RU2747890C1 - Agent for reducing risk and relieving symptoms of beta-coronavirus infection - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: group of inventions relates to the field of medicine, namely to infections diseases, and is intended for the prevention and treatment of infections caused by beta-coronaviruses. Proposed is an agent for reducing the risk of occurrence and/or relieving the symptoms of beta-coronavirus infection in form of an inhalation solution includes glutathione or a composition of glutathione disulfide and glutathione disulfide S-oxide, inosine as a complex active ingredient, potassium chloride and water for injection. Components are used in labeled amount. Also proposed is the use of the said agent for reducing the risk of occurrence and/or relieving symptoms of beta-coronavirus infection. The agent is sprayed with a nebulizer providing a particle size of 4-5 microns with a volume velocity of compressed air of 0.25 ml/min for 3-12 minutes per session, 10-14 days for 2-4 inhalations per day and 4-12-hour periods between sessions.

EFFECT: use of the group of invention allows increasing effectiveness of medicamentous prevention of beta-coronavirus infection and/or relieving its symptoms in persons who are at high risk of contracting this infection.

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Description

The invention relates to the field of chemical-pharmaceutical industry and medicine and concerns new dosage forms of drugs against infections caused by beta-coronaviruses.

Beta coronaviruses are one of four genera (alpha, beta, gamma and delta) coronaviruses, which are positive-strand enveloped RNA viruses that infect mammals. Most clinically relevant to humans are OC43 and HKU1 (which can cause the common cold) line A, SARS-CoV and SARS-CoV-2 (which cause COVID-19 disease) line B, and MERS-CoV line C.

COVID-19 is a disease caused by severe acute respiratory syndrome virus 2, first detected in humans in December 2019 (Zhu N, Zhang D, Wang W, et al. A novel coronavirus from patients with pneumonia in China, 2019. N Engl J Med 2020; 382: 727-733). Until September 29, 2020, more than 30 million cases of infection and 950 thousand deaths have been registered in the world. The most pronounced genetic difference of the SARS-CoV-2 virus is the elongated hycoprotein S, which is approximately 80% identical in nucleotide sequence to the previous SARS-CoV and MERS viruses (Lu R, Zhao X, Li J, et al. Genomic characterization and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding. Lancet 2020; 395: 565-574). Given the similarities and studies of SARS-CoV viruses, the pathogenesis of COVID-19 is considered from the point of view of the inclusion of phases of the infectious process corresponding to the penetration and replication of viral particles in the upper respiratory tract, further migration of the virus along the respiratory tract, followed by a pronounced immune response, causing hypoxia and the development of ARDS (Cao W, Li T. COVID-19: towards understanding of pathogenesis. Cell Research 2020; 30: 367-369).

Despite the ongoing development of new vaccines against COVID-19 in the world, to date, there are no proven effective drugs for the prevention of this disease. According to the Temporary Guidelines of the Ministry of Health of Russia for the drug prevention of COVID-19, it is possible to prescribe intranasal forms of IFN-α (in pregnant women, only intranasal administration of recombinant IFN-α2b is possible) and / or umifenovir.

Health care workers who come into contact with and / or provide COVID-19 patients

care are at a higher risk of infection than the general population. Health workers account for approximately 14% of COVID-19 cases, and up to 35% in some countries, according to WHO data collected as part of WHO's global COVID-19 surveillance. Therefore, reducing the risk of infection is essential to maintain the health of healthcare providers and other people who come into contact with patients and to reduce the spread of COVID-19. However, to date, there are no effective remedies for this category. As a rule, in medical institutions, the prevention of disease in medical workers who have identified a patient consists in treatment with disinfectant and antiseptic solutions. Basically, the research carried out is not about prevention, but about treatment, while the previously known agents that have shown antiviral activity against diseases similar to betavirus infections are being studied.

In particular, a number of studies concern such a group of drugs as antioxidants, the mechanisms of action of which can be aimed at maintaining the ventilation-perfusion functions of the lungs. A striking example is glutathione (γ-L-glutamyl-L-cysteinyl-glycine; GSH) - the main non-protein thiol (Meister A, Anderson ME. Glutathione. Annu Rev Biochem 1983; 52: 711-760), the concentration of which is reduced in alveolar cells when exposed to toxins or respiratory viruses (Mulier B, Rahman I, Watchorn T, Donaldson K, MacNee W, and Jeffery PK. Hydrogen peroxide-induced epithelial injury: the protective role of intracellular nonprotein thiols (NPSH). Eur Respir J 1998; 11 : 384-391). Aerosol delivery of GSH has been shown to restore its level in the fluid of the epithelial lining of the lungs of the lower respiratory tract in vivo (Buhl R, Vogelmeier C, Critenden M, et al. Augmentation of glutathione in the fluid lining the epithelium of the lower respiratory tract by directly administering glutathione aerosol PNAS 1990; 87: 4063-4067).

