CN117430586A - FGFR kinase inhibitor and pharmaceutical application thereof - Google Patents

FGFR kinase inhibitor and pharmaceutical application thereof Download PDF

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CN117430586A
CN117430586A CN202311138918.8A CN202311138918A CN117430586A CN 117430586 A CN117430586 A CN 117430586A CN 202311138918 A CN202311138918 A CN 202311138918A CN 117430586 A CN117430586 A CN 117430586A
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compound
alkyl group
fgfr4
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余洛汀
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Sichuan University
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Abstract

The invention provides an FGFR kinase inhibitor and pharmaceutical application thereof, belonging to the field of drug development. The FGFR kinase inhibitor is a compound shown in a formula I. The compound not only has excellent inhibition effect on FGFR4 kinase, can be used as an FGFR4 kinase inhibitor, but also can effectively inhibit the proliferation of Ba/F3-ETV6-FGFR4-V550L cells and overcome FGFR4 V550L Mutations, which can be used as FGFR4 V550L Mutant inhibitors. Preparation of the compounds of the invention for preventing and/or treating diseases related to FGFR4 kinase activity and FGFR4 V550L The medicine for treating mutation related diseases has wide application prospect.

Description

FGFR kinase inhibitor and pharmaceutical application thereof
Technical Field
The invention belongs to the field of drug development, and in particular relates to an FGFR kinase inhibitor and pharmaceutical application thereof.
Background
Tumor molecular targeted therapy specifically kills tumor cells by using specific structural molecules of tumor tissues or cells as targets, and using drugs capable of specifically binding to these target molecules. Tumor targeted drugs represented by kinase inhibitors have been revolutionized in the treatment of malignant tumors for nearly 10 years due to their rapid efficacy and slight toxic and side effects.
Fibroblast growth factor (Fibroblast growth factor, FGF) is a mitotic signaling molecule comprising more than 20 proteins that plays an important role in controlling cell proliferation, differentiation and survival in many tissues. FGF effects are highly dependent on their affinity for specific receptors, fibroblast growth factor receptors (fibroblast growth factor receptor, FGFR) are the primary receptors for FGF, essentially receptor-type protein Tyrosine Kinases (TKs), and currently known FGFRs mainly include 4 types, namely FGFR1, FGFR2, FGFR3 and FGFR4. Studies have shown that tumor control can be achieved by targeted inhibition of aberrant expression of FGFR. Over-expression of FGFR1 has been reported to cause diseases such as breast cancer and squamous non-small cell lung cancer; FGFR2 mutations can lead to gastric cancer and endometrial cancer, among other conditions; FGFR3 abnormal expression is associated with urothelial cancer and the like; overexpression of FGFR4 can increase the incidence of gastric cancer, liver cancer, prostate cancer, rhabdomyosarcoma, ovarian cancer, colon cancer, cutaneous melanoma, and the like.
Fibroblast growth factor receptor 4 (FGFR 4) is a class of transmembrane tyrosine kinase receptors that are later discovered in the FGFR family. The genetic organization of the FGFR4 gene comprises 18 exons, the C-terminus of half of the igiii domain of which does not vary between the three alternative forms as FGFR 1-3. At present, more research results are achieved for FGFR1-3 inhibitors, but few research reports are reported for specific FGFR4 inhibitors. Many selective FGFR inhibitors have recently begun to be developed clinically, such as AP24534, BGJ398, AZD4547, LY2874455 and JNJ-42756493. These compounds have very high in vitro kinase activity against FGFR1, FGFR2 and FGFR3, but have less than ideal inhibition against FGFR4. For example, N- [5- [2- (3, 5-dimethoxyphenyl) ethyl ] -2H-pyrazol-3-yl ] -4- (3, 5-diethylpiperazin-1-yl) benzamide (AZD 4547) designed by Paul R.Gavined et al has certain therapeutic effects on breast and bone marrow cancers. In cells, AZD4547 effectively inhibited autophosphorylation of FGFR1, 2 and 3 tyrosine kinases by FRS2, plcγ and MAPK, but inhibited FGFR4 cell kinase activity less. The development of a drug having excellent inhibitory activity on FGFR4 is of great importance.
In addition, the first generation FGFR4 inhibitors or some multi-target inhibitors (such as sorafenib and the like) usually generate obvious secondary drug resistance (such as V550L residue misplacement mutation) after 9-12 months of clinical administration, and greatly limit the survival time extension of tumor patients (GAO L, WANG X, TANG Y, et al, FGF19/FGFR4signaling contributes to the resistance of hepatocellular carcinoma tosorafenib [ J)]J ExpClin Cancer Res,2017,36 (1): 8.). But to date can be used to overcome FGFR4 V550L Mutant small molecule inhibitors have also been rarely reported.
