KR101097189B1 - Pharmaceutical compositions comprising dihydroxychromone derivatives as an active ingredient for treating and preventing diseases caused by coronaviruses - Google Patents

Abstract

The present invention relates to a dihydroxychromone derivative, and more particularly to a pharmaceutical composition comprising a dihydroxychromone derivative compound.

The dihydroxychromone derivative of the present invention has an effect of inhibiting the activity of SARS-coronavirus helicase (SCV Hel), which causes Severe Acute Respiratory Syndrome (SARS). Therefore, it can be usefully developed as a pharmaceutical composition for the treatment and prevention of diseases caused by coronaviruses and / or inhibitors of viral helicase activity.

SARS coronavirus (SCV), helicase, inhibitor, dihydroxychromone

Description

PHARMACEUTICAL COMPOSITIONS COMPRISING DIHYDROXYCHROMONE DERIVATIVES AS AN ACTIVE INGREDIENT FOR TREATING AND PREVENTING DISEASES CAUSED BY CORONAVIRUSES}

The present invention relates to novel uses of dihydroxychromone derivatives as therapeutic agents for diseases caused by coronaviruses.

Viral helicases have the ability to hydrolyze NTP and use the energy obtained to unwind viral nucleic acid strands and have already developed new drug candidates targeting helicases of herpes virus (HSV) and hepatitis C virus (HCV). Has been in clinical trials. SARS coronavirus helicase consists of a cysteine-rich N-terminal metal binding domain (MBD) consisting of 100 amino acid residues, a hinge domain and an NTPase / helicase domain. Among them, it is the binding of Zn ^{2 +} ions in the SARS corona virus helicase MBD is known to be important for the antiviral activity. The study of SARS coronavirus inhibitors targeting the NTP binding site of the NTPase / helicase domain is considered to be of sufficient value, but so far not so much research has been done that even the three-dimensional structure of the target is unknown. To date, compounds known to inhibit the activity of SARS coronavirus helicase are bismuth complexes (3.0-11.0 mM) [ Chem . Comm . 2007 , 4413-4415], HE 602 (6.0 mM) [ Chem . Biol . 2004 , 11 , 1293] and vanillinbananin (2.7 mM) [ Chem . Biol. 2005 , 12 , 303].

The inventors of the present invention (Application No. 10-2008-0088348) have demonstrated the ability to inhibit nucleic acid substrate annealing activity of SARS coronavirus helicase by aryl diketosan derivatives with appropriate substituents [ Bioorg . Med . Chem . Lett . 2009 , 19 , 1636-1638] Some of these compounds showed high inhibitory effects. However, aryldiketo acid is difficult to be used as a drug due to strong polarity and chemical instability. Accordingly, in the present invention, as a result of trying to find a new material that can solve the problem of solubility and stability of aryl diketo acid, dihydride having a skeleton similar to the diketo acid of aryl diketo acid [FIG. 1] and proved stability as a kind of flavonoid The invention was completed by verifying the inhibitory effect on the helicase activity of SARS coronavirus after selecting hydroxychromone [FIG. 1] and synthesizing derivatives thereof.

Accordingly, an object of the present invention is to provide a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus including dihydroxychromone derivative as an active ingredient.

It is also an object of the present invention to provide an inhibitor of nucleic acid substrate unwinding activity of coronavirus helicase including dihydroxychromone derivative.

The objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

In order to achieve the above object, the present invention comprises an isomer comprising a compound of formula (1) or a pharmaceutically acceptable salt thereof, a hydrate thereof or a tautomer thereof as an active ingredient, and at least one pharmaceutically acceptable carrier It provides a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, including a diluent or excipient together.

In another aspect, the present invention provides a nucleic acid substrate unwinding activity inhibitor of coronavirus helicase comprising a compound of formula (1).

In Chemical Formula 1,

R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 10; R ₃ to R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₁₀ alkyl, C ₁ -C ₁₀ alkenyl or C ₁ -C ₁₀ alkoxy.

In one embodiment of the invention, R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 6; Preferably, R ₃ to R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₆ alkyl, C ₁ -C ₆ alkenyl or C ₁ -C ₆ alkoxy, and R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 3; More preferably, R ₃ to R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₃ alkyl, C ₁ -C ₃ alkenyl or C ₁ -C ₃ alkoxy. Where R ₁ And R ₂ is hydroxy, or a compound which together form a ring with —O—CH ₂ —O— is most preferred, but is not limited thereto.

In addition, the present invention includes an isomer including a compound of formula (2) or a pharmaceutically acceptable salt thereof, a hydrate thereof, or a tautomer thereof as an active ingredient, and together with at least one pharmaceutically acceptable carrier, diluent or excipient It provides a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus comprising.

In addition, the present invention provides a nucleic acid substrate unwinding activity inhibitor of coronavirus helicase comprising a compound of formula (2).

In Chemical Formula 2,

R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 10; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₁₀ alkyl, C ₁ -C ₁₀ alkenyl or C ₁ -C ₁₀ alkoxy.

In one embodiment of the invention, R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 6; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; Preferably, R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₆ alkyl, C ₁ -C ₆ alkenyl or C ₁ -C ₆ alkoxy, and R ₁ And R ₂ is hydroxy or together forms a ring with —O— (CH ₂ ) nO—, where n is an integer from 1 to 3; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; More preferably, R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₃ alkyl, C ₁ -C ₃ alkenyl or C ₁ -C ₃ alkoxy. Where R ₁ And R ₂ is hydroxy or together forms a ring with —O—CH ₂ —O—; Most preferably, R ₃ is a compound which is a benzyl group unsubstituted or substituted with one or more fluorine or cyano.

"Alkyl" in the present invention includes optionally substituted straight or branched chain hydrocarbons with at least one hydrogen to form radical groups.

"C ₁ -C ₃ alkyl group" is a carbon atom represents a 1-3 alkyl group, "C ₁ -C ₆ alkyl group" is a carbon atom represents a 1-6 alkyl group, "C ₁ -C ₁₀ alkyl group" is carbon The alkyl group of 1-10 atoms is shown.

Alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, l-methylpropyl, pentyl, isopentyl, sec-pentyl, hexyl, heptyl, octyl and the like. Alkyl includes cycloalkyl, such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

"Alkenyl" includes any of the above optionally substituted straight or branched chain hydrocarbon radicals having at least one carbon-carbon double bond (sp ₂ ). Alkenyl is ethenyl (or vinyl), prop-1-enyl, prop-2-enyl (or allyl), isopropenyl (or 1-methylvinyl), but-1-enyl, but-2-enyl , Butadienyl, pentenyl, hexa-2,4-dienyl and the like. Cis and trans or (E) and (Z) forms are included in the present invention.

"Alkoxy" means the group -O-R 'and R' includes an alkyl group of the above definition, ie an optionally substituted straight or branched chain alkyl group with terminal oxygen which bonds the alkyl group to the rest of the molecule. Alkoxy includes methoxy, ethoxy, propoxy, isopropoxy, butoxy, t-butoxy, pentoxy and the like.

