Compounds used as kinase inhibitors and their applications

Abstract

The present invention belongs to the field of heterocyclic compound drug synthesis with nitrogen atoms as heterocyclic atoms, and specifically relates to a class of compounds used as kinase inhibitors and their applications, and also relates to the preparation of the free base crystal form of the compound. The compound used as a kinase inhibitor is a compound as shown in Formula 1, or a deuterated substance thereof, or a pharmaceutically acceptable salt, solvate or prodrug. The above compounds have been confirmed by biological activity experiments to have good inhibitory activity against exon 20 insertion mutations of EGFR and Her2, exon 19 deletion of EGFR, and point mutations of exon 21, and can be used as the original drug of related drugs.

Description

Technical Field

[0001] The invention belongs to the field of drug synthesis of heterocyclic compounds with nitrogen atoms as heterocyclic atoms, and specifically relates to a class of compounds used as kinase inhibitors and applications thereof. Background Art

[0002] The epidermal growth factor receptor belongs to the receptor tyrosine kinase (RTK) family, which includes EGFR / ERBB1, HER2 / ERBB2 / NEU, HER3 / ERBB3 and HER4 / ERBB4. The epidermal growth factor receptor activates its tyrosine kinase activity through homodimerization or heterodimerization, and then phosphorylates its substrate, thereby activating multiple downstream pathways related to it in the cell, such as the PI3K-AKT-mTOR pathway involved in cell survival and the RAS-RAF-MEK-ERK pathway involved in cell proliferation. Mutations or amplifications of the epidermal growth factor receptor will lead to the activation of the epidermal growth factor receptor kinase, leading to the occurrence of various human diseases, such as malignant tumors. For example, in patients with non-small cell lung cancer, about 10% of American patients have EGFR mutations, while the proportion of patients with EGFR mutations in Asian patients can reach nearly 50%. At the same time, the incidence of HER2 mutations in patients with non-small cell lung cancer is about 2-4%.

[0003] EGFR mutations mainly include deletions, insertions, and point mutations, among which exon 19 deletions and exon 21 L858R point mutations account for nearly 90% of EGFR mutations. For patients with tumors with these EGFR mutations, the EGFR-TKIs currently on the market include first-generation Iressa, Tarceva, and Conmena, second-generation afatinib and dacomitinib, and third-generation osimertinib. The other 10% of EGFR mutations mainly involve exons 18 and 20 of EGFR, and the insertion mutation of EGFR exon 20 accounts for about 9% of the entire EGFR mutation. For patients with tumors with HER2 mutations, the most common HER2 mutation is the insertion mutation of HER2 exon 20.

[0004] TAK-788 has a therapeutic effect on exon 20 insertion mutations of EGFR and HER2. The compound has been marketed in the United States. According to the reported clinical trial results, the objective response rate is 43%. Recently, it has been reported that DZD9008 is effective in treating advanced non-small cell lung cancer with EGFR or HER2 mutations. The objective response rate of EGFR exon 20ins is 40%, which is not satisfactory. Summary of the invention

[0005] In order to meet the clinical medication needs of patients with EGFR and HER2 mutations, especially those with EGFR and HER2 exon 20 insertion mutations, WO2021180238 discloses a series of compounds with excellent EGFR and HER2 exon 20 insertion mutation activity and EGFR exon 19 deletion and exon 21 L858R point mutation, which have great potential to be developed into drugs for the treatment of related diseases. Among them, a class of compounds with a tri-pentaamine structure is involved. Further studies have shown that the stereo configuration of the tri-pentaamine structure is very important. When it is converted from trans to cis, its in vivo efficacy will have better efficacy, and it also has a good therapeutic effect in different mutations of related diseases. In subsequent research and development, in order to find a more stable and bioavailable free base compound, the present invention conducts polymorphic research and screening of free base compounds.

[0006] The purpose of the present invention is to provide a class of compounds used as kinase inhibitors, which belong to the cis-tripentaamine structure and have good inhibitory activity against EGFR, HER2 exon 20 insertion mutations, EGFR exon 19 deletions and point mutations of exon 21. The compounds of the present invention have better efficacy and wider applicability than the corresponding trans-structured compounds in patent WO2021180238.

[0007] The second object of the present invention is to provide the use of the above-mentioned compound in the preparation of a drug for treating related diseases caused by EGFR mutation and / or HER2 mutation.

[0008] The present invention also relates to the preparation of the free base crystal form of the above compound, and polymorphic studies are conducted to determine a practical crystal form with high stability and higher bioavailability.

[0009] In order to achieve the above purpose, the technical solution adopted by the present invention is:

[0010] A compound used as a kinase inhibitor, wherein the compound used as a kinase inhibitor is a compound as shown in Formula 1, or a deuterated substance thereof, or a pharmaceutically acceptable salt, solvate or prodrug thereof:

[0011]

[0012] In Formula 1, X is selected from CH, N;

[0013] R 1 Selected from R 5 is H, C1-C3 alkyl, C1-C3 fluoroalkyl;

[0014] R 20 , R 21 , R 22 are each independently selected from methyl or deuterated methyl;

[0015] R 3 Selected from C1-C3 alkyl, C1-C3 haloalkyl;

[0016] R 40 , R 41 , R 42 Each is independently selected from H, D, and F.

[0017] The above compounds have been confirmed through biological activity experiments to have good inhibitory activity against EGFR and HER2 mutations and can be used as the original drugs of related drugs.

[0018] Based on the above original drug, the "pharmaceutically acceptable salt" of the original drug refers to a salt that is pharmaceutically acceptable and has the desired pharmacological activity of the parent compound. Such salts include:

[0019] Acid addition salts formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid, etc., typical inorganic acid salts are selected from hydrochloride, hydrobromide, hydroiodide, sulfate, hydrogensulfate, nitrate, phosphate, acid phosphate. Acid addition salts formed with organic acids, such as formic acid, acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4- Toluenesulfonic acid, camphorsulfonic acid, glucoheptonic acid, 4,4'-methylenebis-(3-hydroxy2-ene-1-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tert-butylacetic acid, dodecylsulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid, etc.; or a salt formed by coordination of an acidic proton present in the parent compound with an organic base (e.g., ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucosamine, etc.). Typical organic acid salts are selected from formate, acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, ethanesulfonate, isethionate, benzenesulfonate, salicylate, picrate, glutamate, ascorbate, camphorate, camphorsulfonate. It is easy to understand that the pharmaceutically acceptable salt is non-toxic.

[0020] Solvates are compounds containing a solvent, such as hydrates, dimethyl sulfoxides, and the like.

[0021] A prodrug refers to a compound that undergoes chemical transformation by metabolism or chemical processes to produce a compound, salt, or solvate of the present invention when used to treat a related disease.

[0022] Preferably, the compound used as a kinase inhibitor is a compound as shown in Formula 2, or a deuterated compound thereof, or a pharmaceutically acceptable salt, solvate or prodrug thereof:

[0023]

[0024] Formula 2.

[0025] Further preferably, in Formula 2, X is selected from CH, N; R 3 Selected from -CH 3 、-CH 2 CH 3 、-CH 2 CF 3 ; R 40 , R 41 Both H, R 42 Select from H or F.

[0026] Preferably, the compound used as a kinase inhibitor is a compound as shown in Formula 3, or a deuterated compound thereof, or a pharmaceutically acceptable salt, solvate or prodrug thereof:

[0027]

[0028] Formula 3;

[0029] In Formula 3, X is selected from CH, N; R 3 -CH 3 、-CH 2 CH 3 、-CH 2 CF 3 ; R 40 , R 41 Both H, R 42 Selected from H or F; R 5 Selected from -CH 3 , -CF 3 .

[0030] Preferably, the compound represented by Formula 1 is:

[0031]

[0032]

[0033] The above-mentioned compound used as a kinase inhibitor is used in a drug for treating related diseases caused by EGFR mutation and / or HER2 mutation.

[0034] The above compounds have good inhibitory activity against exon 20 insertion mutations of EGFR and HER2, and also have good inhibitory activity against EGFR exon 19 deletion and exon 21 point mutations.

[0035] Preferably, the EGFR mutation and / or HER2 mutation include one or a combination of two or more of EGFR exon 20 insertion mutation, HER2 exon 20 insertion mutation, EGFR exon 19 deletion, EGFR exon 20 point mutation, and EGFR exon 21 point mutation. It has been confirmed by biological activity experiments that the above-mentioned compound has a good inhibitory effect on the above-mentioned mutation types.

[0036] Further preferably, the EGFR mutation and / or HER2 mutation is selected from EGFR Del 19 / T790M / C797S mutation and EGFR L858R / T790M / C797S mutation, and the above compound has a good inhibitory effect on the above mutation types.