Disclosure of Invention
The invention aims to provide an FGFR kinase inhibitor and pharmaceutical application thereof.
The invention provides a compound shown in a formula I or a salt, a stereoisomer and a deuterated compound thereof:
wherein the A ring is 5-6 membered saturated cycloalkyl, 5-6 membered saturated heterocyclic group, 5-6 membered aryl or 5-6 membered heteroaryl;
m is an integer of 0 to 5;
R 1 each independently selected fromUnsubstituted or substituted by R 5 The substituted following groups: c (C) 1～6 Alkyl, C 1～6 An alkoxy group; r is R 5 Is hydroxy, amino, halogen, carboxyl, cyano, C 1～6 Alkoxy or C 1～6 An alkyl group; r is R 6 Is C 1～6 An alkyl group;
R 2 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
R 3 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
R 4 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
is->Or-.
Further, the A ring is 5-6 membered saturated cycloalkyl or 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 3;
R 1 each independently selected fromUnsubstituted or substituted by R 5 The substituted following groups: c (C) 1～4 Alkyl, C 1～4 An alkoxy group; r is R 5 Is hydroxy, amino, halogen, carboxyl, cyano, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group;
R 2 is halogen;
R 3 is halogen;
R 4 is C 1～4 An alkyl group.
Further, the structure of the compound is shown as a formula II:
wherein, the A ring is 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 2;
R 1 each independently selected fromUnsubstituted or substituted by R 5 Substituted C 1～4 An alkyl group; r is R 5 Is hydroxy, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group.
Further, the structure of the compound is shown as a formula III-1, a formula III-2 or a formula III-3:
wherein R is 1 Selected from the group consisting ofUnsubstituted or substituted by R 5 Substituted C 1～2 An alkyl group; r is R 5 Is hydroxy, C 1～2 Alkoxy or C 1～2 An alkyl group; r is R 6 Is C 1～2 An alkyl group.
Further, the structure of the compound is shown as a formula IV:
wherein, the A ring is 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 2;
R 1 each independently selected fromUnsubstituted or substituted by R 5 Substituted C 1～4 An alkyl group; r is R 5 Is hydroxy, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group.
Further, the structure of the compound is shown as a formula V-1, a formula V-2 or a formula V-3:
wherein R is 1 Selected from the group consisting ofUnsubstituted or substituted by R 5 Substituted C 1～2 An alkyl group; r is R 5 Is hydroxy, C 1～2 Alkoxy or C 1～2 Alkyl, R 6 Is C 1～2 An alkyl group.
Further, the compound is selected from:
the invention also provides a medicine for treating cancers, which is a preparation prepared by taking the compound or the salt, the stereoisomer and the deuterated compound thereof as active ingredients and adding pharmaceutically acceptable auxiliary materials.
The invention also provides application of the compound or the salt, stereoisomer and deuterated compound thereof in preparing FGFR kinase inhibitor.
Further, the FGFR kinase inhibitor is an FGFR4 kinase inhibitor.
Further, the FGFR4 kinase inhibitor is a drug for preventing and/or treating a disease associated with FGFR4 kinase activity.
Further, the FGFR kinase inhibitor is FGFR4 V550L Mutant inhibitors.
Further, the FGFR4 V550L Mutant inhibitors are prophylactic and/or therapeutic agents and FGFR4 V550L A medicament for mutation-related diseases.
Further, the disease is gastric cancer, liver cancer, prostate cancer, rhabdomyosarcoma, ovarian cancer, colon cancer or cutaneous melanoma.
Experimental results show that the compound has excellent inhibition effect on FGFR4 kinase and can be used as an FGFR4 kinase inhibitor.
The compounds of the invention can act as inhibitors of FGFR4 wild type and mutants.
The compound of the invention can effectively inhibit the proliferation of Ba/F3-ETV6-FGFR4-V550L cells and effectively overcome FGFR4 V550L Mutations, which can be used as FGFR4 V550L Mutant inhibitors.
It is well known to those skilled in the art that inhibition of FGFR4 kinase activity is effective in preventing and treating tumors (including gastric cancer, liver cancer, prostate cancer, rhabdomyosarcoma, ovarian cancer, colon cancer, cutaneous melanoma, etc.) associated with FGFR4 kinase activity.
The compounds of the invention can be used for the preparation of a pharmaceutical composition for the prevention and/or treatment of diseases associated with FGFR4 kinase activity, and FGFR4 V550L The medicine for treating mutation related diseases has wide application prospect.
It should be apparent that, in light of the foregoing, various modifications, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims.
The above-described aspects of the present invention will be described in further detail below with reference to specific embodiments in the form of examples. It should not be understood that the scope of the above subject matter of the present invention is limited to the following examples only. All techniques implemented based on the above description of the invention are within the scope of the invention.
Detailed Description
The raw materials and equipment used in the invention are all known products and are obtained by purchasing commercial products.
