Preparation method of rosaxostat impurity

Abstract

The invention provides a preparation method of a rosaxostat impurity. The preparation method specifically comprises the following steps: S1, carrying out a substitution reaction on a compound 14-hydroxy-7-phenoxy isoquinoline-3-methyl formate and NCS under an acidic condition to generate an impurity A; s2, carrying out methylation reaction on the impurity A and a compound 2-methylboronic acid under an alkaline condition to generate an impurity B; s3, carrying out condensation reaction on the impurity B and a compound 3, namely glycine methyl ester hydrochloride to generate an impurity C. The synthesis method adopted by the invention is simple, the obtained sample is high in purity, and the synthesis method has great significance in process research, impurity analysis and quality control of the rosaxostat.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical chemical synthesis technology, and specifically relates to a method for preparing roxadustat impurities. Background Technology

[0002] Roxadustat, chemically named [(4-hydroxy-1-methyl-7-phenoxyisoquinoline-3-formyl)amino]acetic acid, is a small molecule compound originally developed by FibroGen. It is also the world's first hypoxia-inducible factor (HIF)-prolyl hydroxylase (PHD) inhibitor. This drug can improve iron metabolism disorders by inhibiting prolyl hydroxylase (PHD) to stabilize the HIF pathway under normal oxygen partial pressure. Roxadustat was first approved for marketing in my country in December 2018 under the brand name Roxadustat, primarily for the treatment of anemia associated with chronic kidney disease (CKD), including both dialysis and non-dialysis patients. Its specific structure is as follows:

[0003]

[0004] The study of drug impurities is crucial to the safety, efficacy, and quality control of drugs. Conducting impurity studies ensures the quality of roxadustat raw material and finished products, ultimately ensuring patient safety and efficacy. In 2016, Shanghai Pharmaceuticals Group reported (CN107954931A) a commercial production process route for [(4-hydroxy-1-methyl-7-phenoxyisoquinoline-3-carboxylate)amino]acetic acid, using methyl 4-hydroxy-7-phenoxyisoquinoline-3-carboxylate as the starting material, through halogenation, methylation, condensation, and hydrolysis, as shown below:

[0005]

[0006] In the preparation of roxadustat, the substitution reaction involves multiple reaction sites, generating product a while simultaneously generating impurity A. If these byproducts are not effectively removed, they will sequentially be converted into impurities B, C, and ultimately the API, thus affecting the quality of the active pharmaceutical ingredient (API). Roxadustat impurity C can be used as a reference standard for qualitative and quantitative control of impurities during the production of roxadustat API, thus possessing significant synthetic value. Summary of the Invention

[0007] The purpose of this invention is to provide a method for preparing roxadustat impurities. This process features mild reaction conditions, high product yield and purity, and can meet the requirements for reference standards, thus possessing significant industrial application value.

[0008] The objective of this invention is achieved through the following means:

[0009] A method for preparing roxadustat impurities, the method comprising the following steps:

[0010] S1. Under acidic conditions, compound 1 (methyl 4-hydroxy-7-phenoxyisoquinoline-3-carboxylate) undergoes a substitution reaction with NCS (N-chlorosuccinimide) to generate impurity A;

[0011] S2. Impurity A reacts with compound 2 (methylboronic acid) under alkaline conditions to undergo a methylation reaction to generate impurity B;

[0012] S3. Impurity B undergoes a condensation reaction with compound 3 (methyl glycine hydrochloride) to generate impurity C;

[0013]

[0014] The specific preparation process includes the following steps:

[0015] S1: Compound 1 (methyl 4-hydroxy-7-phenoxyisoquinoline-3-carboxylate) and NCS were stirred and mixed under acidic conditions with dichloroethane as solvent. The chlorination reaction was carried out at 60℃~70℃ under nitrogen protection. The resulting mother liquor was concentrated under reduced pressure, and the solid was collected after crystallization. The solid was separated by column chromatography, and the relevant components were collected to obtain a white to off-white solid. After drying, impurity A was obtained.

[0016] S2: Impurity A, compound 2 (methylboric acid) and palladium catalyst were stirred and mixed under alkaline conditions. Ethylene glycol methyl ether was used as the solvent. Methylation reaction was carried out under nitrogen protection and high temperature reflux. After crystallization and column chromatography separation, the relevant components were collected and dried to obtain a grayish-white solid, namely impurity B.

[0017] S3: Impurity B, compound 3 (methyl glycine hydrochloride) and condensing agent are mixed with tetrahydrofuran as solvent. The mixture undergoes a condensation reaction under reflux. After extraction, washing, and concentration under reduced pressure in the organic phase, the solid is collected after crystallization and dried to obtain a pale yellow solid, which is impurity C.

