JP7672680B2 - Anti-coronavirus agents - Google Patents

Description

The present invention relates to anti-coronavirus agents, etc.

Coronaviruses are a species of virus belonging to the Coronaviridae family, the Coronaviridae subfamily, that are positive-stranded RNA viruses that infect humans and animals and cause respiratory, digestive, vascular, or neurological diseases. Coronaviruses can spread from any infected host animal and cause large-scale epidemics in humans. Examples of coronaviruses in recent years include SARS coronavirus 1 (SARS-CoV-1), which was prevalent in 2002 and 2003, and Middle East Respiratory Syndrome coronavirus (MERS-CoV). Recently, a global epidemic of SARS coronavirus 2 (SARS-CoV-2) has occurred, and since no treatment technology has been established, it continues to cause severe damage in many areas, including medicine and the economy.

The objective of the present invention is to provide an anti-coronavirus agent, in particular an anti-coronavirus agent against SARS-CoV-2.

In view of the above problems, the present inventors have conducted intensive research and have found that the above problems can be solved by an anti-coronavirus agent containing at least one selected from the group consisting of alkyl gallates and their derivatives as an active ingredient. Based on this finding, the present inventors have conducted further research and have completed the present invention. That is, the present invention encompasses the following aspects.

Item 1. An anti-coronavirus agent containing at least one active ingredient selected from the group consisting of alkyl gallates, gallic acid, and derivatives thereof.

Item 2. The anti-coronavirus agent according to Item 1, wherein the coronavirus is SARS-CoV-2.

Item 3. The anti-coronavirus agent according to item 1 or 2, which contains the alkyl gallate as an active ingredient.

Item 4. The alkyl gallate and/or gallic acid is represented by the general formula (1):

[In the formula: R ¹ represents a hydrogen atom or an alkyl group.]
Item 4. The anti-coronavirus agent according to any one of Items 1 to 3, wherein the compound is represented by the formula:

Item 5. The anti-coronavirus agent according to Item 4, wherein R ¹ is a hydrogen atom or an alkyl group having 1 to 7 carbon atoms.

Item 6. The anti-coronavirus agent according to Item 4 or 5, wherein R ¹ is an alkyl group having 1 to 4 carbon atoms.

Item 7. An anti-coronavirus agent according to any one of items 1 to 6, which is used for application to a living body.

Item 8. The anti-coronavirus agent according to Item 7, which is a medicine, a cosmetic, a food composition, a food additive, a disinfectant, or a cleaning agent.

Item 9. An anti-coronavirus agent according to item 7 or 8 for preventing or ameliorating COVID-19.

Item 10. An anti-coronavirus agent according to any one of items 1 to 6, which is used for application to an article.

Item 11. The anti-coronavirus agent according to Item 10, which is a disinfectant or a cleaning agent.

The present invention provides an anti-coronavirus agent, particularly an anti-coronavirus agent against SARS-CoV-2.

The test method (Mix protocol) of Test Example 1 is outlined below. 1 shows the results of Test Example 1. The vertical axis indicates the cell death inhibition rate. The test substances are shown above each graph. The horizontal axis indicates the concentration of the test substance. The test method (post-infection protocol) of Test Example 2 is outlined below. 1 shows the results of Test Example 2. The vertical axis indicates the cell death inhibition rate. The test substances are shown above each graph. The horizontal axis indicates the concentration of the test substance.

In this specification, the expressions "contain" and "include" include the concepts of "contain," "include," "consist essentially of," and "consist only of."

In one aspect, the present invention relates to an anti-coronavirus agent (sometimes referred to as the "agent of the present invention" in this specification) that contains, as an active ingredient, at least one selected from the group consisting of alkyl gallates, gallic acid, and derivatives thereof. This will be described below.

1. Active Ingredient The active ingredient is at least one selected from the group consisting of alkyl gallates, gallic acid, and derivatives thereof.

