Patent Application
20250228814
Small molecule antiviral agents are a diverse class of antiviral therapies, approved for treatment of many viral infections including hepatitis B, hepatitis C, influenza, herpes simplex-1, and HIV. Despite their efficacy, these compounds may often present harmful side effects resulting from toxicity. As such, there exists a need to improve the clinical profile of many antiviral agents.
The present disclosure relates to therapeutic agents and combinations thereof (e.g., pharmaceutical compositions) for the treatment of a viral infection in a subject, tissue or cell. In one aspect, the present disclosure features a method of treating a viral infection (e.g., a coronavirus infection, e.g., Covid-19) in a subject comprising administering a combination of a biflavonoid compound and an antiviral agent (e.g., remdesivir or sofosbuvir) to the subject. In an embodiment, administering a combination of a biflavonoid compound and an antiviral agent to a subject results in a beneficial effect in the subject, e.g., compared with administering the biflavonoid compound and/or the antiviral agent (e.g., remdesivir or sofosbuvir) individually. For example, a combination of a bioflavonoid compound and an antiviral agent may result in the reduced toxicity of the antiviral agent and/or an increase in efficacy of the antiviral agent in the subject.
In one aspect, the present disclosure features a method of treating a viral infection in a subject, comprising providing a combination of a bioflavonoid compound and an antiviral agent to the subject wherein the molar amount of the bioflavonoid compound in the combination is greater than the molar amount of the antiviral agent. In an embodiment, the efficacy of the combination is greater than the efficacy of the biflavonoid compound alone at the molar amount used in the combination. In an embodiment, the efficacy of the combination is greater than the efficacy of the antiviral agent alone at the molar amount used in the combination.
In an embodiment, each of the biflavonoid compound and the antiviral agent is independently formulated as a pharmaceutical composition. In an embodiment, the biflavonoid compound and the antiviral agent are formulated together as a pharmaceutical composition. In an embodiment, each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) concomitantly to the subject. In an embodiment, each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) sequentially to the subject. In an embodiment, the biflavonoid compound is provided (e.g., administered) to the subject prior to the antiviral agent. In an embodiment, the antiviral agent is provided (e.g., administered) to the subject prior to the biflavonoid compound. In an embodiment, the biflavonoid compound is selected from a compound listed in Table 1, or a pharmaceutically acceptable salt thereof. In an embodiment, the antiviral agent is selected from remdesivir or sofosbuvir, or a pharmaceutically acceptable salt thereof.
Described herein are compositions comprising a biflavonoid compound (e.g., a compound of Formula (I) or a pharmaceutically acceptable salt thereof, e.g., ginkgetin or isoginkgetin) and antiviral agent (e.g., remdesivir), as well as methods of using the same to treat a viral infection in the subject, tissue, or cell. In an embodiment, administering a combination of a biflavonoid compound and an antiviral agent to a subject results in a beneficial effect in the subject, e.g., compared with administering the biflavonoid compound and/or the antiviral agent individually. For example, a combination of a biflavonoid compound and an antiviral agent may result in the reduced toxicity of the antiviral agent and/or an increase in efficacy of the antiviral agent in the subject. A description of exemplary embodiments of the disclosure is provided herein.
As used herein, the articles “a” and “an” refer to one or to more than one (e.g., to at least one) of the grammatical object of the article.
“About” and “approximately” shall generally mean an acceptable degree of error for the quantity measured given the nature or precision of the measurements. Exemplary degrees of error are within 20 percent (%), typically, within 10%, and more typically, within 5% of a given value or range of values.
As used herein, the term “acquire” or “acquiring” as the terms are used herein, refer to obtaining possession of a physical entity (e.g., a sample, e.g., blood sample or liver biopsy specimen), or a value, e.g., a numerical value, by “directly acquiring” or “indirectly acquiring” the physical entity or value. “Directly acquiring” means performing a process (e.g., an analytical method) to obtain the physical entity or value. “Indirectly acquiring” refers to receiving the physical entity or value from another party or source (e.g., a third party laboratory that directly acquired the physical entity or value). Directly acquiring a value includes performing a process that includes a physical change in a sample or another substance, e.g., performing an analytical process which includes a physical change in a substance, e.g., a sample, performing an analytical method, e.g., a method as described herein, e.g., by sample analysis of a cell titer or a bodily fluid, e.g., via mass spectroscopy (e.g. LC-MS) or PCR (e.g., RT-PCR).
“Co-administration”, “co-administering”, “co-providing”, “in combination” and “a combination of” as used herein in the context of the administration of a biflavonoid compound and an antiviral agent, refers to administration at the same time or administration of one therapy before (e.g., immediately before, less than about 5, about 10, about 15, about 30, about 45, about 60 minutes, about 1, about 2, about 3, about 4, about 6, about 8, about 10, about 12, about 16, about 20, about 24, about 48, about 72 or more hours before) administration of a secondary therapy. In some embodiments, the therapies to be co-administered are formulated in a single composition. In other embodiments, the therapies to be co-administered are formulated separately.
Numerous ranges, e.g., ranges for the amount of a therapy administered per day, are provided herein. In some embodiments, the range includes both endpoints. In other embodiments, the range excludes one or both endpoints. By way of example, the range can exclude the lower endpoint. Thus, in such an embodiment, a range of 250 to 400 mg/day, excluding the lower endpoint, would cover an amount greater than 250 that is less than or equal to 400 mg/day.
The term “comprise” is intended to mean “include”. Where a term is provided in the singular, it also contemplates aspects of the invention described by the plural of that term. The term “and/or” where used herein is to be taken as specific disclosure of each of the multiple specified features or components with or without another. Thus, the term “and/or” as used in a phrase such as “A and/or B” herein is intended to include “A and B,” “A or B,” “A” (alone), and “B” (alone). Likewise, the term “and/or” as used in a phrase such as “A, B, and/or C” is intended to encompass each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).
A “course” or “course of therapy,” as referred to herein, comprises one or more separate administrations of a therapeutic agent or a combination of therapeutic agents (e.g., a biflavonoid compound (e.g., compound of Formula (I) or a pharmaceutically acceptable salt thereof) and/or an antiviral agent (e.g., remdesivir)). A course of therapy can comprise one or more cycles of a therapeutic agent. In some embodiments, a therapeutic agent is administered to a subject at least once, at least twice, at least three times, at least four times, or more over a course of treatment. A subject may be administered with one or more courses of treatment. In some embodiments, rest periods may be interposed between courses of treatment. For example, a rest period may be about 1, about 2, about 4, about 6, about 8, about 10, about 12, about 16, about 20, or about 24 hours; or about 1, about 2, about 3, about 4, about 5, about 6, or about 7 days; or about 1, about 2, about 3, about 4 or more weeks in length.
A “cycle”, as used herein in the context of a cycle of administration of a therapeutic agent or a combination of therapeutic agents, refers to a period of time for which the therapeutic agent or combination of therapeutic agents is administered to a patient. For example, if a therapeutic agent is administered for a cycle of 4 weeks days, the periodic administration, e.g., daily or twice daily, is given for 4 weeks. A therapeutic agent or combination of therapeutic agents can be administered for more than one cycle. In some embodiments, the first and second or subsequent cycles are the same in terms of one or both of duration and periodic administration. In embodiments, a first and second or subsequent cycle differs in terms of one or both of duration and periodic administration. Rest periods may be interposed between cycles. A rest cycle may be about 1, about 2, about 4, about 6, about 8, about 10, about 12, about 16, about 20, or about 24 hours; or about 1, about 2, about 3, about 4, about 5, about 6, or about 7 days; or about 1, about 2, about 3, about 4 or more weeks in length.
The term “efficacy,” as used herein in the context of a therapeutic agent or combination of therapeutic agents, refers to the ability of a therapeutic agent or a combination of therapeutic agents to effect a desirable treatment outcome, such as (i) the ability to decrease or inhibit viral infection-induced cytotoxicity; (ii) to decrease or inhibit viral replication or infection rate; (iii) to increase the viability of cells infected with, or at risk of infection by, a virus (e.g., a coronavirus, such as SARS-CoV coronavirus, MERS coronavirus, or SARS-CoV-2 coronavirus); (iv) to increase survival rate of a subject or patient infected with, or at risk of infection by, a virus (e.g., a coronavirus, such as SARS-CoV-2); (v) to decrease or alleviate one or more symptoms of viral infection (e.g., Covid-19 symptoms in a subject); or (vi) to inhibit one or more viral enzyme functions (e.g., one or more coronavirus helicases, coronavirus cysteine proteases, or coronavirus replicases).
As used herein, the terms “increasing” and “decreasing” refer to modulating that results in, respectively, greater or lesser amounts of function, expression, or activity of a particular metric relative to a reference. For example, subsequent to administration to a cell, tissue or subject of a combination of a biflavonoid compound and an antiviral agent described herein, the amount of a marker of a metric (e.g., cell viability, virulence) as described herein may be increased or decreased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 98%, 2×, 3×, 5×, 10× or more relative to the amount of the marker prior to administration or relative to the effect of a negative control agent. The metric may be measured subsequent to administration at a time that the administration has had the recited effect, e.g., at least 12 hours, 24 hours, one week, one month, 3 months, or 6 months, after a treatment has begun.
As used herein, the terms “prevent” or “preventing” as used in the context of a disease or disorder described herein (e.g., a viral infection, e.g., a coronavirus infection), refer to administration of a biflavonoid compound in combination with an antiviral agent to a subject, e.g., the administration of a biflavonoid compound (e.g., compound of Formula (I) or a pharmaceutically acceptable salt thereof) and remdesivir, such that the onset of at least one symptom of the disorder or disease is delayed as compared to what would be seen in the absence of administration of said combination.
As used herein, the term “subject” is intended to include human and non-human animals. Exemplary human subjects include a human patient having a disease or disorder, e.g., a disorder described herein (e.g., a viral infection, e.g., a coronavirus infection), or a healthy subject. The term “non-human animals” includes all vertebrates, e.g., non-mammals (such as chickens, amphibians, reptiles) and mammals, such as non-human primates, domesticated and/or agriculturally useful animals, e.g., sheep, dogs, cats, cows, pigs, etc.
As used herein, an amount of a compound, conjugate, or substance effective to treat a disease or disorder (e.g., a viral infection described herein), “therapeutically effective amount,” “effective amount” or “effective course” refers to an amount of the compound or composition which is effective, upon single or multiple dose administration(s) to a subject, in treating a subject, or in curing, alleviating, relieving or improving a subject with a disorder (e.g., a viral infection, e.g., a coronavirus infection) beyond that expected in the absence of such treatment.
As used herein, the terms “treat” or “treating” as used in the context of a disease or disorder described herein (e.g., a viral infection, e.g., a coronavirus infection), refer to administration of a biflavonoid compound in combination with an antiviral agent to a subject, e.g., the administration of a biflavonoid compound (e.g., compound of Formula (I)) and remdesivir, such that at least one symptom of the disorder or disease is cured, healed, alleviated, relieved, altered, remedied, ameliorated, or improved. Treating includes administering an amount effective to alleviate, relieve, alter, remedy, ameliorate, improve or affect the disorder or disease, or the symptoms of the disorder or disease. The treatment may inhibit deterioration or worsening of a symptom of a disorder or disease. In some embodiments, treating includes preventing. In some embodiments, treating does not include preventing.
Described herein are biflavonoid compounds administered in combination with an antiviral compound, e.g., remdesivir, to provide a therapeutic benefit to a cell or subject, e.g., treating a viral infection, reducing the toxicity of an antiviral agent, or decreasing the virulence of a viral infection in a subject.
As described herein, a biflavonoid compound comprises ginkgetin or an analog or variant thereof. Ginkgetin is a biflavonoid, a class of naturally occurring polyphenolic compounds in which two flavinoid units are joined in a pattern through a linker. It is one of the principal active constituents of Ginkgo biloba, also known as the maidenhair tree. Ginkgo biloba contains multiple natural products, including flavonoids, biflavonoids, and terpenoids, which are shown to have antioxidant properties, and improve circulation and memory in mammals.
In an embodiment, the biflavonoid compound is a compound of Formula (I):
In an embodiment, one of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl. In an embodiment, two of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl. In an embodiment, three of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl. In an embodiment, four of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl. In an embodiment, five of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl. In an embodiment, each of R1, R2, R3, R4, R5, and R6 is independently hydrogen or C1-C6 alkyl.
In an embodiment, one of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, two of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, three of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, four of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, five of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, each of R1, R2, R3, R4, R5, and R6 is independently hydrogen. In an embodiment, one of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl. In an embodiment, two of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl. In an embodiment, three of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl. In an embodiment, four of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl. In an embodiment, five of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl. In an embodiment, each of R1, R2, R3, R4, R5, and R6 is independently C1-C6 alkyl.
In an embodiment, R1 is hydrogen or C1-C6 alkyl. In an embodiment, R2 is hydrogen or C1-C6 alkyl. In an embodiment, R3 is hydrogen or C1-C6 alkyl. In an embodiment, R4 is hydrogen or C1-C6 alkyl. In an embodiment, R5 is hydrogen or C1-C6 alkyl. In an embodiment, R6 is hydrogen or C1-C6 alkyl.
In an embodiment, R1 is hydrogen, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is C1-C6 alkyl, and R6 is C1-C6 alkyl. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is hydrogen, and R6 is C1-C6 alkyl. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is hydrogen, and R6 is hydrogen.
In an embodiment, R1 is hydrogen, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is hydrogen, and R6 is C1-C6 alkyl. In an embodiment, R1 is hydrogen, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is hydrogen, and R6 is hydrogen. In an embodiment, R1 is C1-C6 alkyl, R2 is C1-C6 alkyl, R3 is C1-C6 alkyl, R4 is C1-C6 alkyl, R5 is C1-C6 alkyl, and R6 is C1-C6 alkyl. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is hydrogen, R4 is hydrogen, R5 is C1-C6 alkyl, and R6 is hydrogen. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is C1-C6 alkyl, R4 is hydrogen, R5 is C1-C6 alkyl, and R6 is C1-C6 alkyl. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is C1-C6 alkyl, R4 is hydrogen, R5 is hydrogen, and R6 is hydrogen. In an embodiment, R1 is C1-C6 alkyl, R2 is hydrogen, R3 is C1-C6 alkyl, R4 is hydrogen, R5 is hydrogen, and R6 is C1-C6 alkyl. In an embodiment, R1 is hydrogen, R2 is hydrogen, R3 is C1-C6 alkyl, R4 is hydrogen, R5 is hydrogen, and R6 is C1-C6 alkyl.