[0037] Preferably, the disease is cancer caused by the EGFR mutation and / or HER2 mutation. The above compound can also be used in combination with other drugs for the treatment of cancer. Other drugs used in combination can be ERK inhibitors or MEK inhibitors.

[0038] Preferably, the compound is in crystalline form, amorphous form or the solvate; the solvent contained in the solvate is a non-aqueous solvent or a mixed solvent consisting of a non-aqueous solvent and water.

[0039] In order to better improve the stability and activity of the compound, a portion of the compound shown in Formula 1 was selected for crystal form research. Further, the compound of Example 1 in the present invention was subjected to polymorph screening research to find a stable and reliable crystal form, which can not only ensure the stability of the compound quality, but also promote the drug to play a better role in clinical treatment. It was found that the product of Example 1 was a crystal with good crystallinity and was anhydrous crystal form, named free base crystal form I. The compound of Example 1 has a compound as shown in Formula A:

[0040]

[0041] The crystal form is free base crystal form I, and the X-ray powder diffraction pattern of the crystal form has characteristic diffraction peaks at 9.76°±0.2°, 10.45°±0.2°, 16.54°±0.2°, 18.66°±0.2°, 20.07°±0.2°, and 25.90°±0.2°.

[0042] Furthermore, the free base form I has an X-ray powder diffraction pattern with characteristic diffraction peaks at 9.07°±0.2°, 9.76°±0.2°, 10.45°±0.2°, 11.53°±0.2°, 11.80°±0.2°, 12.91°±0.2°, 13.79°±0.2°, 14.67°±0.2°, 15.08°±0.2°, 15.63°±0.2°, 16.54°±0.2°, 17.50°±0.2°, 18.66°±0.2°, 20.07°±0.2°, 21.10°±0.2°, 23.29°±0.2°, 24.16°±0.2°, and 25.90°±0.2°.

[0043] The free base crystalline form I of the compound of Example 1 can be prepared in most solvents by suspension crystallization, anti-solvent precipitation, high and low temperature cycle and evaporative crystallization. The solvents herein include, but are not limited to, one or a mixed solvent of two or more of dichloromethane, 1,4-dioxane, dichloroethane, methyl tert-butyl ether, N-methylpyrrolidone, ethyl acetate, acetone, methanol, dimethyl sulfoxide, isopropyl acetate, butanone, cyclohexane, tetrahydrofuran, water, acetonitrile, isopropanol, ethanol, and n-heptane.

[0044] The present invention found that the compound of Example 1 has two anhydrous crystalline forms (free base crystalline form I and free base crystalline form V) and three solvates, free base crystalline form II is an ethanol solvate, free base crystalline form III is an isopropanol solvate, and free base crystalline form IV is a solvate of dichloroethane and water, and all these free base crystalline forms are characterized. A competitive beating experiment was conducted on the two anhydrous crystalline forms of free base crystalline form I and free base crystalline form V. From the results, it is inferred that free base crystalline form I may be a thermodynamically stable crystalline form, which is more suitable for subsequent development.

[0045] The crystal form of free base crystal form I was evaluated, including dry grinding, wet grinding, tableting (30MPa), stability and hygroscopicity. After 5 minutes of dry grinding, it became amorphous; the crystal form was basically unchanged after 5 minutes of wet grinding with water, and the crystallinity decreased slightly after 5 minutes of wet grinding with ethanol; when the pressure was 30MPa in the tableting test, the crystallinity decreased; in the stability study, the free base crystal form I was relatively stable at different temperatures and humidities, and the liquid phase detection results did not change; a dynamic water vapor adsorption (DVS) test was performed, and the results showed that the water absorption and weight gain were 0.66% at 80% RH, and the free base crystal form I was slightly hygroscopic. In short, the free base crystal form I is a relatively stable anhydrous crystal form with stable solid-state properties and slight hygroscopicity, which can be used for subsequent drug development. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 is the change in tumor volume in the LU0387 model (mm3 );

[0047] Figure 2 Tumor volume change of Ba / F3 EGFR D770_N771 ins SVD model (mm 3 );

[0048] Figure 3 is the single crystal diffraction pattern of the single crystal of Example 1;

[0049] Figure 4 is the XRPD pattern of free base Form I;

[0050] Figure 5 It is the DSC and TGA superposition spectrum of the free base crystal form I;

[0051] Figure 6 This is the XRPD pattern of the amorphous sample obtained by dry grinding;

[0052] Figure 7 is the XRPD pattern of free base Form II;

[0053] Figure 8 It is the DSC and TGA superposition spectrum of the free base crystal form II;

[0054] Fig. 9 is the XRPD pattern of free base Form III;

[0055] Fig.10 It is the DSC and TGA superposition spectrum of the free base form III;

[0056] Fig.11 is the XRPD pattern of free base Form IV;

[0057] Fig.12 It is the DSC and TGA superposition spectrum of the free base crystal form IV;

[0058] Fig.13 is an XRPD overlay of the free base Form V;

[0059] Fig.14 The DSC and TGA superposition of the free base form V. DETAILED DESCRIPTION

[0060] Compared with the compounds with a trans-tripentaamine structure in patent WO2021180238, the present invention confirms that the cis-tripentaamine structure has better in vivo efficacy and also has a good therapeutic effect in different mutations of related diseases.

[0061] The preparation method of the compound of the present invention when X is CH, the reaction route is as follows:

[0062]

[0063] The following steps are involved:

[0064] (1) Compound A and compound B are subjected to Friedel-Crafts reaction in an organic solvent to obtain compound C;

[0065] (2) Compound C and compound D undergo substitution reaction in an organic solvent under acid catalysis to obtain compound E;

[0066] (3) Compound E and compound F undergo a substitution reaction in an organic solvent under base catalysis to obtain compound G;

[0067] (4) Compound G is reduced to obtain compound H;

[0068] (5) Compound H undergoes a condensation reaction in the presence of a base catalyst to obtain a product.

[0069] R 1 for And R 5 For-CF 3 When, the synthesis of the compound shown in Formula 1 can also be carried out according to the following reaction route:

[0070]

[0071] The following steps are involved:

[0072] (1) Compound a and compound b undergo a substitution reaction in an organic solvent to obtain compound c;

[0073] (2) Compound c is hydrolyzed to obtain compound d;

[0074] (3) Compound d undergoes a condensation reaction to obtain compound e;

[0075] (4) deprotecting compound e to obtain compound f;

[0076] (5) Compound f undergoes a ring-closing reaction with trifluoroacetic anhydride in the presence of a base catalyst to obtain compound g;

[0077] (6) Compound g is reduced to obtain compound h;

[0078] (7) Compound h undergoes a condensation reaction in the presence of a base catalyst to obtain a product.

[0079] The preparation method of the compound of the present invention when X is N, the reaction scheme is as follows:

[0080]

[0081] The method comprises the following steps: Compound A' and compound B' undergo substitution reaction to obtain a product.

[0082] Among them, compound A′ was synthesized with reference to WO2021180238; compound B′ was obtained by Friedel-Crafts reaction.

[0083] The implementation process of the present invention is described in detail below in conjunction with specific embodiments.

[0084] 1. Specific Examples of Synthesis of Compounds Used as Kinase Inhibitors

[0085] Example 1

[0086] The compound used as a kinase inhibitor in this example has the following structural formula:

[0087]

[0088] The synthetic route of the compound of this embodiment is as follows:

[0089]

[0090] Synthesis of compound 3: Add compound 1 (2.01 g, 8.5 mmol) to a 250 mL three-necked flask under nitrogen protection, dissolve it completely in 100 mL tetrahydrofuran, add anhydrous aluminum chloride (2.26 g, 17 mmol), and stir at 70°C for 1 hour. Add compound 2 (1.34 g, 10.2 mmol) dropwise, continue to react at 70°C, monitor the reaction with a plate, and the reaction is basically completed after 4 hours. Treat to obtain 1.31 g of the product with a yield of 46.4%.

[0091] Synthesis of compound 5: Compound 3 (1.30 g, 3.9 mmol), compound 4 (0.88 g, 4.68 mmol), and p-toluenesulfonic acid (1.35 g, 7.8 mmol) were added to 65 mL of 1,4-dioxane. 2 The reaction was monitored by a plate, and the reaction was completed and treated to obtain 1.50 g of the product with a yield of 79.7%.