[0018] Preferably, the acidic substance used in the acidic conditions described in S1 is one or more of aluminum trichloride, ferric chloride, and trifluoromethanesulfonate, with ferric chloride being more preferred; the molar ratio of compound 1 to the acidic substance is 1:1.5~2, with 1:1.6~1.8 being more preferred.

[0019] Preferably, the molar ratio of compound 1 to NCS in S1 is 1:2.3~2.6, more preferably 1:2.45~2.55.

[0020] Preferably, the alkaline substance used in the alkaline conditions described in S2 is one or more of potassium phosphate, potassium carbonate, sodium phosphate, and sodium carbonate. More preferably, the alkaline condition is sodium phosphate, and the molar ratio of impurity A to sodium phosphate is 1:3.5.

[0021] Preferably, the molar ratio of impurity A to compound 2 in S2 is 1:2.5~3.5, more preferably 1:3.

[0022] Preferably, the palladium catalyst in S2 is one or more of palladium acetate, tetra(triphenylphosphine)palladium, triphenylphosphine acetate palladium, dichlorodi-tert-butyl-(4-dimethylaminophenyl)phosphine palladium, acetylacetone palladium, (1,3-bis(diphenylphosphine)propane)palladium chloride, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride, [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex, and dichlorodi-tert-butyl-(4-dimethylaminophenyl)phosphine palladium. More preferably, the [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex is used.

[0023] Preferably, the methylation reaction described in S2 uses a high-temperature reflux reaction temperature of 105°C to 115°C.

[0024] Preferably, the molar ratio of impurity B to compound 3 in S3 is 1:1.1 to 1.3. More preferably, it is 1:1.2.

[0025] Preferably, the condensation reaction described in S3 is carried out at a reflux temperature of 35°C to 45°C.

[0026] Preferably, the condensing agent used in the condensation reaction described in S3 is one or more of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, 1-hydroxybenzotriazole, 1H-benzotriazole-1-yloxytripyrrolyl hexafluorophosphate, and N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl)hexafluorophosphate. More preferably, 1-hydroxybenzotriazole and 1H-benzotriazole-1-yloxytripyrrolyl hexafluorophosphate are used.

[0027] Roxadustat impurity C is a conductive impurity. To obtain high yield and high purity of impurity C, specific reaction conditions need to be selected, especially the specific selection of condensing agent and palladium catalyst, which significantly improves yield and conversion. This invention uses specific preparation conditions to further react and purify roxadustat impurity A in the preparation process of roxadustat, thereby obtaining roxadustat impurity C with high purity.

[0028] The advantages of this invention compared with the prior art are as follows:

[0029] The present invention provides a method for preparing roxadustat impurity C, which has the advantages of high product purity, high yield, readily available raw materials and reagents, and simple operation. It can meet the needs of reference standards and has high industrial application value. Attached Figure Description

[0030] Figure 1This is the high-performance liquid chromatogram of roxadustat impurity C in Example 1;

[0031] Figure 2 The high-performance liquid chromatogram of roxadustat impurity C in Comparative Example 1;

[0032] Figure 3 The image shows the high-performance liquid chromatogram of roxadustat impurity C in Comparative Example 2. Detailed Implementation

[0033] The present invention will be further explained and illustrated below through specific embodiments. The starting materials in the following embodiments are: methyl 4-hydroxy-7-phenoxyisoquinoline-3-carboxylate (compound 1) is produced by Jiangsu Weikaier Pharmaceutical Technology Co., Ltd.; methylboronic acid (compound 2) is produced by Weizhi Pharmaceutical Co., Ltd.; and methyl glycine hydrochloride (compound 3) is produced by Nanjing Chemical Reagent Co., Ltd.

[0034] Example 1

[0035] Impurity A: Methyl 1-chloro-4-hydroxy-7-(4-chlorophenoxyisoquinoline)-3-carboxylate; Impurity B: Methyl 1-methyl-4-hydroxy-7-(4-chlorophenoxyisoquinoline)-3-carboxylate; Impurity C: Methyl [[4-hydroxy-1-methyl-7-(4-chlorophenoxyisoquinoline)-3-carbonyl]-amino]-acetate.