The alkyl gallate is an alkyl ester of gallic acid, and is not particularly limited in this respect. Examples of alkyl gallates include methyl gallate, ethyl gallate, propyl gallate, butyl gallate, pentyl gallate, hexyl gallate, heptyl gallate, isopropyl gallate, isobutyl gallate, sec-butyl gallate, tert-butyl gallate, and isoamyl gallate. Among these, preferred are methyl gallate, ethyl gallate, propyl gallate, and butyl gallate, and more preferred are methyl gallate, ethyl gallate, and propyl gallate. Propyl gallate is particularly preferred from the viewpoint of safety, as it is recognized as a food additive.

Alkyl gallates and gallic acids are typically represented by the general formula (1):

[In the formula: R ¹ represents a hydrogen atom or an alkyl group.]
It is a compound represented by the formula:

The alkyl group represented by R ¹ includes any of linear, branched, and cyclic alkyl groups. From the viewpoint of antiviral activity, etc., the alkyl group is preferably a chain alkyl group, and particularly preferably a linear alkyl group. However, it may be a cyclic alkyl group. The number of carbon atoms of the alkyl group (in the case of linear or branched chain) is not particularly limited, and is, for example, 1 to 12. In one embodiment of the present invention, the number of carbon atoms is preferably 1 to 9, more preferably 1 to 7, even more preferably 1 to 5, still more preferably 1 to 4, and particularly preferably 1 to 3. The number of carbon atoms of the alkyl group (in the case of cyclic) is not particularly limited, and is, for example, 3 to 7, preferably 4 to 6. Specific examples of the alkyl group include a methyl group, an ethyl group, an n-propyl group, an isopropyl group, an n-butyl group, an isobutyl group, a tert-butyl group, a sec-butyl group, an n-pentyl group, a neopentyl group, an isopentyl group, an n-hexyl group, a 3-methylpentyl group, an n-heptyl group, and an n-octyl group.

^R1 is preferably an alkyl group.

There are no particular limitations on the alkyl gallates and gallic acid derivatives, so long as they can exert their antiviral effect either directly or after undergoing a reaction such as decomposition.

Examples of derivatives of alkyl gallates and gallic acid include compounds in which one to three hydroxyl groups in the general formula (1) are replaced with -OR (R is an optionally substituted hydrocarbon group or an optionally substituted heterocycle).

In addition, derivatives of alkyl gallates and gallic acid include, for example, glycosides of alkyl gallates or gallic acid. There are no particular limitations on the glycosides of alkyl gallates or gallic acid, so long as they are compounds in which alkyl gallates or gallic acid serve as aglycons and sugars are linked to them by bonds (usually glycosidic bonds). Examples of sugars include glucose, arabinose, pyranose, raffinose, rutinose, apiose, rhamnose, rutinose, primeverose, gentiobiose, digitoxose, galactose, xylose, and the like, and further include sugars in which multiple of these are linked together. The number of monosaccharide residues constituting the sugar is, for example, 1 to 8 or 1 to 6.

In addition, examples of derivatives of alkyl gallates and gallic acid include prodrugs of alkyl gallates or gallic acid. Prodrugs refer to compounds that are converted into the compounds of the present invention in vivo by reactions with enzymes, gastric acid, etc. Prodrugs of alkyl gallates or gallic acid include, for example, compounds in which the hydroxyl group is acylated, alkylated, phosphorylated, or borated (for example, compounds in which the hydroxyl group of an alkyl gallate is acetylated, palmitoylated, propanoylated, pivaloylated, succinylated, fumarylated, alanylated, or dialkyl (e.g., methyl)aminoalkyl (e.g., methyl)carbonylated); Examples of the compounds include compounds in which the carboxy group is esterified or amidated (for example, compounds in which the carboxy group of the compound of the present invention is alkyl (e.g., ethyl) esterified, aryl (e.g., phenyl) esterified, carboxyalkyl (e.g., methyl) esterified, dialkyl (e.g., methyl) aminoalkyl (e.g., methyl) esterified, pivaloyloxyalkyl (e.g., methyl) esterified, 1-{(alkoxy (e.g., ethoxy) carbonyl) oxy} alkyl (e.g., ethyl) esterified, phthalidyl esterified, (5-alkyl (e.g., methyl)-2-oxo-1,3-dioxolen-4-yl) alkyl (e.g., methyl) esterified, 1-{[(cycloalkyl (e.g., cyclohexyl) oxy) carbonyl] oxy} alkyl (e.g., ethyl) esterified, alkyl (e.g., methyl) amidated, etc.). These compounds can be produced by known methods. In addition, the prodrug of the compound of the present invention may be one that changes into the compound of the present invention under physiological conditions such as those described in "Drug Development" Vol. 7, "Molecular Design" pp. 163-198, Hirokawa Shoten, 1990.