[0092] Synthesis of compound 7: Add compound 5 (6.0 g, 12.5 mmol), compound 6 (6.3 g, 100 mmol) and N,N-diisopropylethylamine (3.2 g, 50 mmol) to 120 mL of N,N-dimethylacetamide, react overnight at 90°C, monitor with a plate, and after the reaction of the raw materials is complete, treat to obtain 8.0 g of product. 1H NMR (400MHz, Chloroform-d) δ9.53 (s, 1H), 8.97-8.75 (m, 1H), 7.96 (s, 1H), 7.84 (s, 1H), 7.79 ( d, J=8.0Hz, 1H), 7.38 (d, J=8.2Hz, 1H), 7.29 (d, J=7.2Hz, 1H), 7.20 (t, J=7.6Hz, 1H), 6.81 (s, 1H ), 5.09 (p, J=6.4Hz, 1H), 4.02 (s, 3H), 3.92 (s, 3H), 3.84-3.63 (m, 1H), 3.02 (t, J=6.4Hz, 1H), 2. 84 (s, 3H), 2.70 (s, 3H), 2.43 (s, 2H), 2.32 (s, 1H), 1.27 (d, J = 7.2Hz, 2H), 1.15 (d, J = 6.4Hz, 6H).

[0093] Synthesis of compound 8: Add compound 7 (7.7 g, 41.6 mmol) and palladium carbon (2.4 g, wet palladium carbon 55%) to a mixed solvent of 100 mL of methanol and 100 mL of ethyl acetate, react at room temperature for 4 hours under hydrogen atmosphere, monitor with a spot plate, and after the raw materials react, treat to obtain 7.0 g of product.

[0094] Synthesis of Example 1: Compound 8 (7.00 g, 12.6 mmol) and triethylamine (3.82 g, 37.8 mmol) were dissolved in dichloromethane, cooled to 0°C under nitrogen protection, and then a dichloromethane solution of acryloyl chloride (1.72 g, 18.9 mmol) was added dropwise, and the reaction was maintained at 0°C for 1 hour, and the plate was monitored. After the reaction of the raw materials, they were processed. That is, sodium bicarbonate aqueous solution and dichloromethane were added, stirred and layered, the aqueous phase was extracted once with dichloromethane, the organic phases were combined, dried, spin-dried, and passed through a column. The obtained crude product was first dissolved with a small amount of dichloromethane, and then petroleum ether was added dropwise to precipitate a large amount of solids, filtered, and the filter cake was directly added with methanol for pulping. After drying, 3.5 g of product was finally obtained. [M+H] + : 610.8; 1H NMR (400MHz, Chloroform-d) δ10.38 (s, 1H), 9.60 (s, 1H), 8.87 (s, 1H), 8.61 (s, 1H), 7.89 (s, 1H), 7.58 (s, 1H), 7.32 (d, J=8.2Hz, 1H), 7.20 (t, J=7.6Hz, 1H), 7.13 (t, J=7.6Hz, 1H), 6.80 ( s, 1H), 6.44 (m, 2H), 6.32 (m, 1H), 5.68 (m, 1H), 5.00 (p, J=6.2Hz, 1H), 3.94 (s, 3H), 3.88 (s, 3 H), 3.06 (s, 2H), 2.66 (s, 3H), 2.61 (m, 3H), 2.36 (s, 3H), 1.67 (s, 2H), 1.03 (d, J=6.4Hz, 6H).

[0095] With reference to Example 1, three deuterated products of Examples 2, 3 and 4 were synthesized, as shown in Table 1 below.

[0096] Table 1 Structure and characterization of compounds of Examples 2-4

[0097]

[0098]

[0099] Example 5

[0100] The compound used as a kinase inhibitor in this example has the following structural formula:

[0101]

[0102] The synthetic route of the compound of this embodiment is as follows:

[0103]

[0104] The synthesis process is as follows:

[0105] Synthesis of compound 2: In a 2000mL single-mouth bottle, add compound 1 (54.6g, 303mmol), acetylhydrazine (26.64g, 395mmol), add 1000mL of 1N sodium hydroxide aqueous solution, react at 80°C for 4 hours, and a large amount of solid precipitates. Cool down, add 80mL of concentrated hydrochloric acid, stir at about 0°C for 30 minutes, filter, wash the filter cake with water once, and vacuum dry at 55°C overnight to obtain 32g of product. 1 H NMR (400 MHz, DMSO-d 6 ) δ11.75 (s, 1H), 11.59 (s, 1H), 8.13 (s, 1H), 2.52 (s, 3H).

[0106] Synthesis of compound 3: Add compound 2 (50.0 g, 258 mmol) to a 2000 mL single-mouth bottle, add 1500 mL of toluene, phosphorus oxychloride (237 g, 1550 mmol), N,N-diisopropylethylamine (133 g, 1031 mmol) at room temperature, and white mist is generated. Heat to 80°C, protect with nitrogen, and stir overnight. The next day, cool down to obtain 30 g of product. 1 H NMR (400MHz, Chloroform-d) δ 9.19 (s, 1H), 2.71 (s, 3H).

[0107] Synthesis of compound 5: Add compound 3 (30.0 g, 130 mmol) to a 2000 mL single-mouth bottle, dissolve it in 1500 mL 1,2-dichloroethane, add anhydrous aluminum chloride (29.3 g, 220 mmol), and stir at room temperature for 30 minutes. Cool down to 0°C, add compound 4 (22.1 g, 169 mmol) dropwise, stir at low temperature for 30 minutes, then heat up to 60°C for reaction, and monitor with a plate. After reacting for 4 hours, cool down and treat to obtain 20.0 g of the product. 1 H NMR (400MHz, Chloroform-d) δ 8.84 (s, 1H), 8.02 (dt, J=7.8, 1.0Hz, 1H), 7.97 (s, 1H), 7.40-7.27 (m, 3H), 3.86 (s, 3H), 2.50 (s, 3H).

[0108] Synthesis of compound 7: Compound 5 (2 g, 6.15 mmol), compound 6 (1.37 g, 7.38 mmol), and p-toluenesulfonic acid (2.11 g, 12.3 mmol) were added to 100 mL of dioxane, and the temperature was raised to 80°C under nitrogen protection, and the reaction was allowed to proceed overnight. The reaction was monitored by spot plate, and 0.65 g of product was obtained after treatment.

[0109] Synthesis of compound 9: Compound 7 (950 mg, 2 mmol), compound 8 (1.0 g, 8 mmol) and N,N-diisopropylethylamine (516 mg, 4 mmol) were added to 20 mL of N,N-dimethylacetamide and reacted at 100°C overnight. The plate was monitored and after the reaction of the raw materials was completed, 1.0 g of the product was obtained. 1H NMR (400MHz, Chloroform-d) δ9.56 (s, 1H), 8.88 (s, 1H), 7.92 (d, J = 14.6Hz, 2H), 7.54 (s, 1H), 7.40 (d, J = 8.2Hz, 1H), 7.32 (s, 1H), 7.15 (t, J = 7.6Hz, 1H), 6.84 (s, 1H), 4.06 (s, 3H), 3.93 (s, 3H), 3.04 (t, J=6.0Hz, 1H), 2.88 (s, 3H), 2.72 (s, 3H), 2.44 (s, 2H), 2.38 (s, 3H), 1.28 (s, 4H).

[0110] Synthesis of compound 10: Compound 9 (1.0 g, 1.72 mmol) and palladium carbon (500 mg, wet palladium carbon 55%) were added to 50 mL of methanol and reacted at room temperature for 4 hours under a hydrogen atmosphere. The plate was monitored and after the reaction of the raw materials was completed, 900 mg of the product was obtained.

[0111] Synthesis of Example 5: Compound 10 (900 mg, 1.63 mmol) and triethylamine (495 mg, 4.90 mmol) were dissolved in dichloromethane, cooled to 0°C under nitrogen atmosphere, and then a dichloromethane solution of acryloyl chloride (223 mg, 2.45 mmol) was added dropwise, and the reaction was maintained at 0°C for 2 hours. The plate was monitored, and after the raw materials reacted, they were treated to obtain 200 mg of the product. [M+H] + :606.8.

[0112] With reference to Example 5, three deuterated products of Examples 6, 7 and 8 were synthesized, as shown in Table 2 below.

