A method for synthesizing diastereoisomer impurities of tofacitinib citrate

Abstract

The invention discloses a method for synthesizing tofacitinib citrate diastereomeric impurities, and relates to the technical field of pharmaceutical organic synthesis. The invention uses 3-amino-4-methylpyridine as a starting raw material, and prepares the tofacitinib diastereomeric impurities through quaternary ammonium salt, reduction, methylation, salification, coupling, reduction and condensation reactions. The raw materials of the entire synthetic route are readily available, the reaction conditions are mild, the post-treatment separation and purification operation is simple and easy, the preparation method has good reproducibility, and the tofacitinib diastereomeric impurities with high yield and high purity can be prepared, thereby being suitable for use as an impurity reference substance for tofacitinib citrate.

Description

Technical field:

[0001] The present invention relates to the technical field of pharmaceutical organic synthesis, and in particular to a method for synthesizing tofacitinib citrate diastereoisomer impurities. Background technology:

[0002] Tofacitinib citrate, developed by Pfizer, was first approved by the FDA in November 2012. It is the world's first JAK inhibitor approved for the treatment of rheumatoid arthritis (RA) and the first JAK inhibitor approved in China (Pfizer's 5mg tofacitinib citrate tablets were approved for import on March 10, 2017). The drug is indicated for the treatment of adult patients with moderately to severely active rheumatoid arthritis (RA) who have had an inadequate response or intolerance to methotrexate. It can be used as monotherapy or in combination with methotrexate or other disease-modifying antirheumatic drugs (DMARDs). Furthermore, results from three Phase 3 clinical trials for the treatment of ulcerative colitis have demonstrated that tofacitinib is effective in patients with moderate to severe ulcerative colitis, with responses lasting for over a year in many patients.

[0003] Currently, there are many published methods for preparing tofacitinib citrate, but there is little research on tofacitinib's diastereomeric impurities, and no patent disclosures on tofacitinib's diastereomeric impurities have been found. J. Med. Chem. 2008, 5, 8012-8018 discloses a method for synthesizing tofacitinib's diastereomeric impurities. However, this method involves an 11-step synthesis and the use of Grubss reagent and PtO2. These reagents are expensive and the experimental conditions are harsh, resulting in a poor quality product. Therefore, a simple, mild, and large-scale synthesis method is urgently needed.

[0004] In national drug standards, reference substances can be used for the operation of standard substances such as testing, identification, content determination, and inspection of impurities and related substances. Therefore, impurity reference substances play a key role in drug quality control. The present invention aims to provide a method for synthesizing diastereoisomer impurities of tofacitinib citrate so that they can be used as reference substances to detect and control the content of these impurities in the tofacitinib citrate raw material. Summary of the invention:

[0005] The technical problem to be solved by the present invention is to provide a method for synthesizing diastereomeric impurities of tofacitinib citrate. The purity of the diastereomeric impurities of tofacitinib prepared by this method is higher than 95%, and can be used as a high-purity impurity reference substance for the study of diastereomeric impurities of tofacitinib.

[0006] The technical problem to be solved by the present invention is achieved by adopting the following technical solutions:

[0007] A method for synthesizing a diastereomeric impurity of tofacitinib citrate comprises the following steps: condensing 3-amino-4-methylpyridine with benzyl bromide to obtain an intermediate I; subjecting the intermediate I to a reduction reaction with sodium borohydride to obtain an intermediate II; subjecting the intermediate II to a methylation reaction with paraformaldehyde under the action of sodium methoxide and sodium borohydride to obtain an intermediate III; converting the intermediate III into a hydrochloride in ethanol; filtering the obtained solid to obtain an intermediate IV; adjusting the pH of the obtained mother liquor to 9-10 with an alkaline solution, extracting with an organic solvent, and concentrating under reduced pressure to obtain a crude intermediate V; purifying the crude intermediate V with a diacid by salt formation to obtain a pure intermediate V; subjecting the pure intermediate V to a coupling reaction with 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine under the action of an inorganic base to obtain an intermediate VI; subjecting the intermediate VI to a reduction reaction with a reducing agent and formic acid to obtain an intermediate VII; and subjecting the intermediate VII to a condensation reaction with ethyl cyanoacetate under the action of an organic base to obtain the diastereomeric impurity of tofacitinib citrate.

