WO2021021000A1

WO2021021000A1 - Method for producing avatrombopag

Info

Publication number: WO2021021000A1
Application number: PCT/RU2020/000383
Authority: WO
Inventors: Марат Феликсович ФАЗЫЛОВ; Вадим Юрьевич Балабаньян; Владимир Викторович НЕСТЕРУК
Original assignee: Марат Феликсович ФАЗЫЛОВ
Priority date: 2019-08-01
Filing date: 2020-07-21
Publication date: 2021-02-04
Also published as: RU2709496C1

Abstract

The invention relates to the field of chemistry and the chemical and pharmaceutical industry and concerns a novel method for the synthesis of the substance avatrombopag in the form of a base or a pharmaceutically acceptable salt. Avatrombopag is produced by hydrolysis of an ester produced from 5-chloro-6-(4-(ethoxycarbonyl)piperidin-1-yl)nicotinic acid and 4-(4-chlorothiophen-2-yl)-5-(4-cyclohexylpiperazin-1-yl)thiazol-2-amine. The target product is subjected to additional purification by washing with a series of different solvents, including water, hexane, a mixture of tetrahydrofuran and water, and ethyl acetate.

Description

METHOD FOR OBTAINING AVATROMBOPAG

Technology area

The invention relates to the field of chemistry and the chemical-pharmaceutical industry and relates to a new method for synthesizing the substance of avatrombopag in the form of a base or its pharmaceutically acceptable salt.

Prior art

Thrombocytopenia is a condition characterized by a decrease in the number of platelets, which is accompanied by increased bleeding and problems with stopping bleeding. The goal of treating the disorder is to minimize the risk of hemorrhagic complications by increasing the platelet count to a safe level without causing significant side effects, using minimally toxic treatment options, which is especially important for long-term treatment of patients with refractory or recurrent disease. The main cytokine that stimulates platelet formation is thrombopoietin (TPO).

Currently, the use of thrombopoietin receptor agonists (aTPO-r) represents a new approach to the treatment of patients with thrombocytopenia with the ineffectiveness of previous methods. One such clinically tested agonist is avatrombopag. Chemical name of avatrombopag: 1- [3-chloro-5 - [[[4- (4-chloro-2-thienyl) -5- (4-cyclohexyl- 1-piperazinyl) -2-thiazolyl] amino] carbonyl] -2 pyridinyl] -4-piperidinecarboxylic acid.

Avatrombopag is an orally bioavailable, low molecular weight TPO receptor agonist that stimulates proliferation and differentiation of megakaryocytes from bone marrow progenitor cells, resulting in increased platelet production. The drug does not compete with TPO for binding to the TPO receptor and has an additive effect with TPO on platelet production. However, when taking the drug, a number of side effects appear. Therefore, new methods of obtaining a substance of a higher purity are required.

A known method of producing a compound from international application WO03062233 (A1), publ. 2003-07-31. The document describes methods for the synthesis of derivatives of 2-acylaminothiazole, the general formula of which falls under the formula of avatrombopag.

The initial scheme is: Further reactions depend on the reactive derivatives or the condensing agent. The purification is carried out on silica gel chromatographic columns.

As the closest analogue, the method according to CN 107383000 (A), publ. 2017-11-24. According to this source, the preparation method includes the following steps: obtaining intermediate compound 1 and intermediate compound 2 for carrying out the amidation reaction in the presence of a catalyst in an aprotic solvent; adding alkali to the solvent of intermediate A to carry out the alkaline hydrolysis reaction to obtain the target compound. Intermediate 2 is first synthesized and then reacted with intermediate 1; and the synthetic route of the method is an amplification reaction. An operation such as column chromatography is excluded, the method is suitable for an industrial production process. General scheme:

Disclosure of invention

The object of the present invention is to provide a synthesis of avatrombopag of high purity and product yield while minimizing degradation products.