"Halogen" includes fluorine, chloro, bromo or iodo, preferably chloro or bromo, more preferably chloro, as substituents.

"Hydroxy group" means the group -OH.

In the present invention, a radical refers to a species having one unpaired electron so that the species containing the radical can be covalently bonded to another species. Thus, in this context, radicals are not necessarily free radicals. Rather, radicals represent certain portions of larger molecules. The term "radical" may be used interchangeably with the term "group".

As the pharmaceutically acceptable salt of the compounds of Formula 1 and Formula 2 of the present invention, acid addition salts formed by free acid are useful. Acid addition salts are prepared by conventional methods, for example by dissolving a compound in an excess of aqueous acid solution and precipitating the salt using a water miscible organic solvent such as methanol, ethanol, acetone or acetonitrile. Equivalent molar amounts of the compound and acid or alcohol in water (eg glycol motomethylether) can be heated and the mixture can then be evaporated to dryness or the precipitated salts can be suction filtered.

In this case, organic acids and inorganic acids may be used as the free acid, and hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, and the like may be used as the inorganic acid, and methane may be used as the organic acid. Methane sulfonic acid, p-toluen sulfonic acid, acetic acid, trifluoroacetic acid, citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, propionic acid, citric acid, lactic acid, glycolic acid ), Gluconic acid, galacturon acid, glutamic acid, glutaric acid, glucuronic acid, glucuronic acid, aspartic acid, ascorbic acid (ascorbic acid), carbonic acid, vanillic acid, hydro And the like Io diksan (hydroiodic acid) may be used.

In addition, bases can be used to make pharmaceutically acceptable metal salts. An alkali metal or alkaline earth metal salt is obtained by, for example, dissolving the compound in an excess alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and then evaporating and drying the filtrate. In this case, as the metal salt, it is particularly suitable to prepare sodium, potassium or calcium salt, and the corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (for example, silver nitrate).

Pharmaceutically acceptable salts of the invention include acidic or basic salts which may be present in the compounds of the invention, unless otherwise noted. For example, pharmaceutically acceptable salts include sodium, calcium, and potassium salts of the hydroxy group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, and hydrogen sulfate. sulfate, phosphate, hydrogen sulfate, dihydrogen sulfate, acetate, succinate, citrate, tartrate, lactate ), Methanesulfonate (mesylate, methane sulfonate) and p -toluenesulfonate (tosylate, p- toluen sulfonate) salts, and can be prepared through the preparation or manufacturing process of salts known in the art.

Isomers including the dihydroxychromone derivative compounds of Formula 1 and / or Formula 2, pharmaceutically acceptable salts thereof, hydrates thereof, or tautomers thereof of the present invention can be appropriately adjusted according to the purpose of use, and special pharmaceuticals. Is not.

In addition, the pharmaceutical compositions of the present invention may further comprise one or more pharmaceutically acceptable liquid or solid carriers. It may also be prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, lubricants, surfactants, etc. that are commonly used, but is not limited thereto.

The solid form preparations may be tablets, pills, powders, granules, capsules, pellets, granules or powders, and such solid preparations may be prepared by adding a carrier, excipients and / or diluents to the compound. The carrier, excipient and / or diluent may include lactose, sucrose, dextrose, mannitol, malitol, malitol, sorbitol, xylitol, and erythritol. (erithritol), starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, polyvinyl pyrrolidone polyvinyl pyrrolidone, magnesium stearate, and mineral oil.

The formulations in liquid form may be solutions, suspensions or emulsions, and may include various excipients, for example wetting agents, sweeteners, fragrances, preservatives, etc., in addition to the commonly used simple diluents, water and liquid paraffin. Aqueous suspensions suitable for oral use may be prepared by dispersing the finely divided active component in a viscous material such as natural or synthetic gums, resins, methyl cellulose, sodium carboxymethyl cellulose and known suspending agents. Can be.

Examples of fillers that can be used in the pharmaceutical compositions and dosage forms disclosed herein include talc, calcium carbonate (eg, granules or powders), microcrystalline cellulose, powder cellulose, dexrate, kaolin, mannitol, silicic acid, sorbitol, Starch, pre-gelatin ring starch, and mixtures thereof, including but not limited to. The binder or filler of the pharmaceutical composition of the present invention is generally present in about 50 to about 99 weight percent of the pharmaceutical composition or dosage form.

Disintegrants that can be used in the pharmaceutical compositions and dosage forms disclosed herein are intended to disintegrate when the tablet is exposed to an aqueous environment. Tablets containing too much disintegrant may disintegrate during storage, while tablets containing too little do not disintegrate at the desired rate under the desired conditions. A sufficient amount of disintegrant, therefore not too much or too little, should not be used to form the solid oral dosage form of the invention so as not to be bad for controlling the release of the active ingredient. The amount of disintegrants used varies depending on the type of formulation, and can be readily determined by one of ordinary skill in the art. Typical pharmaceutical compositions contain about 0.5 to about 15 weight percent of disintegrant, preferably about 1 to about 5 weight percent of disintegrant. The disintegrants that can be used in the pharmaceutical compositions and dosage forms of the present invention are agar- Agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, poracryline potassium, sodium starch glycolenmit, potato or tapioca starch, other starch, pre-gelatinized starch, other starch, clay, Other algins, other celluloses, gums, and mixtures thereof.

Glidants that can be used in the pharmaceutical compositions and dosage forms of the present invention are calcium stearate, magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium Lauryl sulfate, talc, hydrogenated vegetable oils (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar , And mixtures thereof. Additional lubricants are, for example, siloid silica (AEROSIL200, manufactured by WR Grace Co., Baltimore, MD), coagulated aerosol of synthetic silica, manufactured by Degussa Co., Plano, TX, CAB- O-SIL (pyrogenic silicon dioxide product, Cabot Co., Boston, Mass.), And mixtures thereof. If used, glidants are generally used in amounts up to about 1 weight percent of the pharmaceutical composition or dosage form in which they are contained.

The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, etc. by various routes, and all modes of administration can be expected. For example, it may be administered orally or parenterally, and if administered parenterally, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, intrauterine dural or intracerebroventricular injection, or the like. Can be. It is preferable that the route of administration of the pharmaceutical composition of the present invention is determined according to the type of the disease to be applied.

Examples of the parenteral administration include injections, drops, sap, ointments, sprays, suspensions, emulsions, suppositories, and the like. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate may be used. As a suppository base, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin, and the like may be used.

As used herein, the terms "treatment" and / or "inhibition" means obtaining the desired pharmacological and / or physiological effect. Such an effect may be a prophylactic effect in terms of completely or partially suppressing a disease or symptoms thereof and / or may be a therapeutic effect in terms of partially or fully curing the adverse effects caused by the disease and / or disease. “Treatment” and / or “inhibition” as used herein encompasses any treatment for diseases of mammals, particularly humans, and (a) in subjects susceptible to disease but not yet diagnosed as diseased. Preventing the development of the disease; (b) inhibiting the disease, ie arresting the onset of the disease; And (c) alleviating the disease, ie causing regression of the disease.