Example 1: synthesis of Compounds 8a-8p
Compounds 8a-8i, 8p were synthesized according to the following scheme:
1. synthesis of Compound 8a
(R)-N-(6-(5-(1-(3,5-dichloropyridin-4-yl)ethoxy)-1H-indazol-3-yl)-2-methyl isoindolin-5-yl)acrylamide(8a)
The first step: synthesis of Compound 2
Compound 1 (13.4 g,100 mmol) and imidazole (10.2 g,150 mmol) were dissolved in DMF (230 mL), cooled to 0deg.C, and t-butyldimethylchlorosilane (22.65 g,150 mmol) was added in portions over 30 minutes and reacted at ambient temperature for 3 hours. After the completion of the reaction, TLC was followed by quenching with water in ice bath and extraction with ethyl acetate three times. The organic phases were combined, washed three times with water, once with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by column chromatography to give compound 2 (orange oily crystal, 22.32g, yield 90%). 1 H NMR(400MHz,DMSO-d 6 ):δ12.89(s,1H),7.92(s,1H),7.41(d,J＝8.8Hz,1H),7.12(d,J＝2.3Hz,1H),6.91(dd,J＝8.8,2.3Hz,1H),0.96(s,9H),0.18(s,6H).ESI-MS m/z 249.1[M+H] + .
And a second step of: synthesis of Compound 3
Compound 2 (12.4 g,50 mmol) was dissolved in dichloromethane (170 mL), cooled to 10℃and N-iodosuccinimide (12.38 g,55 mmol) was added in portions over 30 minutes and reacted at ambient temperature for 3 hours. After the reaction was complete by TLC, it was cooled to 10 ℃ and quenched with water and extracted three times with dichloromethane. The organic phases were combined, washed twice with a saturated sodium thiosulfate solution, once with a saturated saline solution, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give compound 3 (orange solid, 18.51g, yield 99%). 1 H NMR(400MHz,DMSO-d 6 ):δ13.38(s,1H),7.45(d,J＝8.9Hz,1H),7.01(dd,J＝8.9,2.2Hz,1H),6.71(d,J＝2.2Hz,1H),0.97(s,9H),0.19(s,6H).ESI-MS m/z 375.0[M+H] + .
And a third step of: synthesis of Compound 4
Compound 3 (14.97 g,40 mmol) was dissolved in a mixed solution of dichloromethane (100 mL) and tetrahydrofuran (40 mL), cooled to 10℃and after methanesulfonic acid (520. Mu.L, 8 mmol) was added, 3, 4-dihydro-2H-pyran (5.04 g,60 mmol) was added in portions over 30 minutes and reacted at room temperature for 3 hours. After the completion of the reaction, TLC was monitored, cooled to 10 ℃ and quenched with saturated sodium bicarbonate solution and extracted three times with dichloromethane. The organic phases were combined, washed once with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The resulting solid was dissolved in tetrahydrofuran (100 mL), and a 1M tetrabutylammonium fluoride/tetrahydrofuran solution (40 mL,40 mmol) was added and stirred at room temperature for 5 hours. After completion of the reaction, the reaction mixture was concentrated under reduced pressure, the residue was dissolved in ethyl acetate, filtered through celite, the cake was washed with ethyl acetate, and the filtrate was concentrated to dryness. The residue was purified by column chromatography to give compound 4 (8.7 g, yield 63%) as an off-white solid. 1 H NMR(400MHz,DMSO-d 6 ):δ7.65(d,J＝9.0Hz,1H),7.08(dd,J＝8.9,2.3Hz,1H),6.72(d,J＝2.3Hz,1H),5.79(dd,J＝9.8,2.4Hz,1H),3.86(d,J＝11.6Hz,1H),3.71(ddd,J＝11.3,7.9,5.7Hz,1H),2.42-2.25(m,1H),2.04-1.93(m,2H),1.71(d,J＝10.5Hz,1H),1.56(q,J＝7.9,6.2Hz,2H).ESI-MS m/z 345.1[M+H] + .
Fourth step: synthesis of Compound 5
Compound 4 (5.16 g,15 mmol), cesium carbonate (5.38 g,16.5 mmol) was dissolved in acetonitrile (50 mL) to form a suspension, heated to 60 ℃, and (S) -1- (3, 5-dichloropyridin-4-yl) ethyl methanesulfonate (4.66 g,17.25 mmol) was added to the reaction solution and reacted overnight at 60 ℃. After the reaction was completed, it was cooled to room temperature, filtered, the cake was washed with acetonitrile and discarded, and the filtrate was concentrated to dryness. The residue was purified by column chromatography to give 5- ((R) -1- (3, 5-dichloropyridin-4-yl) ethoxy) -3-iodo-1- (tetrahydro-2H-pyran-2-yl) -1H indazole (i.e., compound 5,5.83g, 75% yield). ESI-MS m/z 518.2[ M+H ]] + .
