Agent for drug-induced correction of nitroxydergic disorders. RU patent 2505297.

FIELD: medicine.

SUBSTANCE: what is presented is using 1-(β-phenylethyl)-4-amino-1,2,4-triazolium bromide as an active drug base for the purpose of correcting the nitroxydergic malfunctions.

EFFECT: higher activity of endothelial NO-synthase and high NO production under the action of the declared agent that has higher efficacy as compared to the known analogues, and has no side effects.

2 tbl, 1 ex

The invention relates to the pharmaceutics and medicine, namely to medicines, positively influencing on the state of system.

It is known that malfunction of system, which leads to a deficiency of nitric oxide (NO), and dramatically increases the risk of development of diseases of the respiratory, cardiovascular, hepatobiliary systems, gastrointestinal tract, as well as development of endocrine disorders [1]. In the pathogenesis and clinical diseases such as atherosclerosis, diabetes mellitus, arterial hypertension and its complications, one of the important aspects is considered a violation of the structure and function of the vascular endothelium [1, 3]. Endothelial dysfunction (ED) developed at deficiency of nitric oxide, a violation of expression or transcription of endothelial NO synthase (eNOS), reducing the availability of L-arginine for eNOS, that is in violation of system.

Correction of infringements of a metabolism of nitric oxide, i.e. correction system and restore generate NO, reduces the risk of these diseases, based on endothelial dysfunction.

In the end of XX century it was found that nitric oxide (NO) is one of the universal and necessary regulators of cell metabolism. It turned out that this molecule is shortlived and easily subjected to chemical transformations of free radicals, which continuously enzymatically produced in mammals, affecting a variety of physiological and pathophysiological processes. Study of the role of NO and identified that the nitric oxide messenger, acting as a universal modulator variety of functions, including the regulation of breathing, maintaining cardiovascular homeostasis, the immune status of the organism, the activity of macrophages, gene expression, the plasticity of the nervous tissue, memory, release of neurotransmitters. Nitric oxide plays the role of neuroregulator in the Central nervous system and nerve-muscle synapses.

Opening of various physiological roles of nitric oxide in the body led to the creation, study and application of drugs able to correct system.

However, today known medicines intended for correction of violations of system, not so much, and it should be only drugs that active active substance contain L-arginine (, , ) [2, 3, 5]. In addition, their therapeutic action is not efficient enough, therefore, an urgent task is to expand the Arsenal of preparations for the correction of system, which showed high efficiency and were harmless for patients.

Known biologically active compound bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT), possessing cause cardioprotective, , antihypertensive, anti-oxidant, and action (Russian Federation patent №2404974, C07D 249/08 (2006.01), A61P 9/04 (2006.01), A61P 9/10 (2006.01), A61P 9/12 (2006.01), publ. 27.11.2010; patent of Ukraine №92692, the IPC (2009) A61K 31/41, publ. in bull. «Promyslova і», 2010, №22).

However, nowhere in the literature available to us do not describe a property of the specified substances correct disorders system.

The basis of the invention task of obtaining funds, correcting violations system, which has higher efficiency in comparison with already known means of similar function and has no side-effect.

The problem is solved by the fact that, as an active Foundation of drugs for correction of violations of systems use bromide 1-(beta-phenylethyl)-4-amino-1,2,4-.

The essence of the invention is illustrated by the examples below results of preclinical studies.

Example 1. For research was induced endothelial dysfunction (ED) (with a marked shortage NO, i.e. violation of functioning system) through the daily administration of N-nitro-L-arginine methyl ester (L-NAME) white rats-males weighing 180-190 g once a day, intraperitoneally, in the dose of 25 mg/kg within 8 days. Conducted an assessment of the effectiveness of the correction of violations of system bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT). In the experiment compared the action bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT) and the reference drug L-arginine (EUROBIOPHARM GmgH, Hamburg). Investigational drugs were injected through 30 minutes after administration of L-NAME: L-arginine at doses of 200 mg/kg/day, bromide 1-(beta-phenylethyl)-4-amino-1,2,4- - in a dose of 5 mg/kg/day. Rats were taken from the experiment on the 8th day from the beginning under anesthesia (40 mg/kg), and they for the study were taken heart, which is then homogenized liquid nitrogen and the method of differential centrifugation allocated faction, which contains only endothelial NO synthase (eNOS).

In cytosolic fraction homogenate hearts were determined by biochemical markers of endothelial dysfunction - stable metabolites NO activity and a total of NO-synthase [4]. The level of expression of endothelial nitric oxide synthase (eNOS) was determined in the cytosolic fraction homogenate heart immunoblotting method [4].

The results obtained are presented in table №1.

Impact bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT) and the reference drug L-arginine on indicators of NO in the cytosolic fraction homogenate hearts of rats with endothelial dysfunction with the introduction of the 8 days

Group of animals

NOS, mkmol NADP/min/g protein

Nitrites, mmol/ g of tissue

Concentration of eNOS, $ /g protein

intact (n=10)

12,4±0,63

18,5±0,77

15,7±0,65

ED (control) (n=10)

4,22±0,45

5,81±0,53

3,2±0,17

ED+MT(5 mg/kg) (n=10)

8,7±0,55*1 (+106,2%)

12,2±0,85*1 (+110%)

10,2±0,45*1 (+218,7%)

ED+L-arginine (200 mg/kg) (n=10)

5,00±0,47 (+18,4%)

7,12±0,32* (+22,5%)

3,5±0,27 (+9,3%)

* - changes were significant in relation to the animals of the control group (p<0.05);

1 - changes were significant in relation to the group of animals treated with L- (p<0.05)

As can be seen from table 1, when modeling of endothelial dysfunction (ED) in animals there is a shortage of NO, as evidenced by the significant reduction of its stable metabolite - nitrite-anion by 68.6% against the oppression of activity total NOS 66% decrease in the expression of eNOS 79.6% in the cytosol infarction in comparison with the intact animals.

