Methods for treating veterinary viral diseases

Abstract

1. A method for preventing or treating a viral disease in an animal comprising administering to the animal a pharmaceutical composition comprising the antiviral nucleoside β-DN(4)-hydroxycytidine (NHC) or a prodrug or salt thereof and a pharma- ceutically acceptable carrier, wherein the viral disease is porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency.

Description

Technical Field

[0001] The present invention relates to antiviral nucleoside compounds and pharmaceutical compositions for use in the treatment or prevention of viral infections and viral diseases in animals.

Background Art

[0002] The development of antiviral drugs for veterinary use has been restricted by the need to identify specific viral targets with high selectivity and few side effects. As a result, few antiviral drugs for veterinary use have been developed. See Dal Pozzo & Thiry, Rev. sci. tech. Off. int. Epiz., 2014, 33(3), 791 - 801.

[0003] Porcine reproductive and respiratory syndrome (PRRS) is a viral disease characterized by two overlapping clinical symptoms, reproductive dysfunction or failure in breeding animals, and respiratory diseases in pigs of any age. PRRS is the most economically important disease affecting swine production in the United States. There is no specific treatment for PRRS. Broad - spectrum antibiotics may be useful for controlling secondary infections. Anti - inflammatory products (e.g., aspirin) are generally administered during acute disease. Other useful techniques include early weaning and isolation of piglets, various PRRS vaccination protocols, regular serological monitoring, tests (ELISA, PCR and IFA), and removal of persistent carriers in groups with improved biosecurity and an infection rate of less than 10%. (Accessed on April 22, 2021, Swine Manual, Iowa State University, College of Veterinary Medicine, Swine Manual, https: / / vetmed.iastate.edu / vdpam / FSVD / swine / index - diseases / porcine - reproductive).

[0004] Bovine Respiratory Disease (BRD), also known as "shipping fever," is the most common and costly disease affecting the beef cattle industry in North America. In the broadest sense, BRD refers to any disease of the upper or lower respiratory tract. Bovine BRD is generally associated with lung infections that cause pneumonia in recently weaned or recently arrived feedlot calves (which is why it is often called shipping fever). BRD or shipping fever is most prevalent within the first week of arrival at the feedlot, but can occur in the second half of the feeding period and is also seen in pasture calves. The infectious agents or pathogens required to cause the disease can be broadly classified as viruses, bacteria, and parasites. Viruses known to cause BRD are bovine herpesvirus (IBR); bovine parainfluenza virus (PI-3); bovine respiratory syncytial virus (BRSV); bovine viral diarrhea virus (BVD) and bovine coronavirus (BCV). (See "Bovine Respiratory Disease," Beef Cattle Research Council, revised October 2, 2019, accessed April 22, 2021, http: / / www.beefresearch.ca / research-topic.cfm / bovine-respiratory-disease-38). U.S. Patent Application Publication No. 2017 / 0260147 discloses guanidine derivative compounds that exhibit activity against bovine coronavirus (see Examples 43 and 44). β-D-N(4)-hydroxycytidine (NHC) inhibits the production of cytopathic bovine viral diarrhea virus (BVDV) RNA in a dose-dependent manner at an effective concentration of 90% of 5.4 μM (EC 90 ), and this observation was confirmed by a virus yield assay (EC 90 = 2 μM) (see Stuyver et al., ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Vol. 47, No. 1, Jan. 2003, pp. 244-254).

[0005] Equine influenza is a common, highly contagious respiratory disease of horses with a nearly worldwide distribution. The most common clinical signs of EIV infection in horses are fever, lethargy, anorexia, nasal discharge, and dry cough. Mortality is generally low during EIV outbreaks. Deaths are most common among foals or equids with pre-existing poor health. The risk of EIV infection is not limited to equids; dogs, cats, and humans are also susceptible. This disease has been found to be difficult to control by vaccination because vaccination does not prevent virus shedding. (See Sack et al., EID Journal, Vol. 25(6), June 2019).

[0006] Canine distemper is a highly infectious viral disease caused by a paramyxovirus. It is found worldwide in dogs but can also affect ferrets, raccoons, skunks, gray foxes, and many other animals. Canine distemper affects the gastrointestinal, respiratory, skin, immune, and central nervous systems. There is no treatment for canine distemper. Vaccination is recommended to prevent canine distemper in adult and puppy dogs. (See A. Flowers, "Canine Distemper," revised February 10, 2021, accessed April 23, 2021, https: / / pets.webmd.com / dogs / canine-distemper#1).

[0007] Canine Infectious Respiratory Disease Complex (CIRDC) refers to a syndrome of diseases that can be caused by several different bacterial and viral pathogens. Historically, the most common pathogens associated with CIRDC have been canine parainfluenza virus (CPIV), canine adenovirus type 2 (CAV-2), and Bordetella bronchiseptica. The emergence of novel pathogens, including canine herpesvirus-1 (CHV-1) and canine influenza virus (CIV), has been reported. Treatment of dogs with signs of CIRDC typically includes supportive care. Vaccines are available for many of the common CIRDC pathogens and are recommended for dogs at risk of exposure. However, the available vaccines do not confer sterilizing immunity and there are no specific treatments available for viral CIRDC pathogens. (See K. Reagan & J. Sykes, Vet Clin Small Anim. 50 (2020) 405-418).

[0008] Feline infectious peritonitis (FIP) is a viral disease caused by the feline coronavirus that affects both wild and domestic cats. Feline coronavirus is very common and usually does not cause serious problems except for mild diarrhea. However, when the feline coronavirus mutates into certain strains of coronavirus, FIP can occur. In about 10% of infected cats, the virus multiplies and mutates, causing an infectious disease known as feline infectious peritonitis virus (FIPV), which spreads throughout the cat's body. It can cause extreme inflammatory reactions in the tissues around the abdomen, kidneys, or brain. Although FIP is not considered contagious, it is a very serious disease. Once a cat gets FIP, it is progressive and almost always fatal. FIP has long been considered an incurable disease. Until recently, antiviral drugs have not been introduced to help treat FIP. These drugs have not yet been approved by the Food and Drug Administration (FDA), and their long-term effectiveness remains unknown. FIP vaccines are available, but their effectiveness has not been proven and they are not recommended by the American Association of Feline Practitioners. (See https: / / pets.webmd.com / cats / cat-fip-feline-infectious-peritonitis#1, revised February 12, 2021, accessed April 23, 2021). The nucleoside analogue GS-441524 has been found to strongly inhibit feline infectious peritonitis (FIP) virus in tissue culture and experimental feline infection studies. (See Murphy et al., Veterinary Microbiology 219 (2018) 226-233).

