RU2476224C2 - Drug preparation with cardioprotective activity - Google Patents

Abstract

FIELD: medicine, pharmaceutics.

SUBSTANCE: what is presented is the use of the drug -2-(2-morpholinoethyl)thio-5-ethoxybenzimidazole dihydrochloride (the anxiolytic Afobazole) as a preparation with cardioprotective activity. It is shown that Afobazole (5-10 mg/kg intravenously or intraperitoneally) on various models of myocardial ischemia and heart rhythm disorder exhibits manifested cardioprotective activity observed primarily at the myocardium and may be related to a certain extent to its affinity for sigma1-receptors localised in cardiac myocytes. Antiarrhythmic action of Afobazole is preferentially conditioned by its ability to increase electrical stability of the maycardium directly.

EFFECT: findings entitle recommending the anxiolytic Afobazole to be used in clinical practice as a cardioprotector for the purpose of preventing and treating cardiovascular disorders in the patients suffering cardiophobic and/or asthenic-depressive syndromes burdened by cardiopathies.

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Description

The invention relates to medicine, in particular to pharmacology, and for the use of the known drug 2- (2-morpholinoethyl) thio-5-ethoxybenzimidazole dihydrochloride (anxiolytic afobazole) (RF patent No. 2061682; Seredenin SB, Voronin MV , Expert. And Wedge Pharmacol. - 2009. - T. 72, No. 1. - C.3-11; Radar, 2008. - S.143-144) as a drug with cardioprotective activity.

It is known that a number of derivatives of 2-mercaptobenzimidazole has an antiarrhythmic effect (Chichkanov G.G. et al., Expert. And wedge. Pharmacol. - 1997. - T.60, No. 5. - S.35-39). Previously, when studying the effect of afobazole and its structural analogues on screening models of heart rhythm disturbances at the Research Institute of Pharmacology of the Russian Academy of Medical Sciences, it was shown that these compounds can be assigned to classes I and IV of antiarrhythmic drugs according to the classification of Vaughan Williams (Turilova A.I., Mozhaeva T. I., Expert and wedge. Pharmacol. - 2010. - T. 73, No. 5. - P.8-11) and are effective on models of both atrial and ventricular cardiac arrhythmias (V. Stolyaruk and others. Bulletin of RAMS. - 2010. - No. 4. - P.41-45; Stolyaruk V.N. et al., Bulletin of RAMS. - 2010. - No. 4. - P.49-52).

2- (2-morpholinoethyl) thio-5-ethoxybenzimidazole dihydrochloride - the anxiolytic afobazole - has a complex mechanism of anxiolytic action, which is associated with its ability to prevent the stress-induced decrease in mediator binding in the benzodiazepine region of the GABA receptors (Seredenavit S. B. with ., Vestnik RAMS. - 1998. - No. 11. - S.3-9). It is also shown that afobazole has an affinity for sigma1 and melatonin receptors, inhibits the activity of MAO-A. In addition to anxiolytic afobazole, it also has cytoprotective activity, to a certain extent due to its affinity for sigma1 receptors. The latter are currently considered as an intracellular evolutionary formation, which plays an important role in the physiological protection of cells from pathological influences; in connection with which they can be positioned as a kind of “repair” complex that provides homeostasis of the cell and thereby supports its vital activity (Seredenin SB, Voronin MV, Eksperim. and wedge. pharmacology. - 2009. - T. 72 , No. 1. - C.3-11). In addition to the central nervous system, sigma1 receptors are quite widely represented in the cells of other organs and tissues of the body, including the myocardium (Novakova M. ef al., Gen Physiol Biophys. - 2007. - v.26. - No. 2. - P.110- 7). There is evidence in the literature that sigma1 receptor ligands possess both antiarrhythmic, in particular antifibrillatory {Lishmanov Yu.B. et al., Experiment. and clinic. pharmacology. - 2000. - t. 63, No. 6. - S.24-27; Maslov L.N. et al., Experiment. and clinic. pharmacology. - 1997. - T.60, No. 2. - P.24-26.), As well as anti-ischemic activity (Ai-S.T. et al., Curr. Neurovasc. Res. - 2006. - V. 3, No. 2. - P.89-98.) .

The prevalence of psychosomatic disorders, including cardiophobia, has led to the fact that currently in the framework of psychosomatic medicine a new direction has been identified - psychocardiology (Smulevich AB et al., Psychocardiology. - M .: ed. Med. Inf. Agency, 2005 .-- 784 p.).