Fifth step: synthesis of Compound 6a
Compound 5 (104 mg,0.2 mmol), 2-methyl-5-nitro-6- (4, 5-tetramethyl-1, 3, 2-dioxabenzaldehyde-2-yl) isoindoline (73 mg,0.24 mmol), pdCl 2 (DPPF) (15 mg,0.02 mmol) and Cs 2 CO 3 (130 mg,0.4 mmol) was dissolved in a mixed solution of 1, 4-dioxane (4 mL) and water (1 mL), and after 10 minutes of nitrogen substitution, the reaction was carried out at 85℃overnight under nitrogen protection. After the reaction was completed, the mixture was concentrated under reduced pressure, the concentrate was dissolved in a dichloromethane/methanol (10:1) solution, the mixture was filtered through celite, the cake was washed with a dichloromethane/methanol (10:1) solution and then discarded, and the filtrate was concentrated to dryness. The residue was purified by prep. to give 5- ((R) -1- (3, 5-dichloropyridin-4-yl) ethoxy) -3- (2-methyl-6-nitroindolin-5-yl) -1- (tetrahydro-2H-pyran-2-yl) -1H-indazole (i.e., compound 6a,87.5mg, 77% yield). 1H NMR (400 MHz, chloride-d) delta 8.41 (s, 2H), 7.75 (d, J=4.2 Hz, 1H), 7.55-7.49 (m, 1H), 7.39 (d, J=11.1 Hz, 1H), 7.13 (dt, J=9.1, 2.7Hz, 1H), 6.80 (t, J=3.0 Hz, 1H), 6.00-5.91 (m, 1H), 5.69 (ddd, J=8.8, 6.6,2.9Hz, 1H), 4.03 (d, J=2.5 Hz, 4H), 3.91 (d, J=11.5 Hz, 1H), 3.75-3.64 (m, 1H), 2.66 (s, 3H), 2.46 (d, J=11.6 Hz, 1H), 2.12 (s, 1H), 2.03 (s, 1H), 1.78 (ddd, 6.6,2.9Hz, 1H), 4.03 (d, J=2.5 Hz, 1H), 3.75 (d, 1.9 Hz, 1H) and 3.9M (1H)]+.
Sixth step: synthesis of Compound 7a
The obtained compound 6a (87.5 mg,0.15 mmol) was dissolved in a mixed solvent of ethanol (4 mL) and water (1 mL), ammonium chloride (80 mg,1.5 mmol) was added, the temperature was raised to 60℃and iron powder (42 mg,0.75 mmol) was added, and the temperature was raised to 90℃to reflux for 2 hours. After the reaction was completed, the mixture was concentrated to dryness under reduced pressure, dissolved in a methylene chloride/methanol (10:1) solution, adjusted to pH 7 to 8 with methanolic ammonia, filtered through celite, the cake was washed with a methylene chloride/methanol (10:1) solution, removed, and the filtrate was concentrated to dryness. The residue was purified by prep. to give 2- (5- (1- (3, 5-dichloropyridin-4-yl) ethoxy) -1- (tetrahydro-2H-pyran-2-yl) -1H-indazol-3-yl) -4-fluoroaniline (i.e. compound 7a,71mg, 86% yield). 1 H NMR(400MHz,DMSO-d 6 )δ8.60(d,J＝2.5Hz,2H),7.70(dd,J＝9.1,2.3Hz,1H),7.20(dt,J＝9.1,2.0Hz,1H),7.11(d,J＝10.4Hz,1H),7.03(t,J＝2.5Hz,1H),6.70(s,1H),6.09(dd,J＝6.6,3.3Hz,1H),5.93(s,2H),5.81(t,J＝6.7Hz,1H),4.09(q,J＝5.3Hz,1H),3.88–3.81(m,2H),3.76(d,J＝6.0Hz,2H),3.17(d,J＝5.2Hz,3H),2.35(s,1H),2.00(d,J＝8.9Hz,2H),1.75(d,J＝6.6Hz,3H),1.57(s,2H).ESI-MS m/z 538.2[M+H] + .
