for preparing RIP inhibitor and has better effect than the existing compound.
Test example 2 inhibitory Activity of the Compound of the present invention against TNF-. alpha.induced U937 programmed necrosis
Human histiocyte lymphoma cells in logarithmic growth phase U937 (purchased from ATCC) were collected, the cells were resuspended in 1640 medium, the cell concentration was adjusted to 60 cells/μ L to obtain a cell suspension, and 384 well plates were seeded with 20 μ L of the cell suspension per well plate. Cells were incubated at 37 ℃ with 5% CO2After 24 hours incubation in an incubator, test and control compounds of the invention were diluted with medium to the respective effect concentrations set and added to 384 well plates at 5. mu.l/well. The final concentration of the compound was from 0nM to 1000nM, diluted in 10-fold gradient. AT the same time, programmed necrosis inducer was added AT 5. mu.l/well into 384 wells, and the necrosis inducer components were TNF-. alpha. (100ng/ml), Smac Mimetics (AT-406, 1. mu.M), and z-VAD-FMK (20. mu.M). After addition of the test compound and necrosis inducer, the cells were incubated at 37 ℃ with 5% CO 2Incubate in incubator for 24 hours. Then 30. mu.l of CellTiter-Glo luminescent solution was added to each well, shaken for 2 minutes, allowed to stand at room temperature for 10 minutes, and then absorbance was measured with a microplate reader. The necrosis induction rate and the necrosis inhibition rate are calculated by the following formulas:
necrosis induction rate (%) ═ 100 × (1-B/a); a is a necrosis-free inducer group, and B is a necrosis-inducing agent group
Figure BDA0003459534320000191
A is a necrosis-free inducer group, B is a necrosis-inducing agent group, and X is a compound and necrosis-inducing agent group.
Calculation of inhibition IC of test Compounds on induced apoptosis by GraphPad50The results are given in table 2 below (3 per set of replicates in the experiment):
TABLE 2 inhibitory Activity of the Compounds of the invention IC on TNF-alpha induced U937 programmed necrosis50
Compound (I) IC50(nM)
I-1 2.11
I-2 0.61
I-4 0.55
I-6 0.41
I-7 0.45
I-16 4.87
Control Compound 1 7.81
Control Compound 2 10.36
Control Compound 3 6.92
The test result shows that: IC for inhibiting TNF-alpha induced apoptosis of human histiocytic lymphoma cells by using compound of the invention50The values are significantly lower than for the existing compounds. The compound of the invention can effectively inhibit programmed necrosis of human histiocyte lymphoma cells.
Test example 3 inhibitory Activity of the Compound of the present invention against TNF-. alpha.induced apoptosis in human Colon cancer cells HT-29 and human pancreatic cancer cells PANC-1
Human colon cancer cells HT-29 and human pancreatic cancer cells PANC-1 (purchased from ATCC) in the logarithmic growth phase were collected, the cells were resuspended in 1640 medium, the cell concentration was adjusted to 20 cells/μ L to obtain a cell suspension, and a 384-well plate was seeded with 20 μ L of the cell suspension per well plate. Cells were incubated at 37 ℃ with 5% CO2After 24 hours incubation in an incubator, test and control compounds of the invention were diluted with medium to the respective effect concentrations set and added to 384 well plates at 5. mu.l/well. The final concentration of the compound acting was from 0nM to 1000nM, diluted in 10-fold gradient. AT the same time, programmed necrosis inducer was added AT 5. mu.l/well into 384 wells, and the necrosis inducer components were TNF-. alpha. (100ng/ml), Smac Mimetics (AT-406, 1. mu.M), and z-VAD-FMK (20. mu.M). After addition of the test compound and necrosis inducer, the cells were incubated at 37 ℃ with 5% CO2Incubate in incubator for 24 hours. Then 30. mu.l of CellTiter-Glo luminescent solution was added to each well, shaken for 2 minutes, allowed to stand at room temperature for 10 minutes, and then absorbance was measured with a microplate reader. The necrosis induction rate and the necrosis inhibition rate are calculated by the following formulas:
necrosis induction rate (%) ═ 100 × (1-B/a); a is a necrosis-free inducer group, and B is a necrosis-inducing agent group
Figure BDA0003459534320000201
A is a necrosis-free inducer group, B is a necrosis-inducing agent group, and X is a compound and necrosis-inducing agent group.