[0113] Table 2 Structure and characterization of compounds of Examples 6-8

[0114]

[0115] Example 9

[0116] The compound used as a kinase inhibitor in this example has the following structural formula:

[0117]

[0118] The synthetic route of the compound of this embodiment is as follows:

[0119]

[0120] The synthesis process of the compound of this embodiment is as follows:

[0121] Synthesis of compound 3: Compound 1 (6.0 g, 13.30 mmol), compound 2 (5.03 g, 39.91 mmol) and N,N-diisopropylethylamine (3.43 g, 26.6 mmol) were added to 60 mL of N,N-dimethylacetamide and reacted at 80°C overnight. The plate was monitored and after the reaction of the raw materials was completed, 6.17 g of product was obtained. 1 H NMR (400MHz, Chloroform-d)-δ9.50 (s, 1H), 8.90 (s, 1H), 8.24-7.99 (m, 1H), 7.8 0 (s, 1H), 7.67 (s, 1H), 7.38 (dt, J=8.2, 1.0Hz, 1H), 7.31-7.23 (m, 1H), 7.23-7.16 (m, 1H), 6.65 (s, 1H), 3.95 (s, 3H), 3.94 (s, 3H), 3.70 (s, 3H), 2.95 (s, 3H), 2.89 ( t, J=6.6Hz, 1H), 2.85-2.76 (m, 2H), 2.27 (m, 2H), 2.15 (s, 3H), 1.91-1.81 (m, 2H).

[0122] Synthesis of compound 4: Compound 3 (6.17 g, 11.08 mmol) and lithium hydroxide monohydrate (2.33 g, 55.39 mmol) were added to a mixed solvent of tetrahydrofuran / methanol / water with a volume ratio of 6:3:1, and reacted at 40°C for 16 hours. The raw materials were monitored by spot plate, and after the reaction was completed, 6.02 g of product was obtained.

[0123] Synthesis of compound 5: Compound 4 (6.02 g, 11.08 mmol), tert-butyl carbazate (4.39 g, 33.26 mmol), N, N-diisopropylethylamine (8.58 g, 66.48 mmol) were added to 160 mL of N, N-dimethylformamide, stirred for 15 min, and then 1H-benzotriazol-1-yloxytripyrrolidino hexafluorophosphate (6.90 g, 13.30 mmol) was added, and the mixture was reacted at 25° C. for 16 hours. The reaction was monitored by spot plate, and after the reaction of the raw materials was completed, 1.67 g of product was obtained.

[0124] Synthesis of compound 6: Compound 5 (1.67 g, 2.54 mmol) was dissolved in 15 mL of tetrahydrofuran, slowly added to 15 mL of 4 M hydrogen chloride dioxane solution, and reacted at 40°C for 5 hours under nitrogen protection. Liquid quality monitoring, after the raw material reacted, treated to obtain 1.20 g of product.

[0125] Synthesis of compound 7: Compound 6 (1.0 g, 1.68 mmol) and triethylamine (679 mg, 6.72 mmol) were added to 20 mL of dichloromethane, stirred for 15 minutes, and trifluoroacetic anhydride (1.59 g, 7.58 mmol) was added in batches, and the reaction was completed at 40°C for 8 hours. 521 mg of the product was obtained.

[0126] Synthesis of compound 8: Compound 7 (521 mg, 0.82 mmol) and palladium carbon (156 mg, 10%) were dissolved in 20 mL of methanol. The mixture was stirred at 25°C for 3 hours under hydrogen and the liquid quality was monitored. After the reaction of the raw materials was completed, the mixture was treated to obtain 412 g of product.

[0127] Synthesis of Example 9: Compound 8 (412 mg, 0.68 mmol) and triethylamine (206 mg, 2.04 mmol) were dissolved in 15 mL of dichloromethane, cooled to 0°C under nitrogen, and then a dichloromethane solution of acryloyl chloride (93 mg, 1.02 mmol) was added dropwise, and the reaction was maintained at 0°C for 2 hours. The plate was monitored, and after the reaction of the raw materials was completed, the product was treated to obtain 122 mg. [M+H] + : 660.5; 1 H NMR (400MHz, Chloroform-d) δ 11.36 (m, 1H), 9.87 (m, 1H), 9.09 (m, 1H), 8.92 (s, 1H), 8.37 (m, 1H), 7.95 (m, 1H), 7.33 (m, 1H), 7.18 (m, 1H), 7.01 (m , 2H), 6.80 (m, 1H), 6.46 (d, J=16.6Hz, 1H), 5.74 (d, J=10.2Hz, 1H), 3.92 (d, J=13.5Hz, 6H), 3.29 (m, 2H), 3.04 (m, 1H), 2.78 (m, 8H), 2.21 (m, 2H).

[0128] With reference to Example 9, three deuterated products of Examples 10, 11 and 12 were synthesized, as shown in Table 3 below.

[0129] Table 3 Structure and characterization of compounds of Examples 10-12

[0130]

[0131] Embodiment 13

[0132] The compound used as a kinase inhibitor in this example has the following structural formula:

[0133]

[0134] The synthetic route of the compound of this embodiment is as follows:

[0135]

[0136] Synthesis of compound 2: Add compound 1 (10.00 g, 62.88 mmol) to a 500 mL single-mouth bottle, dissolve it in 100 mL tetrahydrofuran, add cesium carbonate (20.50 g, 62.88 mmol) and trifluoroethanol (6.29 g, 62.87 mmol), add them, and protect them with nitrogen. Monitor with a spot plate, react at 23 ° C for 6 hours until the reaction is basically completed. After treatment, 14.35 g of the product was obtained, with a yield of 95.5%. 1 H NMR (400MHz, Chloroform-d) δ 8.01 (dd, J=9.1, 5.8Hz, 1H), 7.09-6.72 (m, 2H), 4.50 (q, J=7.8Hz, 2H).

[0137] Synthesis of compound 3: Compound 2 (14.00 g, 58.57 mmol) was dissolved in 60 mL of ethanol, and 15 mL of water, ammonium chloride (9.60 g, 179.44 mmol), and reduced iron powder (20.00 g, 357.14 mmol) were added. After the addition was completed, the temperature was raised to 80°C and the reaction was allowed to proceed overnight. The reaction was monitored by a plate. After the reaction was completed, the product was treated to obtain 10.50 g, with a yield of 85.78%. 1 H NMR (400MHz, Chloroform-d) δ 6.89-6.34 (m, 3H), 4.34 (q, J = 8.0Hz, 2H), 3.84-3.27 (m, 2H).

[0138] Synthesis of compound 4: Add compound 3 (10.00 g, 47.83 mmol) to a 250 mL three-necked flask, add 50 mL of concentrated sulfuric acid under stirring, cool to 0°C, add potassium nitrate solid (6.10 g, 60.34 mmol) in batches, and after addition, protect with nitrogen. After reacting at room temperature for 4 hours, the spot reaction is complete, and the product is treated to obtain 8.26 g, with a yield of 67.98%. 1 H NMR (400MHz, Chloroform-d) δ7.44 (d, J=7.2Hz, 1H), 6.66 (d, J=11.5Hz, 1H), 4.46 (q, J=7.7Hz, 2H), 4.01 (m, 2H).

[0139] Synthesis of compound 6: Compound 4 (2.00 g, 6.08 mmol) and compound 5 (2.00 g, 7.87 mmol) were added to 50 mL of acetonitrile, and then p-toluenesulfonic acid monohydrate (0.81 g, 4.26 mmol) was added, and stirred at 80°C overnight. The next day, the plate was monitored and the reaction of the raw materials was basically completed. The temperature was lowered, and solids precipitated in the reaction solution, which was filtered. The filter cake was spin-dried and methanol was added to slurry to obtain 1.67 g of product. 1 H NMR (400MHz, Chloroform-d) δ9.89 (s, 1H), 9.33 (d, J=8.0Hz, 1H), 8.77 (s, 1H), 8.29 (s, 1H), 7.67 (d, J=8.0Hz, 1H), 7.39 (m, 1H), 7 .33 (m, 1H), 7.19 (m, 1H), 6.89 (d, J=11.3Hz, 1H), 5.14 (p, J=6.2Hz, 1H), 4.59 (q, J=7.7Hz, 2H), 3.95 (s, 3H), 1.22 (d, J=6.2Hz, 6H).

[0140] Synthesis of compound 8: Compound 6 (1.52 g, 2.78 mmol), compound 7 (1.05 g, 8.33 mmol) and N,N-diisopropylethylamine (1.05 g, 8.14 mmol) were added to 25 mL of N,N-dimethylacetamide and reacted at 90°C overnight. The plate was monitored and after the reaction of the raw materials was completed, 1.16 g of product was obtained.

[0141] Synthesis of compound 9: Compound 8 (0.59 g, 0.90 mmol) and palladium carbon (400 mg, wet palladium carbon 55%) were added to a mixed solvent of 20 mL methanol and 20 mL ethyl acetate, and reacted at room temperature for 3 hours under a hydrogen atmosphere. The plate was monitored and after the reaction of the raw materials was completed, 520 mg of the product was obtained.

