RU2414235C1 - ADMINISTRATION OF Arg-Pro-Gly-Pro PEPTIDE AS ANTIVIRAL AGENT FOR ANIMALS - Google Patents

Abstract

FIELD: medicine, veterinary science.

SUBSTANCE: invention concerns veterinary science. The invention discloses administration of Arg-Pro-Gly-Pro tetrapeptide as an agent for prevention and treatment of viral diseases. A pharmaceutical composition for prevention and treatment of viral diseases contains an aqueous solution of Arg-Pro-Gly-Pro tetrapeptide in concentration 0.1-1 % and Nipagin in a certain relation of ingredients. A method for prevention of viral diseases involves intranasal, oral or intraperitoneal introduction of said pharmaceutical composition in a dose 0.2-5 mg a day per a mouse for 2 days. A method for treatment of viral diseases involves intranasal, oral or intraperitoneal introduction of said pharmaceutical composition in a dose 0.2-5 mg a day per a mouse for 3 days.

EFFECT: invention enables extending the range of antiviral drugs, providing higher clinical and preventive effectiveness in viral infections, reducing drug cost.

4 cl, 4 tbl, 1 ex

Description

The invention relates to biotechnology and veterinary medicine and relates to the use of the Arg-Pro-Gly-Pro peptide as an antiviral agent for animals.

The development of new antiviral drugs is always relevant in connection with the wide spread of viral diseases. In addition, in recent years there has been a population-related decline in immunity associated with environmental degradation, which also contributes to an increase in viral diseases.

A known method for the prevention and treatment of viral infections, including the use of a medicinal product [Ridostin, Cycloferon, Dalargin, Grippferon, Arbidol-Lance, Acyclovir-Acre]. However, this known method is not in all cases effective.

The Arg-Pro-Gly-Pro peptide is known from the publication [RU 2286166 C1 of 10.27.2006]. However, in this publication, this peptide is used as an agent having anticoagulant-fibrinolytic, antiplatelet and antithrombolytic activity. It does not follow from this publication that the indicated peptide can be used as an antiviral agent. Thus, previously, the prior art not only did not know the use of the indicated peptide as an antiviral agent, but this use did not follow from the prior art. Therefore, the proposed invention is both new relative to the existing level of technology, and corresponding to the condition of the inventive step.

An antiviral agent is known from the prior art, which is a Thr-Lys-Pro-Arg-Pro-Gly-Pro peptide [RU 2337703 C1 of 10.11.2008]. This publication is the closest analogue of the claimed invention. The disadvantage of the closest analogue is the large length of the peptide, which increases the cost of synthesis of the specified peptide. The disadvantage of this tool is the need to enter it intraperitoneally.

The technical result achieved by the implementation of the present invention is to expand the range of antiviral drugs, increase the effectiveness of the prevention and treatment of viral infections, reduce the cost of the drug due to the significant simplification of the synthesis of the proposed tetrapeptide Arg-Pro-Gly-Pro instead of the Selank heptapeptide.

The specified technical result is achieved by the fact that the tetrapeptide Arg-Pro-Gly-Pro is used as a means of prevention and treatment of infectious diseases of viral etiology (influenza, mouse encephalomyocarditis virus).

This technical result is achieved due to the fact that in the method for the prevention and treatment of viral infections, involving the use of a medicinal product, the specified tetrapeptide is used as a medicine by conveniently administering a 0.1-1.1% solution in a daily dose of 0.2 -5 mg / day per mouse for 2-3 days. According to our own studies, a preparation based on this tetrapeptide is able to suppress the reproduction of influenza viruses, mouse encephalomyocarditis virus and can be used for the prevention and treatment of infectious diseases caused by viruses, without causing adverse side effects. The following are examples illustrating the invention.

Example 1

The study of the antiviral activity of the tetrapeptide Arg-Pro-Gly-Pro in an experimental infection caused by encephalomyocarditis virus (VEMK) in vivo

The experiments were performed on mice — white mongrel males weighing 14–16 g. The effect of each peptide was tested on 10 mice. We estimated the survival of mice in each group 14 days after infection with the administration of drugs (10 ^-5 M) intraperitoneally simultaneously with the encephalomyocarditis virus (VEMK).