Studies conducted in this area have shown that the level of phobic anxiety is directly proportional to the risk of fatal and / or nonfatal myocardial infarction, malignant heart rhythm disturbances, and sudden coronary death (Abrams T.E. et al., Circ Cardiovasc Qual Outcomes. - 2009. - v.2. - P.213-220; Ehlens A. et al., Psichosom. Med. - 2000. - v.62, No. 5. - P.693-702; Schulman JK et al., Focus. - 2005. - V.3. - P.208-224. 2005, Walters K. et al., Eur. Heart J. - 2008. - V.29. - P.2981-2988). For example, in a prospective epidemiological study in which 72,359 women took part (follow-up 12 years), a direct correlation was found between the level of phobic anxiety and the risk of malignant heart rhythm disturbances, including sudden coronary death (Albert Ch. M . et al., Circulation. - 2005. - v. 1111. - P.480-487). An analysis of the study, which involved 940 patients hospitalized for diagnostic coronarography, showed a statistically significant relationship between the level of anxiety-phobic disorders and the risk of developing malignant, including fatal, heart rhythm disturbances (Watkins LL et al., Psy-chosom Med . - 2006. - v.68: - P.651-656).

Thus, the introduction into clinical practice of drugs that combine anxiolytic and cardioprotective activity seems to be very relevant.

One of the possible solutions to this problem is the use of domestic highly effective anxiolytic afobazole for these purposes.

Based on this, we systematically studied the cardioprotective activity of afobazole and showed that this activity of afobazole is realized mainly at the level of the heart muscle and, in particular, is associated with the affinity of afobazole for sigma1 receptors localized in cardiomyocytes. The data obtained allow us to recommend the use of afobazole in the clinic as a cardioprotector for the prevention and treatment of cardiovascular disorders in patients suffering from cardiophobic and / or astheno-depressive syndromes, the course of which is aggravated by heart diseases.

Example 1. The study of the antiischemic effect of afobazole

The experiments were carried out on white outbred male rats weighing 180-200 g at a dose of 10 mg / kg once a day for 7 days (group 2). The first administration was carried out immediately after occlusion of the coronary artery. In the control series of experiments (group 1), an equivalent volume of an isotonic sodium chloride solution was introduced according to a similar scheme.

On the eighth day, the animals were anesthetized with urethane (1300 mg / kg, ip), the hearts were removed and fixed in 10% formalin solution. Using a freezing microtome at two levels (top and middle), transverse sections of the heart were made with a thickness of 10-15 microns. Sections were stained according to standard methods - gallocyanin-eosin and picrofuchsin according to Van Gieson. A qualitative visual assessment of micropreparations of the heart was carried out with a description of the pattern of damaged tissue. Quantification was carried out using the Avtandilov grid and a 24-fold magnifier. The area of myocardial infarction, the thickness of the walls of the ventricles and interventricular septum, the diameter of the cavity of the left ventricle was determined on the mid-section of the heart. In addition, in these sections in the conditionally intact zone of the myocardium, to assess the intensity of proliferative processes using the ocular network, we measured (in arbitrary units) the transverse diameter of the nuclei of cardiomyocytes.

The results of the study were statistically processed using the Shapiro-Wilks test, one-dimensional analysis of variance followed by comparison using the Dunnet test (one-way test), the Mann-Whitney-Wilcoxon test, taking into account the multiplicity of comparisons, and the χ ² criterion, taking into account the multiplicity of comparisons.

It was shown that in animals that were injected afobazole at a dose of 10 mg / kg (n = 7) within 7 days after occlusion of the coronary vessel, n = 7, in contrast to control animals (n = 8), the morphological picture of the peri-infarction zone is characterized by high intensity proliferative processes, which, in particular, is evidenced by an increase in the diameter of the nuclei of cardiomyocytes localized in this zone. So, if in the myocardium of control animals the number of cardiomyocyte nuclei with a diameter of more than two arbitrary units is 17.7%, then in rats that received afobazole at a dose of 10 mg / kg, this indicator is statistically significantly higher (P <0.001) and is 65.5%. It can be assumed that it is such a high intensity of proliferative processes that occur in the conditionally intact zone of the myocardium that has led to the fact that in these animals the area of myocardial infarction is statistically significant (P = 0.0266) than in the control.

No less important is the fact that long-term therapy with afobazole statistically significantly prevents post-infarction remodeling of the left ventricle (table 1, figure), in particular, the cavity of the left ventricle of rats receiving afobazole at a dose of 10 mg / kg is 21.5% less than that of animals of the control group (P = 0.0477).

Table 1 The effect of afobazole (10 mg / kg, ip, 7 days) on the morphometric parameters of the heart of rats after experimental myocardial infarction A drug Number of hearts Indicators, M ± m Left ventricular wall thickness, mm The thickness of the interventricular septum, mm The thickness of the wall of the right ventricle, mm The diameter of the cavity of the left ventricle, mm The control 8 0.90 ± 0.19 2.00 ± 0.21 0.88 ± 0.15 5.62 ± 0.41 Afobazole 10 mg / kg 1.67 ± 0.19 2.30 ± 0.21 1.18 ± 0.15 4.40 ± 0.41 7 p = 0.0099 P = 0.2518 P = 0.1463 p = 0.0477 P is indicated with respect to control

The data obtained suggest that afobazole in conditions of acute myocardial infarction shows a pronounced anti-ischemic effect - the drug not only stimulates reparative processes in the heart muscle and reduces the area of ischemic damage, but also prevents postischemic remodeling of the heart muscle. The latter seems to be the most important, since it gives every reason to believe that in animals treated with afobazole, the risk of developing heart failure in the post-infarction period is significantly lower.

