LV14274B - Combination medicinal product comprising meldonium - Google Patents

Abstract

Pharmaceutical composition of gamma-butyrobetaine and Meldonium dihydrate and its use in prevention and/or treatment of atherosclerosis and/or neurological diseases.

Description

DESCRIPTION OF THE INVENTION

FIELD OF THE INVENTION

The present invention relates to the use of a new medical product containing meldonia and a new medical product containing meldonia.

More particularly, the present invention relates to a novel medical use of meldonium or a pharmaceutically acceptable salt thereof in combination with gamma-butyrobetaine or a pharmaceutically acceptable salt thereof, and to a novel medical product containing meldonium succinate in combination with gamma-butyrobetaine succinate. The present invention also relates to a pharmaceutical composition comprising such a medical product and its use in the manufacture of a medicament for the prevention and / or treatment of certain disorders. The present invention also relates to the prevention and / or treatment of known disorders. The new medicinal product containing Meldonium succinate in combination with gammabutyrobetaine succinate relates to the prevention and / or treatment of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease and angina pectoris, myocardial infarction, heart failure, transient and persistent ischemic attack, including cerebrovascular accident and stroke, peripheral arterial occlusive disease. The new medical application for Meldonium or pharmaceutically acceptable salts thereof in combination with gamma-butyrobetaine or its pharmaceutically acceptable salts relates to the prevention and / or treatment of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, non-atherosclerosis, atherosclerosis, , transient and persistent ischemic attacks including cerebrovascular accident and stroke, peripheral arterial occlusive disease. SUMMARY OF THE INVENTION

Currently, atherosclerosis and stroke are among the leading causes of mortality. Elevated levels of low-density lipoprotein (LDL) are generally considered to be high risk factors for atherosclerosis and related cardiovascular pathologies. Because low HDL levels are a risk factor for stroke [Sacco RL, Benson RT, Kargman DE, JAMA 2001; 285: 2729-2735. Wannamethee SG, Shaper AG, Ebrahim S, Stroke 2000; 31: 1882. Sanossian N, Tarlov NE, Curr Treatmt Opt Cardiov Med 2008; 10 (3): 195-206. Rizos E, Mikhailidis DP, Cardiovasc Res 2001; 52 (2), 199-207], New HDL-elevating agents are useful for the prevention and treatment of stroke [Koniukov SG, Liberzon SP, Klin Med (Mosk) 1975; 53 (9): 38-41].

TECHNICAL LEVEL

The use of a combination of meldonium or 3- (2,2,2-trimethylhydrazinium) propionate (also known as Mildronate ™) and gamma-butyrobetaine (GBB) for the treatment of cardiovascular conditions (WO 1997/006794) or for the treatment of impotence in male mammals is known (WO 2003 / 022262).

Meldonium has been described as a clinically useful anti-ischemic and anti-stress agent for the treatment of some cardiovascular diseases and other pathologies caused by tissue ischemia [Karpov RS, Kardiologiya (Rus) 2000; 6: 69-74]. In an experimental model of acute myocardial infarction, meldonium has been shown to reduce the size of myocardial infarction without affecting hcmodynamics [Sēsti et al, J Cardiovasc Pharm 2006; 47 (3): 493-499]. Meldonium is also indicated for the treatment of chronic cerebrovascular insufficiency [Dziak LA, Golik VA, Lik Sprava (Rus) 2003; (5-6): 98-101]. Meldonium has been shown to have antioxidant activity and protect against lipid peroxidation in the clinic [Suslina ZA et al, Eksp Klin Farmakol (Rus) 2003; 66 (3): 32-35]. Known valid activity of meldonium has been found in animal models of alciosclerosis (Veveris M, Smilsaraja B, White JLabAnimSci 2000; 10; 194-199. Veveris M et al, White JLabAnimSci 2002; 12: 116-122. Okunevich IV, Ryzhenkov Eksp, Patol Fizi) Ter (Rus) 2002; (2) 24-7] and observed in a clinic [Karpov RS et al, Ter Arkh (Rus) 1991; 63 (4): 90-3 Luk'yanov YuV, Gorbunov GN, Angiology and vascular surgery (Rus), 1997; (2), 64. Gorbunov GN, Terra Medica 1997; (l): 42-43].

GBB has not been reported to have antiatherosclerotic activity.

OBJECT OF THE INVENTION

The present invention relates to a novel medical use of a known combination product comprising mcldonia or a pharmaceutically acceptable salt thereof and gamma-butyrobetaine or a pharmaceutically acceptable salt thereof for the prophylaxis and / or treatment of a disorder selected from the group consisting of dyslipidemia. , hypo-lipidemia, alciosclerosis, transient and persistent ischemic attacks, including cerebrovascular accident and stroke, and peripheral arterial occlusive disease.

Another object of the invention is a novel combined medical product containing meldonium succinate and gamma-butyrobetaine succinate.

The present invention also relates to a pharmaceutical composition comprising such a combination medical product and its use in the manufacture of a medicament.

The medical product of the present invention is for the treatment and / or prophylaxis of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease, angina pectoris and myocardial infarction, chronic heart failure, transient and ischemic heart disease, and stroke, peripheral arterial occlusive disease.

The term "combined pharmaceutical product" as used herein refers to the simultaneous, sequential or separate administration of the combination agents. A further object of the present invention is a pharmaceutical composition comprising meldonium succinate and gamma-butyrobetaine succinate for said purpose. Other objects of the invention will become apparent later and others will be obvious to the skilled artisan.