The pharmaceutical composition of the present invention is administered in a therapeutically effective amount upon administration for clinical purposes. The term “therapeutically effective amout” means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective amount for a particular patient is age, sex, weight, health condition. , Time of administration, route of administration, rate of excretion, drug mixture and severity of disease. However, for the desired effect, the compound of the present invention may be administered in an amount of 0.0001 to 1000 mg / kg, preferably 0.001 to 100 mg / kg, divided once or several times a day.

The amount of active ingredient of a compound of Formula 1 and / or Formula 2 of the present invention that can be combined with a substance such as a carrier to produce a dosage form can vary depending upon the host to be treated and the particular mode of administration. The composition may contain from 0.001 to 95% of the active ingredient, and a typical pharmaceutical formulation may contain from about 5% to about 95% of the active ingredient (w / w). In another embodiment, the pharmaceutical formulation may contain about 20% to about 80% active ingredient.

Those skilled in the art will readily understand that dosage levels may vary as a function of specific coronavirus helicase inhibitory compounds, the severity of symptoms and the subject's susceptibility to side effects. Preferred dosages of pharmaceutical compositions comprising a given compound of Formula 1 and / or Formula 2 are readily determined by one skilled in the art by a variety of means.

In many embodiments, multiple doses of the pharmaceutical composition of the present invention are administered. For example, about 1 week to about 1 week, about 2 weeks to about 4 weeks, about 1 month to about 2 months, about 2 months to about 4 months, about 4 months to about 6 months, about 6 months after virus exposure. Once a month, twice a month, three times a month, over a period of about 8 months, about 8 months to about 1 year, about 1 year to about 2 years, about 2 years to about 4 years, or more It is administered every other week, once a week, twice a week, three times a week, four times a week, five times a week, six times a week, every other day, every day, twice a day or three times a day .

In the compositions of the present invention the compounds of formula 1 and / or formula 2 should be present in an amount of 0.001 to 50% by weight, preferably 0.01 to 50% by weight, based on the total weight of the total composition.

In many embodiments, the compounds of the present invention inhibit coronavirus replication. For example, the compounds of the present invention may exhibit coronavirus replication at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, as compared to coronavirus replication in the absence of compounds of the present invention, At least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, or at least about 90% or more. Whether the compounds of the present invention inhibit coronavirus replication can be determined using methods known in the art, such as in vitro virus replication assays.

As used herein, the term "coronavirus" includes any member of the family of coronibiridae, including but not limited to any member of the genus Coronavirus and / or any member of the genus Torovirus.

Coronaviridae is a family of viruses characterized by irregular shaped particles of 60-220 nm diameter with a distinctive club-shaped perfluorometer with an envelope on the outside, which gives the virus a crown-like appearance. Derived. Coronovirida includes the genus Coronavirus and Toro virus.

Coronaviruses include infectious bronchitis virus (IBV), bovine coronavirus (BCoV), canine coronavirus (CCoV), feline coronavirus (FeCoV), porcine erythrocyte aggregated encephalomyelitis virus (PEDV), mouse hepatitis virus (MHV), and turkey coronavirus (TCoV), infectious gastrointestinal virus (TGEV), rat coronavirus (RCV), human coronavirus 229E (HCoV-229E), human coronavirus NL63 (NL), human coronavirus OC43 (HCoV-OC43), SARS coronavirus ( SARS-CoV) and the like, but is not limited thereto.

Toro viruses include, but are not limited to, Berne virus and Breda virus.

The present invention provides a pharmaceutical composition for the treatment and prevention of diseases caused by any one of the coronaviruses, and preferably provides a pharmaceutical composition for the treatment and prevention of SARS caused by SARS coronavirus.

Further, the term "coronavirus" includes naturally occurring (eg wild type) coronaviruses; Naturally occurring coronavirus variants; And variants generated by selection; Variants generated by chemical modifications; And genetically modified variants (eg, coronaviruses modified by recombinant DNA methods in the laboratory) and include laboratory generated coronavirus variants.

The present invention provides a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that to inhibit the nucleic acid substrate unwinding activity of the coronavirus helicase comprising a compound of Formula 1 and / or Formula 2 as an active ingredient Preferably, the present invention provides a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized by inhibiting the nucleic acid substrate unwinding activity of SARS coronavirus helicase. In addition, the nucleic acid substrate is characterized in that the DNA or RNA.

In another aspect, the present invention provides a nucleic acid substrate unwinding activity inhibitor of coronavirus helicase comprising a compound of formula (1) and / or formula (2) as an active ingredient, preferably provides an inhibitor of nucleic acid substrate unwinding activity of helicase of SARS coronavirus do. In addition, the nucleic acid substrate is characterized in that the DNA or RNA.

Coronaviruses are commonly composed of a protein consisting of spike (S), envelope (E, envelope), membrane (M) and nucleocapsid (N, nucleocapsid). In case of SARS coronavirus, There is a receptor binding domain in S that mediates attachment of the virus to an intracellular receptor, angiotensin-converting enzyme 2 (ACE2).

Coronaviruses are positive (+)-single stranded RNA viruses.

Coronavirus helicases serve to unwind the duplex RNA into a single stranded form or to solve secondary structures present in the single stranded RNA molecule. The coronavirus genomic RNA molecule, including SARS coronavirus, has a broad secondary structure and can be used as a template strand for coronavirus replication and viral protein synthesis, with the secondary structure unwrapped in a single strand, so the biological activity of helicase is corona. It is a very important part of the viral life cycle.

SARS coronavirus helicase consists of three parts: a cysteine-rich N-terminal metal binding domain (MBD) consisting of 100 amino acid residues, a hinge domain, and an NTPase / helicase domain.

Compounds of the invention appear to act in the NTPase / helicase domain. Binding of the compound to coronavirus helicase in this domain may result in the movement of one or more of the residues in the domain. May cause helicase migration. Migration due to the binding of the compound may cause allosteric migration of coronavirus helicase such that binding of the nucleic acid substrate at the distant site of the helicase is inhibited. In a preferred embodiment, the nucleic acid substrate is DNA or RNA. By inhibiting the binding of the nucleic acid substrate, the activity of coronavirus helicase can be modulated. In a preferred embodiment, the modulation of coronavirus helicase activity is the inhibition of activity. In a preferred embodiment, the regulated coronavirus helicase activity is replication of coronavirus, more preferably replication of SARS coronavirus. In one embodiment, the binding of the compound with the helicase is effective at inhibiting annealing of the nucleic acid substrate (eg, DNA and / or RNA) by the coronavirus helicase, preferably the nucleic acid by the SARS coronavirus helicase It is effective in suppressing loosening of the substrate. Modulation of coronavirus helicase activity can occur in vivo or ex vivo.

In some embodiments of the invention, reducing viral load in a subject exposed to coronavirus (eg, in contact with a subject infected with coronavirus); And / or increased sustained rate of viral response to therapy, reduction in morbidity or mortality in clinical outcomes, and the like.