[0008] The specific synthesis route is as follows:

[0009]

[0010] The inorganic base is at least one of potassium carbonate, sodium hydroxide, and sodium carbonate, and the reaction solvent for the coupling reaction involving the inorganic base is at least one of methanol, ethanol, and water. Preferably, the inorganic base is potassium carbonate and the reaction solvent is water.

[0011] The reducing agent is at least one of palladium carbon, palladium hydroxide carbon, and Raney nickel, and the reaction solvent of the reduction reaction in which the reducing agent participates is at least one of methanol, ethanol, and water. Preferably, the reducing agent is palladium hydroxide carbon, and the reaction solvent is water.

[0012] The organic base is at least one of DBU, triethylamine, and diisopropylethylamine, and the reaction solvent for the condensation reaction in which the organic base participates is at least one of n-butanol, ethanol, and tetrahydrofuran. Preferably, the organic base is DBU and the reaction solvent is ethanol.

[0013] The organic solvent for extraction is one of dichloromethane, ethyl acetate, n-hexane, methyl tert-butyl ether, and diethyl ether. Preferably, the solvent is dichloromethane or n-hexane.

[0014] The selected diacid is one of malonic acid, fumaric acid, maleic acid and tartaric acid. Preferably, the diacid is fumaric acid.

[0015] The molar ratio of the 3-amino-4-methylpyridine to benzyl bromide is 1:(1-2).

[0016] The molar ratio of the intermediate I to sodium borohydride is 1:(2-5).

[0017] The molar ratio of the intermediate II, paraformaldehyde, sodium methoxide and sodium borohydride is 1:(1-2):(1-2):(1-2).

[0018] The molar ratio of the crude intermediate V to the diacid is 1:1.

[0019] The molar ratio of the pure intermediate V, 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine and the inorganic base is 1:(1-1.5):(6-8).

[0020] The amount of the reducing agent used is 10-30% of the mass of the intermediate VI, and the molar ratio of the intermediate VI to formic acid is 1:(2-6).

[0021] The molar ratio of the intermediate VII, ethyl cyanoacetate and the organic base is 1:(1-2):(1-2).

[0022] The beneficial effects of the present invention are as follows: the present invention uses 3-amino-4-methylpyridine as a starting material, and prepares tofacitinib diastereomeric impurities through quaternary ammonium salt, reduction, methylation, salt formation, coupling, reduction and condensation reactions. The raw materials of the entire synthetic route are easily available, the reaction conditions are mild, the post-treatment separation and purification operation is simple and easy, the preparation method has good reproducibility, and the tofacitinib diastereomeric impurities with high yield and high purity can be prepared, thereby being suitable for use as an impurity reference substance for tofacitinib citrate. Description of the drawings:

[0023] Figure 1 is the HPLC chart of tofacitinib;

[0024] Figure 2 is the HPLC profile of diastereomeric impurities;

[0025] Figure 3 HPLC comparison chart of tofacitinib and diastereomeric impurities;

[0026] Figure 4 is the hydrogen spectrum of the diastereomers;

[0027] Figure 5 is the hydrogen spectrum of tofacitinib;

[0028] Figure 6 is the MS spectrum of diastereomers;

[0029] Figure 7 HPLC chart of intermediate V. Specific implementation method:

[0030] In order to make the technical means, creative features, objectives and effects achieved by the present invention easier to understand, the present invention is further described below with reference to specific embodiments and illustrations.

[0031] In the following examples, the concentration of organic solvent and water 1 g / X mL refers to the mass volume ratio relative to the substrate (Intermediate I to Intermediate VII).

[0032] Example 1

[0033] Preparation of 3-amino-1-benzyl-4-methylpyridinium salt (Intermediate I):

[0034] Benzyl bromide was added dropwise to a 1 g / 10 mL solution of 3-amino-4-methylpyridine in dichloromethane at 20-25°C. After complete addition, the mixture was stirred at room temperature for 12 hours and filtered to obtain Intermediate I in a 95% yield. The molar ratio of 3-amino-4-methylpyridine to benzyl bromide was 1:1.02.