DESCRIPTION OF THE INVENTION

The invention relates to a medical product, preferably in unitary dosage form. It does not matter in what form the active ingredient is administered, as long as the desired effect of the invention is achieved. The active ingredient may be in the form of capsules, suspensions, dispersions, elixirs, syrups and the like.

We have unexpectedly found that the combination of meldonium or a pharmaceutically acceptable salt thereof with gamma-butyrobetaine or a pharmaceutically acceptable salt thereof exhibits known beneficial effects that are significantly superior to the meldonium effects. Unexpectedly, meldonium in combination with gamma-butyrobetaine was found to lower triglyceride and low-density lipoprotein (LDL) levels and to increase high-density lipoprotein (HDL) levels better than meldonium alone. We have also surprisingly found that the product of the present invention improves upon experimental ischemia and stroke. This combination of more pronounced beneficial properties makes the combination of Meldonium with gamma-butyrobetaine particularly useful for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, chronic heart failure, transient and persistent ischemia, including cerebrovascular accident and stroke, peripheral arterial occlusive disease. We have found that these beneficial properties are also present in the combination of active ingredients in the form of salts, in particular the combination of meldonium succinate with butyrobetaine succinate. This combination consists of meldonium succinate (WO 2005/012233) and previously unknown gamma-butyrobetaine succinate, substances which were unknown when the pharmacological properties of the combination of meldonium and gamma-butyrobetaine were investigated and disclosed in WO1997 / 006794. This new medicinal product also has outstanding activity in the treatment of known cardiovascular diseases, which significantly exacerbate the activity of the previously discovered combination of meldonium and gamma-butyrobetaine.

The medical product of the present invention may be suitable for oral administration (e.g., as tablets, capsules, aqueous suspensions or dispersible powders or granules), for parenteral administration (e.g., as a sterile aqueous solution for intravenous, subcutaneous or intramuscular administration), or as suppositories for rectal administration. Optimally, the composition of the invention is in a form suitable for oral administration, such as tablets or capsules.

The pharmaceutical composition of the invention may be obtained by conventional procedures using conventional pharmaceutically acceptable diluents or carriers and technologies.

In additional aspects, the present invention provides a method of preventing and / or treating a disorder wherein the disorder is selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease from the group of angina and myocardial infarction, chronic heart failure, transient and persistent heart failure. , including cerebrovascular accident and stroke, peripheral arterial occlusive disease involving the administration of a combination of meldonium succinate with gamma-butyrobetaine.

In another aspect, the present invention provides the use of a novel combined medical product for the manufacture of a medicament for the prevention and / or treatment of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease, angina and myocardial infarction, failure, transient and persistent ischemic attack, including cerebrovascular accident and stroke, peripheral arterial occlusive disease.

The following examples are provided by way of illustration and do not limit the invention.

EXAMPLES

Tests: The pharmacological activity of the compounds tested was investigated by standard methods used in the art. The animals were housed in groups of 6 suitable cages in air-conditioned rooms at 22 ± 1 ° C with a relative humidity of 60 ± 5% and a 12/12 hour light / dark cycle with free access to food and water. All experiments were carried out in accordance with Council Directive 86/609 / EEC of 24 November 1986 on the protection of animals. Every effort was made to minimize animal suffering and reduce the number of animals used.

Substances: Cholesterol (Acros Organics), Meldonium (Grindex), Feed (R 70 Lactamin), Butter (Shop), Na Cholate (Acros Organics). The previously unknown gamma-butyrobetaine succinate (GBBS) was prepared as a white crystalline powder with k.t. 8688 ° C, stable and non-hygroscopic at room conditions. Meldonium succinate (MDS) was prepared by modification of the method described in WO 2005/012233 and was obtained as a colorless crystalline powder, stable and non-hygroscopic under room conditions. All other chemicals are obtained from commercial sources.

ABBREVIATIONS USED

For the sake of brevity, the following abbreviations will be used in the description below:

ATP - adenosine triphosphate

H - Cholesterol

GBB - gamma-butyrobetaine

GBBS - gamma-butyrobetaine succinate, chemically: Addition of 4- (N, N, N-trimethylammonio) butanoate with one molecule of succinic acid, anhydrous HDL - high density lipoproteins

I / R - Ischemia / Reperfusion LDL - Low-density lipoproteins

MD - Meldonium (SNN), chemically: 3- (2,2,2-trimethylhydrazinium) propionate, dihydrate MDS - Meldonium succinate (1: 1), chemically: 3- (N, N, N-trimethylhydrazinium) propionate addition salt with one molecule of succinic acid, anhydrous

TG triglyceride

KJF - Kamban Fibrillation

KT-chamber tachycard

Effect on lutein levels

Example 1: Model of rat hyperlipidemia

The method. The expression method proposed by Arichi et al [Chem Pharm Buli 1982; 30 (5): 1766-1770] was used to induce hyperlipidemia. Wistar male males weighing 220-250 g were used. Rats received a mixture of fat-cholesterol (10% cholesterol and 1% cholic acid in olive oil p.o. at a dose of 10 ml / kg) for 10 days. Solutions of test substances in water or water for intact or cholesterol (H) control groups were administered one hour after the administration of the fat mixture. Blood for analysis was taken on day 11 with cardiac puncture under ether anesthesia.

