RU1835291C - An antiviral means - Google Patents

Abstract

The invention relates to medicine, namely to the pharmacotherapy of infectious diseases. The purpose of the invention is to increase the efficacy of drug resistant viral strains. To do this, trental is administered as an antiviral agent into an organism affected by a viral infection. 5 tab.

Description

The invention relates to the field of virology and can be used to solve the problems associated with the inhibition of viral infections.

The search for antiviral agents among official drugs is particularly relevant today because the development of inhibitors of viral activity from chemical compounds remains a very long process, taking 10-15 years. At the same time, it is possible that this process at a certain stage can be disrupted altogether due to emerging technological, technical, organizational or other difficulties.

The purpose of the invention: the search for a highly effective antiviral agent against drug resistant viral strains.

The goal is achieved by using the trental cardiovascular drug,

Trental is known as a cardiovascular antispasmodic agent. It has a vasodilating effect, improves oxygen supply to tissues, inhibits phosphodiesterase and causes accumulation of cyclic AMP in tissues, inhibits platelet aggregation and reduces blood viscosity. It is used for disorders of peripheral circulation and cerebrovascular pathology (atherosclerotic disorders, ischemic conditions, etc.), for ocular fundus vascular pathology, and for diabetic angiopathies.

The novelty of the invention lies in the discovery of the lost of new biological properties in the form of antiviral action.

The inventiveness of the invention is that it was not possible to propose the presence of a drug widely used in cardiological practice that has a therapeutic effect in cardiovascular disease, a new function - inhibition of viruses.

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The unexpected positive effect is not only the detection of new biological activity to suppress viral reproduction, but also the identification of the high efficiency of inhibition of a wide range of RNA and DNA viruses with effects on drug-resistant viral.

PRI me R 1. The study of inhibitory

Trental activity against

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herpes simplex virus (strain 1C) was performed in vitro on a trypsinized chicken embryo fibroblast culture (FEC). The cell culture grown on Petri dishes (screening test) and the bottom of the penicillin 15 vials (plaque reduction method) was washed from the growth medium and 0.001-0.00001 VOE / virus cell was added. After adsorption at 37 ° C, the virus was removed and the monolayer was coated with a solid nutrient coating prepared on the basis of 199 + 1% agar + 0.005% neutral red. An agar-coated Petri dish was added to glass tubes (0.05 ml). After incubation at 37 ° C, the diameters of the toxicity zones and the suppression of plaque formation (mm) were determined. Using the plaque reduction method, the preparation was added at various concentrations to the solid 30 agar coating. After incubation at 37 ° C for 48 to 72 hours, the infectious titer of the virus was determined both in the presence of the preparation and without it. The criterion for antiviral activity was lower 35 niter of the virus in the presence of the drug in comparison with the control. The minimum active concentration (MAK) was considered to be a decrease in the titer of the virus in comparison with the control by 1.25 Ig BOE / cell.

The toxicity for the FEC culture was determined by incubating the cells at 37 ° C in the presence of various drug concentrations for 96 hours. The highest tolerated concentration (BMD) was considered the 45 highest dose, which did not cause cyto destructive changes in cells upon registration under a small magnification of the microscope. A chemotherapeutic index (CTI) was defined as the IPC / MAC ratio. fifty

As a result of the studies, it turned out that trental. being a low-toxic preparation for cell culture (IPC-800 μg / ml), it had a pronounced inhibitory effect against herpes simplex virus 55 (strain 1C). Thus, at a dose of 100 µg / ml, he completely suppressed the reproduction of the virus in culture (Table 1).

The study of the inhibitory activity of trental in relation to the herpes virus,

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Stent to action of acyc / juvir (ACV strain) was carried out according to the same scheme. Moreover, the official drug acyclovir was taken as a comparative control. The results obtained showed the pronounced ability of trental to suppress the reproduction of ACvR in culture, FEC, which under the same conditions was not inhibited by acyclovir. Using a dose of 100 μg / ml, it was possible to completely inhibit the reproduction of the acyclovir-resistant strain with an infectious titer of 4.64 Ig PFU / ml (Table 1).

Example 2. The study of the antiherpetic activity of trental in the system was carried out in vivo on the model of experimental mouse herpetic meningoencephalitis. Mice (weight 7-10. Grams) were infected with the strain. Koptev intraobdominally at a dose of 100 LDso Trental in tolerant doses was administered intravenously in a volume of 0.2 ml according to the following treatment regimen: 4 hours after infection, then once, for 7 days daily. The results were evaluated by the lethality of the treated animals in comparison with the placebo group received intravenously 0.85% NaC solution according to a similar scheme.