[0009] Feline viral rhinotracheitis (FVR) is a common worldwide respiratory disease of cats caused by feline herpesvirus 1 (FeHV-1). This disease causes a disorder of the lung defense mechanism that makes cats susceptible to secondary bacterial pneumonia or coinfection with feline calicivirus. The virus can also remain latent in ganglia. Most cats that recover from FVR become carriers and shed FeHV-1 spontaneously or after stress. Susceptible animals, especially kittens with low maternal immunity, become infected after exposure to sick cats or carrier cats. Replication of FeHV-1 in the nose, conjunctiva, pharynx, and to a lesser extent the tracheal epithelium causes cell degeneration and exfoliation. (See Alfonso Lopez, Shannon A. Martinson, Pathologic Basis of Veterinary Disease, Elsevier Inc. Sixth Edition, James F. Zachary, Editor, Chapter 9, pp471-560, 2017).

[0010] International Publication No. 2012 / 152317 discloses the use of purine and pyrimidine derivatives in the treatment of feline viral diseases. Feline immunodeficiency virus (FIV) and feline leukemia (FeLV) infections are common in domestic cats (Felis catus) with over 8% infected with FIV and 14% infected with FeLV (see Table 1 of International Publication No. 2012 / 152317). From 2002 to 2017, FIV vaccination was available in the United States and Canada. However, this vaccine provided limited protection, increased the risk of vaccine-site sarcoma, and produced false positives in FIV antibody tests, and has since been discontinued (see "What is FIV and Why is the FIV Vaccine No Longer Available" by N. Stilwell, published on June 21, 2019, updated on July 9, 2020, accessed on April 22, 2021, at https: / / www.petmd.com / cat / care / what-fiv-and-why-fiv-vaccine-no-longer-available).

[0011] U.S. Patent Application Publication No. 2015 / 0018427 demonstrates the use of compounds of the amantadine family for the treatment or prevention of parvovirus infections in humans or animals.

[0012] β-D-N(4)-hydroxycytidine (NHC, 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-(hydroxyamino)pyrimidin-2(1H)-one) has been found to have anti-pestivirus activity and anti-hepatitis virus activity. Antimicrob Agents Chemother, 2003, 47(1):244-54. β-D-N(4)-hydroxycytidine

Chemical formula

[0013] Derivatives and methods for their preparation are shown in PCT International Patent Application No. PCT / US2015 / 066144, published as PCT International Patent Application Publication No. WO2016 / 106050, and U.S. Patent Application No. 15 / 537,087, published as U.S. Patent Application Publication No. US2019 / 0022116, and U.S. Patent Application No. 16 / 921,359, published as U.S. Patent Application Publication No. US2021-0060050A1. NHC is known to tautomerize and is also known as (Z)-1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-(hydroxyimino)-3,4-dihydropyrimidin-2(1H)-one.

Chemical formula

[0014] International Publication No. 2016 / 106050 discloses N4-hydroxycytidine derivatives, compositions, and their use in the treatment and prevention of viral infections.

[0015] International Publication No. WO2019 / 113462 proposes specific N4-hydroxycytidine derivatives and pharmaceutical compositions for use in the treatment or prevention of viral infections such as Eastern, Western, and Venezuelan Equine Encephalitis (EEE, WEE, and VEE respectively), Chikungunya fever (CHIK), Ebola, influenza, RSV, and Zika virus infections.

[0016] Methods for preparing β-D-N(4)-hydroxycytidine (NHC) and its derivatives are disclosed in PCT International Patent Application No. PCT / US2019 / 021168, published as PCT International Patent Application Publication No. WO2019 / 173602, and U.S. Patent Application No. 16 / 755,779, published as U.S. Patent Application Publication No. US2020-0276219A1.

[0017] Antiviral therapies available for treating or preventing Porcine Reproductive and Respiratory Syndrome (PRRS), Bovine Respiratory Disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency in animals are limited or non-existent. Such treatments are needed among livestock producers and pet owners. None of the cited references disclose the use of N4-hydroxycytidine or its prodrugs or salts in the treatment or prevention of these diseases.

[0018] The adenosine nucleoside analog GS-441524 has been used as an antiviral treatment for cats with clinically diagnosed neurological feline infectious peritonitis (see Dickinson et al., J Vet Intern Med. 2020 Jul;34(4):1587-1593.doi:10.1111 / jvim.15780.Epub 2020 May 22, PMID:32441826).

[0019] The efficacy and safety of the nucleoside analogue GS-441524 for the treatment of cats with naturally occurring feline infectious peritonitis were investigated. (See Pedersen et al., J Feline Med Surg. 2019 Apr.;21(4):271-281. doi:10.1177 / 1098612X19825701. Epub 2019 Feb 13. PMID:30755068; PMCID:PMC6435921.)

[0020] The nucleoside analogue GS-441524 has been found to potently inhibit feline infectious peritonitis (FIP) virus in tissue culture and experimental feline infection studies. (See Murphy et al., Vet Microbiol. 2018; Jun;219:226-233. doi:10.1016 / j.vetmic.2018.04.026. Epub 2018 Apr 22. PMID:29778200; PMCID:PMC7117434.)

[0021] Treatment of cats with feline infectious peritonitis with an oral multi-component drug containing GS-441524 has been demonstrated. (Krentz D, Zenger K, Alberer M, Felten S, Bergmann M, Dorsch R, Matiasek K, Kolberg L, Hofmann-Lehmann R, Meli ML, Spiri AM, Horak J, Weber S, Holicki CM, Groschup MH, Zablotski Y, Lescrinier E, Koletzko B, von Both U, Hartmann K. Curing Cats with Feline Infectious Peritonitis with Oral Multi-Component Drug Containing GS-441524. Viruses. November 5, 2021;13(11):2228. doi:10.3390 / v13112228. PMID:34835034; PMCID:PMC8621566.)