 An effective amount of a compound of the invention, and optionally one or more additional antiviral agents, is an amount effective to reduce viral load or achieve a sustained viral response to therapy.

The pharmaceutical composition of the present invention may further comprise one or more antiviral agents.

In an embodiment, the antiviral agent includes but is not limited to nucleotides and nucleotide analogues. Non-limiting examples are azidothymidine (AZT), and analogs and derivatives thereof, 2 ', 3'-dideoxyinosine (DDI). And analogs and derivatives thereof, 2 ', 3'-dideoxycytidine (DDC), and analogs and derivatives thereof, 2', 3'-didehydro-2 ', 3'-dideoxythymidine (D4T) (star Budin), and analogs and derivatives thereof, combivir; Abacavir; Adefovir dipoxyl, sipofovir, ribavirin, ribavirin analogs, viral serine protease inhibitors, viral polymerase inhibitors or interferon-a. Preferably, it may be selected from a viral serine protease inhibitor, a viral polymerase inhibitor, interferon-a or ribavirin.

The pharmaceutical composition of the present invention may further include one or more additives selected from immunomodulators, antioxidants, antibacterial agents or antisense agents.

In some embodiments, the additive may be selected from sillybum marianum, interleukine12, amantadine, ribozyme, thymosin, N-acetyl cysteine or cyclosporin. have.

In some embodiments of the present invention, an effective amount of a compound of Formula 1 and / or Formula 2, and optionally one or more additional antiviral agents, comprises at least about 1000 to about genomic copy number per mL of serum detectable viral titer Effective amount to reduce to 5000, about 500 to about 1000, or about 100 to about 500. In some embodiments, an effective amount of a compound of Formula 1 and / or Formula 2, and optionally one or more additional antiviral agents, is 1.5-log, 2-log, 2.5-log, 3-log, 4- of the virus titer in a subject's serum. It is an amount effective to achieve log, 4, 5 log, or 5-log reduction.

In many embodiments, an effective amount of a compound of Formula 1 and / or Formula 2, and optionally one or more additional antiviral agents, is a synergistic amount. As used herein, a compound of Formula 1 and / or Formula 2; And the "synergistic combination" or "synergistic amount" of additional antiviral agent are (i) Formula 1 and / or when administered at the same dose as monotherapy in the treatment or prophylaxis of a coronavirus infection. Rather than an improved improvement in the therapeutic outcome that can be foreseen or anticipated from a simple additive combination of the therapeutic or prophylactic benefit of the compound of formula 2 and (ii) the therapeutic or prophylactic benefit of an additional antiviral agent when administered at the same dose as a monotherapy. More effective combination dosage.

In some embodiments, selected amounts of compounds of Formula 1 and / or Formula 2; And selected amounts of the additional antiviral agent are effective when used in combination therapy for the disease, but selected amounts of the compounds of Formula 1 and / or Formula 2; And / or selected amounts of additional antiviral agents are not effective when used as monotherapy for a disease. Accordingly, embodiments provide for (1) augmenting the therapeutic benefit of selected amounts of the compounds of Formula 1 and / or Formula 2 when the selected amount of additional antiviral agent is used in combination therapy for the disease, Therapies do not provide a therapeutic benefit when the antiviral agent is used as monotherapy for the disease, (2) A selected amount of the selected amount when the compound of formula 1 and / or formula 2 is used in combination therapy for the disease Therapies that enhance the therapeutic benefit of additional antiviral agents and do not provide therapeutic benefit when the selected amount of the compound of Formula 1 and / or Formula 2 are used as monotherapy for the disease, and (3) the selected amount of formula 1 and / or the compound of formula 2 and selected amounts of additional antiviral agents provide therapeutic benefit when used in combination therapy for the disease. And, the antiviral agent each of the selected amounts of the formula 1 and / or compound of formula (II) and further includes a therapy that does not provide a therapeutic benefit when used in danje therapy against disease. As used herein, the "synergistically effective amount" of a compound of Formula 1, Formula 2 and additional antiviral agents and equivalents thereof includes any of the therapies encompassed by any of (1) to (3) above. Will be understood.

Dihydroxychromone derivatives according to the present invention have a function of inhibiting the nucleic acid substrate annealing activity of SARS coronavirus helicase.

Therefore, since the compounds of the present invention have an effect of inhibiting the activity of helicase of SARS coronavirus, the pharmaceutical composition for the treatment and / or prevention of diseases caused by coronavirus including SARS coronavirus, coronavirus helicase activity It can be effectively used for development as an inhibitor.

Hereinafter, the present invention will be described in detail.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention, it will be apparent to those skilled in the art that the scope of the present invention is not to be construed as being limited by these examples.

Dihydroxychromone derivatives are synthesized from commercially available acetofinone or quercetin, and compounds 5, 10, and 12a are known methods [ Synthesis , 18 : 2768-2770, 2003; Bull . Soc . Chim . Fr. 11 : 124-131, 1979; J. Org. Chem ., 68 : 6842-6845, 2003].

Preparation Example 1 Preparation of 1- (3- Benzyloxy -6- hydroxy -2- methoxy - phenyl ) -ethanone (Compound 6) :

Compound 5 (1.2 g, 6.70 mmol) was dissolved in anhydrous acetone, and potassium carbonate (925 mg, 6.70 mmol) and benzylbromide (0.8 mL, 6.70 mmol) were added, and the mixture was stirred at 60 ^° C. for 2 hours. After the reaction mixture was filtered through a filter paper, the filtrate was concentrated under reduced pressure and purified through silica gel column chromatography (hexane: ethyl acetate = 10: 1) to give compound 6 (1.45 g, 5.33 mmol, 80% yield) as a yellow oil. Got:

¹ H NMR (400 MHz, CDCl ₃ ) d 7.26-7.44 (m. 5H), 7.14 (d, J = 9.0 Hz, 1H), 6.64 (d, J = 9.1 Hz, 1H), 5.04 (s, 2H) , 3.98 (s, 3 H), 2.73 (s, 3 H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 205.03, 157.61, 151.82, 143.92, 136.62, 128.62, 127.89, 127.57, 124.62, 115.26, 112.44, 75.55, 61.00, 32.70.

Manufacturing example  2. Acetic acid  2- acetyl -4- benzyloxy -3- methoxy - phenyl ester  Preparation of (Compound 7):

Compound 6 (1.45 g, 5.33 mmol) was dissolved in anhydrous pyridine (20 mL) and AcCl (1.53 mL, 21.30 mmol) was added and stirred at 60 ^° C. for 12 hours. The reaction was concentrated under reduced pressure and purified via silica gel column chromatography (hexane: ethyl acetate = 10: 1) to give compound 7 (1.49 g, 4.74 mmol, 89% yield) in the form of a yellow oil:

¹ H NMR (400 MHz, CDCl ₃ ) d 7.33-7.45 (m, 5H), 6.98 (d, J = 8.9 Hz, 1H), 6.78 (d, J = 8.9 Hz, 1H), 5.11 (s, 2H) , 3.91 (s, 3 H), 2.52 (s, 3 H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 200.35, 169.71, 149.72, 147.10, 140.30, 136.49, 129.71, 128.76, 128.27, 127.43, 117.99, 115.57, 71.40, 61.83, 31.89, 20.90.