[0142] Synthesis of Example 13: Compound 9 (520 mg, 0.83 mmol) and triethylamine (253 mg, 2.50 mmol) were dissolved in dichloromethane, cooled to 0°C under nitrogen atmosphere, and then a dichloromethane solution of acryloyl chloride (114 mg, 1.25 mmol) was added dropwise, and the reaction was maintained at 0°C for 2 hours. The plate was monitored, and after the reaction of the raw materials was completed, 170 mg of the product was obtained. [M+H] + :606.8. 1HNMR (400MHz, Chloroform-d) δ10.63 (s, 1H), 10.11 (s, 1H), 8.82 (s, 1H), 8.16 (s, 1H), 7.6 6(m, 2H), 7.29(d, J=8.1Hz, 1H), 7.22-7.16(m, 1H), 7.15-7.09(m, 1H), 6.80(s, 1H), 6.37(d d, J=16.9, 2.0Hz, 1H), 5.68 (dd, J=10.1, 2.0Hz, 1H), 5.00 (p, J=6.2Hz, 1H), 4.44 (q, J=8.2H z, 2H), 3.85 (s, 3H), 3.27-2.99 (m, 2H), 2.87-2.51 (m, 6H), 2.11 (m, 2H), 1.18-0.91 (m, 6H).

[0143] Referring to Example 13, Example 14 was synthesized, as shown in Table 4 below.

[0144] Table 4 Structure and characterization of compounds of Example 14

[0145]

[0146]

[0147] Embodiment 15

[0148] The compound used as a kinase inhibitor in this example has the following structural formula:

[0149]

[0150] The synthetic route of the compound of this embodiment is as follows:

[0151]

[0152] Synthesis of Example 15: In a 50mL single-mouth bottle, add compound 2 (112mg, 1.24mmol), 20ml of dichloromethane, and oxalyl chloride (142mg, 1.12mmol), and stir for 2 hours under nitrogen protection. Cool to 0°C, add a dichloromethane solution of compound 1 (438mg, 78.92mmol) and triethylamine (152mg, 1.50mmol) dropwise, keep 0°C for 1 hour, monitor with a plate, and after the raw materials react, treat to finally obtain 160mg of the product. [M+H] + : 628.8; 1H NMR (400MHz, Chloroform-d) δ9.53 (m, 1H), 9.12 (m, 1H), 8.89 (s, 1H), 8.35 (m, 1H), 7 .92 (s, 1H), 7.69 (s, 1H), 7.35 (d, J = 8.1Hz, 1H), 7.23 (t, J = 7.5Hz, 1H), 7.15 (t, J = 7.5 Hz, 1H), 6.84 (s, 1H), 5.84 (dd, J=48.2, 3.4Hz, 1H), 5.28 (dd, J=15.5, 3.4Hz, 1H), 5.0 4(p, J=6.2Hz, 1H), 3.92(s, 6H), 3.49(m, 2H), 2.64(m, 9H), 2.19(m, 2H), 1.09(m, 6H).

[0153] Example 16

[0154] The compound used as a kinase inhibitor in this example has the following structural formula:

[0155]

[0156] The synthetic route of the compound of this embodiment is as follows:

[0157]

[0158] Synthesis of compound 3: Add compound 1 (313 mg, 1 mmol) and 5 mL of acetonitrile to a 100 mL single-mouth bottle, then add the crude product of compound 2 (441 mg). After the addition, nitrogen was used for protection. The plate was monitored and the reaction was basically completed at 80°C overnight. After treatment, 300 mg of the product was obtained.

[0159] Synthesis of compound 4: Compound 3 (1.83 g, 4.54 mmol) was dissolved in 60 mL of methanol, and 1.2 mL of concentrated hydrochloric acid was added. After the addition was complete, the temperature was raised to 60°C and the reaction was allowed to proceed overnight. The reaction was monitored by a plate. After the reaction was completed, the product was treated to obtain 0.89 g. 1 H NMR (400MHz, Chloroform-d) δ7.61 (s, 1H), 4.80 (q, J=8.4Hz, 2H), 3.57 (m, 2H), 3.1 0 (s, 3H), 3.02 (t, J=6.9Hz, 1H), 2.59-2.33 (m, 4H), 2.06 (s, 3H), 1.76-1.51 (m, 2H).

[0160] Synthesis of compound 6: Add compound 4 (73.8 mg, 0.21 mmol) to a 100 mL single-mouth bottle, add compound 5 (56 mg, 0.17 mmol), p-toluenesulfonic acid (48 mg, 0.25 mmol), and then add 5 mL of dioxane, stir at 80°C overnight, and protect with nitrogen. Monitor with a plate, and after the reaction is complete, treat to obtain 50 mg of the product. 1 H NMR (400MHz, Chloroform-d) δ9.76 (s, 1H), 8.90 (s, 1H), 7.94 (s, 1H), 7.80 (d, J=8.0Hz, 1H), 7.56 (s, 1H), 7.39 (d, J=8.2Hz, 1H), 7.33-7.27 (m, 1H), 7.24-7.1 7(m, 1H), 5.10(p, J=6.2Hz, 1H), 4.94(s, 2H), 3.93(s, 3H), 3.78(m, 2H), 3.18( t, J=6.5Hz, 1H), 3.01 (m, 3H), 2.57 (m, 5H), 2.31 (m, 2H), 1.16 (d, J=6.2Hz, 6H).

[0161] Synthesis of compound 7: Compound 6 (98 mg, 0.15 mmol) and palladium carbon (80 mg, wet palladium carbon 55%) were added to a mixed solvent of 3 mL methanol and 2 mL ethyl acetate, and reacted at room temperature for 2 hours under a hydrogen atmosphere. The plate was monitored and after the reaction of the raw materials was completed, 98 mg of the product was obtained.

[0162] Synthesis of Example 16: Compound 8 (300 mg, 0.48 mmol) and triethylamine (150 mg, 1.48 mmol) were dissolved in dichloromethane, cooled to 0°C under nitrogen atmosphere, and then a dichloromethane solution of acryloyl chloride (70 mg, 0.77 mmol) was added dropwise, and the mixture was kept at 0°C for 2 hours. The plate was monitored, and after the reaction of the raw materials was completed, the mixture was treated to obtain 118 mg of the product. [M+H] + :688.8. 1H NMR (400MHz, Chloroform-d) δ10.39 (s, 1H), 9.97 (s, 1H), 8.89 (s, 1H), 8.59 (m, 1H) , 7.59(m, 2H), 7.33(m, 1H), 7.23(m, 1H), 7.14(m, 1H), 6.45(m, 1H), 6.31(m, 1H), 5.7 2(d, J=10.0Hz, 1H), 5.01 (p, J=6.5Hz, 1H), 4.83 (q, J=8.5Hz, 2H), 3.96 (s, 3H), 3.1 4-2.88 (m, 2H), 2.85-2.52 (m, 6H), 2.33 (s, 3H), 1.69 (m, 2H), 1.04 (d, J=6.3Hz, 6H).

[0163] Embodiment 17

[0164] The compound used as a kinase inhibitor in this example has the following structural formula:

[0165]

[0166] The synthetic route of the compound of this embodiment is as follows:

[0167]

[0168] Synthesis of compound 2: Add compound 1 (153 mg, 0.38 mmol) to a 100 mL three-necked flask, add 5 mL of tetrahydrofuran, and then add 4-dimethylaminopyridine (17.8 mg). After the addition is complete, add di-tert-butyl dicarbonate (1.06 g, 4.86 mmol) dropwise under nitrogen protection. Monitor with a plate, react at 80°C for 2 hours, and the reaction is basically complete. After treatment, 104 mg of the product is obtained.

[0169] Synthesis of compound 3: Compound 2 (104 mg, 0.21 mmol) was dissolved in 2 ml of methanol, and 0.05 mL of 5.4 mol / L sodium methoxide methanol solution was added. After addition, the reaction was allowed to proceed for 30 minutes. The reaction was monitored by a spot plate. After completion of the reaction, the product was treated to obtain 30 mg.

[0170] Synthesis of compound 4: Compound 3 (30 mg, 0.065 mmol) and palladium carbon (15 mg, wet palladium carbon 55%) were added to a mixed solvent of 1 mL methanol and 1 mL ethyl acetate, and reacted at room temperature for 2 hours under a hydrogen atmosphere. The plate was monitored and after the reaction of the raw materials was completed, 30 mg of the product was obtained.