Prior Art Documents

Patent Documents

[0022] [Patent Document 1] U.S. Patent Application Publication No. 2017 / 0260147 [Patent Document 2] International Publication No. 2012 / 152317 [Patent Document 3] U.S. Patent Application Publication No. 2015 / 0018427 [Patent Document 4] PCT International Patent Application Publication No. WO2016 / 106050 [Patent Document 5] U.S. Patent Application Publication No. US2019 / 0022116 [Patent Document 6] U.S. Patent Application Publication No. US2021-0060050A1 [Patent Document 7] International Publication No. 2016 / 106050 [Patent Document 8] International Publication No. 2019 / 113462 [Patent Document 9] PCT International Patent Application Publication No. WO2019 / 173602 [Patent Document 10] U.S. Patent Application Publication No. US2020-0276219A1 [Non-Patent Document]

[0023] [Non-Patent Document 1] Dal Pozzo & Thiry, Rev. sci. tech. Off. int. Epiz., 2014, 33(3), 791 - 801 [Non-Patent Document 2] Swine Manual, Iowa State University, College of Veterinary Medicine, Swine Manual, https: / / vetmed.iastate.edu / vdpam / FSVD / swine / index-diseases / porcine-reproductive [Non-Patent Document 3] 「Bovine Respiratory Disease」Beef Catle Research Council, http: / / www.beefresearch.ca / research-topic.cfm / bovine-respiratory-disease-38

Non-Patent Document 4

Non-Patent Document 5

Non-Patent Document 6

Non-Patent Document 7

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Non-Patent Document 11

Non-Patent Document 12

Non-Patent Document 13

Non-Patent Document 14

Non-Patent Document 15

Non-Patent Document 16

Summary of the Invention

Problems to be Solved by the Invention

[0024] A method for preventing or treating a viral disease in an animal, comprising administering to the animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine (NHC) or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency

Brief Description of the Drawings

[0025]

Figure 1

Figure 2

Figure 3A

Figure 3B

Figure 4A

Figure 4B

DETAILED DESCRIPTION OF THE INVENTION

[0026] Viral infections in animals have a significant economic impact on the animal health industry. These infections have traditionally been successfully managed and prevented by vaccination, testing, and culling. Currently, viral infections in animals are not targeted by antiviral small molecule drugs.

[0027] There are several challenges in the use of antiviral drugs in animal health. The treatment or prevention of viral infections in animals is cost-effective. This is particularly true in livestock animals (e.g., cattle, pigs, poultry, and aquaculture). In many jurisdictions, the culling of infected animals is a legal requirement for some viral diseases (e.g., foot-and-mouth disease). Convenience is another challenge for animal health antiviral drugs. For example, in many species, a one-shot dose may be a prerequisite. Additionally, treatment with antiviral drugs should not interfere with vaccination strategies for animals. In the case of food animals, antiviral drugs must be administered to establish minimum residue levels (MRLs) in edible tissues. Finally, antiviral drugs should have a clean toxicity profile for the target animals and be safe for humans administering the drugs.

[0028] Yet another issue is the required level of efficacy. Must an antiviral drug be capable of virus elimination or cure? Is it necessary to simply maintain the viral load, or is it necessary to reduce the viral load to treat or prevent a viral disease? Should viral shedding be reduced or eliminated to prevent disease transmission from one animal to another?

[0029] The synthetic nucleoside derivative N4-hydroxycytidine, as well as its prodrugs and salts, are thought to exert their antiviral action by introducing replication errors during viral RNA replication. As noted above, activity against coronaviruses including SARS, MERS, and SARS-CoV-2 has also been demonstrated. The putative mechanism of action (MOA) is by "error catastrophe" where the replication error rate is greater than the error threshold tolerated to maintain the virus population.

[0030] The synthetic nucleoside derivative N4-hydroxycytidine, as well as its prodrugs and salts, are antiviral drugs effective in treating or preventing viral infections that cause viral diseases, where the viral diseases are considered to be porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency.

[0031] The synthetic nucleoside derivative N4-hydroxycytidine, as well as its prodrugs and salts, are antiviral drugs effective in treating or preventing viral infections that cause viral diseases, where the viral diseases are also considered to be porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency.

[0032] Furthermore, the synthetic nucleoside derivative N4-hydroxycytidine, its prodrugs and salts are antiviral agents effective in stabilizing or reducing the viral load of viral infections in animals, which viral infections are considered to be derived from Porcine Reproductive and Respiratory Syndrome Virus (PRRS-1 / PRRS-2), Bovine Leukemia Virus (BLV), Bovine Coronavirus (BCV), Bovine Respiratory Syncytial Virus (BRSV), Bovine Rotavirus, Bovine Parainfluenza Virus 3, Bovine Viral Diarrhea Virus (BVDV), Equine Influenza Virus, Canine Distemper Virus, Canine Influenza Virus (CIV), Coronaviridae (FIPV), Feline Calicivirus, or Feline Immunodeficiency Virus (FIV).

[0033] The methods or uses described herein also address one or more of the problems of the above-mentioned animal health antiviral therapies. Specifically, the synthetic nucleoside derivative N4-hydroxycytidine, its prodrugs and salts, when administered to animals, maintain or reduce the viral load in animals and reduce or eliminate viral excretion from animals.

[0034] Thereby, the spread of the virus is suppressed and transmission to untreated contact animals is prevented.

[0035] Definitions Certain technical and scientific terms are specifically defined below. Unless specifically defined elsewhere in this specification, all other technical and scientific terms used herein have the meanings commonly understood by those skilled in the art to which this disclosure pertains.

[0036] As used herein, including in the appended claims, the singular forms of words such as "a", "an", and "the" include their corresponding plural references unless the context clearly indicates otherwise.

[0037] The terms "administer" and / or "administering" of a compound are to be understood as including providing the compounds described herein or pharmaceutically acceptable salts thereof, and the aforementioned compositions, to a subject.

[0038] As used herein, the terms "at least one" matter or "one or more" matters each include a single matter selected from a list, as well as a mixture of two or more matters selected from the list.

[0039] The term "pharmaceutically acceptable carrier" refers to any inert substance suitable for use in a formulation for the delivery of a therapeutic agent. The carrier can be an anti-adherent, binder, coating, disintegrant, filler or diluent, lubricant, preservative (such as antioxidant, antibacterial agent, antifungal agent, etc.), sweetening agent, absorption delaying agent, wetting agent, emulsifying agent, buffering agent, etc. Examples of suitable pharmaceutically acceptable carriers include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, etc.), dextrose, vegetable oils (such as olive oil), physiological saline, buffer solutions, buffered physiological saline, and isotonic agents such as sugars, polyalcohols, sorbitol, and sodium chloride.

[0040] The animal can be one or more selected from the group consisting of bovine (e.g., cows), porcine (e.g., pigs), ovine animals (e.g., sheep), caprine (e.g., goats), equine animals (e.g., horses), canine animals (e.g., domestic dogs), feline animals (e.g., domestic cats), lagomorph animals (rabbits), rodents (e.g., rats or mice), Procyon lotor (e.g., raccoons).

[0041] As used herein, the terms "treat" and "treating" refer to any process by which there can be a deceleration, interruption, cessation, control, or termination of the progression of a disease or disorder described herein. These terms do not necessarily indicate the complete elimination of all symptoms of a disease or disorder.