Manufacturing example  3. 6- Benzyloxy -5- methoxy -2- methyl - chromen -4- one  Preparation of (Compound 8):

Compound 7 (1.49 g, 4.74 mmol) was dissolved in tetrahydrofuran and 1M LiHMDS / hexane (19 mL, 18.96 mmol) was added. The mixture was stirred at -78 ^o C for 3 hours and then at room temperature for 3 hours. NH ₄ Cl saturated aqueous solution was added to the reaction to neutralize and washed with water. The organic layer was concentrated under reduced pressure and purified by silica gel column chromatography (hexane: ethyl acetate = 5: 1) to give compound 8 (680 mg, 2.30 mmol, 48% yield) in the form of a yellow oil:

¹ H NMR (400 MHz, CDCl ₃ ) d 7.33-7.46 (m, 5H), 7.16 (d, J = 9.1 Hz, 1H), 7.03 (d, J = 9.1 Hz, 1H), 6.19 (s, 1H) , 5.13 (s, 2H), 3.95 (s, 3H), 2.62 (s, 3H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 160.96, 153.45, 148.91, 148.77, 147.99, 136.95, 129.08, 128.60, 127.86, 118.75, 116.60, 115.33, 112.47, 72.23, 61.70, 23.58.

Preparation Example 4 Preparation of 6- Benzyloxy -5- methoxy -4- oxo- 4H- chromene -2- carbaldehyde (Compound 9) :

Compound 8 (680 mg, 2.30 mmol) was dissolved in bromobenzene (15 mL), selenium dioxide (1.02 g, 9.18 mmol) was added, and the mixture was stirred at 160 ^° C. for 12 hours. The reaction was concentrated under reduced pressure and purified by silica gel column chromatography (hexane: ethyl acetate = 3: 1) to give compound 9 (650 mg, 2.10 mmol, 91% yield) as a yellow powder:

¹ H NMR (400 MHz, CDCl ₃ ) d 7.36-7.45 (m, 5H), 7.31 (d, J = 11.6 Hz, 1H), 7.08 (d, J = 9.1 Hz, 1H), 6.47 (s, 1H) , 5.16 (s, 2 H), 3.94 (s, 3 H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 190.97, 149.63, 148.81, 148.00, 147.69, 136.42, 129.19, 128.84, 127.89, 115.63, 112.76, 72.38, 61.77.

Manufacturing example  5. 2-[(4- Chloro - benzylamino ) - methyl ] -5,6- dihydroxy - chromen -4- one  (compound 2a), 2-[(3- Chloro - benzylamino ) - methyl ] -5,6- dihydroxy - chromen -4- one  (Compound 2b) and  3-{[(5,6- Dihydroxy -4- oxo -4H- chromen -2- ylmethyl ) - amino ] -methyl} -benzonitrile (Compound 2c):

Dissolve the appropriate benzylamine (1.28 mmol) in methanol and adjust to pH 6 by adding glacial acetic acid. Compound 9 (100 mg, 0.32 mmol) and sodium cyanoborohydride (20 mg, 0.32 mmol) were added thereto, followed by stirring at room temperature for 2 hours. The reaction was concentrated under reduced pressure, neutralized with saturated aqueous NaHCO _3, and extracted with ethyl acetate. The organic layer was concentrated under reduced pressure, dissolved in anhydrous dichloromethane, 1M BBr ₃ was added, and the mixture was stirred at room temperature for 12 hours. Methanol was added to the reaction, concentrated under reduced pressure, and recrystallized by addition of dichloromethane to give compound 2a-c (31-38% yield) as a yellow powder:

2a ¹ H NMR (400 MHz, CD ₃ OD) d 7.58 (d, J = 8.3 Hz, 2H), 7.49 (d, J = 8.3 Hz, 2H), 7.11 (d, J = 8.8 Hz, 1H), 6.78 (d, J = 8.8 Hz, 1 H), 6.41 (s, 1 H), 4.53 (s, 2 H), 4.42 (s, 2 H); ¹³ C NMR (MeOD, 100 MHz) d 162.3, 149.3, 147.0, 144.1, 143.3, 137.4, 133.5, 131.1, 130.9, 120.4, 119.0, 108.9, 108.8, 52.8, 51.9;

2b ¹ H NMR (400 MHz, DMSO- d ₆ ) d 7.71 (s, 1H), 7.47-7.66 (m, 3H), 7.14 (d, J = 8.8 Hz, 1H), 6.77 (d, J = 8.8 Hz , 1H), 6.43 (s, 1H), 4.52 (s, 2H), 4.34 (s, 2H); ¹³ C NMR (100 MHz _, DMSO- d ₆ ) d 160.1,151.80, 147.47, 146.40, 143.75, 139.50, 133.48, 128.00, 120.79, 117.10, 113.50, 108.89, 104.60, 51.50, 50.08;

2c ¹ H NMR (400 MHz, DMSO- d ₆ ) d 8.07 (s, 1H), 7.93 (d, J = 8.1 Hz, 1H), 7.91 (d, J = 8.6 Hz, 1H), 7.69 (t, J = 7.8 Hz, 1H), 7.13 (d, J = 8.8 Hz, 1H), 6.78 (d, J = 8.8 Hz, 1H), 6.43 (s, 1H), 4.51 (s, 2H), 4.39 (s, 2H ); ¹³ C NMR (CDCl ₃ , 100 MHz) d 159.3, 146.9, 146.1, 142.8, 141.5, 130.7, 129.0, 119.5, 118.5, 117.8, 116.5, 114.8, 111.5, 107.3, 105.7, 50.1, 49.6.

Manufacturing example  6. 2- Benzo [1,3] dioxol-5- yl -3,5,7- trihydroxy - chromen -4- one  Preparation of (Compound 3b):

Compound 10 (373 mg, 0.82 mmol) was dissolved in anhydrous pyridine, and then piperonyloyl chloride (303 mg, 1.64 mmol) was added thereto and stirred at room temperature for 12 hours. The reaction was neutralized by addition of 1 N hydrochloric acid and extracted with ethyl acetate. The organic layer was concentrated under reduced pressure, dissolved in toluene, and calcium carbonate (456 mg, 3.28 mmol) tetrabutylamoniumbromide (395 mg, 1.23 mmol) was added thereto, and the mixture was stirred at 90 ^° C. for 12 hours. The reaction was concentrated under reduced pressure, dissolved in dichloromethane, and washed with water and brine. The organic layer was concentrated under reduced pressure, dissolved in methanol and anhydrous dichloromethane, and palladium was added thereto. Hydrogen gas was added and stirred for 3 hours at room temperature, and the reaction was filtered through diatomaceous earth. The filtrate was concentrated under reduced pressure and purified by silica gel column chromatography (hexane: ethyl acetate = 3: 1) to give compound 3b (166 mg, 0.53 mmol, 65% yield) as a yellow powder:

¹ H NMR (400 MHz, DMSO- d6 ) d 7.77 (d, J = 8.4 Hz, 1H), 7.70 (s, 1H), 6.97 (d, J = 8.4 Hz, 1H), 6.49 (s, 1H), 6.22 (s, 1 H), 6.14 (s, 2 H); ¹³ C NMR (100 MHz, DMSO- d6 ) d 176.7, 164.8, 161.4, 156.9, 148.2, 125.7, 125.4, 123.4, 109.5, 109.2, 108.2, 103.8, 103.0, 102.4, 98.3, 94.3.