The results are shown in table 1.

Based on the data obtained in in vivo studies, we can conclude that the studied peptides had a pronounced antiviral activity in low concentrations.

Table 1 The survival of mice under the action of the tetrapeptide Arg-Pro-Gly-Pro, administered intraperitoneally simultaneously with infection with the virus encephalomyocarditis (VEMK) No. Peptides % survival one Thr-Lys-Pro-Arg-Pro-Gly-Pro twenty 2 Arg-Pro-Gly-Pro one hundred 3 Pro-gly-pro 80 four Arg-pro 40 5 Gly-pro 60 6 the control twenty

Further, it was of interest to study the effect of the peptide at different concentrations with different methods of administration to mice infected with VEMC. Studies were carried out on white outbred mice weighing 14-16 g - 5 mice per point. Animals were infected with VEMK at a dose of 100 LD 50. Tetrapeptide Arg-Pro-Gly-Pro in concentrations of 1%; 0.5%; 0.2%; 0.1% was administered to animals intraperitoneally, intranasally, orally at 0.2 ml / mouse at the time of infection. As a preservative, the solution contained nipagin with a concentration of 0.09-0.11% solution.

The control group consisted of mice infected with the flu and given a placebo. The observation period is 15 days.

The antiviral effect of the peptide was evaluated by the survival of mice (% survival,% protection, average harmonic lifetime).

The peptide activity was evaluated taking into account:

1) the average harmonic value of the lifetime for each group of mice, which includes information about all individuals, calculated by the formula:

1 / τ = (X ₁ / t ₁ + X ₂ / t ₂ + ... + X _a / t _a ): n, where t is the day of death of animals, X is the number of dead animals on day t, and is the day of observation; n is the number of animals in the group (5 mice);

2) the ratio of protection (KZ) - the ratio of the decrease in the number of sick and dead animals in the experimental group compared with the control, calculated by the formula:

KZ =% dead animals in the control group /% dead animals in the experimental group;

3) the drug effectiveness index (IE), calculated by the formula: IE = [(KZ-1) / KZ] × 100%.

The results of experiments to test the anti-influenza activity of the drugs are presented in tables 2-3.

From the presented data (table 2) it is seen that the tetrapeptide Arg-Pro-Gly-Pro 1%, when administered to mice, intraperitoneally protected 40% of the animals from the action of the encephalomyocarditis virus (VEMK). The use of the peptide at a concentration of 0.5% was effective with all methods of administration. The introduction of the peptide at a concentration of 0.2% - with intraperitoneal administration to animals protected 20% of the mice. The use of the tetrapeptide Arg-Pro-Gly-Pro 0.1% led to the protection of 40% of the animals from infection by oral and intranasal routes of administration. Based on data showing% of surviving animals and the average life expectancy of mice, the most effective methods and doses of tetrapeptide administration were noted:

Arg-Pro-Gly-Pro 1% (w / w);

0.2% (p / os; i / n; w / b);

0.1% (p / os, i / n).

It was shown that the use of the tetrapeptide Arg-Pro-Gly-Pro in 0.2 ml / mouse 0.1-1% solution (0.2-5 mg / mouse) with a preservative in the form of nipagin with a concentration of 0.09-0.11 % with different methods of use contributed to an increase in the life of animals in some cases by 4.5-8.5 days.

table 2 The antiviral activity of the tetrapeptide Arg-Pro-Gly-Pro in different concentrations when administered in vivo in / br, i / n and per / os Peptide Administration Method Peptide concentration % survival % protection τ * Arg-Pro-Gly-Pro per / os one% 0 0 6.17 i / n one% 0 0 6.45 in / br one% 40 40 10.0 per / os 0.5% twenty twenty 8.06 i / n 0.5% twenty twenty 7.41 in / br 0.5% 40 40 8.33 per / os 0.2% 0 0 5.56 i / n 0.2% 0 0 5.49 in / br 0.2% twenty twenty 6.54 per / os 0.1% 40 40 9.80 i / n 0.1% 40 40 8.33 in / br 0.1% twenty twenty 6.25 VEMK CONTROL 100 LD50 0 0 5.35 Note: * τ is the average harmonic value of the life time of animals.