In an embodiment, the compound is a compound of Formula (I-a):
In an embodiment, one of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl. In an embodiment, two of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl. In an embodiment, three of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl. In an embodiment, four of R19, R20, R21, R22, R23, and R243 is independently hydrogen or C1-C6 alkyl. In an embodiment, five of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl. In an embodiment, each of R19, R20, R21, R22, R23 and R24 is independently hydrogen or C1-C6 alkyl.
In an embodiment, one of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, two of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, three of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, four of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, five of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, each of R19, R20, R21, R22, R23, and R24 is independently hydrogen. In an embodiment, one of R19, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl. In an embodiment, two of R19, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl. In an embodiment, three of R19, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl. In an embodiment, four of R19, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl. In an embodiment, five of R9, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl. In an embodiment, each of R19, R20, R21, R22, R23, and R24 is independently C1-C6 alkyl.
In an embodiment, R19 is hydrogen or C1-C6 alkyl. In an embodiment, R20 is hydrogen or C1-C6 alkyl. In an embodiment, R21 is hydrogen or C1-C6 alkyl. In an embodiment, R22 is hydrogen or C1-C6 alkyl. In an embodiment, R23 is hydrogen or C1-C6 alkyl. In an embodiment, R24 is hydrogen or C1-C6 alkyl.
In an embodiment, R19 is hydrogen, R20 is hydrogen, R21 is hydrogen, R22 is hydrogen, R23 is C1-C6 alkyl, and R24 is C1-C6 alkyl. In an embodiment, R19 is hydrogen, R20 is hydrogen, R21 is hydrogen, R22 is hydrogen, R23 is hydrogen, and R24 is hydrogen. In an embodiment, R19 is hydrogen, R20 is C1-C6 alkyl, R21 is hydrogen, R22 is hydrogen, R23 is hydrogen, and R24 is hydrogen.
In an embodiment, the biflavonoid compound is a compound shown in Table 1.
In an embodiment, the biflavonoid compound is ginkgetin or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is isoginkgetin or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is amentoflavone or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is 7,7″-dimethoxyagastisflavone or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is sotestsuflavone or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is sciadopitysin or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is sequoiaflavone or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is bilobetin or a pharmaceutically acceptable salt thereof. In an embodiment, the biflavonoid compound is tetrahydroamentoflavone. In an embodiment, the biflavonoid compound is dioonflavone. In an embodiment, the biflavonoid compound is podocarpusflavone A. In an embodiment, the biflavonoid compound is putraflavone. In an embodiment, the biflavonoid compound is 7L)-Methylsciadopitysin. In an embodiment, the biflavonoid compound is 8-[5-(5,7-Dihydroxy-4-oxochromen-2-yl)-2-hydroxyphenyl]-5-hydroxy-7-methoxy-2-(4-methoxyphenyl)chromen-4-one. In an embodiment, the biflavonoid compound is kayaflavone. In an embodiment, the biflavonoid compound is heveaflavone. In an embodiment, the biflavonoid compound is agathisflavone. In an embodiment, the biflavonoid compound is 5,7-Dihydroxy-8-[5-hydroxy-2-(4-hydroxyphenyl)-7-methoxy-4-oxochromen-6-yl]-2-(4-methoxyphenyl)chromen-4-one.
In an embodiment, the biflavonoid compound is selected from: compound 100, compound 101, compound 102 compound 103, compound 104, compound 105, compound 106, compound 107, compound 108, compound 109, compound 110, compound 111, compound 112, compound 113, compound 114, compound 115, compound 116, and compound 117. In an embodiment, the biflavonoid compound is compound 100. In an embodiment, the biflavonoid compound is compound 101 In an embodiment, the biflavonoid compound is compound 102. In an embodiment, the biflavonoid compound is compound 103. In an embodiment, the biflavonoid compound is compound 104. In an embodiment, the biflavonoid compound is compound 105. In an embodiment, the biflavonoid compound is compound 106. In an embodiment, the biflavonoid compound is compound 107. In an embodiment, the biflavonoid compound is compound 108. In an embodiment, the biflavonoid compound is compound 109. In an embodiment, the biflavonoid compound is compound 110. In an embodiment, the biflavonoid compound is compound 111. In an embodiment, the biflavonoid compound is compound 112. In an embodiment, the biflavonoid compound is compound 113. In an embodiment, the biflavonoid compound is compound 114. In an embodiment, the biflavonoid compound is compound 115. In an embodiment, the biflavonoid compound is compound 116. In an embodiment, the biflavonoid compound is compound 117.
In an embodiment, the biflavonoid compound (e.g., ginkgetin) or a pharmaceutically acceptable salt thereof, is provided as a substantially pure compound, for example, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or at least 99.9% pure, measured, e.g, by HPLC analysis. In embodiments described herein, the biflavonoid compound is provided as ginkgetin (CAS #481-46-9) in the absence of a ginkgolic acid, ginkgolide, or terpenoid that can naturally occur with ginkgetin in Gingko biloba.
A composition useful for the treatment of a viral infection (e.g., as described herein) may contain a single biflavonoid compound (e.g., ginkgetin) or a plurality of biflavonoid compounds. For example, a composition comprising a biflavonoid compound may contain only ginkgetin, e.g., synthetically prepared or extracted from a natural source. In contrast, a composition comprising a biflavonoid compound may also contain a combination of ginkgetin and a closely related analog or variant thereof, e.g., isoginkgetin or amentoflavone.
The biflavonoid compounds provided herein may contain one or more asymmetric centers and thus occur as racemates and racemic mixtures, single enantiomers, individual diastereomers and diastereomeric mixtures. Further, the biflavonoid compounds may exist as one of many tautomeric forms. All such isomeric and tautomeric forms of these compounds are expressly included within the scope. Unless otherwise indicated when a compound is named or depicted by a structure without specifying the stereochemistry and has one or more chiral centers or tautomeric structures, it is understood to represent all possible stereoisomers or tautomers of the compound. The compounds provided herewith may also contain linkages (e.g., carbon-carbon bonds, phosphorus-oxygen bonds, or phosphorus-sulfur bonds) or substituents that can restrict bond rotation, e.g., restriction resulting from the presence of a ring or double bond.
The present disclosure features administration of an antiviral agent to a subject or cell in combination with a biflavonoid compound (e.g., a biflavonoid compound described herein). In an embodiment, administration of the biflavonoid compound in combination with the antiviral agent results in an improvement of the therapeutic window for the antiviral agent, e.g., reducing toxicity of the antiviral agent or reducing the potency of the antiviral agent, e.g., in a subject or cell infected with a viral infection.
An antiviral agent for use in combination with a biflavonoid compound may be any antiviral agent known in the art. In an embodiment, the antiviral agent modulates (e.g., inhibits) a step in the viral life cycle, thus impairing the ability of the virus to replicate and/or propagate. For example, the antiviral agent may be an attachment inhibitor, post-attachment inhibitor, fusion inhibitor, entry inhibitor, uncoating inhibitor, protease inhibitor, polymerase inhibitor, nucleotide reverse transcriptase inhibitor, nucleoside reverse transcriptase inhibitor, non-nucleoside reverse transcriptase inhibitor, and integrase inhibitor. An attachment inhibitor may prevent recognition and/or attachment of a virus particle to a target cell. A post-attachment inhibitor may prevent engagement of the virus particle with the target cell at the site of attachment. A fusion inhibitor or entry inhibitor may prevent a virus particle from fusing with or otherwise entering a target cell. An uncoating inhibitor may prevent release of the contents of a viral particle into the target cell upon entry of the virus particle. Protease inhibitors, polymerase inhibitors, nucleotide reverse transcriptase inhibitors, nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, and integrase inhibitors may prevent successful replication and/or packaging of viral genetic information in the target cell. In some embodiments, the antiviral agent is a polymerase inhibitor. In some embodiments, the polymerase inhibitor is a viral RNA-dependent RNA polymerase inhibitor. In some embodiments, the RNA-dependent RNA polymerase inhibitor is selected from: remdesivir, favipiravir, triazavirin, galidesivir, GS-441524, NHC (EIDD-1931), and EIDD-2801. In some embodiments, the RNA-dependent RNA polymerase inhibitor is remdesivir.
The antiviral agent may function by targeting a specific viral function, such as inhibiting a viral nucleic acid polymerase, viral protease, viral integrase, or viral neuraminidase. In another embodiment, the antiviral agent functions by targeting a host cell function required for successful viral replication, such as viral entry into a host cell, nucleic acid synthesis, protein synthesis, viral capsid assembly, or viral exit from the host cell.
In an embodiment, the antiviral agent is a capsid inhibitor, a secretion inhibitor, a microRNA, an antisense RNA agent, an RNAi agent, or other agent designed to inhibit viral RNA. In an embodiment, the antiviral agent is a small molecule, a lipid, an oligonucleotide, a peptide, or an antibody. In an embodiment, the antiviral agent is a small molecule antiviral agent. The antiviral agent may be a nucleoside analog, a peptide, or a nonribosomal peptide.
In some embodiments, the antiviral agent targets a DNA virus. In some embodiments, the antiviral agent targets an RNA virus. In some embodiments, the antiviral agent has broad spectrum activity against numerous types of viruses, e.g., and is capable of targeting both a DNA virus and an RNA virus.
Exemplary antiviral agents include abacavir, acyclovir, amantadine, ampligen, amprenavir, umifenovir, atripia, alazanavir, biktarvy, baloxavir marboxil, bulevirtide, boceprevir, chloroquine, cidofovir, cobicistat, combivir, daclatasvir, darunavir, delavirdine, descovy, didanosine, docosanol, dolutegravir, doravirine, edoxudine, efavirenz, elvitegravir, emtricitabine, enfuvirtide, entecavir, etravirine, ensitrelvir, famciclovir, favipiravir, fomivirsen, fosamprenavir, foscamet, galidesivir, ganciclovir, hydroxychloroquine, ibacitabine, ibalizumab, idoxuridine, imiquimod, imunovir, ivermectin, indinavir, lamivudine, letermovir, lopinavir, loviride, maraviroc, methisazone, moroxydine, nelfinavir, nexavir, nitazoxanide, norvir, oseltamivir, penciclovir, peramvir, pleconaril, pieconaril, raltegravir, rilpivirine, ribavirin, remdesivir, ritonavir, saquinavir, sofosbuvir, taribavirin, telaprevir, tenofovir, telbivudine, trizivir, triazivirin, tipranavir, truvada, tromantadine, trifluridine, vidarabine, umifenovir, umifenovir, valaciclovir, vicriviroc, vidarabine, zalcitabine, zanamivir, zicovidinr, GS-441524, NHC (EIDD-1931), and EIDD-2801.
In an embodiment, the antiviral agent is a compound of Formula (II):
In an embodiment, the antiviral agent of Formula (II) is remdesivir. In an embodiment, the antiviral agent of Formula (II) is sofosbuvir.
In an embodiment, of Formula (II), ring A is heteroaryl substituted with 1-2 R17.
In an embodiment, of Formula (II), R15 is aryl substituted with 0-1 R17.
In an embodiment, of Formula (II), one of R16a and R16b is hydrogen.
In an embodiment, of Formula (II), one of R16a and R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl.
In an embodiment, of Formula (II), one of R12a and R12b is hydrogen.
In an embodiment, of Formula (II), one of R12a and R12b is —ORA.
In an embodiment, of Formula (II), one of R12a and R12b is halo.
In an embodiment, of Formula (II), one of R13a and R13b is —ORA.
In an embodiment, of Formula (II), one of R13a and R13b is hydrogen.
In an embodiment, of Formula (II), A is heteroaryl substituted with 1 R17; R11 is cyano; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; RA is hydrogen; and R17 is —NH2.
In an embodiment, of Formula (II), A is heteroaryl substituted with 2 R17, R11 is hydrogen; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; and RA is hydrogen; and each R17 is independently oxo.
In an embodiment, the antiviral agent is a compound of Formula (II-a):
In an embodiment, the antiviral agent is a compound of Formula (II-b):
In an embodiment, the antiviral agent is a compound shown in Table 2.
In an embodiment, the antiviral agent is remdesivir or an analog thereof. The antiviral agent (e.g., remdesivir) may be isomerically pure or may be a mixture of various stereoisomers. For example, remdesivir may comprise the (R)-isomer, the (S)-isomer, or a mixture of both. In an embodiment, the antiviral agent is a remdesivir analog, e.g., as described in WO 2021/202907, which is incorporated herein by reference in its entirety.
In an embodiment, the antiviral agent is selected from: compound 118, compound 119, compound 120, compound 121, compound 122, compound 123, compound 124, compound 125, compound 126, and compound 127.
In an embodiment, the antiviral agent is compound 118. In an embodiment, the antiviral agent is compound 119. In an embodiment, the antiviral agent is compound 120. In an embodiment, the antiviral agent is compound 121. In an embodiment, the antiviral agent is compound 122. In an embodiment, the antiviral agent is compound 123. In an embodiment, the antiviral agent is compound 124. In an embodiment, the antiviral agent is compound 125. In an embodiment, the antiviral agent is compound 126. In an embodiment, the antiviral agent is compound 127.
Described herein are methods for treating a viral infection in a subject, tissue or cell. In embodiments, the cell is an isolated cell (e.g., a cell in a cell culture or a cell isolated from a tissue or intact organism). In embodiments, the cell is located in a tissue or organ. The viral infection occurs upon attachment and invasion of a host cell by a virion. Viruses are categorized into various classes, primarily based upon whether the virus is an RNA virus or a DNA virus. In an embodiment, the viral infection is an infection from an RNA virus (e.g., a positive-sense single-stranded RNA virus, a negative-sense single-stranded RNA virus, or a double-stranded RNA virus). In an embodiment, the viral infection is an infection from a DNA virus (e.g., a single-stranded DNA virus or a double-stranded DNA virus).
