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Title:
PHARMACEUTICAL COMPOSITIONS COMPRISING ANTICOAGULANT N-(5-CHLOROPYRIDINE-2-YL)-2-({4-[ETHANIMIDOIL(METHYL)AMINO]BENZOYL}AMINO)-5-METHYLBENZAMIDE
Document Type and Number:
WIPO Patent Application WO/2017/217895
Kind Code:
A1
Abstract:
The invention relates to area of medicine, in particular to pharmacology, namely to novel pharmaceutical compositions for oral use for treatment of diseases associated with increased thrombus formation. The compositions comprise the anticoagulant N-(5-chloropyridine-2-yl)-2-({4-[ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamide or a pharmaceutically acceptable salt thereof, being a factor Xa inhibitor and polyprenol, and are protected from the contact with acidic gastric juice by an enteric coating or capsule. The compositions are of high efficacy and bioavailability and can be used to treat and prevent conditions characterized by undesirable thrombosis.

Inventors:
TOVBIN DMITRY GENNADIEVICH (RU)
TARASOV DMITRY NIKOLAEVICH (RU)
MALAKHOV DMITRY VIKTOROVICH (RU)
Application Number:
PCT/RU2017/050049
Publication Date:
December 21, 2017
Filing Date:
June 13, 2017
Export Citation:
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Assignee:
PHARMADIALL LTD (RU)
International Classes:
A61K9/22; A61K9/52; A61K31/045; A61K31/4425; A61K47/12; A61K47/32; A61K47/38; A61P7/02
Domestic Patent References:
WO2009122431A22009-10-08
Foreign References:
EA201000506A12011-10-31
US5012018A1991-04-30
RU2554735C12015-06-27
Other References:
"Assessment report on Ginkgo biloba L., folium", COMMITTEE ON HERBAL MEDICINAL PRODUCTS (HMPC, 28 January 2014 (2014-01-28), pages 4, 13, XP055464799
Attorney, Agent or Firm:
KOTLOV, Dmitry Vladimirovich et al. (RU)
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Claims:
Claims

1 . The pharmaceutical composition is in the form suitable for oral administration, having an anticoagulant activity and comprising an effective amount of N-(5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamide or a pharmaceutically acceptable salt thereof and polyprenol.

2. The pharmaceutical composition according to claim 1 , wherein the pharmaceutically acceptable salt of the compound N- (5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamide is presented by hydrochloride.

3. The pharmaceutical composition according to claim 1 , wherein it is produced in a solid- dosage form, having an enteric coating or an enteric-coated capsule.

4. The pharmaceutical composition according to claim 3, wherein the enteric coating is a film coating for tablets.

5. The pharmaceutical composition according to claim 4, wherein the enteric coating is a film coating for tablets based on a methacrylic acid copolymer.

6. The pharmaceutical composition according to claim 5, wherein the film coating is a film coating for tablets based on a methacrylic acid copolymer Aquarius™ Control ENA.

7. The pharmaceutical composition according to claim 1 , wherein polyprenol comprises 4-40 isoprene units.

8. The pharmaceutical composition according to claim 1 , wherein further comprises at least one pharmaceutically acceptable excipient.

9. The pharmaceutical composition according to claim 8, wherein the excipients are Prosolv® SMCC 90 and/or magnesium stearate.

10. The pharmaceutical composition according to claim 9, wherein comprises the following composition and ratio of components, wt %:

N-(5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)-5- 1 -80

methylbenzamide hydrochloride.

polyprenol 1 -80,

enteric coating 1 -50,

Prosolv® SMCC 90 0-90,

magnesium stearate 0-50,

1 1 . The pharmaceutical composition according to claim 1 is used for prevention and/or treatment of conditions characterised with undesired thrombosis in mammals.

12. The use according to claim 1 1 , wherein the condition is presented by acute coronary syndrome, myocardial infarction, unstable angina, refractory angina, thrombosis caused by post-thrombolytic therapy or coronary angioplasty, acute ischemic cerebrovascular syndrome, embolic stroke, thrombotic stroke, transient ischemic attacks, venous thrombosis, deep venous thrombosis, pulmonary embolism, coagulopathy, disseminated intravascular coagulation, thrombotic thrombocytopenic purpura, thromboangiitis obliterans, thrombotic disease associated with heparin-induced thrombocytopenia, thrombotic complications associated with extracorporeal circulation and prosthetic devices.

13. The use according to claim 1 1 , wherein the mammal is a human.

Description:
Pharmaceutical compositions comprising anticoagulant

N-(5-chloropyridine-2-yl)-2-({4-[ethanimidoil(methyl)amino]b enzoyl}amino)-5- methylbenzamide Field of the invention

The invention relates to the field of medicine, in particular to pharmacology and deals with prevention and treatment of diseases associated with increased thrombus formation in a human or other mammals using an oral medicinal product of high efficacy and bioavailability.