The problem is solved by the synthesis of Avatrombopag (1- (3-chloro-5 - ((4- (4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1 -yl) thiazol-2-yl) carbomoyl) pyridin-2 -yl) piperidine-4-carboxylic acid (9) according to the following scheme:

The proposed new method of obtaining avatrombopag consists in the fact that Br2 is added dropwise to a solution of 2-acetyl-4-chlorothiophene (1) in diethyl ether, extraction is carried out with ethyl acetate (EYOAc), the residue is dissolved in ethyl alcohol, thiourea is added, boiling, the resulting residue is dissolved in a mixture of EuAc / hexane (1: 1), after cooling to room temperature, the filtered precipitate of 4- (4-chlorothiophen-2-yl) thiazol-2-amine (2) is washed with a mixture of EuAc / hexane and dried in vacuum at room temperature, the dried precipitate is dissolved in dioxane, and upon addition of N-bromosuccinimide, 4- (4-chlorothiophen-2-yl) thiazol-4-bromo-2-amine is obtained (3), dissolve it in acetonitrile, add 1-cyclohexylpiperazine and triethylamine, carrying out the reaction at reflux, the precipitate of 4- (4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1-yl) thiazol-2-amine ( 4) filtered off, washed with water, hexane and diethyl ether and dried, add the resulting product to a solution of 5-chloro-6- (4- (ethoxycarbonyl) piperidin-1-yl) nicotinic acid (7), previously obtained by adding ethyl -piperidine-4-carboxylate (5) in dimethyl sulfoxide of 5,6-dichloronicotinic acid (6) and triethylamine, followed by heating to 150 ° C, crystallization, washing with water, hexane, drying at room temperature and dissolving (7) in pyridine , then phosphorus oxychloride (POC) is added to the cooled solution of the mixture (4) and (7) so that the temperature of the mixture does not exceed 10 ° C, the mixture is heated to room temperature, the crystallization reaction is carried out, the resulting crystalline precipitate of ethyl ether of avatrombopag (8) is washed with water oh and hexane, the powder is dried, dissolved in a mixture of toluene and ethyl acetate, having a ratio by volume of 10 to 1, purified using flash chromatography on silica gel, suspended in a mixture of distilled water and tetrahydrofuran, sodium hydroxide is added at room temperature, stirred, diluted with water , add acetic acid to the resulting solution until pH = 5-5.5, stir at room temperature until the end of crystallization and the formation of a precipitate of avatrombopag (9), the resulting precipitate is filtered off and washed with water, a mixture of tetrahydrofuran-water, then water, hexane and ethyl acetate.

If necessary, a pharmaceutically acceptable salt of avatrombopag, for example, hydrochloric, nitrate, succinate, glutamate, maleate and the like, can be obtained. It is preferred to prepare avatrombopag maleate salt, which is sold under the trade name Doptelet.

The best example of carrying out the invention

The possibility of carrying out the invention is demonstrated by the following examples:

Example 1. Synthesis of avatrombopag

Stage 1. Synthesis of amine 2.

Br (22 g, 0.138 mol) was slowly added dropwise to a solution of 2-acetyl-4-chlorothiophene 1 (20 g, 0.125 mol) in diethyl ether (150 ml) over 20 minutes at 0 ° С. The resulting reaction mixture was stirred at 0 ° C for 2 hours. Upon completion of the reaction (control: TLC, silca gel, toluene, R _f = 0.7), the mixture was diluted with water (300 ml), stirred for 30 minutes, extracted with ethyl acetate (EYOAc) (3x150 ml), washed with water (2x100 ml) and saturated sodium chloride solution (2x50 ml). The solution was dried over anhydrous sodium sulfate and evaporated. The resulting residue was dissolved in ethyl alcohol (150 ml), thiourea (9.5 g, 0.125 mol) was added, and the mixture was refluxed for 1 hour. At the end of the reaction (control: TLC, CHCl3 / EUN = 97: 3, R _f = 0.6), the mixture was evaporated. The resulting residue was dissolved in 300 ml of a mixture of EtOAc / hexane (1: 1) and refluxed for 1 hour. Then the solution was cooled to room temperature, the formed precipitate of amine hydrobromide 2 was filtered off, washed with a mixture of EuAc / hexane (1: 1) (2x50 ml) and dried in a vacuum (10 mm Hg) at room temperature for 5 hours. A suspension of the obtained product in 500 ml of an aqueous solution of K2CO3 (40 g, 0.3 mol) was vigorously stirred for 1 hour at room temperature. Then the precipitate was filtered off, washed with water, and dried in air at room temperature for 2 days. Yellowish, crystalline, 21.9 g (yield 81%, purity 95 +% by NMR), m.p. = 147-150 ° C.

The use of a mixture of hexane and ethyl ester of acetic acid in washing at this stage allows to increase the purification of the intermediate compound.