Among the active ingredients, alkyl gallates are preferred.

The active ingredient may be a single type or a combination of two or more types.

As the alkyl gallate and its derivatives, commercially available products can be used, or products synthesized according to or similar to known methods can be used.

2. Uses The target coronavirus of the agent of the present invention is not particularly limited as long as it is a virus belonging to the Orthocoronavirus subfamily. Coronaviruses include the alphacoronavirus genus, betacoronavirus genus, gammacoronavirus genus, deltacoronavirus genus, etc., among which the betacoronavirus genus is preferred. Betacoronaviruses include SARS-related coronavirus (SARSr-CoV), coronavirus HKU1, MERS coronavirus, etc., among which SARSr-CoV is preferred. SARSr-CoVs include SARS-CoV-2, SARS-CoV-1, etc., among which SARS-CoV-2 is preferred. The agent of the present invention can be particularly suitably used against SARS-CoV-2.

The agent of the present invention can exert an antiviral effect against coronaviruses. Specific examples of the antiviral effect include an effect of suppressing viral infection, an effect of reducing the viral cell infectivity titer, an effect of suppressing viral cell death, a viral inactivation effect, an effect of suppressing viral proliferation, an effect of suppressing viral budding, and an effect of inducing viral resistance. In addition, due to this effect, the agent can also be used as a preventive or ameliorative agent (or a preventive or therapeutic agent) for coronavirus infections (particularly COVID-19).

The agent of the present invention can be widely used in various fields requiring antiviral properties. The agent of the present invention can be used in various fields such as industry, cleaning, medicine, food, and daily necessities. The uses of the agent of the present invention are mainly divided into applications for application to living organisms and applications for application to articles described below.

In one embodiment of the present invention, the agent of the present invention may have a lower content of alkyl gallates having an alkyl group with 8 or more carbon atoms or 5 or more carbon atoms. In the agent of the present invention, the content of alkyl gallates having an alkyl group with 8 or more carbon atoms or 5 or more carbon atoms is, for example, 0 to 50 mass% (or mol%), 0 to 20 mass% (or mol%), 0 to 10 mass% (or mol%), 0 to 5 mass% (or mol%), 0 to 2 mass% (or mol%), 0 to 1 mass% (or mol%), 0 to 0.1 mass% (or mol%), 0 to 0.01 mass% (or mol%), or 0 mass% (or mol%) relative to 100 mass% (or mol%) of alkyl gallates having an alkyl group with 7 or less carbon atoms or 4 or less carbon atoms.

In one embodiment of the present invention, the agent of the present invention may have a smaller content of antiviral components other than alkyl gallates. In the agent of the present invention, the content of antiviral components other than alkyl gallates is, for example, 0 to 50% by mass (or mol%), 0 to 20% by mass (or mol%), 0 to 10% by mass (or mol%), 0 to 5% by mass (or mol%), 0 to 2% by mass (or mol%), 0 to 1% by mass (or mol%), 0 to 0.1% by mass (or mol%), 0 to 0.01% by mass (or mol%), or 0% by mass (or mol%), relative to 100% by mass (or mol%) of the content of alkyl gallates (e.g., alkyl gallates having an alkyl group with 7 or less or 4 or less carbon atoms).