In an embodiment, the viral infection is caused by a double-stranded DNA virus. Exemplary double-stranded DNA viruses are in the Adenoviridae, Herpesviridae, Papillonaviridae, and Poxviridae viral families. For example, the virus may be an adenovirus, herpes simplex type 1 virus, herpes simplex virus type 2, varicella-zoster virus, Epstein-Barr virus, human cytomegalovirus, hepesvirus type 8, papillomavirus, BK virus, JC virus, or smallpox virus. In an embodiment, the viral infection is caused by a single-stranded DNA virus. Exemplary single-stranded DNA viruses are in the Parvoviridae viral family. For example, the virus may be a parvovirus B19 virus. In an embodiment, the viral infection is caused by a double-stranded RNA virus. Exemplary double-stranded RNA viruses are in the Reoviridae family. For example, the virus may be a rotavirus, orbivirus, coltivirus, or banna virus. In an embodiment, the viral infection is caused by a positive-sense single-stranded RNA virus. Exemplary positive-sense single-stranded RNA viruses are in the Astroviridae, Caliciviridae, Coronaviridae, Flaviviridae, Hepeviridae, Matonaviridae, or Picornaviridae viral families. For example, the virus may be a human astrovirus, Norwalk virus, human coronavirus 229E, human coronavirus NL63, human coronavirus OC43, human coronavirus HKU1, Middle East respiratory syndrome-related coronavirus, severe acute respiratory syndrome (SARS) coronavirus, severe acute respiratory syndrome (SARS) coronavirus 2, hepatitis C virus, yellow fever virus, Dengue virus, West Nile virus, TBE virus, Zika virus, hepatitis E, rubella virus, coxsackievirus, hepatitis A virus, poliovirus, or rhinovirus. In an embodiment, the viral infection is caused by a negative-sense single-stranded RNA virus. Exemplary negative-sense single-stranded RNA viruses are in the Arenaviridae, Bunyaviridae, Filoviridae, Orthomyxoviridae, Paramyxoviridae, Pneumoviridae, or Rhabdoviridae viral families. For example, the virus may be a Lassa virus, Crimean-Congo hemorrhagic fever virus, Hantaan virus, Ebola virus, Marburg virus, influenza virus, measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, rabies virus, hepatitis D virus. In an embodiment, the viral infection is caused by a single-stranded RNA retrovirus. Exemplary single-stranded RNA retroviruses are in the Retroviridae viral family. For example, the virus may be a human immunodeficiency virus. In an embodiment, the viral infection is caused by a double-stranded RNA retrovirus. Exemplary double-stranded RNA retroviruses are in the Hepadnaviridae viral family. For example, the virus may be a hepatitis B virus.
In an embodiment, the viral infection is caused by a coronavirus. Coronaviruses are a large family of viruses that are common in people and many different species of animals. Some coronaviruses can cause severe illness in people. Some notable coronaviruses include SARS coronavirus (SARS-CoV), which causes severe acute respiratory syndrome (SARS); MERS coronavirus (MERS-CoV), which causes Middle East respiratory syndrome (MERS); and SARS-CoV-2, which causes coronavirus disease 2019 (COVID-19). Infection with a coronavirus can cause fever, cough and shortness of breath. Infection can be particularly dangerous in older people, people with weakened immune systems, and people with underlying health conditions, such as cardiovascular disease, diabetes, and chronic lung disease, among others. In an embodiment, the viral infection is a SARS-CoV infection. In an embodiment, the viral infection is a MERS-CoV infection. In an embodiment, the viral infection is a SARS-CoV-2 infection.
In an embodiment, the SARS-CoV-2 infection comprises the alpha variant (B.1.1.7 or Q lineage). In an embodiment, the SARS-CoV-2 infection comprises the beta variant (B.1.351 or a descendant lineage). In an embodiment, the SARS-CoV-2 infection comprises the gamma variant (P.1 or a descendant lineage). In an embodiment, the SARS-CoV-2 infection comprises the delta variant (B.1.617.2 or AY lineage). In an embodiment, the SARS-CoV-2 infection comprises the epsilon variant (B.1.427 or B.1.429 lineage). In an embodiment, the SARS-CoV-2 infection comprises the eta variant (B.1.525 lineage). In an embodiment, the SARS-CoV-2 infection comprises the iota variant (B.1.526 lineage). In an embodiment, the SARS-CoV-2 infection comprises the kappa variant (B.1.617.1 lineage). In an embodiment, the SARS-CoV-2 infection comprises the 1.617.3, mu variant (B.1.621 or B.1.621.1 lineage). In an embodiment, the SARS-CoV-2 infection comprises the omicron variant (B.1.1.529, BA.1, BA.1.1, BA.2, BA.3, BA.4, or BA.5 lineage). In an embodiment, the SARS-CoV-2 infection comprises the zeta variant (P.2 lineage).
In an embodiment, the coronavirus comprises one or more (e.g., two, three, four, five, or more) mutations. In an embodiment, the coronavirus comprises between about 1 and 100 (e.g., between about 2 and 95, between about 3 and 90, between about 4 and 80, between about 5 and 75, between about 10 and 50, between about 20 and 40, or about 30) mutations. In an embodiment, the coronavirus comprises one or more mutations selected from R230K, G204R, G251V, L3606F, P4714L, D614G, Q57H, and D614G.
In an embodiment, the coronavirus comprises a nucleotide sequence having at least 65% (e.g., at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or more) sequence identity to a known coronavirus nucleotide sequence (e.g., NC_045512.2, GenBank: PA552564.1, GenBank: PA544053.1, GenBank: OW444422.1, GenBank: OW444299.1, or any other coronavirus nucleotide sequence known in the art). For example, in some embodiments, the coronavirus comprises a nucleotide sequence having at least 70% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 75% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 80% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 85% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 90% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 95% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 96% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 97% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 98% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99.5% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99.9% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having greater than 99.9% sequence identity to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein.
In an embodiment, the coronavirus comprises a polypeptide sequence having at least 65% (e.g., at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or more) sequence identity to a known coronavirus polypeptide sequence (e.g., NC_045512.2, PDB: 7B3D_A, PDB: 6VW1_F, PDB: 6VW1_E, PDB: 7C8J_B, PDB: 7CYP_C, PDB: 7CYP_B, PDB: 7CYP_A, PDB: 7Y9N_B, or any other coronavirus polypeptide sequence known in the art). For example, in some embodiments, the coronavirus comprises a polypeptide sequence having at least 70% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 75% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 80% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 85% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 90% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 95% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 96% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 97% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 98% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 99% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 99.5% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 99.9% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having greater than 99.9% sequence identity to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein.
In an embodiment, the coronavirus comprises a nucleotide sequence having at least 65% (e.g., at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or more) sequence homology to a known coronavirus nucleotide sequence (e.g., NC_045512.2, GenBank: PA552564.1, GenBank: PA544053.1, GenBank: OW444422.1, GenBank: OW444299.1, or any other coronavirus nucleotide sequence known in the art). For example, in some embodiments, the coronavirus comprises a nucleotide sequence having at least 70% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 75% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 80% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 85% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 90% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 95% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 96% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 97% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 98% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99.5% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having at least 99.9% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a nucleotide sequence having greater than 99.9% sequence homology to a known coronavirus nucleotide sequence, e.g., of a coronavirus described herein.
In an embodiment, the coronavirus comprises a polypeptide sequence having at least 65% (e.g., at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.9%, or more) sequence homology to a known coronavirus polypeptide sequence (e.g., NC_045512.2, PDB: 7B3D_A, PDB: 6VW1_F, PDB: 6VW1_E, PDB: 7C8J_B, PDB: 7CYP_C, PDB: 7CYP_B, PDB: 7CYP_A, PDB: 7Y9N_B, or any other coronavirus polypeptide sequence known in the art). For example, in some embodiments, the coronavirus comprises a polypeptide sequence having at least 70% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 75% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 80% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 85% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 90% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 95% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 96% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 97% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 98% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 99% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having at least 99.5% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a sequence having at least 99.9% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein. In some embodiments, the coronavirus comprises a polypeptide sequence having greater than 99.9% sequence homology to a known coronavirus polypeptide sequence, e.g., of a coronavirus described herein.
Described herein are combination therapies comprising a biflavonoid compound and an antiviral agent (e.g., remdesivir) useful for the treatment of a viral infection. The combination therapy may be administered as a single formulation or as separate formulations. In an embodiment, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are administered as a single pharmaceutical composition. In an embodiment, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are administered as separate pharmaceutical compositions. In the case of separate formulations, the biflavonoid compound and the antiviral agent may be administered concomitantly or sequentially. In an embodiment, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are administered concomitantly. In an embodiment, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are administered sequentially. For example, the biflavonoid compound may be administered prior to the antiviral agent (e.g., remdesivir) or subsequent to the antiviral agent (e.g., remdesivir).
In some embodiments, the administration of the biflavonoid compound and the antiviral agent (e.g., remdesivir) has a synergistic or additive effect. For example, the administration of the biflavonoid compound and the antiviral agent (e.g., remdesivir) may have an additive effect, in which the therapeutic effect of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is the total sum of the effects of each of the components individually. In contrast, the administration of the biflavonoid compound and the antiviral agent (e.g., remdesivir) may have a synergistic effect, in which the therapeutic effect of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than the sum of the individual components. The synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) may be 0.1%, 0.25%. 0.5%. 0.75%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more than the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than between 5% and 75% of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually.
In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than 10% of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than 25% of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than 50% of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than 75% of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually.
The synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) may be 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 75-fold, 100-fold, 200-fold, 500-fold greater or more than the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than between 2-fold and 100-fold of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually. In an embodiment, the synergistic effect of the combination of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is greater than between 5-fold and 50-fold of the total sum of the effects of the biflavonoid compound and the antiviral agent (e.g., remdesivir) administered individually.
In one aspect, the amount of the biflavonoid compound and the amount of the antiviral agent are selected such that the molar concentration of the biflavonoid compound is greater than the molar concentration of the antiviral agent (e.g., remdesivir). For example, the molar concentration of the biflavonoid compound is greater than 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, 50-fold, 75-fold, 100-fold, 200-fold, or 500-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is greater than 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is between 2-fold and 20-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is between 5-fold and 10-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is about 5-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is about 10-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir). In an embodiment, the molar concentration of the biflavonoid compound is about 15-fold greater than the molar concentration of the antiviral agent (e.g., remdesivir).
In another aspect, the combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir) results in reducing the toxicity of the antiviral agent (e.g., remdesivir) in a subject or cell. For example, administration of a combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir) may result in reducing the toxicity of the antiviral agent (e.g., remdesivir) in a subject or cell, compared with the toxicity of the antiviral agent (e.g., remdesivir) when administered to the subject or cell individually. In an embodiment, the toxicity of the antiviral agent (e.g., remdesivir) in a subject or cell is reduced upon administration of the antiviral agent (e.g., remdesivir) in combination with a biflavonoid compound, e.g., by about 0.1%, 0.25%. 0.5%. 0.75%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more compared with the toxicity of the antiviral agent (e.g., remdesivir) when administered individually.
In another aspect, the combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir) results in reducing the toxicity of the biflavonoid compound in a subject or cell. For example, administration of a combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir) may result in reducing the toxicity of the biflavonoid compound in a subject or cell, compared with the toxicity of the biflavonoid compound when administered to the subject or cell individually. In an embodiment, the toxicity of the biflavonoid compound in a subject or cell is reduced upon administration of an antiviral agent (e.g., remdesivir) in combination with a biflavonoid compound, e.g., by about 0.1%, 0.25%. 0.5%. 0.75%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9% 10%, 15% 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more compared with the toxicity of the biflavonoid compound when administered individually.
In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 0.1 μM and 500 μM in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 0.5 M and 100 μM in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 1 μM and 100 μM in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 1 μM and 50 μM in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 1 μM and 25 M in a subject or cell. In an embodiment, the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.01 M and 50 μM in a subject or cell. In an embodiment, the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 25 μM in a subject or cell. In an embodiment, the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 10 M in a subject or cell. In an embodiment, the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 M and 5 M in a subject or cell. In an embodiment, the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 2.5 μM in a subject or cell.
In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of between 0.1 μM and 500 μM and the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.01 μM and 50 M in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of to provide a concentration of between 1 μM and 100 μM and the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 M and μM in a subject or cell. In an embodiment, the biflavonoid compound is administered at a dosage to provide a concentration of to provide a concentration of between 1 μM and 25 μM and the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 2.5 μM in a subject or cell.
In another aspect, the combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir) results in reducing the virulence of a virus in a subject. Reducing the virulence may comprise one or more of (i) reducing the infection rate; (ii) reducing the doubling rate, e.g., amount of virus produced by an infected host cell; (iii) reducing the rate of viral DNA/RNA synthesis; (iv) reducing the rate of DNA/RNA mutations by a nucleic acid polymerase; and (v) reducing the rate of virion packaging. In an embodiment, reducing the virulence comprises (i). In an embodiment, reducing the virulence comprises (ii). In an embodiment, reducing the virulence comprises (iii). In an embodiment, reducing the virulence comprises (iv). In an embodiment, reducing the virulence comprises (v).
In an embodiment, the virulence of a virus is reduced in a subject by administering a combination of the biflavonoid compound with an antiviral agent (e.g., remdesivir). In an embodiment, the virulence of a virus is reduced in a subject or cell by about 0.1%, 0.25%. 0.5%. 0.75%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 5%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more by administration of a combination of a biflavonoid compound and an antiviral agent (e.g., remdesivir) compared with administration of one of a biflavonoid compound or an antiviral agent individually.
The present disclosure features methods for treating a subject infected with a viral infection, the methods comprising administering a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1), an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) or a combination thereof.
While it is possible for a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) to be administered alone, it is preferable to administer said compound as a pharmaceutical composition or formulation, where the compounds are combined with one or more pharmaceutically acceptable diluents, excipients or carriers. The compounds according to the disclosure may be formulated for administration in any convenient way for use in human or veterinary medicine. In certain embodiments, the compounds included in the pharmaceutical preparation may be active itself, or may be a prodrug, e.g., capable of being converted to an active compound in a physiological setting. Regardless of the route of administration selected, the compounds of the present disclosure, which may be used in a suitable hydrated form, and/or the pharmaceutical compositions of the present disclosure, are formulated into a pharmaceutically acceptable dosage form such as described below or by other conventional methods known to those of skill in the art.
The amount and concentration of compounds of the present disclosure, e.g., a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) in the pharmaceutical compositions, as well as the quantity of the pharmaceutical composition administered to a subject, can be selected based on clinically relevant factors, such as medically relevant characteristics of the subject (e.g., age, weight, gender, other medical conditions, and the like), the solubility of compounds in the pharmaceutical compositions, the potency and activity of the compounds, and the manner of administration of the pharmaceutical compositions. For further information on Routes of Administration and Dosage Regimes the reader is referred to Chapter 25.3 in Volume 5 of Comprehensive Medicinal Chemistry (Corwin Hansch; Chairman of Editorial Board), Pergamon Press 1990.
Thus, another aspect of the present disclosure provides pharmaceutically acceptable compositions comprising a therapeutically effective amount or prophylactically effective amount of a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir), formulated together with one or more pharmaceutically acceptable carriers (additives) and/or diluents. As described in detail below, the pharmaceutical compositions of the present disclosure may be specially formulated for administration in solid or liquid form, including those adapted for oral or parenteral administration, for example, by oral dosage, or by subcutaneous, intramuscular or intravenous injection as, for example, a sterile solution or suspension. However, in certain embodiments the subject compounds may be simply dissolved or suspended in sterile water. In certain embodiments, the pharmaceutical preparation is non-pyrogenic, i.e., does not elevate the body temperature of a patient.