Biochemistry. Total H, HDL, LDL and TG were detected with COBAS INTEGRA 400 (Roche). The atherosclerosis index (which represents coronary atherosclerosis and is a discriminator of peripheral atherosclerosis) was calculated as follows: Index = LDL / HDL.

Atherosclerosis coefficient (reflecting coronary atherosclerosis) was calculated as follows: Coefficient = Total H / HDL

Statistics. The resulting data were mathematically processed using Microsoft Excel and expressed as mean ± standard deviation (SD). Results from different groups were compared using one-way analysis by ANOVA and Student's t-test. The difference in results was considered significant at P <0.05.

Results. In these experiments, MD was compared to MD + GBB or MDS + GBBS. The results are given in Table 1 below. Rats receiving the fat mixture developed severe hypercholesterolemia and hyperlipidemia. Overall H, LDL and TG values were significantly different from intact control animals. In this model, combined administration of MD and GBB for 10 days lowered TG levels and atherosclerosis index better than administration of MD (P <0.05 vs. H control for MD + GBB and insignificant change from control to MD). In an amazing way

Administration of MDS + GBBS significantly reduced not only TG levels and atherosclerosis index, but also LDL levels and total H / HDL ratios. This fact indicates that the combinations MD + GBB and MDS + GBBS are useful in the prevention and / or treatment of hypercholesterolaemia and hyperlipidemia and may find application in the prevention and / or treatment of atherosclerosis.

Table 1

Effect of MD + GBB and MDS + GBBS on lipid levels in a rat hyperlipidemia model; Mean + VSN, n = 6

Group Total H mg / dl ABL mg / dl LDL mg / dl indix LDL / ABL Coefficient Common H / ABL TG at / dl Intact control 60 + 4.7 · * 45.5 + 4.5 11.2 + 1.8 ·· 0.28 + 0.06 ** 1.31 + 0.08 ·· 57.9 + 6.5 * H control 165 + 14.0 50.6 + 1.8 08.4 + 12.9 2.15 + 0.23 3.18 + 0.18 102.3 + 11.0 MD 146 + 12.7 54.3 + 5.0 88.9 + 13.5 1.70 + 0.37 2.73 + 0.36 78.4 + 9.8 MD + GBB 131 + 14.2 51.0 + 2.6 76.4 + 11.1 1.47 + 0.17 * 2.54 + 0.19 65.5 + 8.7 * MDS + GBBS 133 + 11.9 56 7 + 4 4 73.1 + 8.5 *. 1.32 + 0.22 * 2.37x0.23 * 54.3 + 4.3 *

MD = MD 150 mg / kg

MD + GBB = MD 150 mg / kg + GBB 50 mg / kg MDS + GBB = MDS 150 mg / kg + GBBS 50 mg / 'g * P <0.05 vs. H control ** P <0.005 vs. H control

When investigating MD and GBB salts, it was unexpectedly found that MD and GBB succinic salts exhibit significantly higher activity, especially in the combination of MDS and GBBS. Subsequent experiments with combinations containing MD and GBB salts led to the mouse atherosclerosis model described below.

? ·. example. The mouse; modefis of terosclerosis

The method. The mouse model of atherosclerosis described in the literature [Smith JD, Breslow JL, J Intern Med 1997; 242: 99109] was used. C57BL / 6 female mice used in the experiment, having an average weight of 20.3 g at the start of the experiment, were supplemented with cholesterol and fat in standard feed. In accordance with the recommendations of [Paigen B et al., Atherosclerosis 1987; 68: 231-240 and Nishina P et al., J Lipid Res, 1993; 34: 14131422], cholesterol (1.25%), butter (15%) and sodium cholate (0.5%) were added to the standard feed (except for the intact control group).

The study groups are shown in Table 2. After 22 weeks, the degree of atherosclerotic lesions was assessed by recognized morphological, biochemical, and histological methods and criteria. Final animals were anesthetized (pentobarbital, 60 mg / kg, ip). After laparotomy, blood was removed from the abdominal vein. The blood sample was centrifuged (2000 x min ' ¹ , 15 min) and the resulting serum stored at -20 ° C until biochemical tests were performed. The aortic arch, ascending and descending aorta were isolated and retained for further studies in 4% buffered paraformaldehyde solution. Lipid determination in the aortic base was performed based on the technique developed by Paigen et al. [Paigen B et al., Atherosclerosis 1987; 68: 31-240]. Aortic sections (10 pm) were fixed and stained with Oil red-0 and hematoxylin. The defective area (mean area pm ^{2 in} section) was evaluated by light microscopy at serial intervals of 80 pm starting from the aortic valve. The total area of atherosclerotic changes in the inner surface of the aorta was determined by ss en face technique. Longitudinal open aortic lipids were stained with Sudan IV or Oil red-O. Samples were captured and images analyzed using an Image-Pro Plus 4.5.1 planimetry application. Atherosclerotic changes were expressed as percent lipid infiltrate of total aortic area. Six to eight samples were analyzed per animal group.

Biochemistry and Statistics as described above.

Results. After 22 weeks, all untreated cholesterol control animals had lipid infiltrate in the aorta with an area of 5.62% of the animal's aortic space (Table 2, below). MD treatment significantly reduced the damaged aortic space. The combination of MD and GBB was better than MD in reducing the area of atherosclerotic lesions in the aortic root (P <0.05 versus H in the MD + GBB group, but no significant change in the control in the MD group). The combination of MDS and GBBS resulted in an even greater reduction in the aortic root lesion area, which was significantly better than MD and highly reliable (P <0.005 versus H control and P <0.05 versus MD; Table 2).