Stage 2. Synthesis of amine 4

To a solution of 4- (4-chlorothiophen-2-yl) thiazol-2-amine 2 (21.6 g, 0.1 mol) in dioxane (100 ml) while cooling to 10 ° C with a water bath, N-bromosuccinimide (12.4 g, 0.105 mol) for 1 hour (control of the temperature inside the reaction environment - temperature not higher than 15 ° С). At the end of the reaction (control: TLC, EuAc / hexane = 25:75, R _f = 0.5), the reaction mixture was poured with vigorous stirring into 2 liters of cold (10 ° C) distilled water and stirred for about 1 hour until the crystallization of the precipitate was completed. Then the precipitate that formed was filtered off, washed with water (2x200 ml), hexane (2x100 ml), and dried in air at room temperature for 2 days. Gray. Cryst., ~ 30 g (yield ~ 100%, purity 96 +% by NMR), m.p. = 143-150 ° C. At this stage, a characteristic impurity of 5-bromo-4- (5-bromo-4-chlorothiophen-2-yl) thiazol-2-amine is also formed in a small amount, which then reacts similarly to the target 5-bromo-4- (4 -chlorothiophen-2-yl) thiazol-2-amine.

To a solution of 4- (4-chlorothiophen-2-yl) thiazol-4-bromo-2-amine 3 (29.5 g, 0.1 mol) in acetonitrile (250 ml) were added 1-cyclohexylpiperazine (20.2 g, 0.12 mol) and triethylamine ( 21.5 ml, 0.15 mol) and boiled for 4 hours (control: TLC, EuAc / hexane = 25:75, R _f = 0.1), the reaction mixture was poured into 2 L of water and stirred for 30 minutes at room temperature. Then the precipitate that formed was filtered off, washed with water (2x200 ml), hexane (2x100 ml) and diethyl ether (4x75 ml). The resulting product was dried in air at room temperature for 2 days. Gray, crystal, 28.7 g (yield 80%, purity 95 +% by NMR), m.p. = 172-174 ° C.

Stage 3. Synthesis of acid 7 To a solution of ethyl piperidine-4-carboxylate 5 (18.8 g, 0.12 mol) in dimethyl sulfoxide (100 ml), 5,6-dichloronicotinic acid (19.2 g, 0.1 mol) and triethylamine (25.3 g, 0.25 mol) were added. The mixture was stirred until the reagents were completely dissolved, heated in an oil bath to a temperature of 150 ° C and kept at this temperature for 8 hours (control: TLC, CHCl3 / EUN = 95: 5, R _f = 0.7). Then the reaction mixture was cooled to room temperature, poured into 2 L of water, and acetic acid was added to the resulting solution until pH = 4 (about 0.25 mol). The resulting slurry was stirred for 2 hours at room temperature until final crystallization. The resulting precipitate was filtered off, washed with water (3x500 ml) and hexane (3x300 ml). The resulting product was dried in air at room temperature for 2 days. Yellowish, crystal, 29.7 g (yield 95%, purity 95 +% by NMR), m.p. = 131-133 ° C.

Stage 4. Synthesis of ether 8

This step was carried out as in patent CN107383000A. Because the conditions proposed there are optimal. However, according to the invention, a different method of purification of the final substance 8 is proposed, which allows obtaining a purer product in the next step.

To a solution of 5-chloro-6- (4- (ethoxycarbonyl) piperidin-1-yl) nicotinic acid 7 (3.15 g, 0.01 mol) in pyridine (30 ml) was added 4- (4-chlorothiophen-2-yl) -5 - (4-cyclohexylpiperazin-1 - yl) thiazol-2-amine 4 (3.8 g, 0.01 mol) and cooled to 0 ° C with an ice bath. POC1s (2.25 g, 0.015 mol) was slowly added to the cooled solution with stirring so that the temperature of the mixture did not exceed 10 ° C (about 20 minutes). Then the mixture was heated to room temperature for 2 hours and kept at room temperature for 2 hours (control: TLC, CHC / EUN = 95: 5, R _f = 0.7). Then the reaction mixture was poured into 500 ml of water. The resulting slurry was stirred for 2 hours at room temperature until final crystallization. The resulting bright red precipitate was filtered off, washed with water (3x100 ml) and hexane (2x50 ml). The brick-colored powder was dried in air at room temperature for 2 days.