[0035] Preparation of 1-benzyl-4-methyl-3-amino-piperidine (Intermediate II):

[0036] A 1g / 10mL ethanol solution of Intermediate I was added to a three-necked flask, cooled to 0-5°C, and an aqueous solution of sodium borohydride was added dropwise. After addition, the temperature was raised to 55-60°C and the reaction was allowed to react for 2 hours. The reaction was monitored by TLC. The temperature was then lowered to 0-5°C, and the reaction was quenched with 10% aqueous ammonium chloride. The reaction was extracted three times with 1g / 10mL DCM and concentrated to obtain Intermediate II in a 90% yield. The molar ratio of Intermediate I to sodium borohydride was 1:2.

[0037] Preparation of (1-benzyl-4-methyl-3-piperidinyl)-methylamine (Intermediate III):

[0038] A 1 g / 10 mL methanol solution of Intermediate II was added to a three-necked flask, followed by paraformaldehyde and a 5 M sodium methoxide solution. The mixture was stirred at room temperature for 8 h. Sodium borohydride was added portionwise and stirred for 4 h. The reaction was quenched with 10% aqueous ammonium chloride solution, concentrated, and extracted three times with 1 g / 10 mL DCM. The reaction mixture was concentrated to afford Intermediate III in an 85% yield. The molar ratio of Intermediate II, paraformaldehyde, sodium methoxide solution, and sodium borohydride was 1:2:1.5:1.

[0039] Preparation of (trans-1-benzyl-4-methyl-3-piperidinyl)-methylamine (Intermediate V):

[0040] A 1g / 3ml ethanol solution of (1-benzyl-4-methyl-3-piperidinyl)-methylamine (Intermediate III) was added to a three-necked flask. Concentrated hydrochloric acid was added dropwise. After the addition was complete, the mixture was stirred for 1 hour and filtered. The filtrate was adjusted to a pH of 9-10 with 2N sodium hydroxide solution, extracted three times with 1g / 10mL n-hexane, and concentrated to obtain the crude trans isomer (SR, RS form). The molar ratio of Intermediate III to hydrochloric acid was 1:2.

[0041] The crude trans isomer was dissolved in 1g / 5mL of ethyl acetate. Fumaric acid was then dissolved in ethanol and added dropwise to the system. A solid slowly precipitated with stirring and filtered. The resulting solid was adjusted to a pH of 9-10 with 2N NaOH solution, extracted three times with 1g / 10mL of n-hexane, and concentrated to obtain the pure trans isomer (SR, RS form) in an 80% yield. The molar ratio of fumaric acid to crude trans isomer was 1:1.

[0042] Preparation of N-(trans-1-benzyl-4-methylpiperidin-3-yl)-2-chloro-N-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (Intermediate VI):

[0043] To a three-necked flask, add intermediate V, potassium carbonate, and 1 g / 10 mL of water. Add 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine with mechanical stirring, raise the temperature to reflux with stirring for 18 hours, cool to 20-25°C, filter, add 1 g / 5 mL of acetone solution to the filter cake, raise the temperature to reflux and slurry for 1 hour, cool to room temperature, filter, and dry to obtain intermediate VI in a 92% yield. The molar ratio of intermediate V, potassium carbonate, and 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine is 1:6:1.

[0044] Preparation of N-methyl-N-((trans-4-methylpiperidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (Intermediate VII):

[0045] To a three-necked flask, add Intermediate VI, 1 g / 8 mL of water, and formic acid. Stir until the solution is clear. Add palladium hydroxide on carbon, heat to 50-55°C, and react for 2 h. Monitor by TLC. After completion, filter the filtrate, adjust the pH to 9-10 with 2N sodium hydroxide solution, and extract three times with 1 g / 10 mL of DCM. Combine the organic phases and concentrate to obtain Intermediate VII in a 92% yield. The molar ratio of Intermediate VI to formic acid is 1:2, and the amount of palladium hydroxide on carbon is 10% of the mass of Intermediate VI.

[0046] Preparation of 3-(trans-4-methyl-3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)piperidin-1-yl)-3-oxopropanenitrile:

[0047] In a three-necked flask, intermediate VII was added, followed by a 1 g / 5 mL ethanol solution, ethyl cyanoacetate, and DBU. Under nitrogen protection, the temperature was raised to 40-45° C. and stirred for 12 h. A large amount of solid precipitated from the system, which was filtered and the filter cake was dried to obtain diastereomeric impurities in a yield of 84%.