[0042] As used herein, the terms "prophylaxis" and "antiviral prophylaxis" refer to all processes intended to prevent disease. Prophylaxis can occur either before exposure to a viral infection or after potential exposure to a viral infection.

[0043] The therapeutic agents and compositions used in the methods can be administered via any suitable enteral or parenteral route of administration. The term "enteral route" of administration refers to administration through any part of the gastrointestinal tract. Examples of enteral routes include oral, mucosal, oral and rectal routes, or intragastric routes. Administration by "parenteral route" refers to a route of administration other than the enteral route.

[0044] Administration can be by oral, topical, or injectable routes.

[0045] As used herein, the term "simultaneous administration" with respect to the administration of agents refers to the administration of agents such that the individual agents are present in the subject at the same time. In addition to the simultaneous administration (via the same or alternative routes) of agents, simultaneous administration can include the administration of agents at different times (via the same or alternative routes).

[0046] Variations such as "consists essentially of", "consist essentially of", or "consisting essentially of" as used throughout this specification and the claims indicate the inclusion of any recited element or group of elements that do not substantially change the basic or novel characteristics of the specified dosing regimen, method, or composition, and the inclusion of any other elements of the same or different nature.

[0047] When an embodiment is described in the present specification using the language “comprising,” it is understood that other similar embodiments described using the terms “consisting of” and / or “consisting essentially of” are also provided.

[0048] Unless explicitly stated otherwise, all ranges recited herein are inclusive; that is, the ranges include the upper and lower limits of the range, as well as all values therebetween. All ranges are also intended to include all subranges subsumed therein, even if not necessarily explicitly recited. By way of example, temperature ranges, percentage, equivalence ranges, etc. described herein include the upper and lower limits thereof, as well as any value in the continuum therebetween.

[0049] The numerical values provided herein, and the use of the term “about,” can include variations of ±1%, ±2%, ±3%, ±4%, ±5%, and ±10%, and their numerical equivalents.

[0050] When used to modify a numerically defined parameter (e.g., the dosage of an antiviral nucleoside, or the length of treatment time by combination therapy described herein), “about” means that the parameter can vary by up to about 10% below or above the value recited for that parameter, and where appropriate, the recited parameter may be rounded to the nearest integer. For example, a dosage of about 5 mg / kg can vary between 4.5 mg / kg and 5.5 mg / kg. Further, as used herein, the term “or” indicates alternatives that can be combined where appropriate. That is, the term “or” includes the separately recited options and combinations thereof.

[0051] As used herein, “antiviral nucleoside” means any nucleoside compound that exhibits antiviral activity.

[0052] As used herein, viral load refers to a measure of the amount of virus in an organism, typically in the bloodstream, and is usually expressed as virus particles per milliliter.

[0053] Viral shedding refers to the elimination and release of progeny viruses after successful reproduction during host cell infection.

[0054] Sterilizing immunity means that the immune system can stop pathogens, including viruses, from replicating within an animal.

[0055] As used herein, a prodrug refers to a biologically inactive compound that can be metabolized in the body to produce a drug. The prodrugs of the present invention can have any form suitable for a prescriber, for example, esters, and more specifically alkyl esters, are a non-limiting general prodrug form. Examples of prodrugs of the antiviral nucleoside NHC are N-hydroxycytidine 5'-(2-methylpropanoate) (Compound A):

Chemical formula

Chemical formula

[0056] Salt The compounds of the present invention can be used in the form of pharmaceutically acceptable salts. Those skilled in the art will recognize examples of compounds of the present invention that can form salts. Examples of such compounds are described herein with reference to possible salts. Such references are for illustrative purposes only.

[0057] Pharmaceutically acceptable salts can be used in conjunction with the compounds for treating patients. However, non-pharmaceutical salts can be useful in the preparation of intermediate compounds.

[0058] The term "pharmaceutically acceptable salt" refers to salts (including inner salts such as zwitterions) that have the same efficacy as the parent compound and are not biologically or otherwise undesirable (e.g., not toxic or otherwise harmful to its recipient). Thus, one embodiment of the present invention provides for the use of pharmaceutically acceptable salts of the compounds. As used herein, the term "salt" refers to either acidic salts formed with inorganic and / or organic acids, as well as basic salts formed with inorganic and / or organic bases. The salts of the compounds of the present invention can be formed by methods known to those skilled in the art, for example, by reacting the compounds of the present invention with an acid or base in an amount such as an equivalent in a medium such as a medium in which the salt precipitates or in an aqueous medium, followed by lyophilization.

[0059] Acids generally considered suitable for the formation of pharmaceutically useful salts from basic pharmaceutical compounds are discussed, for example, by P. Stahl et al., Camille G. (eds.), Handbook of Pharmaceutical Salts. Properties, Selection and Use. (2002) Zurich: Wiley-VCH; S. Berge et al., Journal of Pharmaceutical Sciences (1977) 66(1) 1-19; P. Gould, International J. of Pharmaceutics (1986) 33 201-217; Anderson et al., The Practice of Medicinal Chemistry (1996), Academic Press, New York; and The Orange Book (Food & Drug Administration, Washington, D.C. on their website). These disclosures are incorporated herein by reference.

[0060] The present disclosure relates to a method of treating a viral disease as defined herein, the method comprising administering to an animal in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. The present disclosure relates to a method of treating a viral disease, the method comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. Herein, the viral disease is selected from the group consisting of porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency.

[0061] Porcine rotavirus B as a major causative factor for diarrhea occurrence in piglets. Rotavirus (RV) is considered the main cause of acute viral gastroenteritis in young animals. Miyabe et al., Scientifc Reports, (2020) 10:22002, https: / / doi.org / 10.1038 / s41598-020-78797-y.

[0062] Porcine or swine influenza is a respiratory disease of pigs caused by influenza A virus that regularly causes influenza in pigs. In recent years, the major swine influenza viruses prevalent in pigs in the United States are the swine triple reassortant (tr) H1N1 influenza virus, trH3N2 virus, and trH1N2 virus. Centers for Disease Control and Prevention, National Center for Immunization and Respiratory Diseases (NCIRD) https: / / www.cdc.gov / flu / swineflu / keyfacts_pigs.htm (last verified page: August 19, 2014).

[0063] Treatment of such viral diseases results in a reduction of clinical signs in the treated animals and a reduction of mortality in animals showing signs of severe disease. In another embodiment, the treatment promotes recovery from clinical symptoms.