Table 2

Level of atherosclerotic changes in aortic root (A) and inner surface (B); Average + VSN n-6-8

Group Damaged area (A), mkm ¹ Defective Area (B)% Intact control 36 + 15 · ** 0.03 + 0.02 * · H control 5127 + 1109 5.62 + 1.01 MD ISO mg / kg 4294 + 785 3.49 + 0.42 * MD + GBB (150, 50 mg / kg) »474 + 224 * 3.21 + 0.24 * MDS + GBBS (150 + 50 mg / 3) . '«(8'247 ° * 2.70 + 0.36 *

* P <0.05 vs. H control ** P <0.005 vs. H control *** P <0.0005 vs. H control * P <0.05pretMD

Biochemical analysis showed that the combination of MD and GBB significantly reduced the atherosclerosis rate (Table 3 below). The combination of MDS and GBBS was surprisingly even more effective in improving lipid fraction ratios and, in contrast, MD significantly reduced TG and LDL levels as well as the atherosclerosis coefficient (Table 3).

Table 3

Effect of combination of MD and MD5 salts with GBB and GBB on lipid levels; Mean + VSN, n = 6-8

Group TOTAL H mg / dL ABL mg / dl LDL mg / dl Index, LDL / ABL Cot Patient Knpey H / HDL TG mg / dl Intact control 66.5 + 3.2 * ·· 57 + 4.3 9.7 + 1.94 * ·· 0.17 + 0.03 ··· 1.17 + 0.03 * · * 223 + 1.98 *** H control 161 + 8J2 50 + 4.3 111 + 6.9 2.23 + 0.21 3.22 + 0.21 48.1 + 2.2 MD 150 mjks'd 154 + 8.1 55 + 3.3 95 + 5.5 1.72 + 0.10 * 2.81 + 0.19 43.2+ 1.95 MD + GBB 150 + 50 mg / kgfd 152.86 59 + 5.1 93 + 95 1.63 + 0.12 * 2.63 + 0.12 » 43.0 + 2.24 MDS + GBBS 150 + 50 mg / kg / d 142 + 8.2 59 + 5.0 83 + 4.5 * 1.45 + 0.11 *** 2.45 + 0.11 *** 38.8 + 1.68 · *

* P <0.05 vs. H control ** P <0.05 vs. H control *** P <0.005 vs. H control * P <0.05 vs. MD

Effects on the heart and blood circulation

Example 3:

Rat infarction model

The method. Wistar male males weighing 330-360 g were randomized to 3 groups:

1) I / R control (n = 15), animals received p.o. after infarction / reperfusion for 7 days. 2 ml / kg 0.9% NaCl solution

2) MD + GBB group (n = 12), animals received p.o. 150 mg / kg MD and 50 mg / kg GBB

3) MDS + GBBS group (n = 12), animals received p.o. 150 mg / kg MDS and 50 mg / kg GBBS.

One hour after treatment, the animals were anesthetized (pentobarbital sodium 50 mg / kg i.p.) and prepared by mechanical ventilation to block the left coronary artery. The experimental infarction was induced by a 45-minute occlusion of the left coronary artery followed by 2 hours of reperfusion. After an experiment with perfusion staining using triphenyltetrazolium-Evans blue, an ischemic and necrosis zone was determined. The left ventricle was isolated, weighed, and morphological criteria were calculated: ischemic zone in piocentes from left ventricle, necrosis zone in percent of left ventricle, and ratio of necrosis zone to ischemic zone (necrosis index).

Statistics as above.

Results. Myocardial ischemia with subsequent reperfusion resulted in serious cardiac arrhythmias with 6 fatal outcomes in the control group. MD and GBB showed a tendency to reduce the number of animals with severe cardiac arrhythmias (CT and KF), but only the combination of MDS and GBBS resulted in an unexpectedly significant reduction in lethality (Table 4 below).

Table 4

Effects of study substances on rat cardiac arrhythmias and lethality from coronary artery occlusion (45 min) and reperfusion (120 min)

Group n KT KF Spirituality n % n % n % l'R knnlrcle 15th 14th 933 10th 67 6th 40 MD + G.-. Ū 12th 11th 91.7 6th 50 3 25th MDS + OUSS 12th 10th 83 4 40 1 · 83

* P <0.05 vs. I / R control

Morphological analysis of the myocardium showed that MD-GBB significantly reduced the proportion of the necrotic zone in the left ventricle and ischemic zone compared to the I / R control (Table 5). MDS + GBBS had a significantly greater effect on the necrosis index and necrotic area of the left ventricle than MD + GBB (P <0.005 versus P <0.05 vs. I / R control; Table 5).

Table 5

Effect of study substances on rat cardiac morphology after myocardium (45 min) ischemia and reperfusion (120 min); MeanVSN, n = 8-l 1

Group Kreisais Room, mg IZ / QC% ' NZ / MM% ² Necrosis index, ³ % I / R control 803.1 + 253 48,141,73 36,241.96 75.443.67 MD + GBB 8053418.4 48.042.10 25.8 ± 2.77 * 54,345.93 * MDS + GBBS 799.5420.5 48,041.57 24,842.23 * · 51,8,04431 * ·

* P <0.05 versus I / R control ** P <0.005 versus I / R control ¹ IZ / KK = Ischemic zone versus left ventricle x 100 ^J NZ / KK = Necrosis zone versus left ventricle x 100 ³ Necrosis index = Necrosis index zone mass / Ischemic zone mass x 100

Thus, we unexpectedly found that MDS + GBBS significantly better than MD + GBB in protecting against infertility and fatalities (with higher cardioprotective activity).