Seventh step: synthesis of Compound 8a
Compound 7a (71 mg,0.13 mmol) and triethylamine (34. Mu.L, 0.26 mmol) were dissolved in dichloromethane (3 mL), cooled to 0℃and acryloyl chloride (13. Mu.L, 0.16 mmol) was added dropwise, and after the addition was completed, the temperature was raised to room temperature and stirred for 1 hour. After completion of the reaction, the reaction was quenched by addition of methanol, concentrated under reduced pressure, and the residue was purified by preparative plates. The resulting solid was dissolved in dichloromethane (3 mL), trifluoroacetic acid (2 mL) was added and the reaction was allowed to proceed overnight at room temperature. After completion of the reaction by TLC, the solvent was removed by concentration under reduced pressure, pH was adjusted to 7-8 by adding an methanolic ammonia solution, and then concentration under reduced pressure was carried out to dryness, and the residue was purified by a preparative plate to give Compound 8a (off-white solid, 22mg, yield 33%). 1 H NMR(400MHz,DMSO-d 6 ):δ13.40(s,1H),10.78(s,1H),8.60(s,2H),8.25(s,1H),7.55(d,J＝9.0Hz,1H),7.36(s,1H),7.16(dd,J＝9.0,2.3Hz,1H),7.02(d,J＝2.3Hz,1H),6.35-6.15(m,2H),6.07(q,J＝6.6Hz,1H),5.75(dd,J＝9.9,2.0Hz,1H),4.02-3.82(m,4H),2.56(s,3H),1.74(d,J＝6.6Hz,3H).HRMS:calculated for C 26 H 23 Cl 2 N 5 O 2 [(M+H) + ],508.1302；found 508.1301.
2. Synthesis of Compound 8b
(R)-N-(6-(5-(1-(3,5-dichloropyridin-4-yl)ethoxy)-1H-indazol-3-yl)-2-(2-hydr oxyethyl)isoindolin-5-yl)acrylamide(8b)
The synthesis of reference compound 8a differs in that 2-methyl-5-nitro-6- (4, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) isoindol-2-yl) ethanol-1-ol is substituted for 2-methyl-5-nitro-6- (4, 5-tetramethyl-1, 3, 2-dioxabenzaldehyde-2-yl) isoindoline in the fifth step. Compound 8b was finally obtained (white solid, yield 10%). 1 H NMR(400MHz,DMSO-d 6 ):δ13.34(s,1H),10.68(s,1H),8.63(s,2H),8.21(s,1H),7.48(d,J＝9.0Hz,1H),7.38(s,1H),7.20(dd,J＝9.0,2.3Hz,1H),6.96(d,J＝2.3Hz,1H),6.37-6.20(m,2H),6.02(q,J＝6.6Hz,1H),5.78(dd,J＝9.9,2.0Hz,1H),4.08-3.92(m,4H),2.89-2.78(m,3H),2.64(t,J＝6.2Hz,2H),1.76(d,J＝6.6Hz,3H).HRMS:calculated for C 27 H 25 Cl 2 N 5 O 3 [(M+H) + ],538.1407；found 538.1403.
3. Synthesis of Compounds 8 c-8 p
The synthesis method of the reference compound 8a is distinguished in that the 2-methyl-5-nitro-6- (4, 5-tetramethyl-1, 3, 2-dioxabenzaldehyde-2-yl) isoindoline is replaced by a corresponding raw material in the fifth step. Finally, respectively obtaining the compounds 8 c-8 p:
8C white solid, 18% yield, HRMS calculated for C 28 H 27 Cl 2 N 5 O 3 [(M+H) + ],552.1564；found 552.1563.
8d white solid, 22% yield. 1 H NMR(400MHz,DMSO-d 6 ):δ13.38(s,1H),10.63(s,1H),8.57(s,2H),8.15(s,1H),7.54(d,J＝9.0Hz,1H),7.25(s,1H),7.16(dd,J＝9.1,2.2Hz,1H),7.07(d,J＝2.3Hz,1H),6.33-6.15(m,2H),6.09(q,J＝6.5Hz,1H),5.74(dd,J＝9.7,2.0Hz,1H),3.67-3.51(m,2H),2.87(d,J＝6.2Hz,2H),2.66(d,J＝5.8Hz,2H),2.43(s,3H),1.74(d,J＝6.6Hz,3H).HRMS:calculated for C 27 H 25 Cl 2 N 5 O 2 [(M+H) + ],522.1458；found 522.1457.
8e as white solid in 11% yield. 1 H NMR(400MHz,DMSO-d 6 ):δ13.40(s,1H),10.68(s,1H),8.58(s,2H),8.16(s,1H),7.55(d,J＝9.0Hz,1H),7.25(s,1H),7.17(dd,J＝9.0,2.3Hz,1H),7.06(d,J＝2.4Hz,1H),6.29(dd,J＝17.1,9.9Hz,1H),6.19(dd,J＝17.0,2.0Hz,1H),6.10(q,J＝6.6Hz,1H),5.74(dd,J＝10.1,2.1Hz,1H),4.61(s,1H),3.82-3.65(m,3H),3.04(s,1H),2.86(m,4H),2.70(d,J＝10.6Hz,2H),1.74(d,J＝6.6Hz,3H).HRMS:calculated for C 28 H 27 Cl 2 N 5 O 3 [(M+H) + ],552.1564；found 552.1566.