Background of the invention Thromboembolic diseases are one of the main cause of death in developed countries.

Thrombosis is the formation of a thrombus (blood clot) in the lumen of a blood vessel. Thrombus can occur both in veins and arteries; it localizes mostly in veins, especially in veins of lower limbs. The coronary artery thrombosis leads to insufficiency of blood supply to one of the heart segments that can lead to development of acute myocardial infarction. In some cases, the thrombus breaks away from a vessel wall and circulates freely within the bloodstream that can lead to acute blood-vessel occlusion (thromboembolism) with subsequent violation of the local blood supply. Thromboembolism of large branches of the pulmonary artery is especially dangerous: it is a common cause of sudden death. Cerebral vessel occlusion can cause ischemic stroke. The World Health Organization estimates that 17 million people die from cardiovascular diseases each year.

Anticoagulants are the medicinal products used to treat and prevent thromboembolic diseases, in particular with myocardial infarctions and lungs, thrombotic and embolic strokes, thrombophlebitis. Anticoagulants are used to prevent coronary and cerebral atherosclerosis, rheumatic mitral valve diseases. In surgery, anticoagulants are used to prevent the formation of thrombus during operations, in the postoperative period.

According to mechanism of action anticoagulants are classified into several groups: vitamin K antagonists, heparins, direct thrombin inhibitors and direct factor Xa [1 ] inhibitors.

Vitamin K antagonists, among which warfarin is well known, are more convenient to use because they are administered orally (in the form of tablets) whereas heparins are given by injection. However, vitamin K antagonists have a very narrow dosing range, interact with many drug substances and food products and require continuous blood clotting monitoring that imposes a number of limitations for use.

Modern anticoagulants should meet the high-efficacy requirements of preventing blood clotting, low toxicity and low risk of adverse effects, a mainly low risk of spontaneous bleeding.

The latter requirement presents a serious obstacle since the flip side of the blood clotting prevention lies in an increased risk of bleeding. This problem can be solved by a) using the molecules of high affinity and selectivity to proteins involved in the clotting cascade that, in turn, would reduce the used concentration of active substance and consequently the risk of bleeding;

b) creating a dosage form having the properties of maintaining a constant concentration of active ingredient for the longest possible period.

The use of direct factor Xa inhibitors in general practice has several advantages associated with the fact that factor Xa is involved only in the process of blood clotting, and its inhibition results in the fewest adverse effects, that allows to designate them as promising modern anticoagulants.

Most medicinal products reach a consumer in ready-made form or in the form of a so- called medicinal drug. A ready-made medicinal product is a complex consisting of the drug itself and the excipients of a definite form or aggregate state. Very often, the dosage form determines not only the convenience of the drug but also other drug efficacy characteristics, such as bioavailability, time-course etc.

The new class of anticoagulant compounds, including N-(5-chloropyridine-2-yl)-2-({4-

[[etanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzami de, has been described in patent EA 015918 as an effective factor Xa inhibitor. However, the compound N-(5-chloropyridine-2- yl)-2-({4-[ethanimidoil(methyl)amino]benzoyl}amino)-5-methyl benzamide has low bioavailability, which does not allow to apply it in a clinical practice for treatment of the diseases associated with increased thrombotic risk in humans and other mammals. In view of this, the object was set out to develop a dosage form of this compound suitable for oral administration and providing high bioavailability and efficacy of its application in clinical practice.

Summary

The object of this invention lies in the development of pharmaceutical composition, comprising anticoagulant as an active compound, for oral use of such composition for prevention and/or treatment of conditions associated with blood-clotting disorders in humans and other mammals.

The technical result of this invention is the development and production of pharmaceutical composition, comprising anticoagulant as an active compound and having high efficacy and bioavailability, for oral use of such composition for prevention and/or treatment of events associated with blood-clotting disorders in humans and other mammals, in particular for prevention and/or treatment of conditions characterised by undesired thrombosis in humans or other mammals.

The said technical result is achieved by developing and obtaining pharmaceutical compositions in the form suitable for oral administration, comprising an effective amount of N- (5-chloropyridine-2-yl)-2-({4-[ethanoydoil(methyl)amino]benz oyl}amino)-5-methylbenzamide or a pharmaceutically acceptable salt thereof and polyprenol. In other embodiments, a pharmaceutically acceptable salt of the compound N- (5- chloropyridine-2-yl)-2-({4-[ethanimidoil(methyl)amino]benzoy l}amino)-5-methylbenzamide is presented by hydrochloride.