[0171] Synthesis of compound 5: Compound 4 (30 mg, 0.069 mmol) and triethylamine (13 mg, 0.14 mmol) were dissolved in dichloromethane, cooled to 0°C under nitrogen atmosphere, and then a dichloromethane solution of acryloyl chloride (7 mg, 0.077 mmol) was added dropwise, and the reaction was maintained at 0°C for 2 hours. The plate was monitored, and after the reaction of the raw materials was completed, 12 mg of the product was obtained.

[0172] Synthesis of Example 17: Compound 5 (12 mg, 0.024 mmol) and compound 6 (6 mg, 0.018 mmol) and p-toluenesulfonic acid (8 mg, 0.042 mmol) were dissolved in 1 mL of N-methylpyrrolidone and 2 mL of ethylene glycol monomethyl ether, and the temperature was raised to 100°C under a nitrogen atmosphere for 4 hours. The plate was monitored and after the reaction of the raw materials was completed, 3 mg of the product was obtained. [M+H] + :675.8. 1 HNMR (400MHz, Chloroform-d) δ10.42 (s, 0H), 9.97 (s, 0H), 8.91 (s, 0H), 8.70 ( m, 1H), 7.68 (s, 1H), 7.33 (d, J=8.2Hz, 0H), 7.18 (m, 1H), 6.99 (m, 1H), 6.45 (m, 1H), 6.33(m, 1H), 5.73(m, 1H), 5.35(m, 1H), 4.85(m, 2H), 3.99(s, 3H), 3.05(m , 2H), 2.78(m, 1H), 2.62(m, 2H), 2.35(s, 3H), 2.22(s, 3H), 2.10-1.95(m, 2H).

[0173] 2. Biological test evaluation of compounds

[0174] 1. Test of the inhibitory effect of compounds on cell proliferation

[0175] 1.1 Experimental study on the cell proliferation inhibition activity of EGFR and HER2 exon 20 insertion mutations:

[0176] The engineered cells selected in the experiment, Ba / F3-FL-EGFR-V769-D770 ins ASV, Ba / F3-FL-EGFR-D770-N771 ins SVD, Ba / F3-FL-EGFR-H773-V774 ins NPH, Ba / F3-FL-EGFR-A763-Y764 insFQEA, and Ba / F3-HER2-A775-G776 ins YVMA, were provided by Hefei Zhongke Puruisheng Biotechnology Co., Ltd. and were verified before being used in this study.

[0177] Prepare 20× stock solution of the compound to be tested for use, 9 concentrations, 3-fold gradient dilution, starting from 1 μM.

[0178] Take the cell suspension in logarithmic growth phase and inoculate it into a 96-well white cell culture plate, with a volume of 95 μL per well (2000 cells / well). Take 5 μL of 20× test compound and add it to the culture plate containing 95 μL cell suspension according to the plating diagram, and mix well. 37°C, 5% CO 2 Incubate in an incubator for 72 hours. CellTiter-Glo method was used to determine the proliferation inhibitory activity of the compound. SpectraMax Paradigm readings were used to obtain the corresponding fluorescence value RLU per well. The cell proliferation inhibition rate (InhibitionRate) data were processed using the following formula: Inhibition Rate (Inh%) = 100-(RLU compound-RLUblank) / (RLU control-RLU blank)*100%. The cell viability corresponding to different concentrations of the compound was calculated in EXCEL, and then the cell viability curve was drawn using GraphPad Prism software and the IC was calculated. 50 Table 5 shows the names and structures of the reference substances.

[0179] Table 5 Name and structure of reference substances

[0180]

[0181] In the experimental results of Table 6, Table 7 and Table 8, according to IC 50 The values ​​are divided into three categories, A≤30nM, 30nM<B≤100nM, and C>100nM.

[0182] Table 6 Test results of cell proliferation inhibition activity of EGFR exon 20 insertion mutations of different compounds

[0183]

[0184]

[0185] Table 7 Test results of cell proliferation inhibition activity of HER2 exon 20 insertion mutation of different compounds

[0186]

[0187] As can be seen from Table 6, the compounds in the examples all have good inhibitory ability against EGFR exon 20 insertion mutations. They are better than DZD9008 and comparable to the activity of TAK-788. As can be seen from Table 7, the cell activity of Example 5 in HER2 exon 20 insertion mutations is comparable to that of TAK-788 and better than that of DZD9008.

[0188] 1.2 Experimental test of the inhibitory activity of compounds on cell proliferation of cell lines NCI-H1975 and PC9:

[0189] The cell lines NCI-H1975 and PC9 selected in the experiment were provided by Sino-US Crown Biotechnology (Beijing) Co., Ltd. and were verified before being used in this study.

[0190] Prepare 10× stock solution of the compound to be tested for use, 9 concentrations, 4-fold gradient dilution, starting from 10 μM.

[0191] Take the cell suspension in logarithmic growth phase and inoculate it into 96-well culture plates, with a volume of 90 μL per well (2000 cells / well). Take 10 μL of 10× test compound and add it to the culture plates according to the plating diagram and mix well. 37°C, 5% CO 2 The cells were incubated in an incubator. The proliferation inhibition activity of the compounds was determined by the CellTiter-Glo method. The cell proliferation inhibition rate (Inhibition Rate) data was plotted using GraphPad Prism 8.0 software to plot the cell viability curve and calculate the IC 50 The results are shown in Table 8:

[0192] Table 8 Test results of cell proliferation inhibition activity of different compound-related mutations

[0193] Compound DZD9008 TAK-788 Example 1 Example 5 NCI-H1975 A A A A PC9 B A A A

[0194] As can be seen from Table 8, the compounds of the present invention have a good inhibitory effect on the cell proliferation of cell lines NCI-H975 and PC9, and are superior to DZD9008.

[0195] 2. In vivo efficacy studies of compounds

[0196] 2.1 Pharmacodynamic study on lung cancer LU0387 PDX model

[0197] The lung cancer LU0387PDX model is a commonly used xenograft tumor model of Crown Science. The experimental conditions of this model are as follows: After the tumor-bearing mice were euthanized, the tumor was dissected out under sterile conditions, the tumor was cleaned, blood stains, connective tissue and necrotic parts were removed, and the tumor was cut into 2*2*2mm 3Use a transfer needle to inoculate the tumor subcutaneously on the right back of the sterilized experimental mouse. Observe the tumor growth regularly and wait until the tumor grows to an average volume of 100-200mm. 3 The mice were randomly divided into groups according to the tumor size and the weight of the mice for administration. Before the start of administration, all animals were weighed and the tumor volume was measured with a vernier caliper. The mice were randomly divided into groups according to the tumor volume to ensure that the tumor volume between different groups was similar. Each group of 3 mice was orally administered once a day, and the inoculation site was observed after administration. The tumor volume and mouse weight were measured twice a week.

[0198] The experimental design and results are shown in Table 9, wherein PO is oral administration; QD: once a day; TGI (tumor volume inhibition rate) = (1-tumor weight of treatment group / tumor weight of control group) * 100%.

[0199] Table 9 LU0387 model design and results

[0200]

[0201] In the experiment, the tumor volume changed with the number of days of drug administration. Figure 1 shown.

[0202] Depend on Figure 1 It can be seen that at the same dose in the LU0387 model, the efficacy of Example 1 is better than that of TAK-788 and DZD9008. The efficacy of Example 5 is similar to that of TAK-788. As the dose of Example 1 increases, the efficacy is also significantly enhanced.

[0203] 2.2 In vivo pharmacodynamic study of subcutaneous transplant tumor model of Ba / F3-EGFR-D770_N771 ins SVD engineered cell line

[0204] The cell line Ba / F3 EGFR D770_N771 ins SVD was cultured in RPMI1640+10% fetal bovine serum+1% double antibody at 37°C and 5% CO 2 Culture, subculture 2-3 times a week. When the cell saturation is 80-90% and the number reaches the required level, the cells are harvested. 0.2 ml (1*106 cells) of cells are subcutaneously inoculated into the right back of female nude mice aged 6-8 weeks and weighing 18-22 g. The average tumor volume reaches about 150-200 mm 3 The grouping and dosing began at 14:00. Before the start of dosing, all animals were weighed and the tumor volume was measured with a vernier caliper. The mice were randomly divided into groups according to the tumor volume to ensure that the tumor volume between different groups was similar. Each group of 3 mice was orally administered once a day, and the inoculation site was observed after dosing. The tumor volume and mouse weight were measured twice a week.

[0205] Table 10 Ba / F3 EGFR D770_N771 ins SVD model grouping

[0206]

[0207]

[0208] Example 18 and Example 40 in the table are compounds with a trans-tripentaamine structure in patent WO2021180238. The names and structures are shown in Table 11:

[0209] Table 11 Names and structures of trans-structure compounds

[0210]

[0211] In the experiment, the tumor volume changed with the number of days of drug administration. Figure 2 shown.