[0036] (1) Under nitrogen protection, 125 g of dichloroethane was added to the reaction flask, and stirring was started. 20 g of compound 1 (0.067 mol, 1.0 eq) and 19.5 g of ferric chloride (0.121 mol, 1.8 eq) were added, and the temperature was controlled at 0~5℃. 11 g of NCS (0.084 mol, 1.25 eq) was added in batches at 15 min intervals, and the reaction was allowed to proceed for 1 h after the addition was complete. Another 11 g of NCS (0.084 mol, 1.25 eq) was added, and the mixture was stirred for 0.5 h. The temperature was then raised to 60℃~70℃, and the reaction was monitored by TLC until the reaction was complete. The reaction flask was cooled to 10℃~15℃, and purified water was slowly added. The mixture was stirred at 10℃~15℃ for 2 h to induce crystallization. The crystals were filtered and concentrated to obtain 25.81 g of white solid. The solid was separated by column chromatography. The collected eluent was concentrated to dryness to obtain 20.14 g of solid, which was then dried to obtain 16.77 g of white solid, namely impurity A. The yield of this step was 69%.

[0037] (2) Add 24g of impurity A (0.061mol, 1.0eq), 10.9g of compound 2 (0.18mol, 3.0eq), 80.8g of trisodium phosphate (0.21mol, 3.5eq), 0.4g of [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex, 26mL of purified water, and 359.12mL of ethylene glycol methyl ether to the reaction flask. Heat and stir under nitrogen protection, and monitor the reaction by TLC until complete. Filter the reaction solution while hot, and adjust the pH of the filtrate to 2-3 after cooling to room temperature. Repeat the pH test at 15min intervals and adjust to 2-3 twice. Slowly add 300mL of purified water at room temperature and stir to crystallize for 3h. Filter and dry to obtain a light brown solid. Separate by column chromatography. Concentrate the collected eluent and dry to obtain 19.16g of grayish-white solid, i.e., impurity B. The yield of this step is 84%.

[0038] (3) Add 8.7 g of impurity B (0.064 mol, 1.0 eq), 3.85 g of compound 3 (0.031 mol, 1.2 eq), 16.13 g of 1H-benzotriazol-1-yloxytripyrrolyl hexafluorophosphate (0.031 mol, 1.2 eq), 4.19 g of 1-hydroxybenzotriazole (0.031 mol, 1.2 eq), 9.9 g of diisopropylethylamine (0.077 mol, 3.0 eq), and 17.74 g of tetrahydrofuran to the reaction flask. Reflux the mixture at 35℃~45℃ and monitor the reaction by TLC until the reaction is complete (the reaction reaches its endpoint in 1.5 h). Cool the reaction solution to 20℃~30℃, add 72 g of purified water, and maintain the temperature while stirring to allow crystallization for 3 h. Centrifuge until no droplets remain. Dissolve the solid obtained by centrifugation and wash it sequentially with sodium bicarbonate solution, dilute hydrochloric acid, and saturated saline solution. After concentrating the filtrate, n-heptane was added to induce crystallization. The filter cake was then dried under vacuum to obtain 8.96 g of a pale yellow powdery solid, which was impurity C. The yield of this step was 83%, and the overall yield was 48.1%. 1 H-NMR, δ: 13.28 (s, 1H); 8.64 (s, 1H); 7.68~7.61 (d, J=2.1Hz, 1H); 7.54~7.38 ( m, 3H); 7.22~7.19 (m, 2H); 7.17 (s, 1H); 3.68 (s, 2H); 3.54 (s, 3H); 2.79 (s, 3H). )

[0039] In the above embodiments, the chromatographic conditions for sample detection are as follows:

[0040] Chromatographic conditions and system suitability: The column was packed with octadecyl bonded silica gel [Waters XBridge C18 (150 mm × 4.6 mm, 3.5 μm)] or a column with equivalent performance; the mobile phase was 0.1% formic acid aqueous solution; the mobile phase was 100% acetonitrile solution; gradient elution was performed as shown in Table 1; the detection wavelength was 260 nm; the column temperature was 30 °C; the injection volume was 5 μL; and the chromatogram was recorded for approximately 60 min.

[0041] Table 1 Elution gradient

[0042] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0 85 15 5 75 25 20 15 85 50 15 85 51 85 15 60 85 15

[0043] The high-performance liquid chromatogram of the prepared roxadustat impurity is shown in Figure 1. Figure 1 The purity of the main peak of impurity C is 99.67%.

[0044] Comparative Example 1

[0045] The remaining steps are the same as in Example 1, except that in step (3), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is used instead of 1H-benzotriazol-1-yloxytripyrrolidinyl hexafluorophosphate as the condensing agent in the condensation reaction. This step of the reaction reaches its endpoint after 7.5 hours, with a yield of only 43.5% and a purity of 90.43%. Figure 2 High-performance liquid chromatogram of roxadustat impurity C prepared by this process.