The phrases “systemic administration,” “administered systemically,” “peripheral administration” and “administered peripherally” as used herein mean the administration of the compound other than directly into the central nervous system, such that it enters the patient's system and, thus, is subject to metabolism and other like processes, for example, subcutaneous administration.
The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
The phrase “pharmaceutically acceptable carrier” as used herein means a pharmaceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, stabilizing agent, excipient, solvent or encapsulating material, involved in carrying or transporting the subject antagonists from one organ, or portion of the body, to another organ, or portion of the body. Each carrier must be “acceptable” in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials which can serve as pharmaceutically acceptable carriers include, but are not limited to: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) ascorbic acid; (17) pyrogen-free water; (18) isotonic saline; (19) Ringer's solution; (20) ethyl alcohol; (21) phosphate buffer solutions; (22) cyclodextrins such as Captisol®; and (23) other non-toxic compatible substances such as antioxidants and antimicrobial agents employed in pharmaceutical formulations.
As set out above, certain embodiments of the compounds described herein may contain a basic functional group, such as an amine, and are thus capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable acids. The term “pharmaceutically acceptable salts” in this respect, refers to the relatively non-toxic, inorganic and organic acid addition salts of compounds of the present disclosure. These salts can be prepared in situ during the final isolation and purification of the compounds of the disclosure, or by separately reacting a purified compound of the disclosure in its free base form with a suitable organic or inorganic acid, and isolating the salt thus formed. Representative salts include the hydrobromide, hydrochloride, sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate, palmitate, stearate, laurate, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate, napthylate, mesylate, glucoheptonate, lactobionate, and laurylsulphonate salts and the like (see, for example, Berge et al. (1977) “Pharmaceutical Salts”, J. Pharm. Sci. 66:1-19).
In other cases, the compounds of the present disclosure may contain one or more acidic functional groups and, thus, are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable bases. The term “pharmaceutically acceptable salts” in these instances refers to the relatively non-toxic, inorganic and organic base addition salts of the compound of the present disclosure (e.g of a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir)). These salts can likewise be prepared in situ during the final isolation and purification of the compounds, or by separately reacting the purified compound in its free acid form with a suitable base, such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary or tertiary amine. Representative alkali or alkaline earth salts include the lithium, sodium, potassium, calcium, magnesium, and aluminum salts and the like. Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine and the like (see, for example, Berge et al., supra).
Wetting agents, emulsifiers, and lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, release agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the compositions. Examples of pharmaceutically acceptable antioxidants include: (1) water soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite and the like; (2) oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, and the like; and (3) metal chelating agents, such as citric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like. The pharmaceutically acceptable carriers, as well as wetting agents, emulsifiers, lubricants, coloring agents, release agents, coating agents, sweetening, flavoring agents, perfuming agents, preservatives, antioxidants, and other additional components may be present in an amount between about 0.001% and 99% of the composition described herein. For example, said pharmaceutically acceptable carriers, as well as wetting agents, emulsifiers, lubricants, coloring agents, release agents, coating agents, sweetening, flavoring agents, perfuming agents, preservatives, antioxidants, and other additional components may be present from about 0.005%, about 0.01%, about 0.05%, about 0.1%, about 0.25%, about 0.5%, about 0.75%, about 1%, about 1.5%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 85%, about 90%, about 95%, or about 99% of the composition described herein.
Pharmaceutical compositions of the present disclosure may be in a form suitable for oral administration, e.g., a liquid or solid oral dosage form. In some embodiments, the liquid dosage form comprises a suspension, a solution, a linctus, an emulsion, a drink, an elixir, or a syrup. In some embodiments, the solid dosage form comprises a capsule, tablet, powder, dragée, or powder. The pharmaceutical composition may be in unit dosage forms suitable for single administration of precise dosages. Pharmaceutical compositions may comprise, in addition to of a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir), a pharmaceutically acceptable carrier, and may optionally further comprise one or more pharmaceutically acceptable excipients, such as, for example, stabilizers (e.g., a binder, e.g., polymer, e.g., a precipitation inhibitor, diluents, binders, and lubricants.
In some embodiments, the composition described herein comprises a liquid dosage form for oral administration, e.g., a solution or suspension. In other embodiments, the composition described herein comprises a solid dosage form for oral administration capable of being directly compressed into a tablet. In addition, said tablet may include other medicinal or pharmaceutical agents, carriers, and or adjuvants. Exemplary pharmaceutical compositions include compressed tablets (e.g., directly compressed tablets), e.g., comprising one or more of a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) or a pharmaceutically acceptable salt thereof.
Formulations of the present disclosure include those suitable for parenteral administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. The amount of active ingredient which can be combined with a carrier material to produce a single dosage form will vary depending upon the host being treated, the particular mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will generally be that amount of the compound which produces a therapeutic effect. Generally, out of one hundred percent, this amount will range from about 1 percent to about 99 percent of active ingredient, preferably from about 5 percent to about 70 percent, most preferably from about 10 percent to about 30 percent. Pharmaceutical compositions of this disclosure suitable for parenteral administration comprise compounds of the disclosure in combination with one or more pharmaceutically acceptable sterile isotonic aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, or sterile powders which may be reconstituted into sterile injectable solutions or dispersions just prior to use, which may contain antioxidants, buffers, bacteriostats, solutes which render the formulation isotonic with the blood of the intended recipient or suspending or thickening agents.
In some embodiments, a compound of the present disclosure, such as a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1), is provided as a composition in combination with an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir). For example, a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) may be prepared as a fixed dose composition in combination with or the antiviral agent (e.g., a compound of Formula (11), e.g., remdesivir). The fixed dose composition may be formulated for oral administration, e.g., as a solid dosage form or a liquid dosage form. In some embodiments, the liquid dosage form comprises a suspension, a solution, a linctus, an emulsion, a drink, an elixir, or a syrup. In some embodiments, the solid dosage form comprises a capsule, tablet, dragée, or powder.
The combination therapy described herein may involve formulation of the component agents for different routes of administration or for the same route of administration. For example, both the biflavonoid compound and the antiviral agent (e.g., remdesivir) may be formulated for oral administration. In some embodiments, both the biflavonoid compound and the antiviral agent (e.g., remdesivir) may be formulated for intraperitoneal administration. In another embodiment, the biflavonoid compound is formulated for oral administration and the antiviral agent (e.g., remdesivir) is formulated for parenteral administration. In another embodiment, the biflavonoid compound is formulated for oral administration and the antiviral agent (e.g., remdesivir) is formulated for intravenous administration. In another embodiment, the biflavonoid compound is formulated for oral administration and the antiviral agent (e.g., remdesivir) is formulated for intraperitoneal administration. In another embodiment, the biflavonoid compound is formulated for parenteral administration and the antiviral agent (e.g., remdesivir) is formulated for oral administration. In another embodiment, the biflavonoid compound is formulated for intraperitoneal administration and the antiviral agent (e.g., remdesivir) is formulated for parenteral administration. In another embodiment, the biflavonoid compound is formulated for intraperitoneal administration and the antiviral agent (e.g., remdesivir) is formulated for intravenous administration. In another embodiment, the biflavonoid compound is formulated for intraperitoneal administration and the antiviral agent (e.g., remdesivir) is formulated for intraperitoneal administration. In an embodiment, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are formulated as a fixed dose combination (e.g., as a liquid dosage form or solid dosage form, e.g., a capsule or tablet). In some embodiments, the biflavonoid compound and the antiviral agent (e.g., remdesivir) are formulated as a fixed dose combination (e.g., as a liquid dosage form or solid dosage form, e.g., a capsule or tablet) for oral administration.
Examples of suitable aqueous and nonaqueous carriers that may be employed in the pharmaceutical compositions of the disclosure include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.
These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the action of microorganisms may be ensured by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents, such as sugars, sodium chloride, and the like into the compositions. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents that delay absorption such as aluminum monostearate and gelatin. In some cases, in order to prolong the effect of a compound of the present disclosure (e.g., a biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir)) it may be desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material having poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered form of the compound of the present disclosure is accomplished by dissolving or suspending compound in an oil vehicle.
In some embodiments, it may be advantageous to administer the biflavonoid compound (e.g., a compound of Formula (I) or in Table 1) or the antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) of the present disclosure in a sustained fashion. It will be appreciated that any formulation that provides a sustained absorption profile may be used. In certain embodiments, sustained absorption may be achieved by combining a compound of the present disclosure with other pharmaceutically acceptable ingredients, diluents, or carriers that slow its release properties into systemic circulation.
The biflavonoid compounds and antiviral agents, as well as other agents and related compositions thereof used in the methods described herein may be administered to a subject in a variety of forms depending on the selected route of administration, as will be understood by those skilled in the art. Exemplary routes of administration of the compositions used in the methods described herein include topical, enteral, or parenteral applications. Topical applications include but are not limited to epicutaneous, inhalation, enema, eye drops, ear drops, and applications through mucous membranes in the body. Enteral applications include oral administration, rectal administration, vaginal administration, and gastric feeding tubes. Parenteral administration includes intravenous, intraarterial, intracapsular, intraorbital, intracardiac, intradermal, transtracheal, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, intrastemal, intraperitoneal, subcutaneous, intramuscular, transepithelial, nasal, intrapulmonary, intrathecal, rectal, and topical modes of administration. Parenteral administration may be by continuous infusion over a selected period of time.
In certain embodiments of the disclosure, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) are administered orally. In other embodiments of the disclosure, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) are administered intravenously. In certain embodiments of the disclosure, the compositions described herein comprising an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) are administered orally. In other embodiments of the disclosure, the compositions described herein comprising an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) are administered intravenously.
In an embodiment, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered orally in combination with an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir). In an embodiment, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered orally prior to or after oral administration of an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir).
In other embodiments of the disclosure, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered parenterally (e.g., intraperitoneally). In an embodiment, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered parenterally in combination with an antiviral agent (e.g., a compound of Formula (11), e.g., remdesivir). In an embodiment, the compositions described herein comprising a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered parenterally prior to or after oral administration of an antiviral agent (e.g., a compound of Formula (11), e.g., remdesivir).
For intravenous, intraperitoneal, or intrathecal delivery or direct injection, the composition must be sterile and fluid to the extent that the composition is deliverable by syringe. In addition to water, the carrier can be an isotonic buffered saline solution, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyetheylene glycol, and the like), and suitable mixtures thereof. Proper fluidity can be maintained, for example, by use of coating such as lecithin, by maintenance of required particle size in the case of dispersion and by use of surfactants. In many cases, it is preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol or sorbitol, and sodium chloride in the composition. Long-term absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate or gelatin.
The choice of the route of administration will depend on whether a local or systemic effect is to be achieved. For example, for local effects, the composition can be formulated for topical administration and applied directly where its action is desired. For systemic, long-term effects, the composition can be formulated for enteral administration and given via the digestive tract. For systemic, immediate and/or short-term effects, the composition can be formulated for parenteral administration and given by routes other than through the digestive tract.
The compositions of the biflavonoid compounds and antiviral agents as described herein may be formulated into acceptable dosage forms by conventional methods known to those of skill in the art. Actual dosage levels of the active ingredients in the compositions of the present disclosure (e.g., a biflavonoid compound or an antiviral agent) may be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular subject, composition, and mode of administration, without being toxic to the subject. The selected dosage level will depend upon a variety of pharmacokinetic factors including the activity of the particular compositions of the present disclosure employed, the route of administration, the time of administration, the rate of absorption of the particular agent being employed, the duration of the treatment, other drugs, substances, and/or materials used in combination with the particular compositions employed, the age, sex, weight, condition, general health and prior medical history of the subject being treated, and like factors well known in the medical arts. A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the composition required. For example, the physician or veterinarian can start doses of the biflavonoid compounds and/or antiviral agents of the disclosure employed in the composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. In general, a suitable daily dose of a composition of the present disclosure will be that amount of the substance which is the lowest dose effective to produce a therapeutic effect. Such an effective dose will generally depend upon the factors described herein. Preferably, the effective daily dose of a therapeutic composition may be administered as two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms.
Preferred therapeutic dosage levels are between about 0.05 mg/kg to about 1000 mg/kg (e.g., about 0.1 mg/kg, about 0.2 mg/kg, 0.5 mg/kg, 1.0 mg/kg, 1.5 mg/kg, 2 mg/kg, 3 mg/kg, 4 mg/kg, 5 mg/kg, 10 mg/kg, 15 mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg, 45 mg/kg, 50 mg/kg, 60 mg/kg, 70 mg/kg, 80 mg/kg, 90 mg/kg, 100 mg/kg, 125 mg/kg, 150 mg/kg, 175 mg/kg, 200 mg/kg, 250 mg/kg, 300 mg/kg, 350 mg/kg, 400 mg/kg, 450 mg/kg, 500 mg/kg, 600 mg/kg, 700 mg/kg, 800 mg/kg, 900 mg/kg, or 1000 mg/kg) of the composition per day administered (e.g., orally or via injection, e.g., intravenous or intraperitoneal injection) to a subject afflicted with a disease or disorder described herein (e.g., a viral infection). Preferred prophylactic dosage levels are between about 0.05 mg/kg to about 1000 mg/kg (e.g., about 0.1 mg/kg, about 0.2 mg/kg, 0.5 mg/kg, 1.0 mg/kg, 1.5 mg/kg, 2 mg/kg, 3 mg/kg, 4 mg/kg, 5 mg/kg, 10 mg/kg, 15 mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg, 45 mg/kg, 50 mg/kg, 60 mg/kg, 70 mg/kg, 80 mg/kg, 90 mg/kg, 100 mg/kg, 125 mg/kg, 150 mg/kg, 175 mg/kg, 200 mg/kg, 250 mg/kg, 300 mg/kg, 350 mg/kg, 400 mg/kg, 450 mg/kg, 500 mg/kg, 600 mg/kg, 700 mg/kg, 800 mg/kg, 900 mg/kg, or 1000 mg/kg) of the composition per day administered (e.g., orally or via injection) to a subject. The dose may also be titrated (e.g., the dose may be escalated gradually until signs of toxicity appear, such as headache, diarrhea, or nausea). In some embodiments, a loading dose (e.g., an initial high dose) may be administered prior to administration of a maintenance dose (e.g., a comparatively lower dose).