Recently, data was published on the use of high doses of glutathione to treat patients with shortness of breath secondary to COVID-19 pneumonia. It has been concluded that glutathione, glutathione precursors (N-acetyl-cysteine) and alpha-lipoic acid may represent a novel treatment approach to block NF-κB and reverse cytokine storm syndrome and respiratory distress in patients with COVID pneumonia. -19 (https://www.sciencedirect.com/science/article/pii/S22130071203013500). However, only oral and intravenous routes of administration of glutathione have been investigated.

The prior art also discloses solutions for inhalation forms.

In particular, international application WO 2014070769 A1 is known, which discloses an inhalation composition in the form of an aerosol with a gas or a liquid propellant containing glutathione, a pharmaceutically acceptable salt of glutathione or a derivative or prodrug thereof, an organic acid or a pharmaceutically acceptable salt thereof, and bicarbonate. The compositions can be used to treat a disorder or disease of the lungs or respiratory tract, where the disorder is selected from the group consisting of chronic inflammatory lung diseases and / or infections associated with lung transplantation, bronchitis, sinusitis, asthma, bacterial, fungal, parasitic, viral infections, COPD, ARDS, etc. It is noted that prophylactic administration of the composition to the mucosal tissue can establish and / or maintain homeostasis in the mucosal tissue to prevent and / or reduce tissue damage or other pathological damage. However, the possibility of using it for the prevention of beta-coronavirus infections has not been disclosed.

Among the group of peptide drugs for the treatment of coronavirus infections, the inhaled drug dalargin can be noted. In particular, in patent RU 2728939 C1, publ. 08/03/2020, data related to the use of dalargin hexapeptide in the production of medicines for the treatment of coronavirus infection COVID-19 are presented. To prepare an inhalation drug, the hexapeptide diacetate lyophilisate is dissolved in purified water and the resulting solution is poured into vials or ampoules under a nitrogen atmosphere. In the method for treating COVID-19, the resulting solution is administered to a subject in need thereof by inhalation in the form of an aerosol using a nebulizer. However, the possibility of drug prophylaxis by a known agent is not assumed.

Thus, there is a need to find new drugs for effective drug prevention of COVID-19. Such prophylaxis should not only safely protect a person from the high risk of contracting the virus through contact with infected individuals, but should also prevent further development of infection if it enters the body.

The objective of the present invention is to develop an effective drug for the drug prevention of contracting beta-coronavirus infection and / or alleviating its symptoms in individuals who are at high risk of contracting this infection, such as healthcare workers.

When solving this problem, the authors proceeded from the assumption that beta-coronavirus infection as a disease can be prevented by direct maintenance of its own redox, metabolic and dilatation functions of the lung tissue by using drug combinations that have previously shown safety in the aerosol delivery method.

The problem is solved by a new dosage form and its use for pre-exposure prophylaxis and / or alleviation of the symptoms of the disease by using a spray of active substance in low dosages. For inhalation, nebulizers are preferably used that provide a particle size of the sprayed preparation of no more than 4-5 microns.

The proposed agent for reducing the risk of occurrence and / or alleviating the symptoms of beta-coronavirus infection includes therapeutically effective concentrations of glutathione or compositions of glutathione disulfide and glutathione disulfide s-oxide, inosine as a complex active principle. The agent is a composition in the form of a solution for inhalation, including glutathione or a composition of glutathione disulfide and glutathione disulfide of S-oxide, inosine, potassium chloride and water for injection with the following content of components:

Glutathione or glutathione disulfide composition and glutathione disulfide S-oxide 10.0-60.0 mg / ml Inosine 5.0-30.0 mg / ml Potassium 0.020-0.145 mM / ml Water for injections up to 1 ml

The lower limits of the content of the components are the minimum effective concentrations, and the upper ones are selected taking into account the obtained data on the safety of inhalation use. Potassium chloride is taken in an amount selected according to the normotonicity of potassium, to ensure the content of 20-145 millimol / liter of potassium ions (0.020-0.145 mM / ml).

