1H-indazole compound as well as preparation method and application thereof

Abstract

The invention relates to a 1H-indazole compound. The structural formula of the compound is as shown in formula I in the specification. The invention provides a method for preparing the compound and application of the compound in medicines. The 1H-indazole compound provided by the invention can well inhibit the growth of medullary mother cells, and the proliferation of the medullary mother cells can cause the children brain tumor, so that the structural formula I is expected to be further developed into the medicine for treating the children brain tumor. # imgabs0 #

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparation, and in particular relates to a 1H-indazole compound and a preparation method and application thereof. Background Art

[0002] Medulloblastoma (MB) is the most common malignant brain tumor in children, accounting for about 30% of all brain tumors in children. It is highly invasive and difficult to treat, and nearly 35% of patients are considered incurable. Even if the treatment is successful, children may experience serious long-term disability and health problems due to the adverse side effects of radiotherapy and chemotherapy, which will leave a variety of sequelae, including cognitive deficits and endocrine disorders.

[0003]

[0004] MK-886 developed by Merck and Vismodegib developed by Roche showed good effects in inhibiting medulloblastoma in the initial clinical trial stage, but soon developed drug resistance. More importantly, more than half of the patients experienced adverse reactions such as weight loss, fatigue and nausea, muscle cramps, hair loss, loss of taste and joint pain after taking the drugs, leading to serious toxic side effects. Due to these reasons, these two drugs were discontinued in clinical trials and were not approved for the treatment of medulloblastoma, so people are urgently in need of finding new drugs to treat medulloblastoma. Summary of the invention

[0005] The present invention aims to provide a 1H-indazole compound and its preparation method and application, so as to solve the problem that there is a lack of effective drugs for treating medulloblastoma in the market. The 1H-indazole compound provided by the present invention can well inhibit the growth of medulloblastoma and is expected to be developed into a drug for treating medulloblastoma-related diseases.

[0006] In order to achieve the above object, the present invention provides the following technical solutions:

[0007] In one aspect, the present invention provides a 1H-indazole compound, the structural formula of which is shown in Formula I

[0008]

[0009] On the other hand, the present invention also provides a method for preparing a 1H-indazole compound, the method comprising:

[0010] (a) Synthesis of Compound 2: The compound 6-bromo-1H-indazole-4-carboxylic acid methyl ester 1 is subjected to a substitution reaction with isopropyl bromide to obtain the compound 2 shown. The reaction formula is as follows:

[0011]

[0012] The reaction solvent includes but is not limited to DMF, isopropanol, tetrahydrofuran and acetonitrile, and the base includes but is not limited to sodium hydrogen sulfide, potassium carbonate, sodium carbonate, cesium carbonate, potassium tert-butoxide and sodium tert-butoxide, and the reaction temperature can be between 0°C and 60°C;

[0013] (b) Synthesis of compound 3: Compound 2 obtained in step (a) is subjected to esterification reaction with sodium hydroxide solution to obtain compound 3 as shown. The reaction formula is as follows:

[0014]

[0015] The base used for esterification includes but is not limited to sodium hydroxide, potassium hydroxide, potassium carbonate and sodium carbonate, and the reaction temperature can be between 20°C and 60°C;

[0016] (c) Synthesis of compound 4: Compound 3 obtained in step (b) is subjected to a condensation reaction with 3-(aminomethyl)-4,6-dimethylpyridin-2(1H)-one hydrochloride to obtain compound 4 as shown. The reaction formula is as follows:

[0017]

[0018] The reaction condensation agent includes but is not limited to HATU, HOBT, EDCI, BOP and DCC, the reaction solvent includes but is not limited to DMSO, DMF, DCM, acetonitrile, n-butanol, dioxane and tetrahydrofuran, and the base includes but is not limited to triethylamine, diethylamine, diisopropylethylamine, N-methylmorpholine (NMM), potassium carbonate and sodium carbonate.

[0019] (d) Synthesis of compound 5: Compound 4 obtained in step (c) is subjected to coupling reaction with 3-aminophenyl borate to obtain compound 5. The reaction formula is as follows:

[0020]

[0021] The coupling reaction reagent may be boric acid in addition to borate esters, the catalyst used includes but is not limited to tetrakistriphenylphosphine palladium, tert-butylphosphine palladium, PdCl2(dppf), Pd2(dba)3 and palladium acetate, the base used includes but is not limited to sodium carbonate, potassium carbonate, sodium bicarbonate, etc., the reaction solvent includes but is not limited to dioxane-water, DMF-water, ethanol-water, ethylene glycol dimethyl ether-water and toluene-water, etc.