[0212] Depend on Figure 2 It can be seen that in the Ba / F3-EGFR-D770_N771 ins SVD cell transplantation model, at the same dose, the in vivo efficacy of Example 1 and Example 5 is far superior to the compound with a trans-tripentaamine structure in patent WO2021180238, and the tumor inhibition rate is increased by more than 1.6 times.

[0213] In summary, the compounds in this patent have a good inhibitory effect on exon 20 insertion mutations of EGFR or HER2, and also have a good inhibitory effect on cell proliferation of cell lines NCI-H1975 and PC9; for one or more combinations of EGFR exon 20 insertion mutations, HER2 exon 20 insertion mutations, EGFR exon 19 deletions, EGFR exon 20 point mutations, and EGFR exon 21 point mutations, the drug is expected to have a good therapeutic effect on related diseases.

[0214] 3. Structural confirmation of Example 1 and study of the free base crystal form.

[0215]

[0216] 1. Structure confirmation

[0217] Take 214 mg of the compound of Example 1, add 0.5 mL of dichloromethane and 0.5 mL of acetonitrile, filter the solution, put the filtrate into a sample bottle with a sealing film, pierce a hole and place it in a fume hood to evaporate slowly to obtain granular crystals (free base crystal form I). The structure was confirmed by single crystal diffraction, and the results are as follows Figure 3 As shown, the configuration of the compound was confirmed.

[0218] 2. Study on the free base crystal form of Example 1

[0219] The compound obtained in Example 1 was subjected to polymorph screening to find its potential crystal form in order to find an excellent crystal form suitable for subsequent development. The synthesized Example 1 was used as the starting material, and it was detected that the compound was a crystal with good crystallinity and an anhydrous crystal form, named free base crystal form I. Subsequently, an amorphous free base compound was prepared by dry grinding. The above two types were used as starting materials to carry out crystal screening experiments.

[0220] 2.1 Preparation and characterization of starting materials.

[0221] 2.1.1 Characterization of the free base form I of the compound of Example 1

[0222] The raw material of the free base form I of the compound of Example 1 was fully characterized. Polarized light microscopy (PLM) showed that the raw material was irregularly shaped crystals with good crystallinity. Figure 4 is the XRPD pattern of free base form I, Figure 5 The DSC and TGA superimposed graphs of the crystal form. The differential scanning calorimetry (DSC) curve has an endothermic peak at 230°C, which should be the melting peak. The thermogravimetric analysis (TGA) curve has no obvious weight loss before decomposition, indicating an anhydrous crystal form.

[0223] Table 12 X-ray powder diffraction peak data of free base crystal form I

[0224]

[0225]

[0226] 2.1.2 Solubility test of free base crystal form I of Example 1

[0227] The solubility of the free base crystal form I of compound Example 1 at room temperature was roughly determined by visual inspection in 18 solvents. The results are shown in Table 13. The free base crystal form I has high solubility only in dichloroethane and dichloromethane, and low solubility in most of the tested solvents.

[0228] Table 13 Visual observation results of solubility of free base form I of Example 1

[0229] Serial number Solvents Solubility mg / mL Serial number Solvents Solubility mg / mL 1 Dichloromethane ＞107 10 1,4-Dioxane ＜0.27 2 Ethylene dichloride 61.75-123.50 11 Methyl tert-butyl ether ＜0.33 3 N-Methylpyrrolidone 18.25-27.38 12 Ethyl acetate ＜0.29 4 acetone 2.22-4.44 13 Methanol ＜0.40 5 Dimethyl sulfoxide 2.60-2.93 14 Isopropyl acetate ＜0.30 6 Butanone 2.78-4.44 15 Cyclohexane ＜0.33 7 Tetrahydrofuran 2.55-3.19 16 water ＜0.27 8 Acetonitrile 0.93-1.01 17 Isopropyl alcohol ＜0.35 9 Ethanol ＜0.30 18 n-Heptane ＜0.30

[0230] 2.1.3 Preparation and characterization of amorphous raw materials

[0231] 736 mg of the compound of Example 1 (free base crystal form I) was weighed and dry-milled for 5 hours to successfully prepare 669.86 mg of amorphous product. XRPD detection was performed, and the results were as follows Figure 6 shown.

[0232] 2.2. Methods and parameters of physical and chemical testing instruments used in characterization

[0233] 2.2.1 X-ray powder diffraction (XRPD)

[0234] XRPD diffraction patterns were acquired by Bruker D2 Phaser. The samples to be tested were placed on a smooth silicon wafer without background for testing. The measurement parameters are shown in Table 14.

[0235] Table 14 XRPD method parameters

[0236]

[0237]

[0238] 2.2.2 Polarized light microscopy (PLM)

[0239] PLM analysis was performed using an Ott optical microscope BK-Pol. A small amount of sample was placed on a glass slide, a drop of silicone oil was added to disperse it, and then a coverslip was placed on it and observed under a microscope.

[0240] 2.2.3 Differential Scanning Calorimetry (DSC)

[0241] The DSC curve was acquired by TA Instruments DSC 250. The DSC 250 instrument test method is as follows: accurately weigh an appropriate amount of sample into a perforated aluminum crucible, heat it from 25°C to a final temperature of 300°C at a heating rate of 10°C / min, and purge it with nitrogen at a flow rate of 50mL / min.

[0242] 2.2.4 Thermogravimetric analysis (TGA)

[0243] TGA data were acquired by TA Instruments TGA 550. An appropriate amount of sample was placed in a pre-peeled aluminum crucible and heated from room temperature to 300°C at a heating rate of 10°C / min, with the balance chamber purged with 40 mL / min of nitrogen and the sample chamber purged with 25 mL / min of nitrogen.

[0244] 2.3 Study on the free base polymorph of the compound of Example 1.

[0245] The preparation methods mainly include suspension crystallization, anti-solvent precipitation, high and low temperature circulation and evaporative crystallization.

[0246] The suspension crystallization is based on the visual solubility results of the compound in Example 1. At 25°C and 50°C, the free base crystal form I and the amorphous crystal form are used as starting materials, respectively, in a selected single solvent (the solvent is selected from dichloromethane, 1,4-dioxane, dichloroethane, methyl tert-butyl ether, N-methylpyrrolidone, ethyl acetate, acetone, methanol, dimethyl sulfoxide, isopropyl acetate, butanone, cyclohexane, tetrahydrofuran, water, acetonitrile, isopropanol, ethanol, n-heptane) or a mixed solvent to prepare the crystal form. A total of two crystal forms, free base crystal form I and free base crystal form II, are obtained.

[0247] The anti-solvent precipitation was carried out according to the visual solubility results of the compound in Example 1, using dichloromethane and dichloroethane as good solvents, and then adding different anti-solvents to them respectively under stirring conditions at room temperature (~25°C) to prepare the crystal form. A total of two crystal forms were obtained, free base crystal form I and free base crystal form III.

[0248] The high and low temperature cycle is to add a solvent (the solvent is selected from dichloromethane, 1,4-dioxane, dichloroethane, methyl tert-butyl ether, N-methylpyrrolidone, ethyl acetate, acetone, methanol, dimethyl sulfoxide, isopropyl acetate, butanone, cyclohexane, tetrahydrofuran, water, acetonitrile, isopropanol, ethanol, n-heptane) to the compound, and then stir under the temperature cycle condition of 50°C to 5°C to prepare the crystal form. A total of two crystal forms, free base crystal form I and free base crystal form II, were obtained.

[0249] Evaporative crystallization is to add a solvent to the compound, filter the prepared solvent, and then drip the filtrate into the sample bottle. After the sealing film covering the bottle mouth is pierced, it is placed in a fume hood and slowly evaporated. Two crystal forms are obtained, free base crystal form I and free base crystal form IV. Free base crystal form IV is heated to 150°C to remove the solvent and then a new crystal form is prepared, which is named free base crystal form V.

[0250] 2.3.1 Preparation and characterization of free base form I

[0251] The free base crystal form I can be obtained in most solvents. The compound obtained in Example 1 is the free base crystal form I. The characterization results are as follows Figure 4 and Figure 5 shown.

[0252] 2.3.2 Preparation and characterization of free base form II

[0253] The free base crystal form II can be obtained in certain solvent systems containing ethanol. The free base crystal form II can be obtained by suspension crystallization in a single solvent ethanol using an amorphous compound as a raw material. The characterization results are as follows: Figure 7 and Figure 8As shown, the DSC curve of the free base crystal form II has two endothermic peaks at 124°C and 230°C, the TGA curve has a weight loss of 4.48% at 65-150°C, and the free base crystal form II is obtained in an ethanol solvent system. Based on the above, it can be inferred that the free base crystal form II should be an ethanol solvate. The endothermic peak at 124°C on the DSC curve should be a desolvation peak.