In other embodiments, there are provided pharmaceutical compositions in solid-dosage forms, having an enteric coating or an enteric-coated capsule.

In other embodiments, there are provided pharmaceutical compositions in the form of enteric-coated tablets. In a part of the other embodiments, the enteric coating appears to be a film coating for tablets. Preferably, the film coating for tablets is a film coating based on a methacrylic acid copolymer. In particular embodiments, the film coating is a coating for tablets based on a methacrylic acid copolymer Aquarius™ Control ENA.

In other embodiments, the polyprenol comprises 4—40 isoprene units.

In a part of the other embodiments, the pharmaceutical compositions comprise at least one additional pharmaceutically acceptable excipient.

In other embodiments, the pharmaceutical compositions are characterised that comprise excipients are Prosolv® SMCC 90 and/or magnesium stearate.

Additionally, the invention provides pharmaceutical compositions with the following composition and ratio of components, wt %:

N- (5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)

methylbenzamide hydrochloride. polyprenol

enteric coating

Prosolv® SMCC 90

magnesium stearate

Additionally, the invention provides the using of pharmaceutical compositions according to the invention for prevention and/or treatment of conditions associated with undesired thrombosis in humans or other mammals.

In particular embodiments, the pharmaceutical compositions of the invention are designed for prevention and/or treatment of acute coronary syndrome, myocardial infarction, unstable angina, refractory angina, thrombosis caused by post-trombolitic therapy or coronary angioplasty, acute ischemic cerebrovascular syndrome, embolic stroke, thrombotic stroke, transient ischemic attacks, venous thrombosis, deep venous thrombosis, pulmonary embolism, coagulopathy, disseminated intravascular coagulation, thrombotic thrombocytopenic purpura, thromboangiitis obliterans, thrombotic disease associated with heparin-induced thrombocytopenia, thrombotic complications associated with extracorporeal circulation and prosthetic devices. Preferably, the pharmaceutical compositions are used for prevention and/or treatment of disease conditions in a human.

The invention also relates to a method for treating and/or preventing mammal's condition wherein undesired thrombosis comprising the step of administering the pharmaceutical composition which is the subject of the invention.

The invention also includes the production of pharmaceutical compositions of the invention.

Detailed description of the invention

The compound N-(5-chloropyridine-2-yl)-2-({4-

[ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzami de is an effective factor Xa inhibitor (patent EA015918). At the same time, as follows from the carried out animal experiments (experiments were carried out on rats) the compound N-(5-chloropyridine-2-yl)-2- ({4-[ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzam ide has a low bioavailability of (« 15%) that does not allow to use it in clinical practice as an oral medicinal product for prevention and treatment of diseases associated with blood-clotting disorders in humans and other mammals.

To overcome the said mentioned problem, as described in this invention, the pharmaceutical compositions are developed for oral use comprising N-(5-chloropyridine-2-yl)-2- ({4-[ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzam ide or a pharmaceutically acceptable salt thereof and polyprenol protected from the contact with acidic gastric juice by an enteric coating or capsule.

In the course of study, it has been unexpectedly found that the pharmaceutical compositions of the invention comprising N-(5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamide or a pharmaceutically acceptable salt thereof and polyprenol protected from the contact with acidic gastric juice by an enteric coating or capsule, produce a more effective anticoagulant effect than N-(5- chloropyridine-2-yl)-2-({4-[ethanimidoil(methyl)amino]benzoy l}amino)-5-methylbenzamide, or a pharmaceutically acceptable salt thereof, by itself.

Thus, the use of pharmaceutical composition of the invention will significantly improve the efficacy of the anticoagulant N- (5-chloropyridine-2-yl)-2-({4- [ethanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamide, when used orally, and also will allow lowering a dose of the specified compound that, in turn, can decrease the incidence of adverse effects and promote resource saving. Description of the drawings

Figure 1. The influence of the compound No.1 , FDF No.1 , and FDF No.2 administered intragastrically to rats on INR and PR values. The number of animals for each point is n = 8-16. Terms and definitions

As used herein, the term "compound No. 1" refers to N- (5-chloropyridine-2-yl)-2-({4- [ethanoydoil(methyl)amino]benzoyl}amino)-5-methylbenzamide hydrochloride.

The term "polyprenol" in this invention means a class of chemical compounds, polyunsaturated aliphatic branched alcohols of regular structure contained in natural raw materials of plant origin, in particular, in larch needles, usually as a mixture of isoprenols of the gross fo -l, wherein n is equal to 4—40. The chemical formula of polyprenols:

wherein n denotes a number of isoprene units. Polyprenols and their 2,3-dihydro derivatives - dolichols are found in various organs of plant and animal bodies [4].

The adjuvant Prosolv® SMCC 90 is a silicified microcrystalline cellulose.