The resulting powder was dissolved in a mixture of toluene and ethyl acetate (10: 1) and purified by flash chromatography on silica gel using the same mixture as eluent. The fractions containing the product were evaporated on a rotary evaporator at a temperature not exceeding 40 ° C. The resulting red powder (or solid foam) was used further without additional cleaning. 4.7 g (yield 87%, purity ~ 96% by NMR).

Stage 5. Synthesis of avatrombopag 9

This step in CN107383000A was carried out with heating, which leads to the formation of a by-product.

According to the invention, the solvent (tetrahydrofuran instead of alcohol) and the purification method (sequential washing with a series of different solvents) were changed. Moreover, the new process does not include heating.

The starting ester 8 (13.4 g, 0.0198 mol) was suspended in a mixture of distilled water (50 ml) and tetrahydrofuran (150 ml), after which NaOH (2.2 g, 0.055 mol) was added to the suspension. Complete dissolution was observed. The mixture was stirred at room temperature for 15-20 hours, (control: TLC, CHO3 / EUN = 95: 5, R _f = 0.2). The mixture was filtered to remove dust and other foreign impurities, diluted with water (200 ml), and acetic acid was added to the resulting solution until pH = 5-5.5. The resulting emulsion was stirred for about 2 hours at room temperature until the end of crystallization and the formation of a precipitate. The resulting precipitate was filtered off and washed with water (100 ml), a mixture of tetrahydrofuran-water (1 to 1, 2 times 30 ml), water (4 times 100 ml), hexane (2 times 100 ml) and ethyl acetate (3 times 20 ml). The resulting cream-colored crystalline product was dried in air at room temperature for 3 days.

Cream, crystal, 2.75 g (yield 94.5%, purity 98 +% by NMR). T. pl. ~ 265 ° С with decomposition.

As a result of the formation at stage 2 of the characteristic impurity 5-bromo-4- (5-bromo-4-chlorothiophen-2-yl) thiazol-2-amine, as a result of all stages of the synthesis, the final substance of avatrombopag (1 - (3-chloro-5 - ((4- (4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1 -yl) thiazol-2-yl) carbomoyl) pyridin-2-yl) piperidine-4-carboxylic acid) contains trace amounts characteristic impurity 1- (3-chloro-5 - ((4- (5-bromo-4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1 -yl) thiazol-2-yl) carbomoyl) pyridine- 2-yl) piperidine-4-carboxylic acid.

Example 2. Synthesis of the salt of avatrombopag.

Steps are similar to 1-5 in example 1.

Stage 6. Synthesis of Avatrombopag maleate 10

The stage includes dissolving a mixture of avatrombopag 9 and maleic acid in DMF, precipitation of the salt with methylene chloride As a result of the formation at stage 2 of a characteristic impurity 5-bromo-4- (5-bromo-4-chlorothiophen-2-yl) thiazol-2-amine, as a result of all stages of synthesis, the final substance of avatrombopag maleate (salts 1 - (3-chloro -5 - ((4- (4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1 -yl) thiazol-2-yl) carbomoyl) pyridin-2-yl) piperidine-4-carboxylic acid with maleic acid) contains trace amounts of the characteristic impurity salt 1 - (3-chloro-5 - ((4- (5-bromo-4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazin-1 -yl) thiazole-2- yl) carbomoyl) pyridin-2-yl) piperidine-4-carboxylic acid with maleic acid.

These decomposition products are unique features for the claimed method and make it possible to distinguish the final substance of avatrombopag synthesized by the claimed method from similar substances obtained by other methods.

Example 3. Pharmacodynamic studies of the obtained substance for the treatment of mice with idiopathic thrombocytopenic purpura

The tests were carried out on healthy BALB / C mice of SPF level (Specefic pathogen Free), weight 18-22 g, males.

Breakdown of animals into groups: control group, avatrombopag group.

Modeling method: blood was taken from the orbital vein of BALB / C mice, followed by anticoagulation treatment with EDTA-Na2, and then the platelets were separated and washed. Ready the platelet suspension solution was mixed homogeneously with an equal amount of Freund's complete adjuvant and Freund's incomplete adjuvant, respectively. In the first week, the antigen containing Freund's complete adjuvant was injected subcutaneously into the hind paw, back and groin of the guinea pig. Injections were performed 4 times in total, with 5 points for each injection and 100 μl for each point. On week 6, untreated whole blood was collected from the guinea pigs and centrifuged for 10 minutes. at 3000 rpm, obtaining a supernatant that is guinea pig anti-mouse platelet serum (GP-APS). The supernatant was placed in a water bath (56 ° C) for 30 min to inactivate the complement, and then the inactivated supernatant was diluted with normal saline, followed by an agar diffusion antibody titer test and finally stored in a refrigerator at -20 ° C for later. use.