By GraphPad meterCalculation of inhibitory Activity of test Compounds on induced apoptosis IC50The results are given in table 3 below (3 per set of replicates in the experiment):
TABLE 3 inhibitory Activity of the Compounds of the present invention on TNF-alpha induced apoptosis
Compound (I) HT-29IC50(nM) PANC-1IC50(nM)
I-1 44.56 6.01
I-2 37.46 4.77
I-4 43.19 4.98
I-6 23.37 3.07
I-7 24.99 3.32
I-16 89.04 8.95
Control Compound 1 >100 44.10
Control Compound 2 >100 52.87
Control Compound 3 >100 55.12
The experimental results show that: IC of compound of the invention for TNF-induced programmed necrosis of human colon cancer cell HT-29 and human pancreatic cancer cell PANC-150The values are significantly lower than for the existing compounds. The compound can effectively inhibit programmed necrosis of human colon cancer cells and human pancreatic cancer cells.
Test example 4 inhibitory Activity of Compound of the present invention against TNF-. alpha.induced apoptosis of HacaT cells
HacaT cells (purchased from ATCC) in the logarithmic growth phase were collected, and the cells were resuspended in 1640 medium at an adjusted cell concentration of 20 cells/. mu.l to give a cell suspension, which was seeded into 384-well plates, and 20. mu.L of the cell suspension was added to each plate. Cells were incubated at 37 ℃ with 5% CO2After 24 hours incubation in an incubator, test and control compounds of the invention were diluted with medium to the respective effect concentrations set and added to 384 well plates at 5. mu.l/well. The final concentration of the compound was from 0nM to 1000nM, diluted in 10-fold gradient. AT the same time, programmed necrosis inducer was added AT 5. mu.l/well into 384 wells, and the necrosis inducer components were TNF-. alpha. (100ng/ml), Smac Mimetics (AT-406, 1. mu.M), and z-VAD-FMK (20. mu.M). After addition of the test compound and necrosis inducer, the cells were incubated at 37 ℃ with 5% CO 2Incubate in incubator for 24 hours. Then, 30. mu.l of CellTiter-Glo luminescent solution was added to each well, shaken for 2 minutes, and left to stand at room temperatureAnd detecting absorbance by a microplate reader after 10 min. The necrosis induction rate and the necrosis inhibition rate are calculated by the following formulas:
necrosis induction rate (%) ═ 100 × (1-B/a); a is a necrosis-free inducer group, and B is a necrosis-inducing agent group
Figure BDA0003459534320000211
A is a necrosis-free inducer group, B is a necrosis-inducing agent group, and X is a compound and necrosis-inducing agent group.
Calculation of inhibitory Activity IC of test Compounds on induced apoptosis by GraphPad50The results are given in table 4 below (3 per set of replicates in the experiment):
TABLE 4 inhibitory Activity of Compounds of the invention IC on TNF-alpha induced apoptosis of HacaT cells50
Compound (I) IC50(nM)
I-1 6.11
I-2 4.57
I-4 4.62
I-6 3.18
I-7 3.55
I-16 8.84
Control Compound 1 10.4
Control Compound 2 16.8
Control Compound 3 9.9
The test result shows that: IC for inhibition of TNF-alpha induced apoptosis in HacaT cells by compounds of the invention50The value is obviously lower than that of the existing compound, which shows that the compound can effectively inhibit the programmed necrosis of HacaT cells, and further can be used for preventing and treating psoriasis.
In conclusion, the compound has good inhibitory activity on human RIP1 kinase, has better effect than the compound in the prior art, and can be used for preparing RIP1 inhibitors; meanwhile, the inhibition effect of the compound on TNF-induced apoptosis necrosis of pancreatic cancer cells, colon cancer cells, lymph cancer cells and immortalized keratinocytes is obviously superior to that of the existing compound, and the compound can effectively inhibit apoptosis necrosis of cells. Furthermore, the compound can be used for preparing medicines for treating diseases related to RIP1 and apoptosis necrosis, such as various inflammations, psoriasis, tumors and the like, and has wide application prospect.