[0048] The molar ratio of intermediate VII, ethyl cyanoacetate and DBU is 1:1.5:1.2.

[0049] Example 2

[0050] Preparation of 3-amino-1-benzyl-4-methylpyridinium salt (Intermediate I):

[0051] Benzyl bromide was added dropwise to a 1 g / 10 mL solution of 3-amino-4-methylpyridine in dichloromethane at 20-25°C. After the addition was complete, the mixture was stirred at room temperature for 12 h and then filtered to obtain Intermediate I in a 93% yield. The molar ratio of 3-amino-4-methylpyridine to benzyl bromide was 1:1.2.

[0052] Preparation of 1-benzyl-4-methyl-3-amino-piperidine (Intermediate II):

[0053] A 1g / 10mL ethanol solution of Intermediate I was added to a three-necked flask, cooled to 0-5°C, and an aqueous solution of sodium borohydride was added dropwise. After addition, the temperature was raised to 55-60°C and the reaction was allowed to react for 2 hours. The reaction was monitored by TLC, cooled to 0-5°C, and quenched with 300mL of 10% aqueous ammonium chloride. The mixture was extracted three times with 1g / 10mL DCM and concentrated to obtain Intermediate II in an 88% yield. The molar ratio of Intermediate I to sodium borohydride was 1:4.

[0054] Preparation of (1-benzyl-4-methyl-3-piperidinyl)-methylamine (Intermediate III):

[0055] A 1 g / 10 mL methanol solution of Intermediate II was added to a three-necked flask, followed by paraformaldehyde and a 5 M sodium methoxide solution. The mixture was stirred at room temperature for 8 h. Sodium borohydride was added portionwise and stirred for 4 h. The reaction was quenched with 10% aqueous ammonium chloride solution, concentrated, and extracted three times with 1 g / 10 mL DCM. The reaction mixture was concentrated to afford Intermediate III in an 87% yield. The molar ratio of Intermediate II, paraformaldehyde, sodium methoxide solution, and sodium borohydride was 1:1.5:1.2:1.

[0056] Preparation of (trans-1-benzyl-4-methyl-3-piperidinyl)-methylamine (Intermediate V):

[0057] A 1g / 3ml ethanol solution of Intermediate III was added to a three-necked flask, and concentrated hydrochloric acid was added dropwise. After the addition was complete, the mixture was stirred for 1 hour and filtered. The filtrate was adjusted to a pH of 9-10 with 2N sodium hydroxide solution, extracted with 1g / 10mL diethyl ether, and concentrated to obtain the trans isomer (SR, RS form). The molar ratio of Intermediate III to hydrochloric acid was 1:2.

[0058] The crude trans isomer was dissolved in 1g / 5mL of ethyl acetate. The fumaric acid was then dissolved in ethanol and added dropwise to the system. A solid slowly precipitated with stirring and filtered. The resulting solid was adjusted to a pH of 9-10 with 2N NaOH solution, extracted three times with 1g / 10mL of n-hexane, and concentrated to obtain the pure trans isomer (SR, RS form) in an 85% yield. The molar ratio of fumaric acid to crude trans isomer was 1:1.

[0059] Preparation of N-((3S,4S)-1-benzyl-4-methylpiperidin-3-yl)-2-chloro-N-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (Intermediate VI):

[0060] To a three-necked flask, add intermediate V, potassium carbonate, and 1 g / 10 mL of water. Add 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine with mechanical stirring, raise the temperature to reflux with stirring for 18 hours, cool to 20-25°C, filter, add 1 g / 5 mL of acetone solution to the filter cake, raise the temperature to reflux and slurry for 1 hour, cool to room temperature, filter, and dry to obtain intermediate VI in a 94% yield. The molar ratio of intermediate V, potassium carbonate, and 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine is 1:8:1.5.

[0061] Preparation of N-methyl-N-((trans-4-methylpiperidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (Intermediate VII):

[0062] To a three-necked flask, add Intermediate VI, 1g / 8ml of water, and formic acid. Stir until the solution is clear. Add palladium hydroxide on carbon, heat to 50-55°C, and react for 2 hours. Monitor by TLC. After completion, filter the filtrate, adjust the pH to 9-10 with 2N sodium hydroxide solution, and extract three times with 1g / 10mL of DCM solution. Combine the organic phases and concentrate to obtain Intermediate VII in a 95% yield. The molar ratio of Intermediate VI to formic acid is 1:4, and the amount of palladium hydroxide on carbon is 15% of the mass of Intermediate VI.