It can be assumed that the antiischemic effect of afobazole is to some extent related to its affinity for sigma1 receptors, since it is known that agonists of σ ₁ receptors have the ability to reduce the area of ischemic damage (Calvert JW, Lefer DJ, Cardiovasc. Res. - 2009. - v .94, No. 7. - P.805-814.). This effect of σ ₁ receptor agonists is apparently complex and includes the protection of ischemic cells from overload with Ca ^{2+ ions} (Tchedre KT et al., Invest. Ophthalmol. Vis. Sci. - 2007. - v. 48. - E.-Abstract572), a decrease in the tonic effect of the sympathetic nervous system on the myocardium (Zhang H., Cuevas J., J. Neurophysiol. - 2002. - v.10. - P.2867-2879), suppression of the cascade of intracellular reactions initiating apoptosis (Tchedre KT, Yorio T., Invest. Ophalmol. Visual Science. - 2008. - v. 49. - P.2577-2588), including expression of the bcl-2 gene (Yang S. et al., Anesth. Analg. - 2007. - v.104, No. 5. - P.1179-1184), protection of myocardial cells from free-radical aggression (Halliwell B., J. Neurochem. - 2006. - v. 97, No. 6. - P.1634-1658). It cannot also be ruled out that the anti-ischemic effect of afobazole may be due to its ability to inhibit the activity of monoamine oxidase A (Seredenin SB, Voronin MV, Expert. And wedge. Pharmacol. - 2009. - v. 72, No. 1. - C.3-11), since it is known that the hyperactivity of this enzyme in cardiomyocytes, observed under conditions of their ischemic damage, initiates apoptosis of cardiomyocytes and / or damage to their membranes due to oxidative stress (Pchejetski D. et al., Circ. Res. - 2007. -v.100. - P.41-49; Kaludercic N. et al., Circ. Res. - 2010. - v.106, No. 1. - P.193-202).

Example 2. Evaluation of the contribution of central and peripheral mechanisms to the cardioprotective effect of afobazole.

The experiments were performed on anesthetized sodium pentobarbital (40 mg / kg, iv) outbred cats of both sexes weighing 2.9-3.8 kg. Animals were intubated and transferred to artificial lung ventilation with an oxygen-air mixture (1: 1 ratio) at a rate of 130-150 ml / min / kg of animal weight. The threshold of electrical cardiac fibrillation was determined by repeated scanning of the vulnerable period of the cardiac cycle with a series of 20 rectangular DC pulses of increasing strength (stimulus duration 4 ms, frequency 50 pulses / s) before fibrillation occurred. If fibrillation did not stop on its own after 60 seconds, the animals were defibrillated with an electric current discharge (1.5 kV). The minimum current strength, which stably causes ventricular fibrillation (lasting more than 60 sec) upon repeated stimulation, was taken as the fibrillation threshold. An HSE Stimulator II electric stimulator (Hugo Sach Electronik, Germany) was used in the work. Defibrillation was performed using a DI-03 defibrillator (Russia). Heart denervation was performed by removing the right and left stellate ganglia and intersecting the right and left vagus nerves.

Afobazole was administered intravenously (iv) at a dose of 7.5 mg / kg at a constant rate and in a constant volume of 0.9% sodium chloride solution. Control animals were injected with an equivalent amount of 0.9% sodium chloride solution.

It was shown that after denervation of the heart, heart rate statistically significantly (P = 0.011) decreased from 145.8 ± 8.2 beats / min to 110.5 ± 4.4 beats / min, which indicates the cessation of central tonic effects on the heart, since it is well known that the sinoatrial node itself generates heart rate with a lower frequency (Kamkin A., Kamensky A. (ed.) Fundamental and clinical physiology. - M., Academy; 2004. - P.521-551). The threshold of electrical fibrillation of denervated heart decreased by 2-3 times, which, most likely, is associated with a decrease in the electrical stability of cardiomyocytes caused by denervation. So, if electrical cardiac fibrillation in animals with an intact myocardium was caused by a current strength of 1.0-3.0 mA, then against the background of denervation, the fibrillation threshold decreased to 0.5-1.0 mA (Table 2).