[0022] (e) Synthesis of target compound I: Compound 5 obtained in step (d) is subjected to condensation reaction with 2-chloro-4-methylsulfonylbenzoic acid to obtain target compound I. The reaction formula is as follows:

[0023]

[0024] The reaction condensation agent includes but is not limited to HOBT, EDCI, BOP and DCC, the reaction solvent includes but is not limited to DMSO, DMF, DCM and dichloroethane, and the base includes but is not limited to triethylamine, diethylamine, diisopropylethylamine and N-methylmorpholine (NMM).

[0025] The present invention also provides a use of a 1H-indazole compound or a pharmaceutically acceptable salt, solvate or drug complex thereof in the preparation of a drug for treating diseases associated with medulloblastoma.

[0026] Beneficial effects:

[0027] The 6-(3-(2-chloro-4-(methylsulfonyl)benzamide)phenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (compound structural formula I) provided in the present application is a new compound. According to biological tests, it can effectively inhibit the growth of myeloid blasts, and the proliferation of myeloid blasts can cause brain tumors in children. Therefore, a 1H-indazole compound I is expected to be further developed into a drug for treating brain tumors in children. DETAILED DESCRIPTION

[0028] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0029] The experimental methods described in the following examples are conventional methods unless otherwise specified; the reagents and materials described are commercially available unless otherwise specified.

[0030] Embodiment 1:

[0031]

[0032]

[0033] (a) Synthesis of 6-bromo-1-isopropyl-1H-indazole-4-carboxylic acid methyl ester (Compound 2)

[0034] 6-Bromo-1H-indazole-4-carboxylic acid methyl ester (3.5 g, 13.7 mmol) was dissolved in DMF (30 mL). Under ice-water cooling, NaH (0.65 g, 60%, 16.2 mmol) was slowly added. The mixture was stirred for half an hour. 2-Bromopropane (3.35 g, 27.3 mmol) was added. The mixture was warmed to room temperature and stirred overnight. Water (30 mL) was slowly added. The mixture was extracted with ethyl acetate (2x200 mL), washed with water (2x50 mL), washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, concentrated, and purified by rapid silica gel column to obtain an off-white solid 6-bromo-1-isopropyl-1H-indazole-4-carboxylic acid methyl ester (Compound 2, 1.35 g, 4.55 mmol, 33.2% yield).

[0035] 1 H NMR (400MHz, CDCl3): δ (ppm) 1.60 (d, J = 8.0Hz, 6H), 4.00 (s, 3H), 4.83-4.76 (m, 1H), 7.81 (s, 1H), 7.98 (s, 1H), 8.45 (s, 1H).

[0036] (b) Synthesis of 6-bromo-1-isopropyl-1H-indazole-4-carboxylic acid (Compound 3)

[0037] 6-Bromo-1-isopropyl-1H-indazole-4-carboxylic acid methyl ester (1.25 g, 4.2 mmol) was dissolved in a mixture of methanol (24 mL) and THF (6 mL), 3N NaOH (6.7 mL, 20.1 mmol) was added, and the mixture was stirred at room temperature overnight. The mixture was concentrated to dryness under reduced pressure, water (30 mL) was added, and the mixture was stirred. 1N HCl was slowly added dropwise to pH 2-3, and a solid precipitated. The solid was filtered, washed with water, and dried to obtain 6-bromo-1-isopropyl-1H-indazole-4-carboxylic acid (compound 3, 1.05 g, 3.7 mmol, 88.1% yield), a white solid.

[0038] 1 H NMR (400MHz, DMSO-d6): δ (ppm) 1.47 (d, J = 4.0 Hz, 6H), 5.13-5.06 (m, 1H), 7.81 (s, 1H), 8.37 (s, 1H), 8.39 (s, 1H), 13.53 (br, s, 1H).

[0039] MS (ESI): m / z 283.2 [M+H] +

[0040] (c) Synthesis of 6-bromo-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (Compound 4)

[0041] 6-Bromo-1-isopropyl-1H-indazole-4-carboxylic acid (1.0 g, 3.55 mmol), 3-(aminomethyl)-4,6-dimethylpyridin-2(1H)-one hydrochloride (1.0 g, 5.3 mmol), HOBT (0.96 g, 7.1 mmol) and EDCI (1.35 g, 7.1 mmol) were suspended in DMSO (30 mL), N-methylmorpholine (1.44 g, 14.2 mmol) was added, and the mixture was stirred at room temperature overnight. Water (100 mL) was slowly added, and solid precipitated. The mixture was stirred for half an hour, filtered, washed with water, and dried to obtain technetium gray solid 6-bromo-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (compound 4, 1.45 g, 3.48 mmol, 98.0% yield).