Manufacturing example  7. Acetic acid  5- acetoxy -2- benzo [1,3] dioxol-5- yl -7- hydroxy -4-oxo-4H-chromen-3-yl ester  Preparation of (Compound 12b):

Compound 3b (828 mg, 2.64 mmol) was dissolved in anhydrous pyridine, acetic anhydride (2.5 mL, 26.4 mmol) was added, and the mixture was stirred at room temperature for 12 hours. 1 N hydrochloric acid was added to the reaction to neutralize and extracted with dichloromethane. The organic layer was concentrated under reduced pressure and dissolved in 1-methyl-2-pyrrolidone. Imidazole (42 mg, 0.62 mmol) and thiophenol (0.2 mL, 2.06 mmol) were added at 0 ^° C. and stirred at room temperature for 5 hours. 2 N hydrochloric acid was added to the reaction, followed by extraction with ethyl acetate. The organic layer was concentrated under reduced pressure and purified by silica gel column chromatography (hexane: ethyl acetate = 1: 1) to obtain compound 12b (757 mg, 1.90 mmol, 72% yield) as a yellow powder:

¹ H NMR (400 MHz, CDCl ₃ ) d 7.38 (d, J = 8.3 Hz, 1H), 7.28 (s, 1H), 6.89 (d, J = 8.3 Hz, 1H), 6.73 (d, J = 1.9 Hz , 1H), 6.55 (d, J = 2.0 Hz, 1H), 6.05 (s, 2H), 2.40 (s, 3H), 2.32 (s, 3H) ¹³ C NMR (100 MHz, DMSO- d6 ) d 179.7, 178.8 , 177.5, 172.0, 167.8, 164.0, 160.6, 159.9, 157.9, 142.6, 133.1, 133.0, 119.8, 118.9, 118.3, 117.8, 111.7, 110.9, 30.6, 30.1.

Manufacturing example  8. Acetic acid  3- acetoxy -2- benzo [1,3] dioxol-5- yl -7- (4- chloro -benzyloxy) -4-oxo-4H-chromen-5-yl ester  (Compound 13d), Acetic acid  3- acetoxy -2-benzo [1,3] dioxol-5-yl-7- (3-chloro-benzyloxy) -4-oxo-4H-chromen-5-yl ester  (Compound 13e) and Acetic acid  5- acetoxy -2- benzo [1,3] dioxol-5- yl -7- (3- cyano -benzyloxy) -4-oxo-4H-chromen-3-yl ester  Preparation of (Compound 13f):

Compound 12b (120 mg, 0.30 mmol) was dissolved in anhydrous acetone, and potassium carbonate (166 mg, 1.20 mmol) and the appropriate benzylbromide (0.75 mmol) were added thereto, and the mixture was stirred at room temperature for 3 hours. The reaction was filtered through filter paper, the filtrate was concentrated under reduced pressure and purified by silica gel column chromatography (hexane: ethyl acetate = 2: 1) to give compound 13d-f (56-71% yield) as a yellow powder:

13d ¹ H NMR (400 MHz, CDCl ₃ ) d 7.35-7.41 (m, 5H), 7.28 (d, J = 1.6 Hz, 1H), 6.92 (d, J = 8.3 Hz, 1H), 6.87 (d, J = 2.3 Hz, 1H), 6.69 (d, J = 2.3 Hz, 1H), 6.07 (s, 2H), 5.11 (s, 2H), 2.43 (s, 3H), 2.32 (s, 3H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 169.6, 169.1, 167.5, 162.0, 157.5, 154.1, 150.3, 149.7, 147.6, 134.0, 133.3, 132.7, 128.6, 128.4, 122.9, 110.9, 108.6, 108.1, 107.7, 101.4, 99.3, 69.5, 20.7, 20.2;

13e ¹ H NMR (400 MHz, CDCl ₃ ) d 7.44 (s, 1H), 7.35-7.40 (m, 3H), 7.28-7.31 (m, 2H), 6.92 (d, J = 8.3 Hz, 1H), 6.88 (d, J = 2.3 Hz, 1H), 6.70 (d, J = 2.3 Hz, 1H), 6.06 (s, 2H), 5.12 (s, 2H), 2.43 (s, 3H), 2.33 (s, 3H) ; ¹³ C NMR (100 MHz, CDCl ₃ ) d 169.6, 169.1, 167.5, 161.9, 157.5, 154.1, 150.3, 149.7, 147.6, 136.9, 134.3, 132.7, 129.7, 128.2, 127.0, 124.9, 122.9, 122.8, 116.9, 108.6, 108.1, 107.7, 101.4, 99.3, 69.4, 20.6, 20.2;

13f ¹ H NMR (400 MHz, CDCl ₃ ) d 7.75 (s, 1H), 7.68 (d, J = 7.7 Hz, 2H), 7.55 (t, J = 7.9 Hz, 1H), 7.39 (dd, J = 8.3 Hz, 1.6 Hz, 1H), 7.28 (d, J = 1.6 Hz, 1H), 6.92 (d, J = 8.2 Hz, 1H), 6.89 (d, J = 2.2 Hz, 1H), 6.71 (d, J = 2.3 Hz, 1H), 6.07 (s, 2H), 5.18 (s, 2H), 2.44 (s, 3H), 2.33 (s, 3H); ¹³ C NMR (100 MHz, CDCl ₃ ) d 170.2, 169.7, 168.2, 162.2, 158.1, 154.8, 151.1, 150.4, 148.3, 137.1, 133.4, 132.3, 131.7, 130.9, 129.9, 123.5, 123.4, 118.6, 113.3, 111.7, 109.1, 108.8, 108.3, 102.1, 100.0, 69.5, 21.3, 20.9.