The term "patient" encompasses all types of mammals, preferably humans.

The terms "treatment", "therapy" encompass treatment of pathological conditions in mammals, preferably in humans, and include: a) blocking (delaying) the course of disease, b) alleviation of the severity of the disease, i.e. induction of disease regression.

The terms "prophylaxis", "prevention" encompass elimination of risk factors, as well as preventive treatment of subclinical stages of the disease in mammals, preferably in humans, directed at reducing the likelihood of clinical stages of the disease. Patients for preventive therapy are selected on the basis of factors which according to the current data lead to an increased risk of developing clinical stages of the disease as compared with general population. Preventive therapy includes a) primary prevention and b) secondary prevention. Primary prevention is defined as preventive treatment of patients who have not yet had a clinical stage of the disease. Secondary prevention refers to the prevention of repeated onset of the same or similar clinical condition of the disease.

The term "therapeutically effective amount" refers to the amount of compound sufficient for administration to patient for prevention and/or treatment of a condition associated with increased thrombus formation. The precisely required amount may vary from subject to subject depending on the species of mammal, the age and general condition of patient, the severity of disease, the method of drug administration, cotreatment with other drugs, etc. The therapeutically effective amount of the compound which is an active component of the pharmaceutical compositions of the invention administered once or in several separate doses usually lies in the range of 0.01 to 500 mg/kg, preferably from 0.1 to 125 mg/kg, even more preferably from 1 to 100 mg/kg, but is not confined thereto. The compound is usually administered to a patient requiring such treatment at a daily dose of approximately 50 to 2000 mg per patient. The dose can be administered once or several times a day, a week (or any other time interval), or from time to time. Additionally, the compound can be administered to the patient each day for a certain period (for example, 2-10 days), followed by a period without taking any substances (for example, 1 -30 days) or daily up to patient's death.

If the compositions of this invention are used as part of a combined therapy regimen, the dose of each component of the combined therapy is administered within the required period of treatment. The compounds included in the combined therapy can be administered to the patient either simultaneously, in the form of a dosage comprising all components, or in the form of individual doses of components.

The "dosage form" in this document means the state (composition) of a medical product that corresponds to its methods of administration and application and ensures the achievement of the required therapeutic effect.

The term "thrombosis" means the formation or presence of a thrombus, intravascular coagulation that can cause ischemia or infarction of tissues supplied by the vessel.

The term "embolism" means the blockage of a blood or lymphatic vessel by a clot or other foreign matter which was migrated within the bloodstream (lymph) to the site of blockage The term "thromboembolism" means the occlusion of a blood vessel with a thrombotic material brought from the site of origin to another blood vessel.

The mammal's state characterized by undesirable thrombosis herein describes diseases the etiology or pathogenesis of which are associated with thrombosis, such as thrombosis and thromboembolism.

The production of the compound N-(5-chloropyridine-2-yl)-2-({4-

[etanimidoil(methyl)aminolbenzoyl|amino)-5-methylbenzamid e.

Reaction 1 . The production of N-(5-chloropyridine-2-yl)-5-methyl-2-nitro benzamide.

0.8 g of 5-methyl-2-nitrobenzoic acid is boiled in 20 ml of SOCI 2 with a reflux condenser equipped with a calcium chloride tube for 4 hours, thereafter the obtained solution is cooled, evaporated in a rotary evaporator, twice reevaporated with anhydrous THF, the residue is dissolved in 10 ml of THF, the obtained solution is added dropwise for 30 minutes to a stirred solution of 1 .5 g of 2-amino-5-chloropyridine in 20 ml of THF. In 5 hours, the reaction mixture is evaporated, the residue is dissolved in 30 ml of chloroform, rinsed with a saturated aqueous solution of NaHC0 3 , the chloroform extract is evaporated, the residue is applied on a column of 40x150 mm packed with 30-50 μηι of silica gel. The product is eluted with chloroform. Detection is carried out by using an UV cell at a wavelength of 280 nm. The UV-absorbing fractions are collected, the purity of the product is controlled with a thin layer chromatography technique in chloroform. The yield is 650 mg of N-(5-chloropyridine-2-yl)-5-methyl-2-nitrobenzamide The Rf of the product is 0.45, The Rf of the starting compound 2-amino-5-chloropyridine is 0.7. The mass spectrum (MALDI-TOF): M + H 292, M + Na 314.

Reaction 2. The production of 2-amino-N-(5-chloropyridine-2-yl)-5-methylbenzamide.