The results obtained are presented in Table 2. As can be seen, trental showed a pronounced protective effect - a decrease in mortality in the group of treated animals was 66.6 - 75%. At the same time, a high degree of antiviral activity was combined with a wide breadth of the protective effect — the chemotherapy CTI of the drug exceeded 100. The tested doses of the drug were tolerant to mice. When administered intravenously, no toxic effects were noted. Particularly noteworthy is the fact that the drug retained antiviral activity in very low doses, at least 100 times lower than pharmacopeia.

EXAMPLE 3 A study of the inhibitory activity of trental against vaccinia viruses was carried out according to the scheme described in Example 1. The results obtained showed the ability of the drug to significantly suppress the reproduction of vaccinia viruses (BOB) in culture. At the same time, the drug inhibited a laboratory strain sensitive to metisazon (BOB and a resistant strain of the virus (BOBfy, which naturally was not suppressed by methisazone (Table 3).

EXAMPLE 4 A study of the trental inhibitory activity against rotavirus (strain A) was carried out according to the usual scheme already described in Example 1. Studies were performed on a transplanted green monkey kidney cell culture (MA-140). The MPC of the preparation for this culture was 800 µg / ml, which corresponded to that. on FEC cells. As a result of 5 studies, it turned out that trental, used at concentrations of 800, 400, 200, and 100 μg / ml, reduced the infectious titer by more than 1.34 Ig PFU / ml, which, according to the accepted assessment of antiviral effect (7 ), .- testifies to the presence of antirotavirus activity in it (Table 4).

Example 5. The study of the inhibitory properties of trental in relation to the human immunodeficiency virus (IN I-1) was carried out 15 on a culture of MT-4 lymphoblastoid cells. As a result of the studies, it turned out that the drug, being low-current. 400 μg / ml), which is typical for this cell culture), had a pronounced 20 ability to suppress the accumulation of intracellular HIV antigens. Thus, according to immunofluorescence analysis, trental at doses of 4, 20 and 200 μg / ml increased the accumulation of HIV antigens in cells of 25 cells by 55; 63 and 68%, respectively, which, according to the accepted rating system (), allows us to classify it as drugs with anti-HIV activity. At the same doses, trental significantly reduced the infectious titer of the virus. A dose of 200 µg / ml was sufficient to completely suppress HIV reproduction with a titer of 4 Ig CD50 / ml (Table 5}. At the same time, the high degree of antiviral activity of trental was combined with the great breadth of its inhibitory effect - the chemotherapy CTI was 200.

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The study of trental in virological screening using other test viruses revealed, in addition to the examples described, its inhibitory activity against the virus of vesicular stomatitis (Indiana strain),

Thus, the object of the invention has been achieved. A highly effective antiviral agent has been identified that suppresses drug resistant viral strains. As one of examples 1-5, trental, being a low-toxic drug, significantly and sometimes completely suppressed the reproduction of viruses in culture (from 50% to 100%), and also had a protective effect under in vivo experimental infection conditions (66, 6–75.0%), which allows it to be considered (according to the generally accepted assessment system) a highly effective inhibitor of viral activity. The ability of the drug to significantly suppress the reproduction of acyclovir and metisazone-resistant strains allows it to be considered a highly effective inhibitor of drug-resistant viruses.

The positive effect of trental with respect to a whole series of DNA and RNA-containing viruses, herpes simplex, smallpox vaccine, human immunodeficiency, exsicular stomatitis and rotavirus allows attributing it to a wide spectrum of antiviral agents.

FORMULA AND SECTION

The use of trental as an antiviral agent.

Characterization of the inhibitory effect of trental against herpes simplex viruses (HSV)

1 C is a laboratory strain of HSV that is sensitive to the action of acyclovir;

infectious titer 4.77 Ig PFU / ml. ACV E acyclovir-resistant strain of HSV; infectious titer 4.64 Ig PFU / ml

Table 1

The effectiveness of the drug trental on a model of experimental herpetic meningoencephalitis in mice.

o) ++++ - the degree of activity is high

T-a Blitz S Characterization of the inhibitory effect of trental against vaccinia viruses.

VHF - smallpox vaccine-sensitive virus. BOBP - methisazone-resistant vaccinia virus.

Table 4 — Effect of trental on rotavirus replication (strain SA) in MA-140 cell culture.

table 2

Trental inhibitory effect on HIV-1

 Immunofluorescence assay data using human polyclonal antiserum obtained from virus carriers; working dilution 1: 4bO. The initial virus titer was 4 Ig TCDso / ml.

Table 5

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