4, example. Model of rat chronic heart failure

The method. Wistar male males weighing 300-330 g were used. Chronic heart failure was caused by a known method in which heart failure occurred after experimental myocardial infarction (MI), induced by left coronary artery occlusion [Sasaki H et al, JMol Celi Cardiol 2001; 33: 283-294. Batista M et al, JAppl Physiol 2007; 102: 20332039]. Animals were anesthetized with pentobarbital sodium (50 mg / kg i.p.). The surgical field was prepared after the anesthesia and the animal was mechanically ventilated. Under sterile conditions, the chest and pericardium were opened and the left coronary artery, i; inheriting the thread loop at the left ventricular headset level. After 45 minutes of ischemia, the loop was removed and cardiac bleeding restored, reperfusion occurs. Reperfusion is conveniently detected after pre-ischemic cyanotic segment hyperemia. The thoracic wall is closed in layers, administered s.c. analgesics and place the animal on a heated plate until recovery from anesthesia. Control animals (Sham control group) were subjected to identical manipulations except for arterial occlusion. The animals that survived the heart attack were randomly distributed the following day 4:

1) Infarction group (M1 control, n = 8) received water p.o. once a day;

2) Sham control group (n = 8) received water p.o. once a day;

3) MD + GBB group (MD + GBB, n = 8), MD 150 mg / kg and GBB 50 mg / kg p o. once a day;

4) MDS + GBBS group (MDSWBBS, n = 8), MDS 150 mg / kg and GBBS 50 mg / kg p.o. once a day.

The animals were re-anesthetized and switched to mechanical ventilation on day 28 after Ml challenge. A catheter was introduced into the left ventricle and left ventricular pressure and contractility parameters (KK dP / dt max and dP / dt min obtained with EQ-601G differentiator) were recorded with a Polygraph system RM 6000 (Nihon Kohden, Japan). After recording basic contractility parameters (LV dP / dt max and dP / dt min), the left ventricular contractile functional reserve was determined as previously described [Kaga S et al, JMol Celi Cardiol 2006; 40: 138-147]. At the end of the experiment, additional anesthesia was injected and a saturated KC1 solution was administered via a femoral vein to arrest the heart in diastole. The heart was isolated, left ventricular mass was determined, and peak (diastole) volume was determined.

Statistics. Results were expressed as the mean of 6-8 experiments ± VSN. Differences between experimental groups were compared using one-way ANOVA with repeated comparisons (Tukey's test). P <0.05 was considered significant.

Results. Following experimental Ml animals, within 28 days, heart failure was characterized by a decrease in left ventricular systolic pressure (CKDS), increased left ventricular end-diastolic (KKDBS), and decreased left ventricular contractility (systoles + dP / dt and diastoles -dP / dt).

The MDS + GBBS group showed better protection against worsening of cardiac functional parameters after infarction than MD + GBB (MI control parameters KKSS, + pP / dt and -dP / dt were of higher significance, Table 6 below) .. MDS + GBBS better than MD + GBB protected cardiac systolic and diastolic function, maintaining it at the sham group level.

Table 6

Influence of test substances on hemodynamic parameters and left ventricular contractility (+/- dP / dt); Mean ± VSN; n = 6-8

Study group SF * KKSS ^b mmHg KKDBS ' mmHg + dP / dt ^d -dP / drt Sh m control group 397 + 6.5 148 + 5.9 * 3.9 + 0.68 ** 5325 + 201 *** 4435 + 175 ** Mf control 385 + 6.2 128 + 3.8 12.1 + 1.70 3674 + 189 3091 + 221 MI + (MD + GBB) 391 + 3.7 137 + 2.1 63 + 0.84 * 4608 + 173 * 4019 + 230 * MI + (MDS + GBBS) 389 + 5.4 141 + 2.8 * 6.1 + 0.82 * 4705 + 146 * · 4083 + 107 **

* P <0.05 vs. Ml control ** P <0.005 vs. Ml control ♦ ** F <0.0005 for Ml control * SF - heart rate, beats per minute ^b KKSS - ductal ventricular systolic pressure, mm Hg;

'KKDBS - end ventricular end-diastolic pressure, mm Hg; ^d + dP / dt - left ventricular contractility, systolic, mmHg / s' - dP / dt - left ventricular contractility, diastolic, mmHg / s

The stress test showed that Ml animals had significantly lower functional reserve of myocardial contractility compared to Sham control animals (Table 7 below). The mean increase in + dP / dt and -dP / dt during the test was approximately 13% for the M1 control group and 21% for the Sham control group. The MDS + GBBS group had the best results in a stress test similar to the Sham control group (mean + dP / dt and -dP / dt increase during the test was 20.9% and in the Sham group 21%); moreover, the functional reserve of cardiac contractility was relatively better for the MDS + GBBS group than for the MD + GBB group in both systole and diastole (Fig. P <0.005 vs. Ml control and P <0.05 vs. Ml control; Table 7).