0.6 g of N-(5-chloropyridine-2-yl)-5-methyl-2-nitrobenzamide is dissolved in 20 ml of ethyl acetate and mixed with a solution of 4 g of SnCI 2 in 20 ml of water acidified with 0.3 ml of concentrated HCI. The reaction mixture is vigorously stirred for 1 hour, after that it is heated to a boiling point and kept boiling for 3 hours. The reaction mixture is filtered, the aqueous fraction is extracted with CHCI 3 , thereafter 30 ml of 10% aqueous ammonia solution is added to the aqueous fraction with stirring and left standing overnight to form a precipitate. The next day the precipitate is filtered, rinsed with water and chloroform. The aqueous fraction is extracted with chloroform, the extracts are combined and evaporated. The residue is applied on a 30x 150 column packed with 40-60 μηι of silica gel. The product is eluted with chloroform, detection is carried out by using an UV cell at 280 nm, the product purity is controlled by using a TLC technique in chloroform. The Rf is 0.45. The yield of 2-amino-N- (5-chloropyridine-2-yl) -5- methylbenzamide is 350 mg. The mass spectrum (MALDI-TOF): M+H 262, M+Na 284.

Reaction 3. The production of 4-methylaminobenzoic acid trifluoroacetate

1 g of 4-methylamino-benzoic acid is mixed with 3 ml of trifluoroacetic anhydride. In 2 hours, the reaction mixture is evaporated and reevaporated with chloroform. The residue is dissolved in chloroform and applied on a 35x150 column packed with 40-60 μηι of silica gel. The by-products are eluted with chloroform, and the target product is eluted with a 9:1 chloroform/isopropanol mixture added with a 1 % acetic acid. The yield of 4-methylaminobenzoic acid trifluoroacetate is 700 mg. The Rf is 0.2-0.4. The mass spectrum (MALDI-TOF) in the negative ion mode: M "1 246.

Reaction 4. The production of the compound N- (5-chloropyridine-2-yl)-2-[(4-methyl (trifluoroacetyl)aminophenylcarbonyl)amino]-5-methylbenzamid e.

300 mg of 2-amino-N-(5-chloropyridine-2-yl)-5-methylbenzamide, 250 mg of 4- methylaminobenzoic acid trifluoroacetate and 250 mg of EDCI (1 -ethyl-3-(3- dimethylaminopropyl)carbodiimide) are mixed in 1 ml of THF and stirred for 3 days. Then the residue is dissolved in chloroform and applied on a 25x150 column packed with 40-60 μηι of silica gel. The product is eluted with chloroform. The Rf is 0.55. The yield of N- (5- chloropyridine-2-yl)-2-[(4-methyl(trifluoroacetyl)aminopheny lcarbonyl)amino] 5- methylbenzamide is 430 mg. The mass spectrum (MALDI-TOF) in the positive ion mode: M+H 491

Reaction 5. The production of the compound N-(5-chloropyridine-2-yl)-2-[(4- methylamino-phenyl-carbonyl)amino]-5-methylbenzamide. 400 mg of N-(5-chloropyridine-2-yl)-2-[(4-methyl(trifluoroacetyl)amino phenylcarbonyl) amino]5-methylbenzamide is dissolved in 5 ml of isopropanol and added with 2 ml of 10% NaOH The reaction mixture is stirred for 3 hours, then the excess alkali is neutralized with a 5% aqueous solution of HCI, the reaction mixture is evaporated and applied on a column of 25x150 packed with 40-60 μηι of silica gel The product is eluted with chloroform; Rf = 0.4. The yield of N-(5-chloropyridine-2-yl)-2-[(4-methylamino-phenyl-carbonyl) amino]-5-methylbenzamide is 300 mg. Rf=0.5 (chloroform). The mass spectrum (MALDI-TOF): M+H 395, M+Na 417.

Reaction 6. The production of the compound N-(5-chloropyridine-2-yl)-2-[(4-etanimidoil- methyl-aminophenylcarbonyl)amino]-5-methylbenzamide. 250 mg of N-(5-chloropyridine-2-yl)-2-[(4-methylamino-phenyl-carbonyl) amino]-5- methylbenzamide is dissolved with heating in 5 ml of acetonitrile. The obtained solution is cooled with ice and then a dry gaseous HCI is passed therethrough. In 30 min the solution is placed in a refrigerator and allowed to stand at 5°C for 48 hours. Then the reaction mixture is added with NaHC0 3 with vigorous stirring. New portions of NaHC0 3 are added until emission of gases ceases. Usually the procedure requires about 0.5 g of NaHC0 3 . The solution obtained upon neutralizing the excess with HCI is diluted with 5 ml of water and extracted with chloroform three times. The chloroform extracts are combined and evaporated. The residue is dissolved in water and applied on a 20x250 mm column packed with reversed phase of C2 (RP2) The column is rinsed with 100 ml of water; then, the elution with a gradient of ethyl alcohol of from 0 to 50% is carried out against a background of 1 % aqueous acetic acid. The yield of the product is about per 30% of ethyl alcohol. The purity control is carried out with a TLC technique in a 9:1 dioxane/aqueous ammonia system. Rf=0.2. The yield of (5-chloropyridine-2-yl)-2-[(4- etanimidoil-methyl-aminophenylcarbonyl)amino]-5-methylbenzam ide is 130 mg. Rf=0.2. The mass spectrum (MALDI-TOF): M+H 436, M+Na 458.