The frequency of treatment may also vary. The subject can be treated one or more times per day (e.g., once, twice, three, four or more times) or every so-many hours (e.g., about every 2, 4, 6, 8, 12, or 24 hours). The biflavonoid compound and/or antiviral agent concentration can be administered 1 or 2 times per 24 hours. The time course of treatment may be of varying duration, e.g., for two, three, four, five, six, seven, eight, nine, ten, or more days, two weeks, 1 month, 2 months, 4 months, 6 months, 8 months, 10 months, or more than one year. For example, the treatment can be twice a day for three days, twice a day for seven days, twice a day for ten days. Treatment cycles can be repeated at intervals, for example weekly, bimonthly or monthly, which are separated by periods in which no treatment is given. The treatment can be a single treatment or can last as long as the life span of the subject (e.g., many years).
In some embodiments, the dosage of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is between about 5 mg/kg to about 100 mg/kg (e.g., about 5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 30 mg/kg, about 40 mg/kg, about 50 mg/kg, about 60 mg/kg, about 70 mg/kg, about 80 mg/kg, about 90 mg/kg, or about 100 mg/kg). In some embodiments, the dosage of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) between about 10 mg/kg to about 50 mg/kg (e.g., about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 25 mg/kg, about 30 mg/kg, about 35 mg/kg, about 40 mg/kg, about 45 mg/kg, or about 50 mg/kg).
In some embodiments, the dosage of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is about 0.1 mg to about 5 mg (e.g., about 0.1 mg, about 0.2 mg, about 0.3 mg, about 0.4 mg, about 0.5 mg, about 0.6 mg, about 0.7 mg, about 0.8 mg, about 0.9 mg, about 1 mg, about 1.25 mg, about 1.5 mg, about 1.75 mg, about 2 mg, about 2.5 mg, about 3 mg, about 3.5 mg, about 4 mg, about 4.5 mg, or about 5 mg). In some embodiments, the dosage of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is between about 0.01 mg/kg to about 10 mg/kg (e.g., about 0.01 mg/kg, about 0.025 mg/kg, about 0.05 mg/kg, about 0.1 mg/kg, about 0.5 mg/kg, about 1 mg/kg, about 2 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg, or about 10 mg/kg). In some embodiments, the dosage of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is between about 0.1 mg/kg to about 5 mg/kg (e.g., about 0.1 mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.4 mg/kg, about 0.5 mg/kg, about 0.6 mg/kg, about 0.7 mg/kg, about 0.8 mg/kg, about 0.9 mg/kg, about 1 mg/kg, about 1.25 mg/kg, about 1.5 mg/kg, about 1.75 mg/kg, about 2 mg/kg, about 2.5 mg/kg, about 3 mg/kg, about 3.5 mg/kg, about 4 mg/kg, about 4.5 mg/kg, or about 5 mg/kg).
In some embodiments, a course of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is between about 1 day to about 24 weeks. In some embodiments, the course of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered at least weekly (e.g., once a week, twice a week, three times a week, four times a week, five times a week, six times a week, 7 times a week) throughout a course of treatment. In some embodiments, the course of a biflavonoid compound (e.g., a compound of Formula (I) or a compound of Table 1) is administered daily throughout a course of treatment.
In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is between about 5 mg/kg to about 100 mg/kg (e.g., about 5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 30 mg/kg, about 40 mg/kg, about 50 mg/kg, about 60 mg/kg, about 70 mg/kg, about 80 mg/kg, about 90 mg/kg, or about 100 mg/kg). In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is between about 10 mg/kg to about 50 mg/kg (e.g., about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 25 mg/kg, about 30 mg/kg, about 35 mg/kg, about 40 mg/kg, about 45 mg/kg, or about 50 mg/kg). In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is about 50 mg/kg.
In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is about 0.1 mg to about 5 mg (e.g., about 0.1 mg, about 0.2 mg, about 0.3 mg, about 0.4 mg, about 0.5 mg, about 0.6 mg, about 0.7 mg, about 0.8 mg, about 0.9 mg, about 1 mg, about 1.25 mg, about 1.5 mg, about 1.75 mg, about 2 mg, about 2.5 mg, about 3 mg, about 3.5 mg, about 4 mg, about 4.5 mg, or about 5 mg). In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is between about 0.01 mg/kg to about 10 mg/kg (e.g., about 0.01 mg/kg, about 0.025 mg/kg, about 0.05 mg/kg, about 0.1 mg/kg, about 0.5 mg/kg, about 1 mg/kg, about 2 mg/kg, about 3 mg/kg, about 4 mg/kg, about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9 mg/kg, or about 10 mg/kg). In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) is between about 0.1 mg/kg to about 5 mg/kg (e.g., about 0.1 mg/kg, about 0.2 mg/kg, about 0.3 mg/kg, about 0.4 mg/kg, about 0.5 mg/kg, about 0.6 mg/kg, about 0.7 mg/kg, about 0.8 mg/kg, about 0.9 mg/kg, about 1 mg/kg, about 1.25 mg/kg, about 1.5 mg/kg, about 1.75 mg/kg, about 2 mg/kg, about 2.5 mg/kg, about 3 mg/kg, about 3.5 mg/kg, about 4 mg/kg, about 4.5 mg/kg, or about 5 mg/kg).
In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) comprises a loading dose of between about 50 mg to about 300 mg on day 1 followed by maintenance doses of between about 10 to about 100 mg. In some embodiments, the dosage of an antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir) comprises a loading dose of about 200 mg on day 1 followed by maintenance doses of about 100 mg. In some embodiments, the dose schedule does not comprise a loading dose of the antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir). In some embodiments, the dose schedule comprises at least one maintenance dose of the antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir). In some embodiments, the dose schedule comprises more than one maintenance dose of the antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir). In some embodiments, the dose schedule is maintained until the subject experiences alleviation or amelioration of one or more symptoms described herein, as determined by a physician. In some embodiments, the dose schedule comprises daily administration of the antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir). In some embodiments, the dose schedule comprises periodic (e.g., every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, weekly, biweekly, monthly, bimonthly, or yearly) administration of the antiviral agent (e.g., a compound of Formula (II) e.g., remdesivir).
In some embodiments, a course of an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) is between about 1 day to about 24 weeks. In some embodiments, the course of antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) is administered at least weekly (e.g., once a week, twice a week, three times a week, four times a week, five times a week, six times a week, 7 times a week) throughout a course of treatment. In some embodiments, the course of an antiviral agent (e.g., a compound of Formula (II), e.g., remdesivir) is administered daily throughout a course of treatment.
The methods of the present disclosure described herein entail administration of a combination of a biflavonoid compound and an antiviral agent (e.g., remdesivir) for the treatment of a viral infection (e.g., Covid-19). Accordingly, a patient and/or subject can be selected for treatment using a biflavonoid compound and an antiviral agent (e.g., remdesivir) for the treatment of a viral infection (e.g., Covid-19) by first evaluating the patient and/or subject to determine whether the subject is infected with a viral infection (e.g., Covid-19) and determination of the serotypic and genotypic classification of the virus. A subject can be evaluated as infected with a viral infection using methods known in the art. The subject can also be monitored, for example, subsequent to administration of a compound described herein (e.g., a biflavonoid compound and an antiviral agent (e.g., remdesivir)) or a pharmaceutically acceptable salt thereof.
In some embodiments, the subject is a mammal. In some embodiments, the subject is a human. In some embodiments, the subject is an adult. In some embodiments, the subject has an acute form of a viral infection. In some embodiments, the subject has a chronic form of a viral infection. In some embodiments, the subject has been diagnosed with a viral infection.
In some embodiments, the subject has a coronavirus infection (SARS-CoV-2 infection), e.g., Covid-19. In some embodiment, the subject has been diagnosed with a coronavirus infection, e.g., Covid-19. In some embodiments, the subject has Covid-19 and the classification of the Covid-19 infection is known. Exemplary classes of SARS-CoV-2 viral infections include alpha (B.1.1.7 or Q lineage), beta (B.1.351 or a descendant lineage), gamma (P.1 or a descendant lineage), delta (B.1.617.2 or AY lineage), epsilon (B.1.427 or B.1.429 lineage), eta (B.1.525 lineage), iota (B.1.526 lineage), kappa (B.1.617.1 lineage), 1.617.3, mu (B.1.621 or B.1.621.1 lineage), omicron (B.1.1.529, BA.1, BA.1.1, BA.2, BA.3, BA.4, or BA.5 lineage) and zeta (P.2 lineage). In some embodiments, the subject is infected with SARS CoV-2 alpha (B.1.1.7 or Q lineage). In some embodiments, the subject is infected with SARS CoV-2 beta (B.1.351 or a descendant lineage). In some embodiments, the subject is infected with SARS CoV-2 gamma (P.1 or a descendant lineage). In some embodiments, the subject is infected with SARS CoV-2 delta (B.1.617.2 or AY lineage). In some embodiments, the subject is infected with SARS CoV-2 epsilon (B.1.427 or B.1.429 lineage). In some embodiments, the subject is infected with SARS CoV-2 eta (B.1.525 lineage). In some embodiments, the subject is infected with SARS CoV-2 iota (B.1.526 lineage). In some embodiments, the subject is infected with SARS CoV-2 kappa (B.1.617.1 lineage). In some embodiments, the subject is infected with SARS CoV-2 mu (B.1.621 or B.1.621.1 lineage). In some embodiments, the subject is infected with SARS CoV-2 omicron (B.1.1.529, BA.1, BA.1.1, BA.2, BA.3, BA.4, or BA.5 lineage). In some embodiments, the subject is infected with SARS CoV-2 zeta (P.2 lineage).
In some embodiments, the subject is treatment naïve. In some embodiments, the subject has previously been treated for a coronavirus (e.g., SARS CoV-2) infection. For example, the subject may have received a vaccine or other coronavirus therapy. In some embodiments, the subject is suffering from a relapsed coronavirus (e.g., SARS CoV-2) infection. In some embodiments, the subject has been treated with an anti-coronavirus (e.g., SARS CoV-2) agent other than a biflavonoid compound or antiviral agent (e.g., remdesivir) described herein and is suffering from a relapsed coronavirus (e.g., SARS CoV-2) infection. In some embodiments, the subject has been treated with a nucleoside analog, a non-nucleoside antiviral, or an immune enhancer and is suffering from a relapsed coronavirus (e.g., SARS CoV-2) infection. In some embodiments, the subject has been treated with a nucleoside analog, e.g., lamivudine, adefovir dipivoxil, entecavir, telbivudine, clevudine, ribavarin, tenofovir, tenofovir alafenamide, besifovir, or AGX-1009, and is suffering from a relapsed coronavirus (e.g., SARS CoV-2) infection. In some embodiments, the subject has been treated with a non-nucleoside antiviral agent, e.g., NOV-225, BAM 205, Myreludex B, ARC-520, BAY 41-4109, REP 9AC, Alinia (nitazoxanide), Dd-RNAi, NVR-121 (NVR 3-778), BSBI-25, NVP-018, TKM-HBV, or ALN-HBV, and is suffering from a relapsed HBV infection. In some embodiments, the subject has been treated with an immune enhancer, e.g., zadaxin (thymosin alpha-1), GS-4774, CYT107 (interleukin-7), Dv-601, HBV core antigen vaccine, or GS-9620, and is suffering from a relapsed HBV infection.
In some embodiments, the subject has a co-morbidity, such as heart disease, coronary artery disease, a cardiomyopathy, diabetes, obesity, high blood pressure, cancer, cerebrovascular disease, chronic kidney disease, chronic liver disease, cystic fibrosis, an immunodeficiency, and tuberculosis. In some embodiments, the subject has been diagnosed with diabetes (e.g., type 1 diabetes or type 2 diabetes). In some embodiments, the subject has been diagnosed with heart disease, coronary artery disease, a cardiomyopathy, or high blood pressure. In some embodiments, the subject is a smoker. In some embodiments, the subject has a body mass index (BMI) greater than about 30 kg/m2.
In some embodiments, the methods described herein further comprise analyzing or receiving analysis of a sample from the subject at least once prior to the end of treatment. In some embodiments, the blood sample is analyzed for viral load and viral antigen levels. In some embodiments, the blood sample is analyzed for the expression level of a cytokine. In some embodiments, the blood sample is analyzed for the presence of anti-SARS-CoV-2 antibodies.
In some embodiments, the methods described herein further comprise analyzing or receiving analysis of a biopsy specimen from the subject at least once prior to the end of treatment. In some embodiments, the biopsy specimen is analyzed for the levels of viral DNA, viral RNA, and/or viral antigens. In some embodiments, the biopsy specimen is analyzed for the levels of inflammatory biomarkers.
In some embodiments, the Covid-19 vaccine comprises intranasal SARS-CoV-2 vaccine (Altimmune), INO-4800 (Inovio Pharma and Beijing Advaccine Biotechnology Company), APNO1 (APEIRON Biologies), mRNA-1273 vaccine (Moderna and Va CCine Research Center), nucleoside-modified mNRA BNT162b2 todinameran (INN) (Pfizer-BioNTech), adenovirus-based vaccine AZD1222 (recombinant ChAdOx1 adenoviral vector encoding SARS-CoV-2 spike protein antigen, Oxford-AstraZeneca), SARS-CoV-2 Recombinant ChAdOx1 adenoviral vector covishield encoding spike protein antigen (ChAdOx1_nCoV19) (Serum Institute of India), SARS-CoV-2 vaccine (Vero cells), inactivated (lnCoV) (Sinopharm/BIBP), SARS-CoV-2 vaccine (Vero cells), inactivated (Sinovac), recombinant replication-incompetent adenovirus type 26 (Ad26) vector vaccine encoding SARS-CoV-2 spike (S) protein Ad26. COV2. S (Janssen Pharmaceuticals Companies of Johnson & Johnson), adenoviral vector-based Covid-19 vaccine Sputnik V Human (The Gamaleya National Center), recombinant novel coronavirus vaccine Ad5-nCoV (adenovirus type 5 vector) (CanSinoBIO), Peptide antigen vaccine EpiVacCorona (Vector State Research Center of Virology and Biotechnology, Russia), Recombinant Novel Coronavirus Vaccine (CHO) (Zhifei Longcom, China), SARS-CoV-2 vaccine, inactivated (Vero cells) (IMBCAMS, China)), inactivated SARS-CoV-2 vaccine (Vero cells) (Sinopharm/WIBP), Avian coronavirus infectious bronchitis virus (IBV) vaccine (MIGDAL Research Institute), modified horsepox virus vaccine TNX-1800 (Tonix Pharmaceuticals), trimeric S protein (S-trimer)-based recombinant subunit vaccine of SARS-CoV-2 coronavirus (Clover Pharmaceuticals), oral recombinant coronavirus vaccine (Vaxart), (i) whole spike gene of coronavirus or (ii) antigen protein-based linear DNA vaccines of coronavirus proteins (Applied DNA Sciences and Takis Biotech), SARS-Cov-2 coronavirus vaccine NVX-CoV2373 (Novavax), intramuscular vaccine INO-4700 (GLS-5300) (Inovio Pharma and GeneOne Life Science), or any other Covid-19 vaccine known in the art.