8f white solid, 19% yield, 1H NMR (400 MHz, DMSO-d 6)δ13.39(s,1H),10.68(s,1H),8.58(s,2H),8.15(s,1H),7.55(d,J＝9.0Hz,1H),7.22(s,1H),7.17(dd,J＝9.0,2.2Hz,1H),7.06(d,J＝2.3Hz,1H),6.29(dd,J＝17.0,9.9Hz,1H),6.19(dd,J＝17.0,1.9Hz,1H),6.10(q,J＝6.6Hz,1H),5.74(dd,J＝9.9,1.9Hz,1H),3.79–3.63(m,2H),3.61(t,J＝5.7Hz,2H),3.33(s,3H),2.86(d,J＝5.9Hz,2H),2.76(dt,J＝11.5,5.8Hz,4H),1.74(d,J＝6.6Hz,3H).HRMS:calculated for C 29 H 29 Cl 2 N 5 O 3 [(M+H) + ],566.1720；found 566.1719.
8g of white solid, yield 19%, 1 H NMR(400MHz,DMSO-d 6 ):δ13.38(s,1H),10.62(s,1H),8.59(s,2H),8.08(s,1H),7.55(d,J＝9.0Hz,1H),7.26(s,1H),7.16(dd,J＝9.0,2.3Hz,1H),7.03(d,J＝2.3Hz,1H),6.29(dd,J＝17.0,9.9Hz,1H),6.18(dd,J＝17.0,2.0Hz,1H),6.07(q,J＝6.6Hz,1H),5.73(dd,J＝9.9,2.0Hz,1H),3.59-3.47(m,2H),3.03-2.91(m,1H),2.84(dd,J＝16.3,5.9Hz,1H),2.73-2.62(m,2H),2.39(s,3H),1.73(d,J＝6.6Hz,3H).HRMS:calculated for C 27 H 25 Cl 2 N 5 O 2 [(M+H) + ],522.1458；found 522.1456.
8h of white solid, 11% yield, 1 H NMR(400MHz,DMSO-d 6 ):δ13.38(s,1H),10.64(s,1H),8.59(s,2H),8.08(s,1H),7.55(d,J＝9.0Hz,1H),7.25(s,1H),7.17(dd,J＝9.0,2.3Hz,1H),7.03(d,J＝2.3Hz,1H),6.29(dd,J＝17.1,9.9Hz,1H),6.18(dd,J＝17.1,1.9Hz,1H),6.08(q,J＝6.5Hz,1H),5.73(dd,J＝9.9,2.0Hz,1H),4.51(t,J＝5.5Hz,1H),3.71-3.60(m,4H),2.96(t,J＝9.1Hz,1H),2.85-2.73(m,3H),2.62(t,J＝6.2Hz,2H),1.74(d,J＝6.6Hz,3H).HRMS:calculated for C 28 H 27 Cl 2 N 5 O 3 [(M+H) + ]552.1564; found 552.1565.8c-white solid, yield 8%, HRMS: calculated for C 28 H 27 Cl 2 N 5 O 3 [(M+H) + ],552.1564；found552.1563.
8i white solid, 22% yield, 1H NMR (400 MHz, DMSO-d 6) delta 13.39 (s, 1H), 10.65 (s, 1H), 8.60 (s, 2H), 8.09 (s, 1H), 7.55 (d, J=9.0 Hz, 1H), 7.26 (s, 1H), 7.17 (dd, J=9.0, 2.3Hz, 1H), 7.03 (d, J=2.2 Hz, 1H), 6.29 (dd, J=j=)17.0,9.9Hz,1H),6.19(dd,J＝17.1,2.0Hz,1H),6.08(q,J＝6.6Hz,1H),5.74(dd,J＝10.0,1.9Hz,1H),3.73–3.61(m,2H),3.58(t,J＝5.7Hz,2H),3.30(s,3H),3.01–2.91(m,1H),2.81(td,J＝15.2,5.2Hz,3H),2.71(t,J＝5.8Hz,2H),1.74(d,J＝6.5Hz,3H).HRMS:calculated for C 29 H 29 Cl 2 N 5 O 3 [(M+H) + ],566.1720；found566.1722.
8p： 1 H NMR(400MHz,Chloroform-d)δ10.94(s,1H),10.26(s,1H),8.65(s,1H),8.44(s,2H),7.49–7.44(m,2H),7.24(dd,J＝9.0,2.3Hz,1H),7.09(d,J＝2.2Hz,1H),6.37(dd,J＝17.0,1.2Hz,1H),6.22(dd,J＝17.0,10.2Hz,1H),6.04(q,J＝6.7Hz,1H),5.72(dd,J＝10.2,1.2Hz,1H),4.81(m,4H),2.94(s,3H),1.82(d,J＝6.6Hz,3H).