The pharmaceutical compositions suitable for oral administration to mammals, preferably a human, according to the invention can be prepared by the following method:

a) core formation - mixing of active ingredients (N- (5-chloropyridine-2-yl)-2-({4- [ethanimidoil (methyl)amino]benzoyl}amino)-5-methylbenzamide or a pharmaceutically acceptable salt thereof and polyprenol) with pharmaceutically acceptable excipients (for example, microcrystalline cellulose and/or lactose and/or others);

b) tableting of the tablet core and its coating with an enteric coating (for example, Aquarius™ Control ENA of the company Ashland) or placing the ingredients mixed at the preceding step of ingredients into enteric-coated capsule (for example, consisting of gelatin with addition of titanium dioxide);

c) packaging (for example, vials or blisters). Experiments comparing in vivo substance and tableted dosage forms comprising compound No. 1 carried out by using a single oral administration to rats Testing compound Ns1:

Chemical name: N- (5-chloropyridine-2-yl)-2-({4-

[etanimidoil(methyl)amino]benzoyl}amino)-5-methylbenzamid e hydrochloride.

Chemical formula:

Empiric formula: C 23 H 22 CIN 5 0 2 I-ICI.

Molecular weight: 472.37 g/mol.

Description: White or almost white crystalline powder.

Foreign impurities: The content of an individual unspecified impurity is not more than 0.5%, the amount of impurities is not more than 1 .0%.

Water: not more than 1 .0%

Assay: 99.0-101 .0% (calculated on the anhydrous basis)

Microbiological purity: Category 1 .2.B.

Storage conditions: in protected from light place at 2-8 °C. Finished dosage form (FDF No.1) comprising compound No1:

Composition of one tablet:

Constitution of the nucleus:

Active compound:

The compound No1 (FSUE RDC "Pharmzaschyta" of 10 wt%

FMBA of Russia),

calculated on the anhydrous and residual organic solvent- free basis

Auxiliary compounds: Prosolv® SMCC 90 79.5 wt%

Magnesium stearate 0.5 wt%

Coating composition:

Aquarius I M Control ENA White 10 wt%

Weight of a tablet: 10 mg

Method of preparation: The ingredients indicated for the core were mixed, tableted by direct compression to obtain a tablet core, and then the tablet core was coated in a drum type device.

Finished dosage form (FDF No.2) comprising compound No1:

Composition of one tablet:

Constitution of the nucleus:

Active compound:

the compound No1 (FSUE RDC "Pharmzaschyta" of 10 wt%

FMBA of Russia),

calculated on the anhydrous and residual organic solvent- free basis

Auxiliary compounds:

Polyprenol (the purity is 98%), obtained from larch needles 10 wt%

(Representative Office of Solagran Ltd in Russia)

Prosolv® SMCC 90 69.5 wt%

Magnesium stearate 0.5 wt%

Coating composition:

Aquarius™ White 10 wt%

Weight of a tablet: 10 mg

Method of preparation: The ingredients indicated for the core were mixed, tableted by direct compression to obtain a tablet core, and then the tablet core was coated in a drum type device.

Rats are standard objects of preclinical studies focused on the specific pharmacological activity of potential medicinal products - anticoagulants. Animal experiments were carried out according to the legal and ethical norms for handling of animals in accordance with the rules adopted by the European Convention for the Protection of Vertebrate Animals used for experimental and other scientific purposes [2].

The testing substance- compound No.1 in the form of a tablet (FDF No.1 ), in the form of a tablet with polyprenol (FDF No.2) or simply substance of the compound No.1 - were administered intragastrically (intragastric administration). The individual dose administered to each animal was calculated on the basis of body weight and corrected after each weighing. Compound administration: single dosing.

Intragastric administration was used as this route of drug administration is identical to the planned one in a clinical practice.

The conventional animals were used in the experiments, the total number was 48 Wistar rats.

Animal suppliers: rats - the branch "Andreevka", SCBT RAMS (Scientific Center for Biomedical Technology of the Russian Academy of Medical Sciences) The animals were purpose-bred and not used in studies before. The manufacturer has provided the latest data of animal health monitoring (veterinary certificate).

Weight of animals: the animals used in the study were preferably of the same weight. The weight of rats is 280-320 g.