2-1. Uses applied to living organisms Examples of uses applied to living organisms include medicines, cosmetics, food compositions (including health foods, health enhancers, and nutritional supplements (supplements, etc.)), food additives, disinfectants, cleaning agents, etc. In this case, the subjects of application are not particularly limited, and examples include various mammals such as humans, monkeys, mice, rats, dogs, cats, rabbits, pigs, horses, cows, sheep, goats, and deer.

By applying the agent of the present invention to a living body, the active ingredient can exert an antiviral effect at the site of contact.

The form of the agent of the present invention is not particularly limited, and may be a form that is commonly used for each application depending on the application of the agent of the present invention.

In terms of forms, when the application is medicinal, for example, injections, drips, gargles, inhalants, patches (plasters, tapes such as plasters (reservoir type, matrix type, etc.), poultices, patches, microneedles, etc.), ointments, liquids for external use (liniments, lotions, etc.), sprays (aerosols for external use, pump sprays, etc.), creams, gels, eye drops, eye ointments, nasal drops, suppositories, semi-solid rectal preparations, enemas, and other preparation forms suitable for parenteral intake; These include tablets (including orally disintegrating tablets, chewable tablets, effervescent tablets, lozenges, jelly drops, sublingual tablets, etc.), pills, granules, fine granules, powders, hard capsules, soft capsules, dry syrups, liquids (including drinks, suspensions, syrups), jellies, and other formulations suitable for oral intake (oral formulations), tube feeding, enteral feeding, nasal catheters, esophageal fistula catheters, gastrostomy catheters, etc.

When used as a cosmetic product, the formulation may be, for example, a liquid, gel, cream, ointment, spray, or stick.

In the case of use as a food composition, examples of the form include liquid, gel, or solid foods, such as beverages such as juice, soft drinks, tea, soup, and soy milk, salad oil, dressing, yogurt, jelly, pudding, furikake, infant formula, cake mix, dairy products (e.g., powder, liquid, gel, solid, etc.), bread, and confectionery (e.g., cookies, etc.). In the case of use as a food additive, health enhancer, or nutritional supplement (supplement, etc.), examples of the form include tablets (including orally disintegrating tablets, chewable tablets, effervescent tablets, lozenges, jelly drops, etc.), pills, granules, fine granules, powders, hard capsules, soft capsules, dry syrups, liquids (including suspensions and syrups), and jellies.

When the application is a disinfectant or cleaning agent, the form may be any of liquids (solutions, emulsions, suspensions, sprays, etc.), semisolids (gels, creams, pastes, etc.), and solids (tablets, particulates, capsules, films, kneaded materials, molten solids, waxy solids, elastic solids, etc.). For example, when applied to the oral cavity, more specific examples include dentifrices (toothpaste, liquid dentifrice, liquid dentifrice, powder dentifrice, etc.), mouthwashes, liniments, patches, mouth fresheners, and foods (e.g., chewing gum, tablets, candies, gummies, films, lozenges, etc.). When applied to the nasal cavity, more specific examples include spray-type nasal drops, etc. When applied to the skin, examples include soap, body wash, shampoo, rinse, sprays, etc.

The agent of the present invention may further contain other components as necessary. The other components are not particularly limited as long as they are components that can be blended into, for example, medicines, cosmetics, disinfectants, cleaning agents, etc., and examples thereof include bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, thickeners, moisturizers, colorants, fragrances, chelating agents, etc.

There are no particular limitations on the buffer, and any suitable buffer may be used depending on the application. Preferred examples include phosphate buffer, acetate buffer, etc.

The content of the active ingredient in the agent of the present invention depends on the type of active ingredient, the purpose, the mode of use, the subject to which it is applied, and the condition of the subject to which it is applied, and is not limited thereto, but can be, for example, 0.000001 to 100% by weight, and preferably 0.01 to 50% by weight.