In an embodiment, the Covid-19 vaccine is Vaxzevria or Covishield, e.g., as disclosed in Patent Publication No. PCT/GB2012/000467, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Comirnaty, e.g., as disclosed in Patent Publication Nos. WO2021213924 and PCT/US2021/023267, each of which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Johnson & Johnson COVID-19 Vaccine (also known as COVID-19 Vaccine Janssen), e.g., as disclosed in U.S. Patent No. U.S. Ser. No. 11/498,944B2, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Spikevax, e.g., as disclosed in U.S. Patent Publication No. WO2021154763, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is the Sinopharm BIBP COVID-19 vaccine (also known as BBIBP-CorV). In an embodiment, the Covid-19 vaccine is Sputnik V COVID-19 vaccine, e.g., as disclosed in U.S. Patent Publication No. US20220305111A1, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is CoronaVac. In an embodiment, the Covid-19 vaccine is Nuvaxovid or Covovax, e.g., as disclosed in U.S. Patent No. U.S. Ser. No. 11/541,112B2, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Covaxin, e.g., as disclosed in Patent Publication No. WO2021224946A1, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is VLA2001, e.g., as disclosed in U.S. Patent Publication No. US20230038284A1, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is VidPrevtyn Beta. In an embodiment, the Covid-19 vaccine is Sputnik Light. In an embodiment, the Covid-19 vaccine is Convidecia, e.g., as disclosed in Patent Publication No. CN111218459B2, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is the Sinopharm WIBP COVID-19 vaccine. In an embodiment, the Covid-19 vaccine is Abdala. In an embodiment, the Covid-19 vaccine is EpiVacCorona. In an embodiment, the Covid-19 vaccine is Zifivax, e.g., as disclosed in Chinese Patent No. CN106928326B, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Soberana 02. In an embodiment, the Covid-19 vaccine is CoviVac. In an embodiment, the Covid-19 vaccine is QazCovid-in. In an embodiment, the Covid-19 vaccine is Minhai COVID-19 vaccine, e.g., as disclosed in Chinese Patent No. CN112225782B, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is MVC-COV1901, e.g., as disclosed in U.S. Patent Publication No. US20230038284A1, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Corbevax. In an embodiment, the Covid-19 vaccine is COVIran Barekat. In an embodiment, the Covid-19 vaccine is the Chinese Academy of Medical Sciences COVID-19 vaccine. In an embodiment, the Covid-19 vaccine is ZyCoV-D. In an embodiment, the Covid-19 vaccine is FAKHRAVAC or MIVAC. In an embodiment, the Covid-19 vaccine is COVAX-19. In an embodiment, the Covid-19 vaccine is Razi Cov Pars. In an embodiment, the Covid-19 vaccine is Turkovac. In an embodiment, the Covid-19 vaccine is the Sinopharm CNBG COVID-19 vaccine, e.g., as disclosed in Chinese PatentNo. CN113817029B, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Soberana Plus. In an embodiment, the Covid-19 vaccine is CoVLP (also known as Covifenz). In an embodiment, the Covid-19 vaccine is Noora. In an embodiment, the Covid-19 vaccine is SKYCovione. In an embodiment, the Covid-19 vaccine is Walvax, e.g., as disclosed in Patent Publication No. CN113215178A, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is iNCOVACC or BBV154, e.g., as disclosed in Patent Publication No. WO2022038642A1, which is hereby incorporated by reference in its entirety. In an embodiment, the Covid-19 vaccine is Gemcovac or GEMCOVAC-19. In an embodiment, the Covid-19 vaccine is V-01. In an embodiment, the Covid-19 vaccine is IndoVac.
In an embodiment, the subject has received a Covid-19 vaccine (e.g., a Covid-19 vaccine described herein). In some embodiments, the subject has received a Covid-19 vaccine (e.g., a Covid-19 vaccine described herein) about 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 1 year, 2 years, or 3 years prior to administration of a hypericin compound and an antiviral agent described herein. In some embodiments, the subject has received at least one dose (e.g., at least two doses, at least three doses, at least four doses, at least five doses, or more) of a Covid-19 vaccine. In some embodiments, the subject has received multiple doses of the same Covid-19 vaccine. In some embodiments, the subject has received one or more doses of multiple Covid-19 vaccines (e.g., one dose of Covishield, two doses of Spikevax, and/or one dose of Comirnaty).
In some embodiments, additional therapeutic agents may be administered with compositions of the present disclosure for the treatment of a viral infection, such as a coronavirus infection (e.g., Covid-19), or any symptom or associated condition thereof. When combination therapy is employed, the additional therapeutic agent(s) can be administered as a separate formulation or may be combined with any of the compositions described herein.
For example, any of the methods described herein may further comprise the administration of a therapeutically effective amount of an additional agent in conjunction with a biflavonoid compound or an antiviral agent (e.g., remdesivir). Exemplary additional agents include an immune therapy, a vaccine, an anti-inflammatory agent, a pain reliever, a mucolytic agent, a cancer therapy, an antiviral agent, an antifungal agent, an antibacterial agent, a bronchodilator, or a vasodilator.
In some embodiments, the additional agent is an anti-inflammatory agent. For example, the anti-inflammatory agent may be an angiotensin-converting enzyme 2 (ACE-2) inhibitor (e.g., lisinopril, benazepril, captopril, enalapril, fosinopril, moexipril, perindopril, or quinapril), a corticosteroid (e.g., cortisone, prednisone, prednisolone, methylprednisolone, dexamethasone, betamethasone, or hydrocortisone) or a non-steroidal anti-inflammatory drug (NSAID) (e.g., ibuprofen, naproxen, diclofenac, celecoxib, mefenamic acid, etoricoxib, or indomethacin).
In some embodiments, the additional agent is a cancer therapy. In some embodiments, the cancer therapy agent is selected from methotrexate, 5-fluorouracil, doxorubicin, vincristine, bleomycin, vinblastine, dacarbazine, toposide, cisplatin, epirubicin, and sorafenib tosylate.
In some embodiments, the additional agent is a second antiviral agent. In some embodiments, the antiviral agent comprises an interferon, a nucleoside analog, a non-nucleoside antiviral, or a non-interferon immune enhancer. In some embodiments, the additional antiviral agent comprises penciclovir, peramivir, oseltamivir, lamivudine, adefovir dipivoxil, entecavir, telbivudine, clevudine, ribavarin, tenofovir, besifovir, AGX-1009, NOV-225, BAM 205, Myrcludex B, ARC-520, BAY 41-4109, REP 9AC, Alinia (nitazoxanide), Dd-RNAi, NVR-121 (NVR 3-778), BSBI-25, NVP-018, TKM-HBV, ALN-HBV, zadaxin (thymosin alpha-1), GS-4774, CYT107 (interleukin-7), Dv-601, or GS-9620.
In some embodiments, the additional agent is a nucleoside analog, a non-nucleoside antiviral, or an immune enhancer. In some embodiments, the nucleoside analog is selected from lamivudine, adefovir dipivoxil, entecavir, telbivudine, clevudine, ribavarin, tenofovir, tenofovir alafenamide, besifovir, and AGX-1009. In some embodiments, the additional agent is a non-nucleoside antiviral agent. In some embodiments, the non-nucleoside antiviral agent is selected from NOV-225, BAM 205, Myrcludex B, ARC-520, BAY 41-4109, REP 9AC, Alinia (nitazoxanide), Dd-RNAi, NVR-121 (NVR 3-778), BSBI-25, and NVP-018. In some embodiments, the additional agent is an immune enhancer. In some embodiments, the immune enhancer is selected from zadaxin (thymosin alpha-1), GS-4774, CYT107 (interleukin-7), and Dv-601.
In some embodiments, the additional agent may be an agent that is used to treat symptoms of a co-morbidity. For example, in some embodiments, the additional agent is an agent that is used to treat symptoms of a co-morbidity selected from heart disease, coronary artery disease, a cardiomyopathy, diabetes, obesity, high blood pressure, HIV infection, cancer, cerebrovascular disease, chronic kidney disease, chronic liver disease, chronic lung disease, cystic fibrosis, an immunodeficiency, neuropathy, neurodegenerative disease, a pulmonary condition, an autoimmune disease, and tuberculosis. In an embodiment, the chronic lung disease may comprise asthma, bronchiectasis, bronchopulmonary dysplasia, chronic obstructive pulmonary disease (COPD), emphysema, chronic bronchitis, interstitial lung disease (e.g., idiopathic pulmonary fibrosis), pulmonary embolism, and pulmonary hypertension. In some embodiments, the additional agent is an agent that is known to treat a co-morbidity described herein. In some embodiments, the additional agent is an agent that is not commonly used to treat a co-morbidity described herein, but is determined by a skilled practitioner to be useful for treating a symptom of a co-morbidity described herein.
In some embodiments, the additional agent is an agent that is used to treat symptoms of other conditions or disorders, including but not limited to digestive system conditions (e.g., GI distress such as abdominal pain, diarrhea, nausea, vomiting, or loss of appetite), respiratory issues, memory loss, and abnormal blood clotting.
In some embodiments, the additional agent is an agent that is used to treat Covid-19. In some embodiments, the agent that is used to treat Covid-19 is a vaccine described herein. In some embodiments, the agent that is used to treat Covid-19 is an oral antiviral Covid-19 treatment (e.g., Paxlovid (nirmatrelvir and ritonavir), Lagevrio (molnupiravir), Olumiant (baricitinib), or Evusheld (tixagevimab and cilgavimab)). In some embodiments, the agent that is used to treat Covid-19 is a subcutaneous Covid-19 treatment (e.g., Anakinra (Kineret)). In some embodiments, the agent that is used to treat Covid-19 is an intravenous Covid-19 treatment (e.g., Gohibic (Vilobelimab), bebtelovimab, bamlanivimab and etesevimab, REGEN-COV (casirivimab and imdevimab), or sotrovimab). In some embodiments, the agent that is used to treat Covid-19 is a corticosteroid. In some embodiments, the additional agent comprises a known combination of agents that is used to treat Covid-19, e.g., a combination of remdesivir and dexamethasone, a combination of remdesivir and baricitinib, a combination of remdesivir and tocilizumab, a combination of dexamethasone and baricitinib, a combination of dexamethasone and tocilizumab, a combination of heparin and dexamethasone, a combination of heparin and remdesivir, or a combination of one or more antiviral Covid-19 agents listed above.
Administration in combination can proceed by any technique apparent to those of skill in the art including, for example, separate, sequential, concurrent, and alternating administration.
The disclosure herein is further presented as a non-limiting list of numbered embodiments.
1. A method of treating a viral infection in a subject, comprising providing a combination of a biflavonoid compound and an antiviral agent to the subject,
2. The method of embodiment 1, wherein the efficacy of the combination is greater than: (i) the efficacy of the biflavonoid compound alone at the molar amount used in the combination; or
3. The method of any one of embodiments 1-2, comprising (i).
4. The method of any one of embodiments 1-3, comprising (ii).
5. The method of any one of the preceding embodiments, wherein the efficacy of the combination is at least X1-fold greater than the efficacy of the biflavonoid compound alone at the molar amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
6. The method of any one of the preceding embodiments, wherein the efficacy of the combination is at least X1-fold greater than the efficacy of the antiviral agent alone at the molar amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
7. The method of any one of the preceding embodiments, wherein providing the combination comprises administering the combination to the subject.
8. The method of any one of the preceding embodiments, wherein the molar amount of the biflavonoid compound in the combination comprises the molar concentration of the biflavonoid compound in the combination.
9. The method of any one of the preceding embodiments, wherein the molar amount of the antiviral agent in the combination comprises the molar concentration of the antiviral agent in the combination.
10. The method of any one of the preceding embodiments, wherein each of the biflavonoid compound and the antiviral agent is independently formulated as a pharmaceutical composition.
11. The method of any one of the preceding embodiments, wherein the biflavonoid compound and the antiviral agent are formulated together as a pharmaceutical composition.
12. The method of any one of the preceding embodiments, wherein each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) concomitantly to the subject.
13. The method of any one of embodiments 1-11, wherein each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) sequentially to the subject.
14. The method of embodiment 13, wherein the biflavonoid compound is provided (e.g., administered) to the subject prior to the antiviral agent.
15. The method of embodiment 13, wherein the antiviral agent is provided (e.g., administered) to the subject prior to the biflavonoid compound.
16. The method of any one of the preceding embodiments, wherein the biflavonoid compound is a compound of Formula (I):
17. The method of embodiment 16, wherein R1 is hydrogen or C1-C6 alkyl.
18. The method of any one of embodiments 16-17, wherein each of R2 and R3 is independently hydrogen or C1-C6 alkyl.
19. The method of any one of embodiments 16-18, wherein each of R4 and R5 is independently hydrogen or C1-C6 alkyl.
20. The method of any one of embodiments 16-19, wherein R6 is hydrogen or C1-C6 alkyl.
21. The method of any one of embodiments 1-15, wherein the biflavonoid compound is a compound of Formula (I-a):
22. The method of embodiment 21, wherein one of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
23. The method of any one of embodiments 21-22, wherein two of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
24. The method of any one of embodiments 21-23, wherein three of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
25. The method of any one of embodiments 21-24, wherein four of R19, R20, R21, R22, R23, and R243 is independently hydrogen or C1-C6 alkyl.
26. The method of any one of embodiments 21-25, wherein five of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
27. The method of any one of embodiments 21-26, wherein each of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
28. The method of any one of the preceding embodiments, wherein the biflavonoid compound is prepared synthetically or is extracted from a natural source (e.g., Ginkgo biloba).
29. The method of embodiment 28, wherein the biflavonoid compound is prepared synthetically.
30. The method of embodiment 28, wherein the biflavonoid compound is extracted from a natural source (e.g., Ginkgo biloba).
31. The method of any one of the preceding embodiments, wherein the biflavonoid compound is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.
32. The method of embodiment 31, wherein the biflavonoid compound is substantially pure.
33. The method of any one of embodiments 31-32, wherein the biflavonoid compound is provided as a pharmaceutical composition and the pharmaceutical composition comprises less than about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% of another compound in listed in Table 1 or a pharmaceutically acceptable salt thereof.