[0254] Table 15 X-ray powder diffraction peak data of free base crystal form II

[0255]

[0256] 2.3.3 Preparation and characterization of free base form III

[0257] The free base crystal form III can be obtained in certain solvent systems containing isopropanol. When the crystal form is prepared by anti-solvent precipitation, the free base crystal form III is found in the dichloromethane / isopropanol or dichloroethane / isopropanol system. The characterization results are as follows Fig. 9 and Fig.10 As shown, the DSC curve of the free base crystal form III has two endothermic peaks at 121°C and 230°C, the TGA curve has a weight loss of 9.57% at 75-145°C, and after the free base crystal form III is heated to 150°C for desolvation, the XRPD transforms into the free base crystal form I. The free base crystal form III is obtained in the solvent system of isopropanol. Based on the above, it can be inferred that the free base crystal form III should be a solvate of isopropanol, and the endothermic peak at 121°C on the DSC curve should be a desolvation peak.

[0258] Table 16 X-ray powder diffraction peak data of free base form III

[0259]

[0260]

[0261] 2.3.4 Preparation and characterization of free base form IV

[0262] Free base crystal form IV can be obtained in some solvent systems containing dichloroethane. Weigh 92.28 mg of raw material, add it to a sample bottle, add 5.0 mL of dichloroethane to dissolve, cover the sample bottle with a sealing film, pierce a hole and place it in a fume hood to evaporate slowly, so that crystal form IV is prepared by evaporation and crystallization. Its characterization results are as follows Fig.11 and Fig.12 As shown, the DSC curve of the free base crystal form IV has two endothermic peaks at 108°C and 231°C, and the TGA curve has a weight loss of 10.41% at 25-150°C. 1The H NMR results showed that only 6.14% of dichloroethane solvent remained, which indicated that the sample may contain 4.27% of water. Based on the above, it can be inferred that the free base crystal form IV may be a solvate of dichloroethane and water.

[0263] Table 17 X-ray powder diffraction peak data of free base crystal form IV

[0264]

[0265] 2.3.5 Preparation and characterization of free base form V

[0266] After heating to 150°C to remove the solvent, the free base crystal form IV was characterized by XRPD, showing that it was transformed into a new crystal form, named free base crystal form V. The characterization results are as follows Fig.13 and Fig.14 As shown, the DSC curve of the free base crystal form V has an endothermic peak at 231°C, and the TGA curve has no obvious weight loss before decomposition. In summary, it can be inferred that the free base crystal form V should be an anhydrous crystal form.

[0267] Table 18 X-ray powder diffraction peak data of free base form V

[0268]

[0269] 3. Crystalline evaluation of free base form I

[0270] In this screening process, only two anhydrous crystalline forms, free base form I and free base form V, were found. Saturated solutions were prepared with acetone and acetonitrile, respectively, and then competitive slurrying was performed on the two at 25°C or 50°C. From the results, it was inferred that free base form I may be the thermodynamically stable crystalline form of the compound. In addition, free base form V was not directly obtained in the screening experiment, but only obtained by desolvation of free base form IV. Therefore, only free base form I was evaluated for crystalline form, including dry grinding, wet grinding, tableting (30MPa), stability and hygroscopicity studies.

[0271] Through grinding experiments, it was found that the free base crystal form I turned into an amorphous state after 5 minutes of dry grinding; after 5 minutes of wet grinding with water, there was no obvious change in the crystallinity; after 5 minutes of wet grinding with ethanol, the crystallinity decreased slightly.

[0272] In the tableting experiment, when the pressure was 30 MPa, the crystallinity decreased slightly.

[0273] In the stability experiment, it was found that when the free base crystal form I was placed open at 80°C for 3 days, 60°C for 7 days, 25°C / 60% RH for 7 days, 40°C / 75% RH for 7 days, and 25°C / 90±5% RH for 7 days, samples were taken for XRPD detection. The results showed that the free base crystal form I did not change significantly, and the liquid phase detection results of the drug did not change.

[0274] The free base crystal form I was subjected to a dynamic vapor sorption (DVS) test, and the results showed that the free base crystal form I had a slightly hygroscopic property, with a water absorption weight gain of 0.66% at 80% RH.

[0275] In summary, the spatial configuration of this type of compound as shown in Formula 1 was determined, and in particular, the spatial structure of Example 1 was accurately determined. In the crystal screening study of the compound of Example 1, a total of 2 anhydrous crystal forms (free base crystal form I and free base crystal form V) and 3 solvent compounds (free base crystal form II, free base crystal form III and free base crystal form IV) were found. In the experiment, it was found that free base crystal form I is a relatively stable anhydrous crystal form with stable solid-state properties and slightly hygroscopic properties, which can be used for subsequent drug development.

Claims

1. A class of compounds useful as kinase inhibitors, It is characterized in that The compound used as a kinase inhibitor is a compound as shown in Formula 1, or a pharmaceutically acceptable salt thereof: In Formula 1, X is selected from CH, N; R 1 Selected from R 5 is H, C1-C3 alkyl, C1-C3 fluoroalkyl; R 20 , R 21 , R 22 Each is independently selected from methyl or deuterated methyl; R 3 Selected from C1-C3 alkyl, C1-C3 haloalkyl; R 40 , R 41 , R 42 Each is independently selected from H, D, and F.

2. The compound for use as a kinase inhibitor according to claim 1, It is characterized in that It is a compound as shown in Formula 2, or a pharmaceutically acceptable salt:

3. The compound for use as a kinase inhibitor as claimed in claim 2, It is characterized in that In Formula 2, X is selected from CH, N; R 3 Selected from -CH 3 、-CH 2 CH 3 、-CH 2 CF 3 ; R 40 , R 41 Both H, R 42 Select from H or F.

4. The compound for use as a kinase inhibitor according to claim 1, It is characterized in that It is a compound as shown in Formula 3, or a pharmaceutically acceptable salt:

5. The compound for use as a kinase inhibitor as claimed in claim 4, It is characterized in that In Formula 3, X is selected from CH, N; R 3 -CH 3 、-CH 2 CH 3 、-CH 2 CF 3 ; R 40 , R 41 Both H, R 42 Select from H or F, R 5 Selected from -CH 3 , -CF 3 .

6. The compound for use as a kinase inhibitor according to claim 1, It is characterized in that The compound shown in formula 1 is:

7. A compound for use as a kinase inhibitor according to any one of claims 1 to 6, It is characterized in that The compound is crystalline and amorphous.

8. The compound for use as a kinase inhibitor as claimed in claim 7, It is characterized in that The compound whose structure is shown in formula A: The crystal form is free base crystal form I, and the X-ray powder diffraction pattern of the crystal form has characteristic diffraction peaks at 2θ of 9.76°±0.2°, 10.45°±0.2°, 16.54°±0.2°, 18.66°±0.2°, 20.07°±0.2°, and 25.90°±0.2°.

9. The compound for use as a kinase inhibitor as claimed in claim 8, It is characterized in that The X-ray powder diffraction pattern of the free base form I has characteristic diffraction peaks at 2θ of 9.07°±0.2°, 9.76°±0.2°, 10.45°±0.2°, 11.53°±0.2°, 11.80°±0.2°, 12.91°±0.2°, 13.79°±0.2°, 14.67°±0.2°, 15.08°±0.2°, 15.63°±0.2°, 16.54°±0.2°, 17.50°±0.2°, 18.66°±0.2°, 20.07°±0.2°, 21.10°±0.2°, 23.29°±0.2°, 24.16°±0.2° and 25.90°±0.2°.

10. Use of the compound used as a kinase inhibitor according to any one of claims 1 to 9 in the preparation of a medicament for treating diseases related to EGFR mutation and / or HER2 mutation.

11. The use according to claim 10, It is characterized in that The EGFR mutation and / or HER2 mutation includes one or a combination of two or more of EGFR exon 20 insertion mutation, HER2 exon 20 insertion mutation, EGFR exon 19 deletion, EGFR exon 21 point mutation, and EGFR exon 20 point mutation.

12. The use according to claim 11, It is characterized in that The EGFR mutation and / or HER2 mutation is selected from EGFRDel 19 / T790M mutation and EGFR L858R / T790M mutation.

13. The use according to claim 10, 11 or 12, It is characterized in that The disease is cancer caused by the EGFR mutation and / or HER2 mutation.

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