Cages for keeping animals: animals were kept in polypropylene cages of 460x300x160 mm, 4 rats in each, wood shavings were used as a litter for laboratory animals.

Feed: rats were fed with standard granulated complete combined feed (extruded) PK-

120, according to GOST R 51849-2001 R.5 Feed was placed ad libitum in the deepening of the cage or in the food bowl.

Water: Tap water was given ad libitum in 500 ml glass drinking bowls.

Environmental parameters: Animals were kept under controlled environmental conditions: at a temperature of 18-22°C and 50-70% relative humidity.

Daily temperature and humidity measurements in each experimental room were being recorded by hand. Lighting in rooms - natural-artificial (12 hours - day, 12 hours - night).

Quarantine/Acclimatization: The branch "Andreevka" of FPFIS SCBT FMBA of Russia (Federal Publicly Funded Institution of Science Scientific Center for Biomedical Technology of the Federal Medical Biological Agency of Russia) is safe for especially dangerous and quarantinable diseases. The animals were kept for 30 days in quarantine before shipment. There were no violations of transportation. Quarantine at the laboratory facilities was reduced to 4 days to enable animal adaptation. During this period, the rats were healthy and did not have any behavioural disorders.

Identification of animals: each animal in the cage was assigned an individual number marked on its wool. The cages were provided with labels indicating the study, its type, sex and group of animals, time frame for study.

Allocation to groups: Before the beginning of the study, the required number of animals meeting the inclusion criteria for the study were allocated to groups by the method of randomization depending on the body weight.

Design of experiment: Table 1 Dosing regimen of the substance and tableted dosage forms comprising compound No.1 for rats.

48 rats were used as models.

12 hours prior to experiments and during the whole experiment the rats were not given any feed. Before and during the experiments, rats were given water ad libitum.

Experimental procedure:

The rats were weighed. They were allocated to groups by the method of randomization depending on their body weight.

2 ml of aqueous solution comprising compound No.1 in the calculation of 15 mg of the active compound No.1 per 1 kg of live weight was administered intragastrically to the first test group of 16 rats;

a tablet of FDF No.1 in the calculation of 15 mg of the active compound No.1 per 1 kg of live weight was administered by gavage using a metal probe to the second test group of 16 rats; a tablet of FDF No.2 in the calculation of 15 mg of the active compound No.1 per 1 kg of live weigh was administered by gavage using a metal probe to the 3rd test group of 16 rats;

The blood sampling was carried out without anaesthesia from the tail vein, preheating the rat's tail in a water bath at 44° C for 10 minutes.

For blood sampling, each group of animals was divided into two identical subgroups (i. e two subgroups of 8 animals in each), wherein the blood sampling was carried out before the beginning of the experiment in all animals and after 0.25 h, 0.5 h, 1 h, 2 h, 3 h in eight animals of the first subgroup, and also after 4h, 6h, 8h, 10h and 24h following the administration of the substance to the remaining eight animals of the second subgroup. The blood sampling was carried out using a 23G butterfly needle into plastic microtubes (such as Eppendorf microtubes) comprising 50 μΙ of 0.1 1 M sodium citrate solution to the volume of 0.5 ml in the way that the ratio of sodium citrate solution to blood was 1 : 9.

Within 30 minutes after the collection, the blood was centrifuged at 1000 rpm (240 g) for 7 minutes for obtaining platelet-poor plasma, the plasma was transferred to another tube and recentrifuged at 3000 rpm (1200 g) for 15 minutes at 20°C.

The obtained plasma was poured into plastic tubes (such as Eppendorf tubes) and frozen at -70°C. Not later than in 1 week, the anticoagulant activity of the plasma being thawed at 37° for 1 hour was measured together with prothrombin time (PT). Determination of prothrombin time (PT).

The prothrombin time was determined by the method described in the instructions to the Renamplastin reagent kit made by RENAM Research-and-Production Association [3].

Prothrombin time is a highly sensitive and simple screening test that can be used to determine blood clotting/coagulation pathway disorders (sensitive to concentration changes of the factors II, V, VII, and X).

Principle of the method: When adding a tissue thromboplastin excess and calcium ions to citrate plasma, the time of fibrin clot formation depends only on the activity of the external and general coagulation pathway factors: Factors I, II, V, VII, X. The time from the moment of addition to the thromboplastin plasma with calcium until the formation of the fibrin clot is measured.

Carrying-out of an analysis: 50 μΙ of citrated plasma was transferred into a cuvette of the analyzer and incubated at 37°C for exactly 1-2 minutes. Then, 100 μΙ of renamplastin was added and the coagulation time in seconds was recorded on the coagulation analyzer.

The obtained results were expressed in the International Normalized Ratio (INR).