[0046] Comparative Example 2

[0047] The remaining steps are the same as in Example 1, except that in step (2), dichlorodi-tert-butyl-(4-dimethylaminophenyl)phosphine palladium is used instead of the [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex as the palladium catalyst for the Suzuki reaction. The product conversion rate in this step is only 21.4%, and the purity of the main peak is 98.76%. Figure 3 High-performance liquid chromatogram of roxadustat impurity C prepared by this process.

Claims

1. A method for preparing roxadustat impurities, characterized in that, The method includes the following steps: S1. Under acidic conditions, methyl 14-hydroxy-7-phenoxyisoquinoline-3-carboxylate undergoes a substitution reaction with NCS to generate impurity A; S2. Impurity A reacts with compound 2-methylboric acid under alkaline conditions to undergo a methylation reaction to generate impurity B; S3. Impurity B undergoes a condensation reaction with compound 3 glycine methyl ester hydrochloride to generate impurity C.

2. The method for preparing roxadustat impurities according to claim 1, characterized in that, Specifically, S1 involves mixing methyl 14-hydroxy-7-phenoxyisoquinoline-3-carboxylate and NCS under acidic conditions with dichloroethane as the solvent, and conducting a chlorination reaction at 60°C–70°C under nitrogen protection. The resulting mother liquor is concentrated under reduced pressure, and the solid is collected after crystallization. The solid is then separated by column chromatography, and the relevant components are collected to obtain a white to off-white solid. After drying, impurity A is obtained.

3. The method for preparing roxadustat impurities according to claim 1, characterized in that, The acidic substance used in the acidic conditions described in S1 is one or more of aluminum trichloride, ferric chloride, and trifluoromethanesulfonate, preferably ferric chloride; the molar ratio of compound 1 to the acidic substance is 1:1.5~2, preferably 1:1.6~1.8; the molar ratio of compound 1 to NCS is 1:2.3~2.6, preferably 1:2.45~2.

55.

4. The method for preparing roxadustat impurities according to claim 1, characterized in that, Specifically, S2 involves mixing impurity A, compound 2-methylboric acid, and palladium catalyst under alkaline conditions, using ethylene glycol methyl ether as the solvent, and subjecting the mixture to a high-temperature reflux reaction under nitrogen protection. After crystallization and column chromatography separation, the relevant components are collected and dried to obtain a grayish-white solid, i.e., impurity B. The high-temperature reflux reaction used in the methylation reaction is 105℃~115℃.

5. The method for preparing roxadustat impurities according to claim 4, characterized in that, The palladium catalyst described in S2 is one or more of the following: palladium acetate, tetra(triphenylphosphine)palladium, triphenylphosphine acetate palladium, dichlorodi-tert-butyl-(4-dimethylaminophenyl)phosphine palladium, acetylacetone palladium, (1,3-bis(diphenylphosphine)propane)palladium chloride, [1,1′-bis(diphenylphosphine)ferrocene]palladium dichloride, and [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex; preferably [1,1'-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex.

6. The method for preparing roxadustat impurities according to claim 1, characterized in that, The alkaline conditions described in S2 use one or more of potassium phosphate, potassium carbonate, sodium phosphate, and sodium carbonate, with sodium phosphate being the preferred alkaline condition. The molar ratio of impurity A to sodium phosphate is 1:3.

5.

7. The method for preparing roxadustat impurities according to claim 1, characterized in that, The molar ratio of impurity A to compound 2 in S2 is 1:2.5~3.5, preferably 1:

3.

8. The method for preparing roxadustat impurities according to claim 1, characterized in that, Specifically, S3 involves mixing impurity B, compound 3, and a condensing agent in tetrahydrofuran as the solvent. The mixture undergoes a condensation reaction under reflux, followed by extraction, washing, and concentration of the organic phase under reduced pressure. After crystallization, the solid is collected and dried to obtain a pale yellow solid, which is impurity C. The reflux reaction temperature is 35°C to 45°C.

9. The method for preparing roxadustat impurities according to claim 1, characterized in that, The molar ratio of impurity B to compound 3 in S3 is 1:1.1~1.3; preferably 1:1.

2.

10. The method for preparing roxadustat impurities according to claim 1, characterized in that, The condensation reaction described in S3 uses one or more of the following condensing agents: 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, 1-hydroxybenzotriazole, 1H-benzotriazole-1-yloxytripyrrolidinyl hexafluorophosphate, and N,N,N′,N′-tetramethyl-O-(7-azabenzotriazole-1-yl)hexafluorophosphate urea; preferably 1-hydroxybenzotriazole and 1H-benzotriazole-1-yloxytripyrrolidinyl hexafluorophosphate.

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