Glutathione affects the intracellular processes of thiol metabolism, the redox state of cells, showing an immunomodulatory and systemic cytoprotective effect. It has a high affinity for the cells of the central organs of the immune system and the lymphoid tissue system; stimulates the processes of erythropoiesis, lymphopoiesis and granulocyto-monocytopoiesis; activates the phagocytosis system (including under conditions of immunodeficiency states), promotes the restoration of the level of neutrophils, monocytes, lymphocytes in the peripheral blood and the functional activity of tissue macrophages, initiates the cytokine system.

Glutathione disulfide (or oxidized glutathione, GSSG) is a dimer of glutathione tripeptide, γ-glutamylcysteinylglycine, in which two molecules of said tripeptide are linked to each other by a covalent disulfide bond between cysteine residues. It exhibits antioxidant, anti-inflammatory, hepatoprotective properties and also affects, like glutathione, the cytokine system.

Glutathione S-oxide disulfide (also called glutathione thiosulfinate, or GS (O) SG) or sulfone), has similar pharmacokinetic properties to oxidized glutathione, and is also a negative regulator of oxidized glutathione degradation enzymes, thus acting as a prolonging agent of GSSG, optimizing it pharmacokinetics, potentiates the biological effects of oxidized glutathione, which optimizes the pharmacodynamics of GSSG and makes it possible to use smaller doses of GSSG to obtain the desired therapeutic effect. A composition consisting of glutathione disulfide and S-oxide glutathione disulfide is described, for example, in patent RU 2659161 C1.

Inosine is a purine derivative that can be considered a precursor of ATP. It has antihypoxic, metabolic and antiarrhythmic effects. Increases the energy balance of the myocardium, improves coronary circulation, promotes metabolic activation under conditions of hypoxia and in the absence of ATP. Reduces platelet aggregation, activates tissue regeneration. In combination with glutathione, it exhibits a more effective cyto- and hepatoprotective effect.

The choice of excipients for creating an inhalation form is of great importance, they provide the necessary osmolarity, viscosity of the dosage form, stability of medicinal substances. Typically, most of these forms include solvents, propellants, surfactants, film formers, flavoring agents, antimicrobial preservatives, antioxidants. However, some ingredients can cause mucous membrane irritation and coughing. The inventive form has a minimum of excipients, while safely and effectively provides deep penetration to the lungs.

The product is intended for use to reduce the risk of onset and / or alleviate the symptoms of beta-coronavirus infection by nebulizing it with a nebulizer providing a spray particle size of 4-5 microns at a compressed air volumetric velocity of 0.25 ml / min for 3-12 minutes per session, 10-14 days, 2-4 inhalations per day and 4 - 12-hour periods between sessions.

Preferably, the beta coronavirus infection is SARS-CoV infection.

The possibility of carrying out the invention can be illustrated by the following examples.

Example 1

Composition of glutathione disulfide and glutathione S-oxide disulfide 10 mg, Inosine 5 mg, Potassium chloride to provide a potassium ion content of 0.020 mM / ml (1.49 mg), water for injection up to 1 ml (pH 5.5-7.2).

The agent is prepared under aseptic conditions by separately dissolving the active principle in water and potassium chloride in water, followed by mixing them.

Example 2

Glutathione 60 mg, Inosine 30 mg, Potassium chloride to provide a potassium ion content of 0.145 mM / ml (10.8 mg), water for injection up to 1 ml (pH 5.5-7.2).

Example 3 Study of pharmacological activity.

The study group was defined as individuals providing medical care and services to patients with COVID-19 (healthcare workers, MR), including doctors, nurses, orderlies, technicians and other personnel, using the necessary personal protective equipment according to established recommendations. An aerosol drug combination (ALA) was prepared ex tempore for one 5-minute inhalation session. The solutions of examples 1 and 2 were tested. Participants applied ALA on their own using personal nebulizers with a compressed air volumetric flow rate of 0.25 ml / min. The included study participants underwent prophylaxis medication as instructed for 14 days, four inhalations per day and 4-hour intervals between sessions.

Data on adherence to ongoing prophylactic treatment and adverse events were recorded on the 7th and 14th days of the study; additional information was requested before the 28th day. The main indicator of the study is cases of infection with the SARS-CoV-2 virus, detected according to the data of genetic or immunological tests during the period of the study. A comparison group of medical workers not included in the study was analyzed retrospectively at the same hospital, including only those with negative results of genetic and serological tests and who continued to work until the date of the end of the study. To detect SARS-CoV-2 by RT-PCR, the gene for RNA-dependent RNA polymerase (RNA-dependent RNA polymerase; RdRp) was used in the sequence structure of the open reading frame1ab (English - open reading frame1ab; ORF1ab) and the envelope gene (E) using a set Polivir SARSCoV-2 (Litekh, Moscow, Russia) and an amplifier CFX-96 Touch system (Bio-Rad, Hercules, USA). The result was considered positive if the cut-off value was 35 for both genes. Serum antibodies were tested using diagnostic kits approved in the Russian Federation for the qualitative detection of immunoglobulins to SARS-CoV-2 in human plasma according to the manufacturer's instructions: SARS-CoV-2-IgMIFA BEST (Vector-Best, Novosibirsk, Russia) and SARS-CoV -2 – IgG (Litekh, Moscow, Russia). Antibodies were analyzed spectrophotometrically for optical density at 450 nm using a HumaReader HS microtiter plate reader (HUMAN, Wesbaden, Germany).