Mice were injected with guinea pig anti-mouse platelet serum (APS) at a dose of 100 μl / 20 g of body weight by intraperitoneal injection, in the 1st, 3rd, 5th, 7th, 9th, 11th and 13th day of the experiment.

Observation of developed subcutaneous purpura:

Class I, light bleeding is observed at injection sites and hemorrhagic patches spread to other sites (40% of mice);

Class II, overt bleeding is observed at injection sites, and ecchymosis and petechiae are observed elsewhere (35%); Class III, severe bleeding is observed at the injection sites, a large number of ecchymosis and petechiae appeared on the skin, and ulceration and blackening are noted on the skin (25%).

Avatrombopag was administered at a dose of 20 mg / kg. Platelet counts were measured over a 2-week period.

Platelet count (average) before modeling:

Control - 748.2 x10 ⁹ / l

Avatrombopag group - 749.0 x10 ⁹ / l

Platelet count on day 14:

Control - 382.4 x ^{10 9} / l

Avatrombopag group - 582.8 x 10 ⁹ / l.

The platelet count in mice has shown that avatrombopag can effectively limit the decrease in platelet count and be used in thrombopenia.

The substance according to the invention has a minimal manifestation of side effects, in particular, there is practically no hyperthermia and edema.

Industrial applicability

The invention is applicable in the field of chemistry and the chemical-pharmaceutical industry and allows the synthesis of the substance of avatrombopag in the form of a base or its pharmaceutically acceptable salt.

Claims

CLAIM

A method for producing avatrombopag, characterized in that Br2 is added dropwise to a solution of 2-acetyl-4-chlorothiophene (1) in diethyl ether, extraction is carried out with ethyl acetate of acetic acid (EYOAc), the residue is dissolved in ethyl alcohol, thiourea is added, subjected to boiling, the resulting residue is dissolved in a mixture of EuAc / hexane having a volume ratio of 1: 1, the filtered precipitate of 4- (4-chlorothiophen-2-yl) thiazol-2-amine (2) formed after cooling to room temperature is washed with a mixture of EuAc / hexane and dried in vacuo at room temperature, dissolved in dioxane, and adding N-bromosuccinimide gives 4- (4-chlorothiophen-2-yl) thiazol-4-bromo-2-amine, as well as a characteristic impurity 5-bromo-4- (5 -bromo-4-chlorothiophen-2-yl) thiazol-2-amine, which then reacts similarly to (3), dissolve it in acetonitrile, add 1-cyclohexylpiperazine and triethylamine, reacting under reflux, precipitate 4- (4-chlorothiophen-2-yl) -5- (4-cyclohexylpiperazine-1 -yl) thiazol-2-amine (4) is filtered off, washed with water, hexane and diethyl ether and dried, the resulting product is added to a solution of 5-chloro-6- (4- (ethoxycarbonyl) piperidin-1 - yl) nicotinic acid (7 ), previously obtained by adding 5,6-dichloronicotinic acid (6) and triethylamine to a solution of ethyl piperidine-4-carboxylate (5) in dimethyl sulfoxide, followed by heating to a temperature 150 ° C, crystallization, washing with water, hexane, drying at room temperature and dissolving (7) in pyridine, then phosphorus oxychloride (POC1s) is added to the cooled solution of mixture (4) and (7) so that the temperature of the mixture does not exceed 10 ° C, heated to room temperature, a crystallization reaction is carried out, the obtained crystalline precipitate of ethyl ester of Avatrombopag (8) is washed with water and hexane, the powder is dried, dissolved in a mixture of toluene and ethyl acetate, having a ratio by volume of 10 to 1, purified using flash chromatography on silica gel, suspended in a mixture of distilled water and tetrahydrofuran, add sodium hydroxide at room temperature, stir, dilute with water, add acetic acid to the resulting solution until pH = 5-5.5, stir at room temperature until crystallization and formation of avatrombopag precipitate (9) , the resulting precipitate was filtered off and washed with water, with a mixture of tetrahydrofuran with water, then again with water. d, hexane and ethyl acetate.