Claims (10)

1. A compound represented by the formula (I), or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof:
Figure FDA0003459534310000011
wherein, the first and the second end of the pipe are connected with each other,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
x is selected from O, CH2；
R4Selected from the S configuration-CH3Or R configuration-CH3；
The A ring is selected from five-membered heterocyclic rings.
2. The compound according to claim 1, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, wherein:
a ring is selected from
Figure FDA0003459534310000012
Figure FDA0003459534310000013
3. The compound according to claim 1, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, wherein: the compound is represented by the formula (II):
Figure FDA0003459534310000014
wherein the content of the first and second substances,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
R4selected from the S configuration-CH3Or R configuration-CH3；
The A ring is selected from five-membered heterocycle;
preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure FDA0003459534310000021
Figure FDA0003459534310000022
4. The compound according to claim 3, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, wherein: the compound is represented by the formula (III):
Figure FDA0003459534310000023
wherein the content of the first and second substances,
R1selected from hydrogen, methyl, deuterated methyl;
R2、R3each independently selected from hydrogen, halogen;
the A ring is selected from five-membered heterocycle;
preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure FDA0003459534310000024
Figure FDA0003459534310000025
5. The compound according to claim 4, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, characterized in that: the compound is represented by a formula (IV):
Figure FDA0003459534310000026
Figure FDA0003459534310000031
wherein, the first and the second end of the pipe are connected with each other,
R1selected from hydrogen, methyl, deuterated methyl;
the A ring is selected from five-membered heterocycle;
preferably, the first and second electrodes are formed of a metal,
a ring is selected from
Figure FDA0003459534310000032
Figure FDA0003459534310000033
6. The compound according to any one of claims 1 to 5, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, wherein: the salt is mesylate, hydrochloride, sulfate, and organic acid salt.
7. The compound according to any one of claims 1 to 5, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, wherein: the compound is one of the following compounds:
Figure FDA0003459534310000034
Figure FDA0003459534310000041
8. use of a compound according to any one of claims 1 to 7, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, in the preparation of a RIP inhibitor;
preferably, the RIP inhibitor is a RIP1 inhibitor;
more preferably, the RIP inhibitor is a drug that inhibits apoptosis.
9. Use of a compound of any one of claims 1 to 7, or a salt thereof, or a stereoisomer thereof, or a solvate thereof, or a prodrug thereof, in the manufacture of a medicament for the treatment of a disease associated with apoptosis;
Preferably, the medicament is an anti-inflammatory, tumor-preventing and/or treating medicament, psoriasis-preventing and/or treating medicament;
more preferably, the medicament is a medicament for preventing and/or treating pancreatic cancer, colon cancer, lung cancer, liver cancer, breast cancer, lymph cancer;
and/or, the medicament is a medicament for preventing and/or treating rheumatoid arthritis and ulcerative colitis.
10. A medicament, characterized by: the compound is prepared by taking the compound or the salt thereof, the stereoisomer thereof, the solvate thereof or the prodrug thereof as an active ingredient and adding pharmaceutically acceptable auxiliary materials or auxiliary ingredients.
CN202210015083.6A 2021-01-07 2022-01-07 Compound serving as RIP inhibitor and preparation method and application thereof Active CN114716427B (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105121432A (en) * 2013-02-15 2015-12-02 葛兰素史密斯克莱知识产权发展有限公司 Heterocyclic amides as kinase inhibitors
CN109071504A (en) * 2016-02-05 2018-12-21 戴纳立制药公司 The inhibitor of receptor interacting protein kinases 1

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105121432A (en) * 2013-02-15 2015-12-02 葛兰素史密斯克莱知识产权发展有限公司 Heterocyclic amides as kinase inhibitors
CN109071504A (en) * 2016-02-05 2018-12-21 戴纳立制药公司 The inhibitor of receptor interacting protein kinases 1

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