[0042] 1 H NMR (400MHz, DMSO-d6): δ (ppm) 1.49 (d, J = 8.0Hz, 6H), 2.15 (s, 3H), 2.23 (s, 3H), 4.37 (d, J = 4.0Hz, 2H), 5 .11-5.04(m,1H),5.91(s,1H),7.72(s,1H),8.22(s,1H),8.39(s,1H),8.62-8.60(m,1H),11.53(s,1H).

[0043] MS (ESI): m / z 417.4 [M+H] +

[0044] (d) Synthesis of 6-(3-aminophenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (Compound 5)

[0045] 6-Bromo-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (200 mg, 0.48 mmol), 3-aminophenylboronic acid ester (157 mg, 0.72 mmol), Pd(PPh3)4 (55.5 mg, 0.048 mmol), sodium carbonate (152.6 mg, 1.44 mmol) and dioxane-water (10 mL-3 mL), Reflux for 7 hours under nitrogen protection, cool, concentrate to dryness, add water (50 mL), shake and filter to obtain a solid, purify by silica gel flash column (elution with dichloromethane: methanol = 20: 1), and concentrate to obtain an off-white solid 6-(3-aminophenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (Compound 5, 130 mg, 0.30 mmol, 62.5% yield).

[0046] 1 H NMR (400MHz, DMSO-d6): δ (ppm) 1.53 (d, J = 8.0Hz, 6H), 2.15 (s, 3H), 2.24 (s, 3H), 4.42 (d, J = 4.0Hz, 2H), 5.18-5.13 (m, 3H), 5.91 (s, 1H), 6 .64(d,J=8.0Hz,1H),7.00-6.97(m,2H),7.18-7.14(t,J=8.0Hz,1H),7.80(s,1H),7.97(s,1H),8.37(s,1H),8.61(s,1H),11.53(s,1H).

[0047] MS (ESI): m / z 430.5 [M+H] +

[0048] (e) Synthesis of 6-(3-(2-chloro-4-(methylsulfonyl)benzamido)phenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (target compound I)

[0049] 6-(3-Aminophenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (60 mg, 0.14 mmol), 2-chloro-4-(methylsulfonyl)benzoic acid (40 mg, 0.17 mmol), HOBT (37.8 mg, 0.28 mmol) and EDCI (53.7 mg, 0.28 mmol) were suspended in DMSO (2.5 mL) and N-methylbenzene was added. morpholine (70.8 mg, 0.7 mmol), stirred at room temperature overnight, water (50 mL) was slowly added, solid precipitated, stirred for half an hour, filtered, washed with water, and dried to obtain technetium gray solid 6-(3-(2-chloro-4-(methylsulfonyl)benzamido)phenyl)-N-((1,2-dihydro-4,6-dimethyl-2-oxopyridin-3-yl)methyl)-1-isopropyl-1H-indazole-4-carboxamide (target compound I, 45 mg, 0.07 mmol, 50.0% yield).

[0050] 1 H NMR (400MHz, DMSO-d6): δ (ppm) 1.53 (d, J = 8.0Hz, 6H), 2.15 (s, 3H), 2.26 (s, 3H), 3.39 (s, 3H), 4.43 (d, J = 4.0Hz, 2H), 5.20-5.17 (m, 1H), 5.91 (s, 1H) ,7.66-7.53(m,2H),7.86(s,2H),7.95(d,J=8.0Hz,1H),8.07(d,J=8.0Hz ,3H),8.18(s,1H),8.38(s,1H),8.64(s,1H),10.85(s,1H),11.54(s,1H).

[0051] MS (ESI): m / z 646.4 [M+H] +

[0052] Example 2: Effect of Compound I on Myeloid Cell Proliferation

[0053] 1. Cell culture

[0054] After primary medulloblastoma was dissected from the brain, it was digested by papain and centrifuged through Percoll to obtain a single cell suspension, and 2x10^5 cells were plated on a 24-well plate slide. After two hours of complete cell adhesion, culture medium (B27 medium containing 1mM sodium pyruvate, 2mM L-glutamine, and 1% penicillin / streptomycin) was added to 500 microliters per well and cultured at 37°C and 5% carbon dioxide for 48 hours. 96-well plates were seeded with 3x 10^5 cells in 100 microliters of culture medium. After 4-6 hours of complete cell adhesion, the original culture medium was removed, and 100 microliters of drug culture medium with different concentrations (prepared in "2. Dosing Scheme") were added, and culture was continued for 48 hours.