[0064] The present disclosure relates to a method of providing antiviral prophylaxis, comprising administering to a subject in need thereof a composition comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. Here, the viral disease is selected from the group consisting of porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis, or feline immunodeficiency. The antiviral prophylaxis method is particularly useful when the antiviral nucleoside NHC or prodrug or a salt thereof is administered at the peak of viral replication but before the onset of viral shedding.

[0065] Furthermore, the present disclosure relates to a method of treating a viral infection, comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. The viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), equine influenza virus, canine distemper virus, canine influenza virus (CIV), coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0066] Furthermore, the present disclosure relates to a method of providing antiviral prophylaxis, the method comprising administering to a subject in need thereof a composition comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. Here, the viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), equine influenza virus, canine distemper virus, canine influenza virus (CIV), coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0067] Products provided for treatment and prophylaxis may include compositions containing an antiviral nucleoside NHC or a prodrug or a salt thereof in the composition. The treatment may also include one or more additional therapeutic agents. One or more additional active agents may be administered together with (co-administered) the antiviral nucleoside, or may be administered separately from the antiviral nucleoside in a different dosage form. That is, the additional active agent may be administered in a single dosage form together with the antiviral nucleoside, or the additional active agent may be administered in a dosage form separate from the dosage form containing the antiviral nucleoside. The treatments disclosed herein may be used in combination with one or more other active agents, including, but not limited to, antiviral agents used for the prevention, treatment, control, amelioration, or reduction of risk of a particular disease or condition (e.g., viral infections). In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof disclosed herein is combined with one or more other antiviral agents for use in the prevention, treatment, control, amelioration, or reduction of risk of a particular disease or condition for which the antiviral nucleoside NHC or prodrug or a salt thereof disclosed herein is useful. Such other active agents may be administered simultaneously or sequentially with the compounds of the present disclosure, by the route and in the amounts commonly used therefor. When the treatments disclosed herein are used in combination with one or more other active agents, the antiviral nucleoside may be administered simultaneously with the one or more other active agents, or before or after the one or more other active agents. The antiviral nucleoside can be administered separately, by the same or different routes of administration, or together in the same pharmaceutical composition with the other agent.

[0068] The dosage of the antiviral nucleoside NHC or prodrug or a salt thereof can vary and depends on the therapeutically effective dose of each agent. Generally, the therapeutically effective dose of each is used. Combinations comprising at least one antiviral nucleoside and other active agents generally include the therapeutically effective dose of each active agent. In such combinations, the antiviral nucleosides and other active agents disclosed herein can be administered separately or in combination. Further, the administration of one element can be before, simultaneous with, or after the administration of the other agent. In one embodiment, the present disclosure provides an antiviral nucleoside and at least one other active agent as a combination preparation for simultaneous, separate or sequential use in therapy. In one embodiment, the therapy is for the treatment of a viral disease, and the viral disease is selected from the group consisting of porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency. In one embodiment, the therapy is antiviral prophylaxis such as for a potential infection, either before or after exposure to a viral infection, and the viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency. In another embodiment, the therapy is for the treatment of a viral disease, and the virus is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), equine influenza virus, canine distemper virus, canine influenza virus (CIV), coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0069] Additional active agents can be one or more agents selected from the group consisting of antiviral compounds, antigens, adjuvants, anti-cancer agents, CTLA-4 agonists, LAG-3 agonists, PD-1 pathway antagonists, lipids, liposomes, peptides, cytotoxic agents, chemotherapeutic agents, immunomodulatory cell lines, checkpoint inhibitors, vascular endothelial growth factor (VEGF) receptor inhibitors, topoisomerase II inhibitors, smoothening inhibitors, alkylating agents, antibiotics, antimetabolites, retinoids, steroids, and immunomodulators including, but not limited to, antiviral vaccines. It will be understood that the description of the additional active agents above and those listed below may overlap. It is also understood that the treatment combinations are subject to optimization and that the best combinations for the use of antiviral nucleosides and one or more additional active agents will be determined based on the needs of the individual animal.

[0070] Vaccine therapies that can be used in combination with the treatments disclosed herein include, but are not limited to, inactivated vaccines, live attenuated vaccines, recombinant vaccines, replication-deficient viral vector vaccines, mRNA-based vaccines, DNA vaccines, nanoparticle vaccines, non-replicating viral vectors, self-replicating RNA vaccines, self-amplifying RNA vaccines, protein subunit vaccines, Ii-Key peptide COVID-19 vaccines, gp96-based vaccines, intranasal vaccines, and mRNA lipid nanoparticle (mRNA10 LNP) vaccines.

[0071] The present disclosure further relates to a method of treating a viral disease, the method comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. Here, the antiviral nucleoside is administered once a day for 1 to 10 days, such as 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, or 10 days. In certain embodiments, the antiviral nucleoside NHC or a prodrug or a salt thereof is administered once a day for 3 to 5 days, such as 3 days, 4 days, or 5 days.

[0072] The present disclosure further relates to a method of treating a viral disease, the method comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. The antiviral nucleoside is administered twice a day for 1 to 10 days, such as 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days or 10 days. In certain embodiments, the antiviral nucleoside is administered twice a day for 3 to 5 days, such as 3 days, 4 days or 5 days.

[0073] In embodiments of the treatment methods disclosed herein, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day as a single dose in an amount of about 50 mg to about 1600 mg, such as about 100 mg to about 1400 mg, about 200 mg to about 1200 mg, about 300 mg to about 1000 mg, or about 400 mg to about 800 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day as a single dose in an amount of about 50 mg, about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg, about 800 mg, about 1000 mg, about 1200 mg, about 1400 mg, or about 1600 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day as a single dose in an amount of about 200 mg. In aspects of such embodiments, the antiviral nucleoside is administered once a day as a single dose in an amount of about 300 mg. In aspects of such embodiments, the antiviral nucleoside is administered once a day as a single dose in an amount of about 400 mg. In aspects of such embodiments, the antiviral nucleoside is administered once a day as a single dose in an amount of about 500 mg. In aspects of such embodiments, the antiviral nucleoside is administered once a day as a single dose in an amount of about 600 mg. In aspects of such embodiments, the antiviral nucleoside is administered once a day as a single dose in an amount of about 700 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day as a single dose in an amount of about 800 mg.

[0074] In embodiments of the treatment methods disclosed herein, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as individual doses in an amount of about 50 mg to about 1600 mg, such as about 100 mg to about 1400 mg, about 200 mg to about 1200 mg, about 300 mg to about 1000 mg, or about 400 mg to about 800 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as individual doses in an amount of about 50 mg, about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg, about 800 mg, about 1000 mg, about 1200 mg, about 1400 mg, or about 1600 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as an individual dose in an amount of about 200 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as an individual dose in an amount of about 300 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as an individual dose in an amount of about 400 mg. In aspects of such embodiments, the antiviral nucleoside is administered twice daily as a single dose in an amount of about 500 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as an individual dose in an amount of about 600 mg. In aspects of such embodiments, the antiviral nucleoside is administered twice daily as a single dose in an amount of about 700 mg. In aspects of such embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered twice daily as an individual dose in an amount of about 800 mg.