Afobazole (7.5 mg / kg, iv) in animals with denervated myocardium, as well as in animals with intact myocardium, in all experiments, statistically significantly (P <0.05) increases the threshold of cardiac electrical fibrillation - fibrillation failed cause electric pulses with a current ranging from 0.5 to 100 mA (table 2).

table 2 The effect of afobazole (7.5 mg / kg, iv) on the threshold of electrical fibrillation of denervated heart in cats. N experience Io, mA Idenerv, mA Ipr, mA one 1,0 0.5 > 100 2 2.0 1,0 > 100 3 1,5 1,0 > 100 four 3.0 1,0 > 100 5 2.0 1,0 > 100 6 3.0 1,0 > 100 Designations: Io - initial fibrillation threshold (intact myocardium); Idenerv - fibrillation threshold after denervation of the heart; Ipr - threshold for fibrillation of a denervated heart on the background of afobazole.

Since the intensity of the anti-fibrillatory effect of afobazole did not differ in animals with an intact and denervated myocardium, there is every reason to believe that the antiarrhythmic effect of the drug is mainly due to its ability to directly increase the electrical stability of the heart muscle.

Example 3. The study of the possible mechanisms of the cardioprotective action of afobazole

As noted above, afobazole is a sigma1 receptor agonist. Since it is known that sigma1 receptor ligands have an antiarrhythmic effect, an attempt was made to evaluate the possible contribution of sigma1 receptors to the antiarrhythmic effect of afobazole. For this purpose, the antiarrhythmic effect of afobazole was evaluated against the background of the preliminary administration of the sigma1 haloperidol receptor antagonist. Although haloperidol is considered as a non-selective antagonist of sigma receptors, there is evidence that its affinity for sigma1 receptors is 170 times higher than for sigma2 receptors (Klouz A. et al., Therapie. - 2001. - v. 56, No. 5. - P.557-562).

The experiments were carried out on anesthetized (urethane, 1300 mg / kg, iv) outbred male rats weighing 350-400 g. The threshold of electrical fibrillation of the heart was determined in the same way as in example 2. The minimum current strength was taken as the threshold for ventricular fibrillation, causing double repetition of ventricular fibrillation. Only animals in which ventricular fibrillation occurred at a current strength of not more than 6 mA were selected for the experiment. Afobazole was administered intravenously (iv) at a dose of 7.5 mg / kg at a constant rate and in a constant volume of 0.9% sodium chloride solution. Haloperidol (Gideon Richter) was administered iv in a dose of 0.5 mg / kg at a constant rate and in a constant volume of a 0.9% sodium chloride solution.

The results were statistically processed using analysis of variance for repeated changes with further use of the Newman-Cales multiple comparison method, Student's test and Wilcoxon's paired test.

It was shown that against the background of preliminary administration of haloperidol at a dose of 0.5 mg / kg, the antifibrillatory effect of afobazole, administered at the rate of 7.5 mg / kg, is not realized (Table 3). In addition, it was noted that if intact animals against the background of afobazole there is a decrease in heart rate, then in animals treated with haloperidol, the pulse-inhibitory effect of afobazole is not realized. The differences are statistically significant (P = 0.046).

Table 3 The effect of afobazole on the threshold of electrical fibrillation of the ventricles of rat hearts with prior administration of haloperidol Experience number Threshold Background Haloperidol (0.5 mg / kg, w / w) Afobazole (7.5 mg / kg, iv) Threshold Heart rate before administration Heart rate after administration Threshold one 3 0.5 416 416 0.5 2 3 one 400 400 one 3 one 0.5 375 375 0.5 four 2 one 261 261 one 5 2 four 366 366 four 6 3 3 300 300 3 7 2 one 333 333 one 8 one 0.5 316 250 3

Thus, the obtained results suggest that the antiarrhythmic effect of afobazole is to some extent related to its affinity for sigma1 receptors localized in cardiomyocytes, since the drug does not exhibit antiarrhythmic activity against the background of their blockade.

The data presented indicate that the anxiolytic afobazole has a pronounced cardioprotective effect, which is realized at the level of cardiomyocytes and may be associated with the affinity of the drug for sigma1 receptors. Based on this, afobazole can be recommended for use in the clinic as a cardioprotector for the prevention and treatment of cardiovascular disorders in patients suffering from cardiophobic and / or asthenic-depressive syndromes, which are aggravated by heart diseases.

Description of the picture.

The figure shows cross sections of the left ventricle of the heart of rats with experimental myocardial infarction: A - heart of an intact animal; B - the heart of the animal 7 days after myocardial infarction (control); B - the heart of an animal with myocardial infarction receiving afobazole at a dose of 5.0 mg / kg / day, ip, 7 days; G - the heart of an animal with myocardial infarction receiving afobazole at a dose of 10.0 mg / kg / day, ip, 7 days.

Claims (1)

The use of 2- (2-morpholinoethyl) thio-5-ethoxybenzimidazole dihydrochloride as a cardioprotective agent.