[0055] 2. Dosing plan

[0056] The test sample was dissolved in a small amount of DMSO, and then a 100 μM stock solution was prepared with culture medium (B27 medium containing 1 mM sodium pyruvate, 2 mM L-glutamine, and 1% penicillin / streptomycin), which was then diluted into five concentrations of 500 nM, 1 μM, 5 μM, 10 μM, and 20 μM, with three replicate wells for each concentration.

[0057] At the same time, the control group was added with an equal volume of DMSO, and the blank control group was added with only an equal volume of culture medium and no cells.

[0058] 3. CCK-8 absorbance detection

[0059] After 48 hours of cell culture, 10 μl of CCK-8 was added to each well of the 96-well plate and placed in an incubator for 3 hours. The absorbance was then measured at a wavelength of 450 nm. The inhibition rate was measured and the concentration required to inhibit 50% cell growth was calculated. 50 .

[0060] 4. Data analysis: All data were analyzed using GraphPad Prism 6.0.

[0061] 5. Test results

[0062] Table 1 IC of different drugs inhibiting myeloid blasts 50

[0063] <![CDATA[IC 50 ]]> Compound I 0.89uM MK-886 6.68uM

[0064] The results are shown in Table 1. The IC 50 The IC value of the reference substance MK-886 is 0.89uM. 50 The test results show that the compound I provided by the present invention has a good effect of inhibiting the growth of myeloid blasts, and its activity of inhibiting myeloid blasts is 7.5 times that of the reference substance MK-886.

[0065] The above-mentioned embodiments only express several implementation methods of the present invention, and the description is relatively specific and detailed, but it cannot be understood as limiting the scope of the invention patent. It should be pointed out that for ordinary technicians in this field, several modifications and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention.

Claims

1. A 1H-indazole compound, characterized in that: The structural formula of the compound is shown in Formula I.

2. A method for preparing the 1H-indazole compound according to claim 1, characterized in that: The steps include: (a) Synthesis of Compound 2: Compound 6-bromo-1H-indazole-4-carboxylic acid methyl ester 1 undergoes a substitution reaction with isopropyl bromide to obtain Compound 2. The reaction formula is as follows: The reaction solvent comprises at least one of DMF, isopropanol, tetrahydrofuran and acetonitrile, the base comprises at least one of sodium hydrogen sulfide, potassium carbonate, sodium carbonate, cesium carbonate, potassium tert-butoxide and sodium tert-butoxide, and the reaction temperature is between 0° C. and 60° C.; (b) Synthesis of compound 3: Compound 2 synthesized in step (a) is subjected to esterification reaction with sodium hydroxide solution to obtain compound 3, as shown in the following reaction formula: The base used for esterification comprises at least one of sodium hydroxide, potassium hydroxide, potassium carbonate and sodium carbonate, and the reaction temperature is between 20° C. and 60° C.; (c) Synthesis of compound 4: Compound 3 obtained in step (b) is subjected to a condensation reaction with 3-(aminomethyl)-4,6-dimethylpyridin-2(1H)-one hydrochloride to obtain compound 4 as shown. The reaction formula is as follows: The reaction condensation agent comprises at least one of HATU, HOBT, EDCI, BOP and DCC, the reaction solvent comprises at least one of DMSO, DMF, DCM, acetonitrile, n-butanol, dioxane and tetrahydrofuran, and the base comprises at least one of triethylamine, diethylamine, diisopropylethylamine, N-methylmorpholine, potassium carbonate and sodium carbonate; (d) Synthesis of compound 5: Compound 4 obtained in step (c) is subjected to coupling reaction with 3-aminophenyl borate to obtain compound 5. The reaction formula is as follows: The coupling reaction reagent is a borate or boric acid, the catalyst comprises at least one of tetrakistriphenylphosphine palladium, tert-butylphosphine palladium, PdCl2(dppf), Pd2(dba)3 and palladium acetate, the base comprises at least one of sodium carbonate, potassium carbonate and sodium bicarbonate, and the reaction solvent comprises at least one of dioxane-water, DMF-water, ethanol-water, ethylene glycol dimethyl ether-water and toluene-water; (e) Synthesis of target compound I: Compound 5 obtained in step (d) is subjected to condensation reaction with 2-chloro-4-methylsulfonylbenzoic acid to obtain target compound I. The reaction formula is as follows: The reaction condensation agent includes at least one of HOBT, EDCI, BOP and DCC, the reaction solvent includes at least one of DMSO, DMF, DCM and dichloroethane, and the base includes at least one of triethylamine, diethylamine, diisopropylethylamine and N-methylmorpholine.

3. Use of the 1H-indazole compound or a pharmaceutically acceptable salt, solvate or drug complex thereof according to claim 1 in the preparation of a drug for treating diseases related to medulloblastoma.

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