Characterization data for compound 8j is the same as compound 8a, characterization data for compound 8k is the same as compound 8b, characterization data for compound 8l is the same as compound 8d, characterization data for compound 8m is the same as compound 8f, characterization data for compound 8n is the same as compound 8g, and characterization data for compound 8o is the same as compound 8 h.
TABLE 1 Structure of Compounds 8a-8p
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The following experiments prove the beneficial effects of the invention.
Experimental example 1 inhibition Activity of the Compounds of the invention against FGFR4 kinase and against FGFR4 kinase
Experiment of Ba/F3-ETV6-FGFR4-V550L cell proliferation inhibition
1. Kinase activity test method:
the method for testing the inhibitory effect of the compounds on the kinase activity of FGFR4 is as follows.
(1) Compound preparation:
test compounds were dissolved in 100% DMSO and prepared into 10mM stock solutions and stored in a nitrogen cabinet protected from light.
(2) Kinase reaction process:
(1) Preparing a 1 XKinase buffer;
(2) Preparing a compound concentration gradient: test compounds were tested at 100nM and diluted in 384source plates to 100-fold final concentration in 100% DMSO and 8-10 concentrations using precision 3-fold dilution. Transferring 250nL of 100 times the final concentration of the compound into the destination plate 384-well-plate using a dispenser Echo 550;
(3) Preparing a Kinase solution with a final concentration of 2.5 times by using a 1 XKinase buffer;
(4) Adding 10 mu L of kinase solution with 2.5 times of final concentration to each of the compound well and the positive control well; add 10. Mu.L of 1 XKinase buffer to the negative control wells;
(5) Centrifuging at 1000rpm for 30 seconds, shaking and mixing the reaction plate uniformly, and incubating for 10 minutes or 60 minutes at room temperature;
(6) A mixed solution of ATP and Kinase substrate22 was prepared at 25/15 times the final concentration using a 1 XKinase buffer;
(7) Adding 15 mu L of a mixed solution of ATP and a substrate with 5/3 times of the final concentration, and initiating a reaction;
(8) Centrifuging the 384-well plate at 1000rpm for 30 seconds, shaking and uniformly mixing, and incubating at room temperature for corresponding time;
(9) Adding 30 mu L of stop detection solution to stop kinase reaction, centrifuging at 1000rpm for 30 seconds, and shaking and mixing uniformly;
(10) The conversion was read with Caliper EZ Reader.
(3) Data analysis:
the calculation formula is as follows:
wherein: inhibition rate: % Inhibition; conversion% _sample: conversion reading of the sample; convertion% _min: negative control Kong Junzhi, representing conversion reading without enzyme wells; convesion% _max: positive control Kong Junzhi, represents a conversion reading without compound inhibition wells.
Fitting dose-response curve:
the log value of the concentration is taken as an X axis, the percent inhibition rate is taken as a Y axis, and a log (inhibitor) vs. response-Variable slope fit quantitative effect curve of analysis software GraphPad Prism 5 is adopted, so that the IC of each compound on the enzyme activity is obtained 50 Values.
The calculation formula is Y=bottom+ (Top-Bottom)/(1+10 ((LogIC) 50 -X)*HillSlope))
2. Cell proliferation assay
a) All cell lines were cultured in complete medium at 37 ℃,5% CO 2;
b) Cells in the logarithmic growth phase were harvested and counted using a platelet counter. Detecting the cell activity by trypan blue exclusion method, and ensuring the cell activity to be more than 90%;
c) The cell density is adjusted by using a complete culture medium, and then the cells are inoculated in a 96-well cell culture plate, and 90 mu L of cells are inoculated in each well, wherein the total number of the cells is 3000;
d) Cells in 96-well plates were placed at 37℃with 5% CO 2 Culturing under the condition;
e) Preparing a 10-time drug solution, wherein the highest concentration of the compound to be tested is 5 mu M,9 concentrations are detected, 3 times dilution is carried out, then the compound to be tested is transferred into corresponding experimental holes of a 96-hole cell plate with 10 mu L of each serial diluted compound, and three compound holes are arranged for each drug concentration;
f) Cells in the dosed 96-well plates were placed at 37℃with 5% CO 2 Culturing under the condition for 72 hoursAfter that, CTG analysis is performed;
g) Thawing CTG reagent and equilibration of cell plates to room temperature for 30 min;
h) Adding an equal volume of CTG solution to each well;
i) Vibrating on an orbital shaker for 5 minutes to lyse cells;
j) The cell plates were left at room temperature for 20 min to stabilize the luminescence signal;
k) The luminescence value is read and the data is collected.
3. Data analysis:
analysis of data using GraphPad Prism 7.0 software, fitting data to derive dose-response curves using nonlinear S-curve regression, and calculating IC therefrom 50 Values.
Cell viability (%) = (Lum) Drug to be tested -Lum Culture broth control )/(Lum Cell control -Lum Culture broth control )×100％.