[0075] In embodiments of the treatment methods disclosed herein, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in a dosage in an amount of about 1 mg / kg body weight of the animal to about 500 mg / kg body weight of the animal, or about 10 mg / kg body weight of the animal to about 100 mg / kg body weight of the animal, or about 1 mg / kg body weight of the animal to about 10 mg / kg body weight of the animal, or about 10 mg / kg body weight of the animal to about 50 mg / kg body weight of the animal, or about 20 mg / kg body weight of the animal to about 40 mg / kg body weight of the animal, or about 50 mg / kg body weight of the animal to about 100 mg / kg body weight of the animal, or about 100 mg / kg body weight of the animal to about 500 mg / kg body weight of the animal.

[0076] The present disclosure further relates to a method of providing antiviral prophylaxis, the method comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or prodrug or a salt thereof. Here, the antiviral nucleoside is administered once a day for 1 to 42 days, such as for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 26 days, 27 days, 28 days, 29 days, 30 days, 31 days, 32 days, 33 days, 34 days, 35 days, 36 days, 37 days, 38 days, 39 days, 40 days, 41 days or 42 days. In certain embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day for 1 to 21 days, such as for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, or 21 days. In certain embodiments, the antiviral nucleoside NHC or prodrug or a salt thereof is administered once a day for 3 to 14 days, such as for 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, or 14 days.

[0077] The present disclosure further relates to a method of providing antiviral prophylaxis, the method comprising administering to a subject in need thereof a treatment comprising an antiviral nucleoside NHC or a prodrug or a salt thereof. Here, the antiviral nucleoside is administered twice a day for 1 to 42 days, such as for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, 21 days, 22 days, 23 days, 24 days, 25 days, 26 days, 27 days, 28 days, 29 days, 30 days, 31 days, 32 days, 33 days, 34 days, 35 days, 36 days, 37 days, 38 days, 39 days, 40 days, 41 days or 42 days. In certain embodiments, the antiviral nucleoside NHC or a prodrug or a salt thereof is administered twice a day for 1 to 21 days, such as for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 15 days, 16 days, 17 days, 18 days, 19 days, 20 days, or 21 days. In certain embodiments, the antiviral nucleoside NHC or a prodrug or a salt thereof is administered twice a day for 3 to 14 days, such as for 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, or 14 days.

[0078] In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered orally. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in the form of an oral pill or tablet. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in liquid form. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in the animal's feed. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in the animal's drinking water. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered topically. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered to oral tissues. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in the form of a nasal spray. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered by injection. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered in the form of an implant. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered to the animal's eye. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered to the animal's ear.

[0079] In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered to all animals in a group or collection of animals including some infected animals and some non-infected animals. In one embodiment, the antiviral nucleoside NHC or prodrug or a salt thereof is administered to animals that have been or potentially been exposed to the virus but have not yet developed symptoms of the viral disease.

[0080] One embodiment of the present invention is a method for preventing or treating a viral disease in an animal, comprising administering to the animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency.

[0081] In another embodiment, the administration is by oral, topical, or injection.

[0082] In another embodiment, the animal is a pig and the disease is porcine reproductive and respiratory syndrome (PRRS). In another embodiment, the animal is a bovine animal and the disease is bovine respiratory disease (BRD). In another embodiment, the animal is a horse and the disease is equine influenza. In another embodiment, the animal is a dog and the disease is canine distemper or canine respiratory disease. In another embodiment, the animal is a cat and the disease is feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency.

[0083] In another embodiment, the pharmaceutical composition is administered in a single dose or in multiple doses administered multiple times.

[0084] An alternative embodiment is a method for preventing or treating a viral disease in an animal, comprising administering to the animal a pharmaceutical composition comprising β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is caused by SARS-CoV-2 infection in the animal and the animal is a pig, a cow, a horse, a dog or a cat.

[0085] In another embodiment, the administration of the pharmaceutical composition to the animal does not interfere with the animal's vaccination strategy.

[0086] An alternative embodiment is a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier for inducing an antiviral response in an animal, wherein the animal is a pig, cow, horse, dog or cat, the animal is suffering from or is susceptible to a viral disease, and the viral disease is porcine reproductive and respiratory syndrome (PRRS), bovine respiratory disease (BRD), equine influenza, canine distemper, canine respiratory disease, feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency disease, a pharmaceutical composition.

[0087] In another embodiment, the animal is a pig and the disease is porcine reproductive and respiratory syndrome (PRRS). In another embodiment, the animal is a bovine animal and the disease is bovine respiratory disease (BRD). In another embodiment, the animal is a horse and the disease is equine influenza. In another embodiment, the animal is a dog and the disease is canine distemper or canine respiratory disease. In another embodiment, the animal is a cat and the disease is feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency disease.

[0088] In another embodiment, the antiviral response is that the amount of virus in the animal remains substantially the same.

[0089] In another embodiment, the antiviral response is that the viral load of the animal decreases.

[0090] In another embodiment, the antiviral response is that the viral load of the animal is substantially eliminated.

[0091] In another embodiment, the administration is oral.

[0092] In another embodiment, the oral administration is in the form of tablets or liquid.

[0093] In another embodiment, the oral administration is carried out in the drinking water of the animal.

[0094] In another embodiment, oral administration is carried out in the animal's feed.

[0095] In another embodiment, the administration is local.

[0096] In another embodiment, the administration is by injection.

[0097] An alternative embodiment is a method for treating a viral infection, comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier. Here, the viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), equine influenza virus, canine distemper virus, canine influenza virus (CIV), coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0098] An alternative embodiment is a method for treating a viral infection, comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier. Here, the viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), bovine influenza virus, canine distemper virus, coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0099] In another embodiment, the viral infection is porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2). In another embodiment, the viral infection is bovine leukemia virus (BLV). In another embodiment, the viral infection is bovine coronavirus (BCV). In another embodiment, the viral infection is bovine respiratory syncytial virus (BRSV). In another embodiment, the viral infection is bovine rotavirus. In another embodiment, the viral infection is bovine parainfluenza virus 3. In another embodiment, the viral infection is bovine viral diarrhea virus (BVDV). In another embodiment, the viral infection is equine influenza virus. In another embodiment, the viral infection is canine distemper virus. In another embodiment, the viral infection is canine influenza virus (CIV). In another embodiment, the viral infection is Coronaviridae (FIPV). In another embodiment, the viral infection is feline calicivirus. In another embodiment, the viral infection is feline immunodeficiency virus (FIV).