34. The method of any one of the preceding embodiments, wherein the biflavonoid compound is ginkgetin or a pharmaceutically acceptable salt thereof.
35. The method of embodiment 27, wherein the ginkgetin or a pharmaceutically acceptable salt thereof is provided as a pharmaceutical composition, and the pharmaceutical composition comprises less than about 90%, 95%, 99%, or 99.9% of another compound in listed in Table 1 or a pharmaceutically acceptable salt thereof.
36. The method of any one of the preceding embodiments, wherein the antiviral agent comprises a small molecule, antibody, peptide, or oligonucleotide.
37. The method of any one of the preceding embodiments, wherein the antiviral agent is an agent that modulates a step in the viral life cycle.
38. The method of any one of the preceding embodiments, wherein the antiviral agent is selected from the group consisting of an attachment inhibitor, post-attachment inhibitor, fusion inhibitor, entry inhibitor, uncoating inhibitor, protease inhibitor, polymerase inhibitor, nucleotide reverse transcriptase inhibitor, nucleoside reverse transcriptase inhibitor, non-nucleoside reverse transcriptase inhibitor, and integrase inhibitor.
39. The method of any one of the preceding embodiments, wherein the antiviral agent comprises a nucleoside analog or a non-ribosomal peptide.
40. The method of any one of the preceding embodiments, wherein the antiviral agent is compound of Formula (II):
41. The method of embodiment 40, wherein the antiviral agent of Formula (II) is remdesivir.
42. The method of embodiment 40, wherein the antiviral agent of Formula (II) is sofosbuvir.
43. The method of embodiment 40, wherein ring A is heteroaryl substituted with 1-2 R′?.
44. The method of any one of embodiments 40-43, wherein R15 is aryl substituted with 0-1 R17.
45. The method of any one of embodiments 40-44, wherein one of R16a and R16b is hydrogen.
46. The method of any one of embodiments 40-45, wherein one of R16A and R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl.
47. The method of any one of embodiments 40-46, wherein one of R12a and R12b is hydrogen.
48. The method of any one of embodiments 40-47, wherein one of R12a and R12b is —ORA.
49. The method of any one of embodiments 40-48, wherein one of R12a and R12b is halo.
50. The method of any one of embodiments 40-49, wherein one of R13a and R13b is —ORA.
51. The method of any one of embodiments 40-50, wherein one of R13a and R13b is hydrogen.
52. The method of any one of embodiments 40-51, wherein A is heteroaryl substituted with 1 R17; R11 is cyano; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; RA is hydrogen; and R17 is —NH2.
53. The method of any one of embodiments 40-52, wherein A is heteroaryl substituted with 2 R17, R11 is hydrogen; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; and RA is hydrogen; and each R17 is independently oxo.
54. The method of any one of embodiments 1-39, wherein the antiviral agent is a compound of Formula (II-a):
55. The method of any one of embodiments 1-39, wherein the antiviral agent is a compound of Formula (II-b):
56. The method of any one of the preceding embodiments, wherein the antiviral agent is a compound selected from Table 2 or a pharmaceutically acceptable salt thereof.
57. The method of any one of the preceding embodiments, wherein the antiviral agent targets an RNA virus.
58. The method of any one of the preceding embodiments, wherein the antiviral agent is selected from remdesivir, sofosbuvir, or a pharmaceutically acceptable salt thereof.
59. The method of any one of the preceding embodiments, wherein the antiviral agent is selected from remdesivir or a pharmaceutically acceptable salt thereof.
60. The method of any one of the preceding embodiments, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 0.1 M and 500 μM in the subject or a cell.
61. The method of any one of the preceding embodiments, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 1 M and 25 μM in the subject or a cell.
62. The method of any one of the preceding embodiments, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.01 μM and 25 μM in the subject or a cell.
63. The method of any one of the preceding embodiments, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 5 μM in the subject or a cell.
64. The method of any one of the preceding embodiments, wherein the ratio of the amount of biflavonoid compound to the antiviral agent in the combination is between 200:1 to 1:1.
65. The method of embodiment 64, wherein the ratio of the amount of biflavonoid compound to the antiviral agent in the combination is between 50:1 to 1:1.
66. The method of any one of the preceding embodiments, wherein:
67. The method of any one of the preceding embodiments, further comprising administration of an additional agent.
68. The method of any one of the preceding embodiments, wherein the subject is a mammal (e.g., a human).
69. The method of any one of the preceding embodiments, wherein the viral infection is present within a virally infected organ, tissue, or cell in a subject.
70. The method of embodiment 69, wherein the virally infected organ is selected from the group consisting of the brain, spinal cord, eye, skin, lung, heart, pancreas, large intestine, small intestine, stomach, liver, gall bladder, kidney, or spleen.
71. The method of embodiment 69, wherein the virally infected tissue is selected from the group consisting of lung tissue, tracheal tissue, intestinal tissue, skin tissue, pancreatic tissue, vascular tissue, mucosal tissue, kidney tissue, brain tissue, nervous tissue, or cardiac tissue.
72. The method of embodiment 69, wherein the virally infected cell comprises an angiotensin-converting enzyme 2 (ACE2) receptor on the cell surface.
73. The method of embodiment 69, wherein the virally infected cell is selected from the group consisting of an epithelial cell or an endothelial cell.
74. The method of embodiment 69, wherein the virally infected cell is a basal cell, luminal cell, secretory cell, or ciliated cell.
75. The method of any one of embodiments 69, wherein the virally infected cell is a bronchial cell, renal cell, enterocyte, goblet cell, skin cell, islet cell, neuronal cell, glial cell, or heart cell.
76. The method of any one of the preceding embodiments, wherein the viral infection is a coronavirus infection.
77. The method of any one of the preceding embodiments, wherein the viral infection is a SARS coronavirus (SARS-CoV) infection, MERS coronavirus (MERS-CoV) infection, or SARS-CoV-2 infection.
78. The method of any one of the preceding embodiments, wherein the viral infection is a SARS CoV-2 infection.
79. A method of treating a SARS CoV-2 infection in a subject, comprising providing a combination of (i) ginkgetin or a pharmaceutically acceptable salt thereof and (ii) remdesivir or a pharmaceutically acceptable salt thereof to the subject,
80. A method of reducing the toxicity of an antiviral agent in a cell or subject, comprising administering a combination of a biflavonoid compound and the antiviral agent to the subject, wherein the molar amount of the biflavonoid compound in the combination is greater than the molar amount of the antiviral agent, thereby reducing the toxicity of the antiviral agent in the cell or subject.
81. The method of embodiment 80, wherein the efficacy of the combination is greater than:
82. The method of embodiment 81, wherein the reducing the toxicity is relative to the toxicity of the antiviral agent in the cell or subject when provided in the absence of the hypericin compound.
83. The method of embodiment 82, comprising (i).
84. The method of embodiment 82, comprising (ii).
85. The method of any one of embodiments 80-84, wherein the efficacy of the combination is at least X1-fold greater than the efficacy of the biflavonoid compound alone at the molar amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
86. The method of any one of embodiments 80-85, wherein the efficacy of the combination is at least X1-fold greater than the efficacy of the antiviral agent alone at the molar amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
87. The method of any one of embodiments 80-86, wherein the toxicity of the antiviral agent in the cell or subject is reduced upon administration of the antiviral agent in combination with a biflavonoid compound between 1% and 50%, e.g., compared with the toxicity of the antiviral agent when administered in the absence of the biflavonoid compound.
88. The method of any one of embodiments 80-87, wherein the molar amount of the biflavonoid compound in the combination comprises the molar concentration of the biflavonoid compound in the combination.
89. The method of any one of embodiments 80-88, wherein the molar amount of the antiviral agent in the combination comprises the molar concentration of the antiviral agent in the combination.
90. The method of any one of embodiments 80-89, wherein each of the biflavonoid compound and the antiviral agent is independently formulated as a pharmaceutical composition.
91. The method of any one of embodiments 80-90, wherein the biflavonoid compound and the antiviral agent are formulated together as a pharmaceutical composition.
92. The method of any one of embodiments 80-91, wherein each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) concomitantly to the subject.
93. The method of any one of embodiments 80-91, wherein each of the biflavonoid compound and the antiviral agent is provided (e.g., administered) sequentially to the subject.
94. The method of embodiment 93, wherein the biflavonoid compound is provided (e.g., administered) to the subject prior to the antiviral agent.
95. The method of embodiment 93, wherein the antiviral agent is provided (e.g., administered) to the subject prior to the biflavonoid compound.
96. The method of any one of embodiments 80-95, wherein the biflavonoid compound is a compound of Formula (I):
97. The method of embodiment 96, wherein R1 is hydrogen or C1-C6 alkyl.
98. The method of any one of embodiments 96-97, wherein each of R2 and R3 is independently hydrogen or C1-C6 alkyl.
99. The method of any one of embodiments 96-98, wherein each of R4 and R5 is independently hydrogen or C1-C6 alkyl.
100. The method of any one of embodiments 96-99, wherein R6 is hydrogen or C1-C6 alkyl.
101. The method of any one of embodiments 80-95, wherein the biflavonoid compound is a compound of Formula (I-a):
102. The method of embodiment 101, wherein one of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
103. The method of any one of embodiments 101-102, wherein two of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
104. The method of any one of embodiments 101-103, wherein three of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
105. The method of any one of embodiments 101-104, wherein four of R19, R20, R21, R22, R23, and R243 is independently hydrogen or C1-C6 alkyl.
106. The method of any one of embodiments 101-105, wherein five of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
107. The method of any one of embodiments 101-106, wherein each of R19, R20, R21, R22, R23 and R24 is independently hydrogen or C1-C6 alkyl.
108. The method of any one of embodiments 80-107, wherein the biflavonoid compound is prepared synthetically or is extracted from a natural source (e.g., Ginkgo biloba).
109. The method of embodiment 108, wherein the biflavonoid compound is prepared synthetically.
110. The method of embodiment 108, wherein the biflavonoid compound is extracted from a natural source (e.g., Ginkgo biloba).
111. The method of any one of embodiments 80-110, wherein the biflavonoid compound is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.
112. The method of embodiment 111, wherein the biflavonoid compound is substantially pure.
113. The method of any one of embodiments 111-112, wherein the biflavonoid compound is provided as a pharmaceutical composition and the pharmaceutical composition comprises less than about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 99.9% of another compound in listed in Table 1 or a pharmaceutically acceptable salt thereof.
114. The method of any one of embodiments 80-113, wherein the biflavonoid compound is ginkgetin or a pharmaceutically acceptable salt thereof.
115. The method of embodiment 114, wherein the ginkgetin or a pharmaceutically acceptable salt thereof is provided as a pharmaceutical composition, and the pharmaceutical composition comprises less than about 90%, 95%, 99%, or 99.9% of another compound in listed in Table 1 or a pharmaceutically acceptable salt thereof.
116. The method of any one of embodiments 80-115, wherein the antiviral agent is a small molecule, antibody, peptide, or oligonucleotide.
117. The method of any one of embodiments 80-116, wherein the antiviral agent is an agent that modulates a step in the viral life cycle.
118. The method of any one of embodiments 80-117, wherein the antiviral agent selected from an attachment inhibitor, post-attachment inhibitor, fusion inhibitor, entry inhibitor, uncoating inhibitor, protease inhibitor, polymerase inhibitor, nucleotide reverse transcriptase inhibitor, nucleoside reverse transcriptase inhibitor, non-nucleoside reverse transcriptase inhibitor, and integrase inhibitor.
119. The method of any one of embodiments 80-118, wherein the antiviral agent is a nucleoside analog or a non-ribosomal peptide.
120. The method of any one of embodiments 80-119, wherein the antiviral agent is compound of Formula (II):
121. The method of embodiment 120, wherein the antiviral agent of Formula (II) is remdesivir.
122. The method of embodiment 120, wherein the antiviral agent of Formula (II) is sofosbuvir.
123. The method of embodiment 120, wherein ring A is heteroaryl substituted with 1-2 R17.
124. The method of any one of embodiments 120-123, wherein R15 is aryl substituted with 0-1 R17.
125. The method of any one of embodiments 120-124, wherein one of R16a and R16b is hydrogen.
126. The method of any one of embodiments 120-125, wherein one of R16a and R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl.
127. The method of any one of embodiments 120-126, wherein one of R12 and R12b is hydrogen.
128. The method of any one of embodiments 120-127, wherein one of R12a and R12b is —ORA.
129. The method of any one of embodiments 120-128, wherein one of R12a and R12b is halo.
130. The method of any one of embodiments 120-129, wherein one of R13a and R13b is —ORA.
131. The method of any one of embodiments 120-130, wherein one of R13a and R13b is hydrogen.
132. The method of any one of embodiments 120-131, wherein A is heteroaryl substituted with 1 R17; R11 is cyano; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; RA is hydrogen; and R17 is —NH2.
133. The method of any one of embodiments 120-132, wherein A is heteroaryl substituted with 2 R17, R11 is hydrogen; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R1a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; and RA is hydrogen; and each R17 is independently oxo.
134. The method of any one of embodiments 80-119, wherein the antiviral agent is a compound of Formula (II-a):
135. The method of any one of embodiments 80-119, wherein the antiviral agent is a compound of Formula (II-b):
136. The method of any one of embodiments 80-135, wherein the antiviral agent is a compound selected from Table 2 or a pharmaceutically acceptable salt thereof.
137. The method of any one of embodiments 80-136, wherein the antiviral agent targets an RNA virus.
138. The method of any one of embodiments 80-137, wherein the antiviral agent is selected from remdesivir, sofosbuvir, or a pharmaceutically acceptable salt thereof.
139. The method of any one of embodiments 80-138, wherein the antiviral agent is selected from remdesivir or a pharmaceutically acceptable salt thereof.
140. The method of any one of embodiments 80-139, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 0.1 μM and 500 μM in a subject or cell.
141. The method of any one of embodiments 80-140, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 1 μM and 25 μM in a subject or cell.
142. The method of any one of embodiments 80-141, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.01 μM and 25 μM in a subject or cell.
143. The method of any one of embodiments 80-142, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 5 μM in a subject or cell.
144. The method of any one of embodiments 80-143, wherein the ratio of the amount of biflavonoid compound to the antiviral agent in the combination is between 200:1 to 1:1.
145. The method of embodiment 144, wherein the ratio of the amount of biflavonoid compound to the antiviral agent in the combination is between 50:1 to 1:1.
146. The method of any one of embodiments 80-145, wherein:
147. The method of any one of embodiments 80-146, further comprising administration of an additional agent.
148. The method of any one of embodiments 80-147, wherein the subject is a mammal (e.g., a human).
149. The method of any one of embodiments 80-148, wherein the cell comprises an angiotensin-converting enzyme 2 (ACE2) receptor on the cell surface.