Manufacturing example  9. 7- (4- Chloro - benzyloxy ) -2- (3,4- dihydroxy - phenyl ) -3,5-dihydroxy-chromen-4-one (Compound 4a), 7- (3- Chloro - benzyloxy ) -2- (3,4- dihydroxy - phenyl ) -3,5-dihydroxy-chromen-4-one (Compound 4b) and  3- [2- (3,4- Dihydroxy - phenyl ) -3,5-dihydroxy-4-oxo-4H-chromen-7-yloxymethyl] -benzonitrile (Compound 4c):

Compound 12a (380 mg, 0.81 mmol) was dissolved in anhydrous acetone, and potassium carbonate (447 mg, 3.24 mmol) and appropriate benzylbromide (2.03 mmol) were added thereto, and the mixture was stirred at room temperature for 3 hours. The reaction was filtered through a filter paper, and the filtrate was concentrated under reduced pressure, 2M ammonia methanol solution was added, and stirred at 0 ^° C. for 3 hours. The reaction was concentrated under reduced pressure and recrystallized by adding dichloromethane to give compound 4a-c (25-29% yield):

4a ¹ H NMR (400 MHz, DMSO- d6 ) d 7.72 (d, J = 1.9 Hz, 1H), 7.56 (dd, J = 8.4, 1.9 Hz, 1H), 7.50 (m, 4H), 6.90 (d, J = 8.5 Hz, 1H), 6.80 (d, J = 1.8 Hz, 1H), 6.44 (d, J = 1.9 Hz, 1H), 5.25 (s, 2H); ¹³ C NMR (100 MHz, Acetone- d6 ) d 177.1, 165.7, 162.1, 157.9, 148.9, 148.0, 146.3, 136.7, 134.7, 130.6, 129.8, 123.9, 121.8, 116.4, 116.1, 105.5, 100.6, 99.2, 94.0, 70.0;

4b ¹ H NMR (400 MHz, DMSO- d6 ) d 7.72 (d, J = 1.9 Hz, 1H), 7.57 (d, J = 1.9 Hz, 1H), 7.55 (s, 1H), 7.45 (m, 3H) , 6.90 (d, J = 8.5 Hz, 1H), 6.81 (d, J = 2.0 Hz, 1H), 6.50 (d, 2.0 Hz, 1H), 5.26 (s, 2H); ¹³ C NMR (100 MHz, Acetone- d6 ) d 178.7, 165.1, 161.6, 157.4, 148.2, 147.3, 145.6, 139.7, 134.6, 130.9, 128.7, 128.1, 126.6, 123.4, 121.2, 115.9, 115.6, 104.9, 98.7, 98.6, 70.0;

4c ¹ H NMR (400 MHz, DMSO- d6 ) d 7.96 (s, 1H), 7.84 (t, J = 8.0 Hz, 2H), 7.75 (d, J = 2.0 Hz, 1H), 7.65 (t, J = 7.9 Hz, 1H), 7.56 (dd, J = 8.4, 2.0 Hz, 1H), 6.90 (d, J = 8.4 Hz, 1H), 6.82 (d, J = 2.0 Hz, 1H), 6.47 (d, J = 2.1 Hz, 1H), 5.31 (s, 2H); ¹³ C NMR (100 MHz, Acetone- d6 ) d 176.6, 165.2, 157.6, 148.3, 147.4, 145.7, 139.3, 137.0, 132.9, 132.6, 131.8, 130.7, 123.7, 121.5, 121.4, 119.1, 116.2, 115.8, 113.5, 105.1, 98.9, 93.7, 69.9.

Manufacturing example  10. 2- Benzo [1,3] dioxol-5- yl -7- (4- chloro - benzyloxy ) -3,5-dihydroxy-chromen-4-one (compound 4d), 2- Benzo [1,3] dioxol-5- yl -7- (3- chloro - benzyloxy ) -3,5-dihydroxy-chromen-4-one (Compound 4e) and  3- (2- Benzo [1,3] dioxol-5- yl Preparation of -3,5-dihydroxy-4-oxo-4H-chromen-7-yloxymethyl) -benzonitrile (Compound 4f):

Compound 13d-f was dissolved in 2M ammonia methanol solution. After stirring for 5 hours at room temperature and concentrated under reduced pressure to give the compound 4d-f (90 ~ 92% yield) as a yellow powder:

4d ¹ H NMR (400 MHz, DMSO- d6 ) d 7.82 (d, J = 8.3 Hz, 1H), 7.74 (s, 1H), 7.47-7.52 (m, 4H), 7.13 (d, J = 8.4 Hz, 1H), 6.90 (s, 1H), 6.45 (s, 1H), 6.14 (s, 2H), 5.24 (s, 2H); ¹³ C NMR (100 MHz, DMSO- d6 ) d 181.9, 164.2, 160.8, 147.9, 137.0, 135.6, 130.0, 128.9, 125.4, 125.0, 123.2, 108.9, 107.8, 104.6, 102.1, 98.5, 98.0, 93.5, 92.4, 92.1 , 69.5;

4e ¹ H NMR (400 MHz, DMSO- d6 ) d 7.82 (d, J = 8.4 Hz, 1H), 7.74 (s, 1H), 7.56 (s, 1H), 7.45 (s, 3H), 7.13 (d, J = 8.3 Hz, 1H), 6.91 (s, 1H), 6.47 (s, 1H), 6.14 (s, 2H), 5.26 (s, 2H); ¹³ C NMR (100 MHz, DMSO- d6 ) d 179.6, 171.8, 164.1, 160.8, 156.4, 149.8, 149.1, 148.0, 139.1, 133.5, 130.9, 128.4, 127.8, 126.7, 125.6, 123.2, 108.9, 107.8, 102.1, 98.8 , 98.4, 93.5, 69.3;

4f ¹ H NMR (400 MHz, DMSO- d6 ) d 7.96 (s, 1H), 7.82-7.86 (m, 3H), 7.74 (s, 1H), 7.65 (t, J = 7.7 Hz, 1H), 7.13 ( d, J = 8.2 Hz, 1H), 6.93 (s, 1H), 6.48 (s, 1H), 6.15 (s, 2H), 5.30 (s, 2H); ¹³ C NMR (100 MHz, DMSO- d6 ) d 172.1, 162.7, 156.8, 153.6, 148.4, 147.0, 138.6, 133.3, 132.6, 131.9, 131.7, 130.6, 125.7, 125.3, 123.6, 119.3, 112.3, 109.2, 108.1, 105.1 , 102.5, 98.8, 93.8, 69.4.

Example  One. Dihydroxychromone  SARS coronavirus of derivatives Helicase   Validation of activity inhibitory capacity:

Cloning and purification of SARS coronavirus helicase is known in the art [ Angew . Chem . Int. Ed . 2007, 46 , 6464] and the ability of the dihydroxychromone derivatives to inhibit SARS coronavirus helicase activity was assessed by determining the ability of the compound to inhibit DNA unwinding. The DNA loosening inhibitory ability is based on fluorescence resonance energy transfer phenomena (FRET-based assay) [ Biochem . Biophys . Res . Commun . 2008, 366, 738].