[0063] Preparation of 3-(trans-4-methyl-3-(methyl(7H-pyrrolo[2,3-d]pyrimidin-4-yl)amino)piperidin-1-yl)-3-oxopropanenitrile:

[0064] To a three-necked flask, add Intermediate VII, a 1g / 5mL ethanol solution, ethyl cyanoacetate, and DBU. Under nitrogen, heat to 40-45°C and stir for 12 hours. A large amount of solid precipitates, which is filtered and the filter cake dried to afford diastereomeric impurities in an 86% yield. The molar ratio of Intermediate VII, ethyl cyanoacetate, and DBU is 1:2:2.

[0065] The basic principles, main features, and advantages of the present invention are shown and described above. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention. Such changes and modifications are intended to fall within the scope of the present invention. The scope of protection claimed in the present invention is defined by the appended claims and their equivalents.

Claims

1. A method for synthesizing diastereoisomer impurities of tofacitinib citrate, characterized in that: 3-amino-4-methylpyridine and benzyl bromide undergo a condensation reaction to obtain intermediate I, intermediate I undergoes a reduction reaction with sodium borohydride to obtain intermediate II, intermediate II undergoes a methylation reaction with paraformaldehyde in the presence of sodium methoxide and sodium borohydride to obtain intermediate III, intermediate III is converted into a hydrochloride in ethanol, and the resulting solid is filtered to obtain intermediate IV, the resulting mother liquor is adjusted to a pH of 9-10 with an alkaline solution, extracted with an organic solvent, and concentrated under reduced pressure to obtain a crude intermediate V, which is purified by salt formation with a diacid to obtain a pure intermediate V, which is then reacted with 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine in the presence of an inorganic base to obtain intermediate VI through a coupling reaction, intermediate VI undergoes a reduction reaction with a reducing agent and formic acid to obtain intermediate VII, and intermediate VII undergoes a condensation reaction with ethyl cyanoacetate in the presence of an organic base to obtain a diastereoisomer impurity of tofacitinib citrate; The specific synthesis route is as follows:

2. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, wherein: The inorganic base is at least one of potassium carbonate, sodium hydroxide and sodium carbonate, and the reaction solvent of the coupling reaction involving the inorganic base is at least one of methanol, ethanol and water.

3. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, characterized in that: The reducing agent is at least one of palladium carbon, palladium hydroxide carbon, and Raney nickel, and the reaction solvent of the reduction reaction in which the reducing agent participates is at least one of methanol, ethanol, and water.

4. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, wherein: The organic base is at least one of DBU, triethylamine, and diisopropylethylamine, and the reaction solvent of the condensation reaction in which the organic base participates is at least one of n-butanol, ethanol, and tetrahydrofuran.

5. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, wherein: The organic solvent for extraction is one of dichloromethane, ethyl acetate, n-hexane, methyl tert-butyl ether and diethyl ether.

6. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, wherein: The selected diacid is one of malonic acid, fumaric acid, maleic acid and tartaric acid.

7. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, characterized in that: The molar ratio of the 3-amino-4-methylpyridine and benzyl bromide is 1:(1-2); the molar ratio of the intermediate I and sodium borohydride is 1:(2-5).

8. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, wherein: The molar ratio of the intermediate II, paraformaldehyde, sodium methoxide and sodium borohydride is 1:(1-2):(1-2):(1-2).

9. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, characterized in that: The molar ratio of the crude intermediate V to the diacid is 1:1; the molar ratio of the pure intermediate V, 2,4-dichloro-7H-pyrrolo[2,3-D]pyrimidine, and the inorganic base is 1:(1-1.5):(6-8).

10. The method for synthesizing diastereoisomer impurities of tofacitinib citrate according to claim 1, characterized in that: The amount of the reducing agent used is 10-30% of the mass of the intermediate VI, the molar ratio of the intermediate VI to formic acid is 1:(2-6); the molar ratio of the intermediate VII, ethyl cyanoacetate and the organic base is 1:(1-2):(1-2).

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