4. Experimental results
TABLE 2 IC of the inventive compounds for inhibiting FGFR4 kinase activity and inhibiting Ba/F3-ETV6-FGFR4-V550L cell proliferation 50 Value of
The results are shown in Table 2. The compound has excellent inhibition effect on FGFR4 kinase, can be used as an FGFR4 kinase inhibitor, can effectively inhibit the proliferation of Ba/F3-ETV6-FGFR4-V550L cells, and can effectively overcome FGFR4 V550L Mutations, which can be used as FGFR4 V550L Mutant inhibitors.

Claims (10)

1. A compound of formula I or a salt, stereoisomer, deuterated compound thereof:
wherein the A ring is 5-6 membered saturated cycloalkyl, 5-6 membered saturated heterocyclic group, 5-6 membered aryl or 5-6 membered heteroaryl;
m is an integer of 0 to 5;
R 1 each independently selected fromUnsubstituted or substituted by R 5 The substituted following groups: c (C) 1～6 Alkyl, C 1～6 An alkoxy group; r is R 5 Is hydroxy, amino, halogen, carboxyl, cyano, C 1～6 Alkoxy or C 1～6 An alkyl group; r is R 6 Is C 1～6 An alkyl group;
R 2 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
R 3 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
R 4 is halogen, C 1～6 Alkyl or C 1～6 An alkoxy group;
is->
2. The compound or salt, stereoisomer, deuterated compound according to claim 1 wherein: the ring A is 5-6 membered saturated cycloalkyl or 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 3;
R 1 each independently selected fromUnsubstituted or substituted by R 5 The substituted following groups: c (C) 1～4 Alkyl, C 1～4 An alkoxy group; r is R 5 Is hydroxy, amino, halogen, carboxyl, cyano, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group;
R 2 is halogen;
R 3 is halogen;
R 4 is C 1～4 An alkyl group.
3. The compound or salt, stereoisomer, deuterated compound according to claim 2 wherein: the structure of the compound is shown as a formula II:
wherein, the A ring is 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 2;
R 1 each independently selected fromUnsubstituted or substituted by R 5 Substituted C 1～4 An alkyl group; r is R 5 Is hydroxy, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group;
preferably, the structure of the compound is shown as a formula III-1, a formula III-2 or a formula III-3:
wherein R is 1 Selected from the group consisting ofUnsubstituted or substituted by R 5 Substituted C 1～2 An alkyl group; r is R 5 Is hydroxy, C 1～2 Alkoxy or C 1～2 An alkyl group; r is R 6 Is C 1～2 An alkyl group.
4. The compound or salt, stereoisomer, deuterated compound according to claim 2 wherein: the structure of the compound is shown in a formula IV:
wherein, the A ring is 5-6 membered saturated heterocyclic group;
m is an integer of 0 to 2;
R 1 each independently selected fromUnsubstituted or substituted by R 5 Substituted C 1～4 An alkyl group; r is R 5 Is hydroxy, C 1～4 Alkoxy or C 1～4 An alkyl group; r is R 6 Is C 1～4 An alkyl group;
preferably, the structure of the compound is shown as formula V-1, formula V-2 or formula V-3:
wherein R is 1 Selected from the group consisting ofUnsubstituted or substituted by R 5 Substituted C 1～2 An alkyl group; r is R 5 Is hydroxy, C 1～2 Alkoxy or C 1～2 Alkyl, R 6 Is C 1～2 An alkyl group.
5. The compound or salt, stereoisomer, deuterated compound according to claim 1 wherein: the compound is selected from:
6. a medicament for treating cancer, characterized in that: a preparation prepared by taking the compound or the salt, the stereoisomer and the deuterated compound thereof as an active ingredient and adding pharmaceutically acceptable auxiliary materials.
7. Use of a compound according to any one of claims 1 to 5, or a salt, stereoisomer, deuterated compound thereof, for the preparation of an FGFR kinase inhibitor.
8. Use according to claim 7, characterized in that: the FGFR kinase inhibitor is an FGFR4 kinase inhibitor;
preferably, the FGFR4 kinase inhibitor is a medicament for preventing and/or treating a disease associated with FGFR4 kinase activity.
9. Use according to claim 7, characterized in that: the FGFR kinase inhibitor is FGFR4 V550L Mutant inhibitors;
preferably, the FGFR4 V550L Mutant inhibitors are prophylactic and/or therapeutic agents and FGFR4 V550L A medicament for mutation-related diseases.
10. Use according to any one of claims 8 or 9, characterized in that: the disease is gastric cancer, liver cancer, prostate cancer, rhabdomyosarcoma, ovarian cancer, colon cancer or skin melanoma.
CN202311138918.8A 2022-11-09 2023-09-05 FGFR kinase inhibitor and pharmaceutical application thereof Pending CN117430586A (en)

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