Two oligo DNAs that were fluorescently labeled at the ends were synthesized (with 25 complementary base pairs and 20 bases at the 5 ends are single stranded) and separated by PAGE purification:

5′-20T25Tam: 5′-TTTTTTTTTTTTTTTTTTTTGAGCGGATTACTATACTACATTAGA (TAMRA) -3 ′ (SEQ ID NO: 1)

3′-0T25Flu: 5 ′-(Fluorescein) TCTAATGTAGTATAGTAATCCGCTC-3 ′ (SEQ ID NO: 2)

To prepare double-stranded DNA substrates, the two oligo DNAs were mixed at a ratio of 1.2: 1 (5′-20T25Tam: 3′-0T25Flu, 3′-0T25Flu at 5.0 mM) and heated to 95 ^o C and 37 ^o After an hour of cooling in C, annealing is performed. Uncoupling reaction of double-stranded DNA produced 80 ml of mixture (150 nM SCV Hel, 1 mM MgCl ₂ , 9 mM ATP, 5 mM DTT, synthetic compound in various concentrations, 20 mM HEPES, pH 7.4) at room temperature. Begins. The reaction mixture is incubated for 10 minutes and then 20 nM double stranded DNA substrate (20 ml) is added. The reaction solution was reacted at 37 ^° C. for 2 minutes and then purified by 100 ml of reaction solution (0.1 M EDTA, 0.4 mM trap DNA, 20 mM HEPES (pH 7.4)). Fluorescence was measured using a fluorophotometer (Ex = 485 nm / Em = 535 nm).

As a result of verifying the inhibitory ability of dihydroxychromone derivatives against SARS coronavirus helicase, Compound 2a-c showed inhibitory effect at the concentration of 50 μM or higher, and Compound 3b also showed the inhibitory effect at the concentration of 50 μM or higher. . IC ₅₀ values of compounds 3a and 4a-f were between 2.7 and 15.4 μM, and compound 4c showed the highest inhibitory effect among the compounds used in the experiment [FIG. 3].

The specific parts of the present invention have been described in detail above, and for those skilled in the art, these specific descriptions are merely preferred embodiments, and the scope of the present invention is not limited thereto. Will be obvious. Thus, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

Figure 1 shows the structural similarity of aryl diketo acid (ADK) and dihydroxychromone.

FIG. 2 shows the structures of compounds 2a-c , 3a-b, 4a-f .

FIG. 3 shows IC ₅₀ values of dihydroxychromone derivatives.

4 shows the method for synthesizing Compounds 2a-c .

5 shows a method for synthesizing compound 3b .

FIG. 6 shows the synthesis of Compounds 4a-f .

<110> Konkuk University Industrial Cooperation Corp <120> PHARMACEUTICAL COMPOSITIONS COMPRISING DIHYDROXYCHROMONE          DERIVATIVES AS AN ACTIVE INGREDIENT FOR TREATING AND PREVENTING          DISEASES CAUSED BY CORONAVIRUSES <130> P09-E221 <160> 2 <170> KopatentIn 1.71 <210> 1 <211> 45 <212> DNA <213> Artificial Sequence <220> <223> oligonucleotide <400> 1 tttttttttt tttttttttt gagcggatta ctatactaca ttaga 45 <210> 2 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> oligonucleotide <400> 2 tctaatgtag tatagtaatc cgctc 25  

Claims (32)

delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete It is caused by a coronavirus comprising a compound of the formula (2), a pharmaceutically acceptable salt thereof, an isomer thereof, or a tautomer thereof, as an active ingredient, together with a pharmaceutically acceptable carrier, diluent or excipient. Pharmaceutical compositions for the treatment and prevention of diseases: [Formula 2]

In Chemical Formula 2, R ₁ and R ₂ are hydroxy or together form a ring in the structure —O— (CH ₂ ) nO—, where n is an integer from 1 to 10; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₁₀ alkyl, C ₁ -C ₁₀ alkenyl or C ₁ -C ₁₀ alkoxy. The method of claim 17, R ₁ and R ₂ are hydroxy or together form a ring in the structure —O— (CH ₂ ) nO—, where n is an integer from 1 to 6; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₆ alkyl, C ₁ -C ₆ alkenyl or C ₁ -C ₆ alkoxy compounds of the disease caused by coronavirus, characterized in that Pharmaceutical compositions for the treatment and prophylaxis. The method of claim 17, R ₁ and R ₂ are hydroxy or together form a ring in the structure —O— (CH ₂ ) nO—, where n is an integer from 1 to 3; R ₃ is a benzyl group unsubstituted or substituted with one or more halogen or cyano or a phenyl group unsubstituted or substituted with one or more halogen or cyano; R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₃ alkyl, C ₁ -C ₃ alkenyl or C ₁ -C ₃ alkoxy compounds characterized by Pharmaceutical compositions for the treatment and prophylaxis. The method of claim 17, R ₁ and R ₂ are hydroxy or together form a ring with the structure -O-CH ₂ -O-; R ₃ is a benzyl group unsubstituted or substituted with one or more fluorine or cyano; R ₄ and R ₅ are each independently hydrogen, hydroxy, C ₁ -C ₃ alkyl, C ₁ -C ₃ alkenyl or C ₁ -C ₃ alkoxy compounds characterized by Pharmaceutical compositions for the treatment and prophylaxis. The method according to any one of claims 17 to 20, The coronavirus is infectious bronchitis virus (IBV), bovine coronavirus (BCoV), canine coronavirus (CCoV), feline coronavirus (FeCoV), porcine erythrocyte agglutination encephalomyelitis virus (PEDV), mouse hepatitis virus (MHV), turkey corona Virus (TCoV), infectious gastrointestinal virus (TGEV), rat coronavirus (RCV), human coronavirus 229E (HCoV-229E), human coronavirus NL63 (NL), human coronavirus OC43 (HCoV-OC43) and SARS Pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that any one selected from coronavirus (SARS-CoV). The method according to any one of claims 17 to 20, Said disease is SARS induced by SARS coronavirus (SARS), characterized in that the pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus. The method according to any one of claims 17 to 20, The compound is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that to inhibit the release of the nucleic acid substrate of the coronavirus helicase. 24. The method of claim 23, The coronavirus helicase is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that SARS coronavirus helicase. 24. The method of claim 23, The nucleic acid substrate is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that the DNA or RNA. The method according to any one of claims 17 to 20, The pharmaceutical composition is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that it further comprises one or more other antiviral agents. The method of claim 26, The antiviral agent is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that selected from viral serine protease inhibitors, viral polymerase inhibitors, interferona or ribavirin. The method according to any one of claims 17 to 20, The pharmaceutical composition is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus, characterized in that it further comprises one or more additional additives selected from immunomodulators, antioxidants, antibacterial agents and antisense agents. The method of claim 28, The additive is characterized in that it is selected from silybum marianum, interleukine 12, amantadine, amntadine, ribozyme, thymosin, N-acetyl cysteine or cyclosporin Pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus. Pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus comprising a dihydroxychromone derivative represented by the following formula (2) for inhibiting the nucleic acid substrate unwinding activity of the coronavirus helicase. [Formula 2]

31. The method of claim 30, The nucleic acid substrate is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus comprising a dihydroxychromone derivative, characterized in that DNA or RNA. The method of claim 30 or 31, The coronavirus is a pharmaceutical composition for the treatment and prevention of diseases caused by coronavirus comprising a dihydroxychromone derivative, characterized in that SARS coronavirus.

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