Table 3 Evaluation of the effectiveness of the tetrapeptide Arg-Pro-Gly-Pro in influenza Peptide administration schedule Peptide dose KZ * IE **% Arg-Pro-Gly-Pro per / os one% 1,0 0 i / n one% 1,0 0 in / br one% 1,67 40 per / os 0.5% 1.25 twenty i / n 0.5% 1.25 twenty in / br 0.5% 1,67 40 per / os 0.2% 1,0 0 i / n 0.2% 1,0 0 in / br 0.2% 1.25 twenty per / os 0.1% 1,67 40 i / n 0.1% 1,67 40 in / br 0.1% 1.25 twenty Virus control 100LD50 1,0 0 Note: * KZ - protection ratio; ** IE - drug effectiveness index.

The calculation of the efficacy index (IE) of the peptide for influenza showed (Table 3) that with the introduction of the tetrapeptide Arg-Pro-Gly-Pro IE = 40% when using 1% and 0.5% with i.v. and also 0.1% with per / os and i / n, it also effectively protected animals.

Determination of the antiviral activity of the tetrapeptide Arg-Pro-Gly-Pro with different schemes of administration to animals

The study of the antiviral activity of peptides in experimental infection caused by the mouse encephalomyocarditis virus (VEMK) in vivo.

Studies were conducted on the effectiveness of a number of peptides in infections caused by the mouse encephalomyocarditis virus (VEMC) in vivo with the introduction of the peptide to mice according to the schemes:

prevention -

1) 24 hours before infection,

2) 2 consecutive days 24 and 48 hours before infection;

treatment -

1) 24 hours after infection,

2) 24 and 48 hours after infection,

3) 3 times in a row after 24, 48 and 72 hours after infection.

Arg-Pro-Gly-Pro 0.1% i / n.

Infection was performed with mouse encephalomyocarditis virus (VEMK) at a dose of 100 LD50.

The results of the study of the antiviral activity of the tetrapeptide with different regimens for the administration of influenza virus in the experimental model of VEMC infection in mice are presented in Table 4. The tetrapeptide was shown to protect animals from the action of VEMK. There were up to 50% more surviving animals with the prophylactic tetrapeptide administration scheme compared to the control and up to 60% with the tetrapeptide used to treat mice.

Table 4 The survival of mice under the influence of the tetrapeptide Arg-Pro-Gly-Pro with different administration schemes for infection of mice with encephalomyocarditis virus (VEMK) Peptide % survival Arg-Pro-Gly-Pro prophylaxis single administration 60 Arg-Pro-Gly-Pro prophylaxis 2 single administration 70 Arg-Pro-Gly-Pro Treatment 1 single administration 60 Arg-Pro-Gly-Pro Treatment 2 times a day 70 Arg-Pro-Gly-Pro Treatment 3 times a day 80 the control twenty

The data obtained showed that the greatest antiviral effect against VEMC was achieved with the introduction of the tetrapeptide twice before infection or within three days after infection of mice.

Claims (4)

1. The use of the tetrapeptide Arg-Pro-Gly-Pro as a means for the prevention and treatment of viral diseases. 2. A pharmaceutical composition for the prevention and treatment of viral diseases, containing an aqueous solution of the tetrapeptide Arg-Pro-Gly-Pro, where the composition contains the tetrapeptide Arg-Pro-Gly-Pro in a concentration of 0.1-1% and nipagin as a preservative in the following contents components, g / l:
tetrapeptide Arg-Pro-Gly-Pro 1.0-10.0 nipagin 0.9-1.1 distilled water up to 1 l 3. A method for the prevention of viral diseases, including the introduction of the pharmaceutical composition according to claim 2 intranasally, orally or intraperitoneally at a dose of 0.2-5 mg per day per mouse for 2 days. 4. A method of treating viral diseases, comprising administering a pharmaceutical composition according to claim 2 intranasally, orally or intraperitoneally at a dose of 0.2-5 mg per day per mouse for 3 days.