Table 7

Effect of MD and MDS on left-arm contraction load in the stress test; Average * SVN; n = 6-8

Group + dP / di, c 'koinCJA + dP / dt, (else -dP / dt; original + dP / dt; in the test Sham control group 5325 * 201 6274 * 197 "· 4435 * 175 * · 5322 * 219 ·· * Ml control 3674i1X9 4266 * 243 3091 * 221 3386 * 237 MI + (MD + GBB) 4608 ± 1/3 * 5462 * 206 * 4019 * 230 * 4754 * 182 * M1 + (MDS + GBBS) 4705 * 146 ** 5749 * 154 ** 4083 * 107 ** 4835 * 120 · *

* P <0.05 for Ml control ** P <0.005 for Ml control *** P <0.0005 for Ml control

Analysis of morphometric data showed that animals developed cardiac and left ventricular hypertrophy and enlargement within 28 days of MI (Table 8).

Table 8

Effect of study substances on cardiac morphometric parameters; Mean ± SVN; n = 6-8

Group KM *, g SM ^k , mg KK.rng SM / KM ⁴ KK / SM ⁴ KKDT ¹ , ml Sham control group 467 * 12 1176 * 72 790x57 2.52 * 0.08 · 0.68 * 0.01 * 0.587 * 0.037 ** Ml control 447 * 16 1325 * 96 970 * 91 2.96 * 0.15 0.73 * 0.01 0.813 * 0.040 MIHMD + GBB) 463 * 16 1260 * 71 8ι5 ^: 62 2.71 * 0.08 0.68 * 0.01 * 0.692 * 0.017 * MI + (MDS + GBBS) 455 * 13 1215 * 54 817 * 42 2.67 * 0.08 0.67 * 0.01 * 0.657 * 0.016 "

'KM - body mass ^b SM - heart' weight 'KK - left ventricular mass ^d SM / KM - heart mass versus body mass x 1000' KK / SM - left ventricular mass versus heart mass ^f KKDT - left ventricular volume in diastole, ml * P <0.05 vs. Ml control • * P <0.005 vs. Ml control

In the rat chronic heart failure model, MDS + GBBS administration had a more significant protection against CK expansion than MD + GBB administration (Table 8, P <0.005 MDS + GBBS versus M1 control and P <0.05 MD + GBB versus M1 control). These results suggest that MDS + GBBS has a significant cardioprotective and adaptogenic activity in the rat model of chronic heart failure. This is evident from the maintenance of systolic and diastolic functions and significantly higher functional backup under stress.

These data suggest the clinical use of MDS + GBBS for the attenuation and prevention of post-infarction complications, including the treatment of cardiac systole and diastolic insufficiency [Dickstein K et al, European Heart 72008; 29: 2388-2442].

Example 5. Detection of antihypoxic and anti-ischemic effects in the brain The purpose of these tests was to investigate the anti-hypoxic and anti-ischemic effects of MD and GBB and their salts in experimental models of cerebral ischemia and infarction, alone and in combination.

Model of mouse brain hypoxia

The method. ICR male males weighing 21-25 g were randomized to 4 groups. The animals received test substances in aqueous solution repeatedly for 7 days p.o. The control group received the same volume of water (10 ml / kg). The last dose was given 1 hour before the experiment. The groups were:

1) MD group received 150 mg / kg MD

2) MD + GBB group, received 1 JO mg / kg MD + 50 mg / kg GBB

3) MDS + GBBS group, received 150 mg / kg MDS + 50 mg / kg GBBS

4) Control group, received 10 ml / kg water.

Brain hypoxia induced by experimental hemorrhage was induced with MgCl ₂ [Berga P et al, Arzneimittelforschung 1986; 36 (9): 1314-1320] administered over a period of 3 s in a tail vein (2% MgCl 2, 200 mg / kg). Time to last respiratory failure was defined as survival time.

Statistics. Data are presented as mean ± SD of 6 to 8 individual experiments. Differences between experimental groups were compared using one-way ANOVA with repeated comparisons (Tukcy test). P <0.05 was considered significant.

Results. The combination of MDS + GBBS showed significantly higher antihypoxic activity than MD or MD + GBB as seen by the increase in survival time (Fig. P <0.005 MDS + GBBS versus control and P <0.05 MD or MD1-GBB versus control). table).

Table 9

Effects of study substances on survival time; Mean ± VSN, n = 6-8.

Group Survival Time, s Control 25.3 + 1.07 MD (150 mg / kg / d) x 7 28.4 ± 0.83 * MD + GBB (150 mg / kg / d + 50 mg / kg / d) x 7 30.3 + 1.93 * MDS + GBBS (150 mg / kg / d + 50 mg / kg / d) x 7 32.6 + 1.21 **

* P <0.05 vs. control ** P <0.005 vs. control

Pcjii middle cerebral artery obstruction mot? J; s

The method. ICR male mice weighing 21-26 g were randomized to 5 groups. The groups were:

1) MD group, received 150 mg / kg MD;

2) MD + GBB group, received 150 mg / kg MD + 50 mg / kg GBB;

3) MDS + GBBS group, received 150 mg / kg MDS + 50 mg / kg GBBS;

4) Control group, received 10 ml / kg water;

5) Sham control group, received 10 ml / kg water.

A prophylactic (once daily for 7 days) treatment protocol was used.