[0100] An alternative embodiment is a method of providing antiviral prophylaxis, comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier. Here, the viral infection is selected from the group consisting of porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2), bovine leukemia virus (BLV), bovine coronavirus (BCV), bovine respiratory syncytial virus (BRSV), bovine rotavirus, bovine parainfluenza virus 3, bovine viral diarrhea virus (BVDV), equine influenza virus, canine distemper virus, canine influenza virus (CIV), Coronaviridae (FIPV), feline calicivirus, or feline immunodeficiency virus (FIV).

[0101] Another embodiment is a method of providing antiviral prophylaxis, the method comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral infection is transmitted via the respiratory system or the upper respiratory tract. In another embodiment, the viral infection is an infection transmitted via the air. In yet another embodiment, the viral disease is transmitted via the gastrointestinal tract.

[0102] In another embodiment, the viral infection is porcine reproductive and respiratory syndrome virus (PRRS-1 / PRRS-2). In another embodiment, the viral infection is bovine leukemia virus (BLV). In another embodiment, the viral infection is bovine coronavirus (BCV). In another embodiment, the viral infection is bovine respiratory syncytial virus (BRSV). In another embodiment, the viral infection is bovine rotavirus. In another embodiment, the viral infection is bovine parainfluenza virus 3. In another embodiment, the viral infection is bovine viral diarrhea virus (BVDV). In another embodiment, the viral infection is equine influenza virus. In another embodiment, the viral infection is canine distemper virus. In another embodiment, the viral infection is canine influenza virus (CIV). In another embodiment, the viral infection is feline infectious peritonitis virus (FIPV). In another embodiment, the viral infection is feline calicivirus. In another embodiment, the viral infection is feline immunodeficiency virus (FIV).

[0103] An embodiment of the present invention is a method of preventing or treating a viral disease in an animal, the method comprising administering to the animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is caused by an RNA viral infection in the animal.

[0104] In another embodiment, the animal is a pig, cow, horse, dog or cat.

[0105] In another embodiment, the animal is a pig and the disease is porcine reproductive and respiratory syndrome (PRRS).

[0106] In another embodiment, the animal is a bovine and the disease is bovine respiratory disease (BRD).

[0107] In another embodiment, the animal is a horse and the disease is equine influenza.

[0108] In another embodiment, the animal is a dog and the disease is canine distemper or canine respiratory disease.

[0109] In another embodiment, the animal is a cat and the disease is feline infectious peritonitis, feline viral rhinotracheitis or feline immunodeficiency disease.

[0110] In another embodiment, the RNA virus is a single-stranded RNA (ssRNA) virus.

[0111] In another embodiment, the RNA virus is a double-stranded RNA (dsRNA) virus.

[0112] In another embodiment, the RNA virus is a positive-strand RNA virus.

[0113] In another embodiment, the positive-strand RNA virus is from the family Alphavirus, Astroviridae, Caliciviridae, Coronaviridae, Flaviviridae, Hepadnaviridae, Nodaviridae, Picornaviridae, Togaviridae or Togaviridae.

[0114] In another embodiment, the RNA virus is a negative-strand RNA virus.

[0115] In another embodiment, the RNA virus is an ambisense RNA virus.

[0116] In another embodiment, the negative-strand RNA virus is from the family Arenaviridae, Bornaviridae, Bunyaviridae, Filoviridae, Nyamiviridae, Orthomyxoviridae, Paramyxoviridae, Pneumoviridae, or Rhabdoviridae.

[0117] One embodiment of the present invention is a method for preventing or treating a viral disease in an animal, comprising administering to the animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is caused by a porcine influenza virus, porcine rotavirus, feline infectious peritonitis virus (FIPV) or feline calicivirus.

[0118] In another embodiment, the administration is by an oral, topical or injection route, preferably oral.

[0119] In a preferred embodiment, the animal is a pig and the disease is caused by a porcine influenza virus or porcine rotavirus.

[0120] In a preferred embodiment, the animal is a pig and the disease is caused by a porcine influenza virus.

[0121] In a preferred embodiment, the animal is a pig and the disease is caused by a porcine rotavirus.

[0122] In a preferred embodiment, the animal is a cat and the disease is caused by a feline infectious peritonitis virus or feline calicivirus.

[0123] In a preferred embodiment, the animal is a cat and the disease is caused by a feline infectious peritonitis virus.

[0124] In a preferred embodiment, the animal is a cat and the disease is caused by a feline calicivirus.

[0125] In another embodiment, the pharmaceutical composition is administered in a single dose or in multiple doses administered multiple times.

[0126] Another embodiment of the present invention is a method for preventing or treating a viral disease in an animal, comprising administering to the animal a pharmaceutical composition comprising β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is caused by SARS-CoV-2 infection in the animal, and the animal is a pig, cow, horse, dog or cat.

[0127] Another embodiment of the present invention is a pharmaceutical composition for use in preventing or treating a viral disease in an animal, comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the animal is a pig or a cat, and the animal is suffering from, or is susceptible to, a viral disease caused by porcine influenza virus, porcine rotavirus, feline infectious peritonitis virus, or feline calicivirus.

[0128] Another embodiment of the present invention is a pharmaceutical composition for inducing an antiviral response in an animal, comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the animal is a pig or a cat, the animal is suffering from, or is susceptible to, a viral disease, and the viral disease is caused by porcine influenza virus, porcine rotavirus, feline infectious peritonitis virus or feline calicivirus.

[0129] In another embodiment, the antiviral response is that the amount of virus in the animal remains substantially the same.

[0130] In another embodiment, the antiviral response is that the amount of virus in the animal decreases.

[0131] In another embodiment, the antiviral response is the substantial elimination of the amount of virus in the animal.

[0132] Another embodiment of the present invention is a method for treating a viral infection, the method comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral infection is selected from the group consisting of porcine rotavirus, porcine influenza virus, feline infectious peritonitis virus (FIPV), or feline calicivirus.

[0133] Another embodiment of the present invention is a method for providing antiviral prophylaxis, the method comprising administering to an animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral infection is selected from the group consisting of porcine rotavirus, porcine influenza virus, feline infectious peritonitis virus (FIPV), or feline calicivirus.