The amount of application (e.g., administration, ingestion, inoculation, etc.) of the agent of the present invention is not particularly limited as long as it is an effective amount that produces the desired effect, and is usually 0.1 to 1000 mg/kg body weight per day in terms of the weight of the active ingredient. The above dosage is preferably administered once a day or in two or three divided doses, and can be increased or decreased as appropriate depending on the age, pathology, and symptoms.

2-2. Applications for use in articles Examples of applications for use in articles include disinfectants, cleaning agents, etc. In this case, the application targets are not particularly limited, and include industrial products and their raw materials used in various fields.

Specific examples of items include masks, face masks, gloves, etc. Other examples include endotracheal intubation tubes, bite blocks, laryngoscopes, electrodes, measuring instruments, infusion devices, oxygen masks, nasal catheters, tubes, catheters, lab coats, gowns, caps, protective clothing, protective shields, surgical instruments and other medical equipment, operating tables, beds, stretchers, wheelchairs, etc. Other examples include office automation equipment, home appliances, air conditioning equipment, vacuum cleaners, desks, chairs, sofas, benches, windows, straps, handles, seats, automatic ticket gates, automatic ticket vending machines, vending machines, doors, fences, handrails, tableware, cooking utensils, packaging film, packaging bags, bottles, bottles, packaging packs, sinks, toilets, stationery, books, shelves, toothbrushes, mirrors, air conditioning filters, masks, coats, jackets, trousers, skirts, dress shirts, knit shirts, blouses, sweaters, cardigans, nightwear, undergarments, diapers, supporters, socks, tights, stockings, hats, scarves, mufflers, collars, stoles, gloves, clothing linings, clothing interlinings, clothing padding, work clothes, uniforms, school uniforms, and other clothing, curtains, screen doors, futon fabrics, futon batting, futon covers, pillowcases, sheets, mats, carpets, towels, handkerchiefs, wall cloth, band-aids, bandages, etc.

By applying the agent of the present invention to an article, the agent can exert an antiviral effect at the site where the active ingredient comes into contact. The site where the active ingredient comes into contact is not limited to the site on the article, but also includes sites inside the body when the article is applied to a living body.

The dosage form of the agent of the present invention is not particularly limited and can be appropriately selected depending on the application. Examples of dosage forms include liquids such as solutions, emulsions, suspensions, dispersions, sprays, and aerosols; and solids or semi-solids such as wettable powders, powders, granules, microgranules, and flowables. The agent can be applied or coated onto various articles.

The agent of the present invention may further contain other components as necessary. The other components are not particularly limited as long as they are components that can be blended into, for example, cleaning agents and disinfectants for articles, and examples thereof include bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, thickeners, moisturizers, colorants, fragrances, chelating agents, etc.

There are no particular limitations on the buffer, and any suitable buffer may be used depending on the application. Preferred examples include phosphate buffer, acetate buffer, etc.

The content of the active ingredient in the agent of the present invention depends on the type of active ingredient, the purpose, the mode of use, the subject to which it is applied, and the condition of the subject to which it is applied, and is not limited thereto, but can be, for example, 0.000001 to 100% by weight, and preferably 0.001 to 50% by weight.

The present invention will be described in detail below based on examples, but the present invention is not limited to these examples.

Test Example 1. Antiviral effect evaluation test (Mix protocol)
The outline of this test is shown in Figure 1. Methyl gallate (general formula (1): R ¹ = methyl group), propyl gallate (general formula (1): R ¹ = n-propyl group), or butyl gallate (general formula (1): R ¹ = n-butyl group) was dissolved in filtered sterilized water to a concentration of 100 mM to obtain a sample solution.