Statistical Analysis Chi-square test was used to compare the level of infection with SARS-CoV-2 virus between the test group and the control group; the value was considered statistically significant at p <0.05. Statistical analysis was performed using SAS software.

The study involved 367 people: 99 health workers - healthy volunteers with negative results of genetic and immunological analysis for SARS-CoV-2; 268 medical workers made up the comparison group.

The average age of the participants was 27.0 years (95% CI: 25.3 - 28.7), 69% were female and 52% were nurses (51/99). The demographic parameters of the group of participants and the comparison group did not differ significantly from each other (Table 1).

Table 1

Characteristics of medical personnel in the hospital for the treatment of COVID-19.

Characteristic Study participants
with ALA (n = 99) Comparison group
(n = 268) Demographic data Median age - years (BO) 25 (5) 25 (11) Age category - No. (%) <40 88 (89) 228 (85) 40 to 65 years old 11 (11) 40 (15) Male gender - No. (%) 31 (31) 106 (40) Professional data Doctors - No. (%) 38 (38) 99 (37) Nurses - No. (%) 51 (52) 140 (52) Others - No. (%) 10 (10) 29 (11)

* VO - probable deviation. ALK - aerosol drug combination

As a result of the study, it was found that pathogenetically justified pre-exposure drug prophylaxis was accompanied by a significant decrease in the rate of infection with the SARS-CoV-2 virus in medical workers providing medical care and services to patients with COVID-19 to 2% (composition 1) and 1.2% (composition 2) in comparison with the level of infection with the SARS-CoV-2 virus up to 9% in medical workers who did not use prophylactic treatment in the same hospital during the study period.

A significant decrease in the incidence of SARS-CoV-2 in a study in medical workers who used ALA confirms the assumed significance of virus-induced ACE2 / Ang II regulatory imbalance in the lungs, followed by increasing pulmonary hypoxia and peripheral ventilation perfusion disorders in the clinical picture of COVID-19. Moreover, the results obtained confirm the overriding importance of these processes in the pathogenesis of COVID-19 in comparison with various variants of the immune response and viral load per se.

Example 4

In the group with a positive result for SARS-CoV-2 (43 patients), 18 had an asymptomatic course, in the rest of the group (25 patients), the drug significantly reduced the existing symptoms of infection, contributing to the acceleration of recovery. At the same time, on the basis of biochemical analyzes, the study of the physical condition and the data of immunological tests, an improvement in microcirculation and rheological properties of blood, an increase in tissue saturation with oxygen, an improvement in the energy supply of cells, in particular, the heart, brain and liver, the stability of their membranes, suppression of RNA replication, were noted. viruses, an increase in the activity of macrophages and the production of interferons.

Thus, the proposed agent in inhalation form can be used both to prevent and to facilitate the course of infections caused by beta-coronaviruses, including COVID-19.

Claims (4)

1. An agent for reducing the risk of occurrence and / or alleviating the symptoms of beta-coronavirus infection in the form of a solution for inhalation, characterized in that it includes glutathione or a composition of glutathione disulfide and glutathione S-oxide disulfide, inosine as a complex active principle, potassium chloride and water for injection with the following content of components: Glutathione or glutathione disulfide composition and glutathione disulfide S-oxide 10.0-60.0 mg / ml Inosine 5.0-30.0 mg / ml Potassium 0.020-0.145 mM / ml Water for injections up to 1 ml 2. The use of the agent according to claim 1 to reduce the risk of occurrence and / or alleviate the symptoms of beta-coronavirus infection by spraying it with a nebulizer providing a spray particle size of 4-5 microns with a compressed air volumetric velocity of 0.25 ml / min for 3-12 minutes per session, 10-14 days, 2-4 inhalations per day and 4-12-hour periods between sessions. 3. Use according to claim 2, wherein the beta coronavirus infection is SARS-CoV infection.