150. The method of any one of embodiments 80-149, wherein the cell is a cell present in the respiratory system, nervous system, gastrointestinal system, circulatory system, endocrine system, immune system, or excretory system.
151. The method of any one of embodiments 80-150, wherein the cell is a bronchial cell, renal cell, enterocyte, goblet cell, skin cell, islet cell, neuronal cell, glial cell, or heart cell.
152. The method of any one of embodiments 80-151, wherein the cell or subject is infected with a viral infection.
153. The method of embodiment 152, wherein the viral infection is a SARS coronavirus (SARS-CoV), MERS coronavirus (MERS-CoV), or SARS-CoV-2 infection.
154. The method of any one of embodiments 152-153, wherein the viral infection is a SARS CoV-2 infection.
155. A method of reducing the toxicity of remdesivir in a cell or subject, comprising providing a combination of (i) ginkgetin or a pharmaceutically acceptable salt thereof and (ii) remdesivir or a pharmaceutically acceptable salt thereof to the cell or subject,
156. A method of reducing the virulence of a virus in a subject, comprising administering a combination of a biflavonoid compound and remdesivir to the subject,
157. The method of embodiment 156, wherein reducing the virulence comprises one or more of:
158. The method of any one of embodiments 156-157, wherein the efficacy of the combination is greater than:
159. The method of embodiment 158, comprising (i).
160. The method of embodiment 158, comprising (ii).
161. The method of any one of embodiments 156-160, wherein the efficacy of the combination is at least X1 greater than the efficacy of the biflavonoid compound alone at the amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
162. The method of any one of embodiments 156-161, wherein the efficacy of the combination is at least X1 greater than the efficacy of the antiviral agent alone at the amount used in the combination, wherein X1 is 1, 1.25, 1.5, 1.75, 2, 2.5, or greater.
163. The method of any one of embodiments 156-162, wherein the biflavonoid compound is a compound of Formula (I):
164. The method of embodiment 163, wherein R1 is hydrogen or C1-C6 alkyl.
165. The method of any one of embodiments 163-164, wherein each of R2 and R3 is independently hydrogen or C1-C6 alkyl.
166. The method of any one of embodiments 163-165, wherein each of R4 and R5 is independently hydrogen or C1-C6 alkyl.
167. The method of any one of embodiments 163-166, wherein R6 is hydrogen or C1-C6 alkyl.
168. The method of any one of embodiments 156-162, wherein the biflavonoid compound is a compound of Formula (I-a):
169. The method of embodiment 168, wherein one of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
170. The method of any one of embodiments 168-169, wherein two of R19, R20, R21, R22, R23 and R24 is independently hydrogen or C1-C6 alkyl.
171. The method of any one of embodiments 168-170, wherein three of R19, R20, R21, R22, R23 and R24 is independently hydrogen or C1-C6 alkyl.
172. The method of any one of embodiments 168-171, wherein four of R19, R20, R21, R22, R23, and R243 is independently hydrogen or C1-C6 alkyl.
173. The method of any one of embodiments 168-172, wherein five of R19, R20, R21, R22, R23, and R24 is independently hydrogen or C1-C6 alkyl.
174. The method of any one of embodiments 168-173, wherein each of R19, R20, R21, R22, R23 and R24 is independently hydrogen or C1-C6 alkyl.
175. The method of any one of embodiments 156-174, wherein the biflavonoid compound is prepared synthetically or is extracted from a natural source (e.g., Ginkgo biloba).
176. The method of embodiment 175, wherein the biflavonoid compound is prepared synthetically.
177. The method of embodiment 175, wherein the biflavonoid compound is extracted from a natural source (e.g., Ginkgo biloba).
178. The method of any one of embodiments 156-177, wherein the biflavonoid compound is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.
179. The method of any one of embodiments 156-178, wherein the biflavonoid compound is ginkgetin or a pharmaceutically acceptable salt thereof.
180. The method of any one of embodiments 156-179, wherein the antiviral agent is a small molecule, antibody, peptide, or oligonucleotide.
181. The method of any one of embodiments 156-180, wherein the antiviral agent selected from an attachment inhibitor, post-attachment inhibitor, fusion inhibitor, entry inhibitor, uncoating inhibitor, protease inhibitor, polymerase inhibitor, nucleotide reverse transcriptase inhibitor, nucleoside reverse transcriptase inhibitor, non-nucleoside reverse transcriptase inhibitor, and integrase inhibitor.
182. The method of any one of embodiments 156-181, wherein the antiviral agent is a nucleoside analog or a non-ribosomal peptide.
183. The method of any one of embodiments 156-182, wherein the antiviral agent is compound of Formula (II):
184. The method of embodiment 183, wherein the antiviral agent of Formula (II) is remdesivir.
185. The method of embodiment 183, wherein the antiviral agent of Formula (II) is sofosbuvir.
186. The method of embodiment 183, wherein ring A is heteroaryl substituted with 1-2 R17.
187. The method of any one of embodiments 183-186, wherein R15 is aryl substituted with 0-1 R17.
188. The method of any one of embodiments 183-187, wherein one of R16a and R16b is hydrogen.
189. The method of any one of embodiments 183-188, wherein one of R16a and R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl.
190. The method of any one of embodiments 183-189, wherein one of R12a and R12b is hydrogen.
191. The method of any one of embodiments 183-190, wherein one of R12a and R12b is —ORA.
192. The method of any one of embodiments 183-191, wherein one of R12a and R12b is halo.
193. The method of any one of embodiments 183-192, wherein one of R13a and R13b is —ORA.
194. The method of any one of embodiments 183-193, wherein one of R13a and R13b is hydrogen.
195. The method of any one of embodiments 183-194, wherein A is heteroaryl substituted with 1 R17; R11 is cyano; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; RA is hydrogen; and R17 is —NH2.
196. The method of any one of embodiments 183-195, wherein A is heteroaryl substituted with 2 R17, R11 is hydrogen; X is O; Z is O; R12a is hydrogen; R12b is —ORA; R13a is hydrogen; R13b is —ORA; R15 is aryl; R16a is hydrogen; R16b is —C1-C6 alkylene-C(O)O—C1-C6 alkyl; and RA is hydrogen; and each R17 is independently oxo.
197. The method of any one of embodiments 156-182, wherein the antiviral agent is a compound of Formula (II-a):
198. The method of any one of embodiments 156-182, wherein the antiviral agent is a compound of Formula (II-b):
199. The method of any one of embodiments 156-198, wherein the antiviral agent is a compound selected from Table 2 or a pharmaceutically acceptable salt thereof.
200. The method of any one of embodiments 156-199, wherein the antiviral agent targets an RNA virus.
201. The method of any one of embodiments 156-200, wherein the antiviral agent is selected from remdesivir, sofosbuvir, or a pharmaceutically acceptable salt thereof.
202. The method of embodiment 201, wherein the antiviral agent is selected from remdesivir or a pharmaceutically acceptable salt thereof.
203. The method of any one of embodiments 156-202, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 0.1 μM and 500 μM in a subject or cell.
204. The method of any one of embodiments 156-203, wherein the biflavonoid compound is administered at a dosage to provide a concentration of between 1 μM and 25 μM in a subject or cell.
205. The method of any one of embodiments 156-204, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.01 μM and 25 μM in a subject or cell.
206. The method of any one of embodiments 156-205, wherein the antiviral agent (e.g., remdesivir) is administered at a dosage to provide a concentration of between 0.1 μM and 5 μM in a subject or cell.
207. The method of any one of embodiments 156-206, wherein:
208. The method of any one of embodiments 156-207, wherein each of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is independently formulated as a pharmaceutical composition.
209. The method of any one of embodiments 156-207, wherein the biflavonoid compound and the antiviral agent (e.g., remdesivir) are formulated together as a pharmaceutical composition.
210. The method of any one of embodiments 156-209, wherein each of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is administered concomitantly.
211. The method of any one of embodiments 156-209, wherein each of the biflavonoid compound and the antiviral agent (e.g., remdesivir) is administered sequentially.
212. The method of embodiment 211, wherein the biflavonoid compound is administered prior to the antiviral agent (e.g., remdesivir).
213. The method of embodiment 211, wherein the antiviral agent (e.g., remdesivir) is administered prior to the biflavonoid compound.
214. The method of any one of embodiments 156-213, further comprising administration of an additional agent.
215. The method of any one of embodiments 156-214, wherein the subject is a mammal (e.g., a human).
216. A composition for use in treating a viral infection in a subject, comprising a combination of a biflavonoid compound and an antiviral agent, wherein the molar amount of the biflavonoid compound in the combination is greater than the molar amount of the antiviral agent.
217. A composition for use in treating a SARS CoV-2 infection in a subject, comprising a combination of (i) ginkgetin or a pharmaceutically acceptable salt thereof and (ii) remdesivir or a pharmaceutically acceptable salt thereof,
218. A composition for use in reducing the toxicity of an antiviral agent in a cell or subject, comprising a combination of a biflavonoid compound and the antiviral agent,
219. A composition for use in reducing the toxicity of remdesivir in a cell or subject, comprising a combination of (i) ginkgetin or a pharmaceutically acceptable salt thereof and (ii) remdesivir or a pharmaceutically acceptable salt thereof,
220. A composition for use in reducing the virulence of a virus in a subject, comprising a combination of a biflavonoid compound and remdesivir,
The following examples are provided to further illustrate some embodiments of the present disclosure but are not intended to limit the scope of the disclosure. It will be understood by their exemplary nature that other procedures, methodologies, or techniques known to those skilled in the art may alternatively be used.
This example demonstrates the ability of biflavonoid compounds disclosed herein to act in combination with known an exemplary antiviral agent (e.g., remdesivir) to decrease viral infection-induced cytotoxicity and viral replication of the novel SARS-CoV-2 coronavirus in a model cell line. In particular, the ability of the biflavonoid compounds to synergize with remdesivir at a sub-therapeutic dose of remdesivir (0-2.5 M) to achieve an increase in cell viability over time in cells infected with SARS-CoV-2.
Vero E6 cells were seeded at 25,000 cells/well in 96-well plates and SARS-CoV-2 was added into the wells at a MOI of 0.01. After 1 hour, ginkgetin (Cayman Chemical, >98% purity) was added into the wells at final concentrations of 1.5625-100 μM or the vehicle only control (DMSO), followed by the addition of remdesivir (Novation Chemicals, 98% purity) at final concentrations of 0.3125-5 μM or the vehicle only control (DMSO), and incubated at 37° C.
After 72 hours of incubation, host cell viability was assessed using CellTiter-Glo® Luminescent Cell Viability Assay (Promega) according to the manufacturer's instructions. Results were normalized to the highest dose of remdesivir alone (5 μM) and are summarized below in Table 2. As shown, remdesivir (5 μM) alone dose-dependently improved host cell viability of SARS-CoV-2-infected Vero E6 cells as expected. Similarly, ginkgetin (25-50 μM) alone enabled modest improvements in host cell viability.
However, ginkgetin (1.5625-6.25 μM) demonstrated combinatorial activity with remdesivir. For example, ginkgetin (1.5625-6.25 μM) combined into remdesivir (1.25 or 2.5 μM) improved host cell viability up to >90%, which is approximately 3- or 2.5-fold compared to remdesivir (1.25 or 2.5 μM) alone which only had host cell viability of 29 or 39%, respectively (Table 2). This demonstrates that it is possible to nearly achieve the efficacy of 5 μM remdesivir, with half (2.5 μM remdesivir) or one-third (1.25 μM remdesivir) of the dose by combining ginkgetin.
Similarly, remdesivir can increase the efficacy of ginkgetin. Ginkgetin (12.5-100 μM) alone enabled modest improvements in host cell viability up to 40% at 100 μM. For example, remdesivir (0.3125-5 μM) can combine with ginkgetin (1.5625-6.25 μM) to increase viability to 88-93%, compared to the 18-24% viability seen at 1.5625-6.25 μM of ginkgetin alone.
Next, median tissue culture infectious dose (TCID50) assay was used to assess the viral titer in treated cells. Remdesivir alone dose-dependently reduced viral titers of SARS-CoV-2 to zero TCID50/mL, compared to control conditions without Remdesivir which maintained viral titers around 10≢TCID50/mL (
B6.Cg-Ceslctm1.1Loc/J mice were infected with 1×10{circumflex over ( )}5 particles of SARS-CoV-2 virus [B.1.351] via intranasal administration. Mice were treated daily with vehicle or with remdesivir (5, 10 or 25 mg/kg) intraperitoneally starting day 1 post infection. At day 4 post infection, all mice were euthanized. At that time point, lungs were collected and stored at −80° C. Lung tissues were processed for determination of infectious titers (plaque forming unit per mg of tissues). The results are summarized in
In this example, the efficacy and safety/tolerability of ginkgetin and remdesivir were investigated in mice infected with SARS CoV-2 virus. In brief, mice (10-12 weeks old) were infected with 1×10{circumflex over ( )}5 particles of SARS-CoV-2 virus [B.1.351] via intranasal administration. For mice receiving ginkgetin, the mice were pre-treated with ginkgetin starting two days prior to infection. Post-infection, the mice were administered remdesivir or other treatment starting day 1 post infection daily via TP as follows: (1) negative vehicle control (saline), (2) Remdesivir (25 mg/kg), (3) Remdesivir (10 mg/kg), (4) Ginkgetin (20 mg/kg), (5) Ginkgetin (20 mg/kg)+Remdesivir (10 mg/kg). Each group contained 7 mice. Survival, body weight, temperature, and scoring (symptoms, disease features, etc.) of mice were tracked and monitored daily. At day 4 post infection, all mice were euthanized. At that time point, tissues (lung) as well as serum were collected and stored at −80° C. Pieces of lung, heart, and brain tissues were also fixed in in PFA (4%). Lung tissues were processed for determination of infectious titers (plaque forming unit per 10 mg of tissues).
The results of the experiments are summarized in
The entire contents of all patents, published applications, and references cited herein are incorporated by reference in their entirety. While example embodiments have been particularly shown and described, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the embodiments encompassed by the appended claims.
The present application claims priority to U.S. Provisional Application No. 63/354,493, filed Jun. 22, 2022; and U.S. Provisional Application No. 63/394,184, filed Aug. 1, 2022; the contents of each of which are incorporated herein by reference in their entirety.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US2023/026026 | 6/22/2023 | WO |
| Number | Date | Country | |
|---|---|---|---|
| 63354493 | Jun 2022 | US | |
| 63394184 | Aug 2022 | US |