Course administration to animals with endothelial dysfunction drug MT at a dose of 5 mg/kg/day resulted in increases NO production in the cytosol infarction 110%. MT increased physiological path of NO synthesis, which increases the activity of endothelial NO-synthase. Thus, in the group of animals with ED treated MT of activity total of NO synthase in the cytosol increased by 106.2 percent, and noted the increased expression of endothelial NO-synthase to be 218.7% compared to the untreated animals. L-Arginine influence only on education NO, significantly increasing the level of nitrites by 22.5%, however, he did not exert significant influence on the activity and expression of NO-synthase in the myocardium.

Example 2. To assess the effectiveness of the correction of violations of system bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT) were used 30 rats of both sexes weighing 180-200 g with spontaneous arterial hypertension ISIAH (hereditarily stress arterial hypertension, a shortage NO) and 8 normotensive control animals lines WKR (normotensive Wistar-Kyoto rats), weight 180-220, MT administered intraperitoneally daily dose of 5 mg/kg within 15 days. L-arginine (EUROBIOPHARM GmgH, Hamburg) was administered at a dose of 200 mg/kg according to the same scheme. On 15 day of the experiment the animals were taken from the experiment under anesthesia (40 mg/kg), and their heart was taken, which is then homogenized liquid nitrogen and the method of differential centrifugation allocated faction, which contains only endothelial NO synthase (eNOS). In cytosolic fraction homogenate hearts were determined by biochemical markers of endothelial dysfunction - stable metabolites NO activity and a total of NO-synthase [4]. The level of expression of endothelial nitric oxide synthase (eNOS) was determined in the cytosolic fraction homogenate heart immunoblotting method [4]. Research results are presented in table №2.

The impact of MT -drug L-arginine on indicators of NO in the cytosolic fraction homogenate hearts of rats with spontaneous arterial hypertension in the introduction for 15 days

Group of animals

NOS, mkmol NADP/min/g protein

Nitrites, mmol/ g of tissue

Concentration of eNOS, $ /g protein

() (n=8)

11,75±0,75

17,23±0,84

15,2±0,73

SHR (control) (n=10)

5,15±0,64

7,76±0,67

4,1±0,21

SHR+MT (5 mg/kg) (n=10)

9,71±0,65*1 (+88,5%)

15,15±0,77*1 (+95,2%)

12,3±0,65*1 (+200%)

SHR+L-arginine (200 mg/kg) (n=10)

6,20±0,55 (+20,4%)

9,87±0,44* (+27.2%)of

4,77±0,53 (+16,3%)

* - changes were significant in relation to the animals of the control group (p<0.05);

1 - changes were significant in relation to the group of animals treated with L-arginine (p<0.05)

As table 2 shows, in ISIAH rats, a decrease of NO education, as evidenced by a significant reduction of its stable metabolite - nitrite-anion by 55.0% against the oppression of activity total NOS 56.2% and decrease in the expression of eNOS by 73.0% in the cytosol infarction compared with rats.

Course purpose rats the drug MT at a dose of 5 mg/kg/day resulted in increases NO production in the cytosol infarction 95%. MT increased physiological path of NO synthesis, which increases the activity of endothelial NO-synthase. Thus, in the group rats receiving MT of activity total of NO synthase in the cytosol increased by 88% on the background of increase of expression of endothelial NO-synthase by 200% compared to the untreated animals. L-Arginine did not exert significant influence on the activity and expression of NO-synthase in the myocardium, and influenced only on education NO, significantly increasing the level of nitrites by 27.2%.

Thus, a known drug - bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT), we identified a new property is to be corrective effect in relation to system. On the activity of MT significantly exceeded the drug comparison - L-arginine. The toxicity of the drug MT belongs to class IV (low toxicity). Thus, the use of the invention will allow in the future to create high-performance harmless medicines containing as active active substance bromide 1-(beta-phenylethyl)-4-amino-1,2,4- (MT)for the correction of system.

Literature

1. A new look at the correction of endothelial dysfunction / M.V., Pokrovsky M.V., Kochkarov V.I. and others // Russian journal of immunology. - 2006. - Vol.9, supp.3. - P.59-63.

2. M.B., artyushkova E.B. Principles of pharmacological correction of endothelial dysfunction // Kuban medical scientific Bulletin. - 2007. - №1-2. - P.146-150.

3. Diagnosis and treatment of arterial hypertension. Russian recommendations (third revision). - M, 2008.

4. Preclinical study of specific activity of potential drugs: method, recommendations of SPC MOH Ukraine / Chekman I.S, SCI Gubsky, I.PH. Б. - K., 2010. - 81 C.

5. Compendium. Drugs and medications. - K.: Morion, 2010. - S. L-418, L-1505.

Application bromide 1-(beta-phenylethyl)-4-amino-1,2,4- as an active Foundation of drugs for correction of malfunctions system.