The sample solution was diluted in two stages with filtered sterilized water. 200 μl of SF DMEM (MS) or 200 μl of SARS-CoV-2 (5 X 10^5 TCID50/200 μl) was added to 2 μl of sample solution of each concentration and left to stand at room temperature for 1 minute. The day before, VeroE6/TMPRSS2 cells were seeded at 5 X 10^4/100 μl/well and cultured for 24 hours in a 96-well plate. The supernatant was removed from the plate and 100 μl of the above virus/sample solution was added (MOI 5). After 1 hour of adsorption, the virus solution was removed and MS was added at 100 μl/well and cultured for 24 hours. The viability of the cells was quantified using the following method.

The medium was removed from the plate and washed with PBS. Cell Count Reagent SF (Nacalai) was mixed with phenol red-free medium at 20%, and 50 μl/well of the mixture was added to the cells and incubated for 60 minutes. As a blank, wells to which cells were not incubated were also prepared to which only the reagent mixture was added. After the reaction was completed, 20 μl/well of 10% SDS was added to inactivate any virus remaining in the wells. The absorbance at a wavelength of 450 nm was measured using a plate reader. Measurements were performed in tetraplicates. The %Inhibition of cell death was calculated based on the following formula: %Inhibition of cell death = (OD((Virus + Alkyl gallate)-OD(Virus only))/(OD(Alkyl gallate only)-OD(Virus only)) x 100.

<Results>
The results are shown in Figure 2. When added to SARS-CoV-2, methyl gallate, propyl gallate, and butyl gallate all suppressed cell death caused by virus infection in a concentration-dependent manner.

Test Example 2. Antiviral effect evaluation test (Post-infection protocol)
The outline of this test is shown in Figure 3. Methyl gallate (general formula (1): R ¹ = methyl group) or propyl gallate (general formula (1): R ¹ = n-propyl group) was dissolved in filtered sterilized water to a concentration of 250 mM. The solution was then treated in a thermostatic bath at 78°C for 5 minutes to obtain a sample solution.

The sample solution was diluted in two steps with 0.5% FBS DMEM (MS). VeroE6/TMPRSS2 cells were seeded at 5 x 10^4/100μl/well in a 96-well plate and cultured for 15 hours, after which the supernatant was removed from the cells and SARS-CoV-2 was added at an MOI of 5 and allowed to adsorb for 1 hour. The virus solution was removed and sample solution or MS of each concentration was added at 100μl/well and cultured for 24 hours. Quantification of cell viability and calculation of % inhibition of cell death were performed in the same manner as in Test Example 1 above.

<Results>
The results are shown in Figure 4. When added to cells to which SARS-CoV-2 had been adsorbed after washing, both methyl gallate and propyl gallate concentration-dependently suppressed cell death caused by virus infection.

Claims (8)

An anti-coronavirus agent comprising at least one active ingredient selected from the group consisting of alkyl gallates, gallic acid, and derivatives thereof ,
The alkyl gallate and/or gallic acid has the general formula (1):
[In the formula: R ¹ represents a hydrogen atom or an alkyl group.]
A compound represented by the formula:
R ¹ is a hydrogen atom or an alkyl group having 1 to 7 carbon atoms,
the coronavirus is SARS-CoV-2,
The derivative is a glycoside or a prodrug in which one of the hydroxyl groups in the general formula (1) is substituted with -OR (R is an optionally substituted hydrocarbon group or an optionally substituted heterocycle).
Anti-coronavirus agent . The anti-coronavirus agent according to claim 1 , comprising the alkyl gallate as an active ingredient. The anti-coronavirus agent according to claim 1 or 2 , wherein R ¹ is an alkyl group having 1 to 4 carbon atoms. The anti-coronavirus agent according to any one of claims 1 to 3 , which is used for application to a living body. The anti-coronavirus agent according to claim 4 , which is a medicine, a cosmetic, a food composition, a food additive, a disinfectant or a cleaning agent. The anti-coronavirus agent according to claim 4 or 5 , for preventing or ameliorating COVID-19. The anti-coronavirus agent according to any one of claims 1 to 3 , which is applied to an article. The anti-coronavirus agent according to claim 7 , which is a disinfectant or a cleaning agent.

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