INR= PR ISI ,

wherein ISI denoted International Sensitivity Index of Renamplastin, which is indicated in the attached test-system data sheet. PR - prothrombin ratio.

PR = PT B / PT 100% ,

wherein PT B denotes the prothrombin time of the testing rat plasma (in seconds), PT 100% denotes the prothrombin time of the same rat plasma collected before the beginning of the experiment (in seconds).

To evaluate the efficacy of FDF No1 and FDF No2 intragastrically administered to rats, the influence of the administration of the compound No1 , FDFNol and FDF No2 on PT (INR) values was studied.

The dependence of INR and PT on time of post administration of the compound No.1 and FDF No1 and FDF No2 in rat plasma is shown in Table 2 and in Figure 1 . Table 2 The PT values with intragastric administration of the compound No.1 , FDF No.1 and FDF No.2 to rats.

22.20 26.10 31 .10 51 .10 44.30 15.90

22.10 45.00 41 .90 28.20 33.90 20.60

24.10 33.80 38.10 40.30 23.20 21 .10

21 .70 44.00 35.90 32.20 35.60 17.90

18.90 28.40 37.40 37.10 34.00 17.20

20.40 38.20 47.20 45.60 37.80 22.10

FDF No2

Post administration time, h

0 0.25 0.5 1 2 3

PT, sec.

20.1 21 .3 26.1 24.9 39.8 41 .4

Dose- 21 .8 20.3 25.4 23.4 48.4 48.2

15 mg/kg 19.9 21 .6 19.7 21 .6 39.0 47.0

18.0 20.5 20.7 23.7 45.2 57.3

22.2 21 .0 18.9 30.6 42.3 54.4

18.6 22.8 26.1 33.2 39.8 45.1

19.8 21 .3 22.8 28.1 32.3 29.7

18.6 22.0 19.8 19.9 43.2 39.8

FDF No2

Post administration time, h

0 4 6 8 10 24

PT, sec.

18.4 42.5 39.0 52.2 38.5 18.4

Dose- 17.9 55.4 53.9 56.3 39.9 23.3

15 mg/kg 17.5 45.6 46.8 79.5 56.0 19.7

20.2 56.6 79.2 36.6 34.9 24.1

22.4 38.8 49.0 48.1 40.3 18.5

22.6 71 .0 52.2 47.0 48.9 23.7

23.2 58.6 63.8 50.1 34.4 16.2

20.5 64.3 69.1 64.7 47.2 27.7

The experiment results have shown that the compound No.1 administered intragastrically to rats increased INR, and in 4-10 hours after administration and up to 24 hours, the INR decreased again to the initial value. The efficacy of FDF No.1 at the same dose is in 2- 2.5 times higher than that of pure compound No.1 that is obviously due to the fact that the compound No.1 reaching the stomach is partially destroyed there by gastric juice, and the FDF No.1 is protected from destruction by coating. The efficacy of FDF No.2 at the same dose is in 5-7 times higher than that of pure compound No. 1 , that is obviously due to the presence of polyprenol in this dosage form. The obtained data indicate that the use of FDF No.2 will reduce the dose of the active substance and this, in turn, can lead to a reduction of incidence of adverse effects

Thus, the pharmaceutical compositions according to the invention, in particular the pharmaceutical compositions comprising compound No.1 and polyprenol protected by an enteric coating or capsule from the contact with acidic gastric juice are more efficient as an anticoagulant than the compound N-(5-chloropyridine-2-yl)-2-({4-

[ethanoydoil(methyl)amino]benzoyl}amino)-5-methylbenzamid e or a pharmaceutically acceptable salt, in particular, the compound No.1 by itself, and are promising for the treatment of disorders associated with blood-clotting disorder.

Despite the fact that the invention has been described with reference to the forms of invention embodiment, it should be obvious to the persons skilled in the art that the specific, detailed described experiments are shown for the purpose of illustrating this invention only, and should not be considered as those that in any way confine the scope of the invention. It should be understood that the embodiment of various modifications are possible without deviation from the main point of this invention.

References:

1 Hellenic J Cardiol. Parenteral Anticoagulants in Acute Coronary Syndromes and Interventional Cardiology// A Consensus Document. - 2015. - Sep-Oct - V56(5). - P.379- 393.

2 European Convention for the Protection of Vertebrate Animals Used for Experimental and other Scientific Purposes (ETS 123). - Strasbourg. - 1986.

3 http://www.renam.ru

4 Swiezewska E., Sasak W., Mankowski T., Jankowski W., Vogtman T., Krajewska I., Hertel J.,

Skoczylas E., Chojnacki T. The search for plant polyprenols // Acta Biochim. Polon. 1994. V. 41 . P. 221 -26.