The onset of the middle cerebral artery (VSA) was obstructed by the intraluminal filament technique according to a known method [Longa EZ et al., Stroke 1989; 20: 84-91] with adaptation in mice according to Iwai M et al. \ Circulation 2004; 110: 843-848]. The animals were anesthetized with chloral hydrate (400 mg / kg i.p.). Body temperature was maintained at 37 ± 0.3 ° C with an electronically controlled heated plate. Following the incision of the midline of the neck, the right carotid and external carotid arteries were isolated and temporarily ligated. A nylon monofilament suture 6/0 (Ethicon) with a dull end and a silicone resin overlay (Xantopren; Bayer Dental) was inserted into the right internal carotid artery and inserted approximately 10 mm distal to the carotid artery bifurcation to block the onset of the middle cerebral artery. The Sham control group underwent identical surgery except for arterial occlusion. Postoperative pain was alleviated by s.c. tramadol (20 mg / kg) was administered ampicillin (100 mg / kg) to prevent infection. Neurological deficits indicative of general sensory motor dysfunction were measured 24 hours after VSA occlusion [Wei EQ et al, Acta Physiol Sin 2003; 55: 742-747]: 0, no deficit; 1, flexion of the contralateral foreleg, lifting the animal behind the tail; 2, rotation to the contralateral side; 3, falling to the contralateral side; 4, no spontaneous motor activity.

Following neurological assessment, the animals were overdosed on pentobarbital sodium, isolated the brain, and cut into 6 layers of approximately 1.5 mm thickness.

Cuts 15 min. was stained with 2% triphenyltetrazolium sodium at 37 ° C and analyzed. The third cranial section (at the level of the chiazma opticum) was taken as the most suitable for calculating the area of the ischemic brain area, since it is completely supplied by blood from the middle cerebral artery.

Statistics. Data are presented as mean ± SD of 7 to 8 individual experiments (animals). Differences between experimental groups were compared using one-way ANOVA with repeated comparisons (Tukey's test). Differences in neurological endpoints for saliva groups were analyzed by an odd Student's t-test. P <0.05 was considered significant.

Results. All 8 control group animals had varying degrees of neurological impairment (1 to 4 points) 24 hours after VSA blockage, with an average of 2.75 points. Only one animal in the Sham control group had a minor impairment (0.17 points, P <0.0005; Table 10).

Table 10

Effects of study substances on neurological stool 24 hours after VSA blockage. Medium + VSN; n = 7-8

Group Noology, points Control 2.75 + 1.04 Sham group 0.17 + 0.17 · ** MD 1.65 + 0.26 * MD + GBB 1.43 + 0.29 * MDS + GBBS 1.31 + 0.21 ··

* P <0.05 vs. control ** P <0.005 vs. control *** P <0.0005 vs. control

Prophylactic treatment with MD or MD + GBB prevented worsening of neurological status after 24 hours of SVA blockage. The combination MDS + GBBS showed a further increase in protective activity against ischemic-induced neurological disorders (Fig. P <0.005 MDS + GBBS versus control and P <0, G5 MD or MD + GBB versus control. Table 10).

II. table

Effect of 7-day application of study substances on the size of the lesion area of the brain NCnnjas-infarki.i; Mean ± SVN; n = 7-8

Group Ischemic damage area,% Control 23.7 + 2.35 MD 17.2 + 2.07 MD + GBB 14.9 + 1.74 * MDS + GBBS 13.7 + 1.70 »

* P <0.05 vs. control

Morphological analysis showed that MD + GBB and MDS + GBBS are relatively better than MD in protecting against brain tissue damage caused by SVA blockage (Fig. P <0.05 MD + GBB or MDS + GBBS versus control and insignificant MD versus control).

Table 11).

Overall conclusions

We unexpectedly found that the efficacy of meldonium in the treatment of elevated triglycerides and LDL levels, as well as in the normalization of HDL levels, can be significantly increased when used in combination with gamma-butyrobetaine. We also found that this combination is much better than Meldonia alone in treating brain hypoxia, ischemia and stroke in experimental animals.

We have clearly shown that the efficacy of the combination of Meldonium and GBB is surprisingly even more pronounced when Meldonium and GBB are used as salts of succinic acid. We found that the combination of these salts significantly outweighs the activity previously discovered in combinations of Meldonium with GBB dihydrate in the treatment of cardiovascular and cerebrovascular diseases, such as myocardial infarction, cardiac dysfunction, and brain injury caused by hypoxia and stroke. Therefore, we concluded that the administration of meldonium and GBB 'in the form of both dihydrates and succinates is promising for the treatment of dyslipidemia, hyptrlipidemia, atherosclerosis and cardiovascular disease, as well as for brain hypoxia, ischemia and stroke.

Ways of implementing the invention

Tennin, "combination medical product" as used herein, refers to the simultaneous, sequential or separate administration of components of a combination. The invention thus described provides a combination medical product comprising MD or its faimaeeitbki acceptable salt and GBB or a pharmaceutically acceptable salt thereof for simultaneous, sequential or separate use in the treatment or prevention of a disorder wherein the disorder is selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, chronic heart failure, transient and persistent ischemic attack, including cerebrovascular accident and stroke, peripheral arterial occlusive disease. The combination medical product of the invention described may be administered in the form of a pharmaceutical composition. According to this aspect of the invention there is provided a pharmaceutical composition comprising MD or a pharmaceutically acceptable salt thereof and GBB or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable diluent or carrier.

In another aspect, the present invention provides a combined medical product comprising MDS and GBBS for simultaneous, sequential or separate use for the treatment and / or prophylaxis of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease. angina pectoris and myocardial infarction, chronic heart failure: transient and persistent ischemic attack, including cerebrovascular accident and stroke, peripheral arterial occlusive disease.

The combination medical product of the invention described may be administered in the form of a pharmaceutical composition. According to this aspect of the invention there is provided a pharmaceutical composition comprising MDS and GBBS together with a pharmaceutically acceptable diluent or carrier.

The pharmaceutical composition according to the present invention also includes separate compositions, including a pinno composition comprising MD or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable diluent or carrier, and a second composition comprising GBB or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable diluent or carrier. particularly preferred is a composition comprising MDS and GBBS as ingredients.

Such a composition provides sequential or discrete application. Because the treatment or prophylaxis of said disorders involves the long-term use of a medical product, the most optimal embodiment of the invention is achieved by an oral dosage form such as tablets or capsules. The amount of each active ingredient in the pharmaceutical composition of the described combined medicinal product depends on the condition being treated. Specialists treating patients with the aforementioned disorders can readily select the appropriate amounts of each active ingredient and the appropriate dosage regimen.

According to a further aspect of the invention there is provided the use of a combination medical product or pharmaceutical composition as described herein for the manufacture of a medicament for the treatment of metabolic-related disorders or for simultaneous, sequential or separate administration.

Claims (15)

Claims A combination medical product comprising meldonia or a pharmaceutically acceptable salt thereof in combination with gamma-butyrobetaine or a pharmaceutically acceptable salt thereof, for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, chronic heart disease. seizure disorders, transient and persistent ischemic attacks, including cerebrovascular accident and stroke and peripheral arterial occlusive disease. 2. The product of claim 1, wherein the meldonium is in the form of a quick-acting, slow-acting or sustained-release dosage form. 3. Combined medicinal product containing Meldonium succinate (1: 1) and gamma-butyrobetaine succinate (1: 1) for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease angina pectoris and myocardial infarction, chronic heart failure, transient and persistent ischemic attack, including cerebrovascular accident and stroke and peripheral arterial occlusive disease. 4. A pharmaceutical composition comprising meldonia or its faunaeically acceptable salt and gamma-butyrobetaine or a pharmaceutically acceptable salt thereof, in association with a pharmaceutically acceptable diluent or carrier, for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia. , atherosclerosis, chronic heart failure, transient and persistent ischemic attack, including cerebrovascular accident and stroke and peripheral arterial occlusive disease. The composition of claim 4, wherein the meldo-aluminum is in the form of a quick-acting, decisive or sustained-release drug. 6. A pharmaceutical composition for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, atherosclerosis, hyperlipidemia, coronary heart disease, angina pectoris and myocardial infarction, chronic heart failure, transient and persistent ischemic attack, including cerebral ischemic attack, and peripheral arterial occlusive disease comprising Meldonium succinate (1: 1) and gamma-butyrobetam succinate (1: 1) in association with a pharmaceutically acceptable diluent or carrier. 7. The pharmaceutical composition of claim 4, wherein the ratio of meldonium or pharmaceutically acceptable salt thereof to gamma butirobetam or pharmaceutically acceptable salt thereof is from 3: 1 to 1: 3. The pharmaceutical composition of claim 6, wherein the ratio of meldonium succinate (1: 1) to gamma-butyrobetaine succinate (1: 1) is from 3: 1 to 1: 3. 9. A method of treating and / or preventing a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, chronic heart failure, transient and persistent ischemic attack, including cerebrovascular accident and occlusive disease of the peripheral artery, an effective, simultaneous, sequential or separate administration of a pharmaceutically acceptable salt of meldonium or la and gamma-butyrobetaine or a pharmaceutically acceptable salt thereof. 10. The method of claim 9, wherein the meldonium is in the form of a fast-acting, slow-acting, or sustained-acting drug. The method of claim 9, wherein the combination is administered as two separate pharmaceutical compositions, wherein the first composition comprises meldonium or a pharmaceutically acceptable salt thereof together with a pharmaceutically acceptable diluent or carrier, and the second composition si.ur guino-butyrobetaine or a pharmaceutically acceptable salt thereof. together with a pharmaceutically acceptable diluent or carrier. 12. A method of treating and / or preventing a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease, angina pectoris and myocardial infarction, chronic heart failure, transient and persistent ischemic attack, and cerebrovascular accident. peripheral arterial occlusive disease, which involves administering to the patient to be treated an effective amount of meldonium succinate (1: 1) and gamma-butyrobetaine succinate (1: 1) simultaneously, sequentially or separately. The method of claim 12, wherein the combination is administered as two separate pharmaceutical compositions, wherein the first composition comprises Meldonium succinate (1: 1) together with a pharmaceutically acceptable diluent or carrier, and the second composition contains gamma-butyrobetaine succinate (1: 1). together with a pharmaceutically acceptable diluent or carrier. Use of a combination medical product according to claim 1 for the manufacture of a medicament for the treatment and / or prevention of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, chronic heart failure, transient and persistent ischemic attack, including cerebrovascular accident. events and stroke and peripheral arterial occlusive disease. Use of a combined medical product according to claim 3 for the manufacture of a medicament for the treatment and / or prophylaxis of a disorder selected from the group consisting of dyslipidemia, hyperlipidemia, atherosclerosis, coronary heart disease from the group of angina pectoris and myocardial infarction, chronic heart failure , transient and persistent ischemic attack, including cerebrovascular accident and occlusive disease of the stroke and peripheral arteries.