[0134] Another embodiment of the present invention is a method for preventing or treating a viral disease in an animal, the method comprising administering to the animal a pharmaceutical composition comprising an antiviral nucleoside β-D-N(4)-hydroxycytidine or a prodrug or salt thereof and a pharmaceutically acceptable carrier, wherein the viral disease is caused by a coronavirus.

[0135] In another embodiment, the animal is Feline or Porcine.

[0136] Porcine means related to pig or swine.

[0137] Feline means related to cat or other members of the cat family.

[0138] Further embodiments of the present disclosure include the pharmaceutical compositions, combinations, uses, and methods described above, and it should be understood that each embodiment can be combined with one or more other embodiments as long as such combinations are consistent with the description of the embodiments. It should be further understood that the embodiments provided above include all embodiments, including embodiments that result from combinations of the embodiments.

[0139] [Examples] [Example 1] CPE antiviral assay of feline infectious peritonitis virus (FIPV) in Crandell-Rees feline kidney (CrFK) cells Crandell-Rees feline kidney (CRFK, ATCC CCL-94) cells were cultured in DMEM medium containing 2% FBS, GlutaMax, D-glucose, sodium pyruvate, non-essential amino acids, and penicillin / streptomycin. Cells were harvested for virus infection by trypsinization. 50 ml of CRFK cells (1.5×10 5 cells / ml) were infected with feline infectious peritonitis (FIPV) strain 79-1146. The infection conditions were optimized for the CPE assay window 4 days after infection. FIPV-infected CRFK cells were mixed with the NHC compound in serial 1 / 3 dilutions from a final concentration of 10 μM to 0.000508 μM. Thereafter, each dilution was plated repeatedly at 100 μl / well in a 96-well plate (Costar3903 or 3904). Cells were incubated for 96 hours at 37°C and 5% CO2 in a humidified incubator. The cytopathic effect (CPE) was determined using the CellTiterGlo Luminescent Cell Viability Assay (Promega) in a Tecan Infinite M1000 PRO plate reader according to the manufacturer's protocol. The effective concentration (EC 50 ) of NHC that reduces FIVP replication by 50% was calculated using GraphPad Prism8 in a four-parameter non-linear regression model.

[0140] The results of this test indicate the concentration of NHC that effectively inhibits FIP virus in CRFK cells. See Figure 1 and Table 1 below. [Table 1]

[0141] MK-0608 is a potent and orally bioavailable inhibitor of HCV replication in vitro (Carroll et al., ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Mar. 2009, p. 926-934).

[0142] GC-376 is a 3C-like protease inhibitor known to treat various forms of feline infectious peritonitis (Pedersen et al. J. of Feline Med Surg. 2018 Apr;20(4):378-392).

[0143] Remdesivir is a prodrug of adenosine triphosphate (ATP) analog and has potential antiviral activity against various RNA viruses. Retrieved from https: / / pubchem.ncbi.nlm.nih.gov / compound / Remdesivir on July 21, 2021 by APA National Center for Biotechnology Information (2021). PubChem Compound Summary for CID121304016, Remdesivir.

[0144] NHC is β-D-N(4)-hydroxycytidine as described above.

[0145] [Example 2] NHC's CrFK cell-based antiviral activity against feline calicivirus (FCV) 5% FCS and 0.15% NaHCO3 in 2×10 5A monolayer of Crandell-Reese feline kidney (CrFK) cells grown in 96-well plates (Greiner catalog number 655090) with M6B8 medium supplemented at a density of cells / ml and 100 μl / well was infected with feline calicivirus (FCV) wild-type strain 255 at multiplicities of infection of 0.01 and 0.001. NHC was added to the infected cells at 1 / 3 serial dilutions to obtain a test concentration range of 10 μM to 0.000508 μM, and the cultures were incubated at 37 °C and 5% CO2 for up to 48 hours. The degree of virus replication was evaluated by determining the degree of FCV-induced cytopathic effect (CPE) by adding the reagent 1:1 (v / v) to the culture plates equilibrated at room temperature for 30 minutes using the CellTiterGlo® Luminescent Cell Viability Assay (Promega catalog number G7571). The plates were incubated at room temperature for 10 minutes and then luminescence was read using a Biotek Cytation5. The effective concentration (EC 50 ) of NHC that reduced FCV replication by 50% was calculated in a four-parameter non-linear regression model using GraphPad Prism8. The ED 50 of NHC against feline calicivirus is 3.8 μM (Figure 2).

[0146] [Example 3] Antibovine rotavirus activity of β-D-N(4)-hydroxycytidine (NHC) The antiviral activity of NHC against bovine rotavirus was tested in a cell culture-based assay. MA-104 (African green monkey kidney tissue) cells were cultured and harvested according to standard procedures. The harvested MA-104 cell suspension was seeded into 96-well plates (Greiner CELLSTAR® black, Sigma-Aldrich® M0562) at a density of 3×10 4 cells / mL in M6B8 medium containing 5% FBS at 200 μl / well. After 4 days of culture, the monolayer was washed and 100 μl / well of 3630 TCID 50Cells were infected with rotavirus A by adding porcine rotavirus A strain at / mL. One hour after infection, NHC was added at serial 1 / 3 dilutions to obtain a test concentration range of 270 μM to 123 nM, and the cells were cultured for 3 days at 37 °C under 5% CO2. Inhibition of rotavirus A strain replication by NHC was quantified using a mouse anti-rotavirus A strain-specific monoclonal antibody (Ingenasa, #1JF10), and then quantified in an immunofluorescence assay using an FITC fluorescent dye-conjugated goat anti-mouse antibody (Nordic) and DAPI (Sigma Aldrich) nuclear staining. The degree of immunofluorescence was quantified as relative fluorescence units (RFU) at 485 nm excitation and 528 nm emission using a Cytation5 cell imaging multimode reader and Gen5 software version 3.10 (Agilent BioTek). The inhibitory concentration (IC 50 ) of NHC that reduces rotavirus replication by 50% was calculated by a four-parameter non-linear regression model using GraphPad Prism8. The results demonstrate that NHC can inhibit the replication of porcine rotavirus strains G5P7 and G9P7. The effective concentration (EC 50 ) of NHC at which porcine rotavirus replication is halved is 2.6 μM for the G5P7 strain (Figure 3a) and 1.9 μM for the G9P7 strain (Figure 3b).

[0147] [Example 4] Anti-swine influenza virus A activity of β-D-N(4)-hydroxycytidine (NHC) The antiviral activity of NHC against swine influenza virus was tested in a cell culture-based assay. An MDCK (Madin-Darby canine kidney) cell suspension was seeded at a density of 6×10 4 cells / mL in a 96-well plate (Greiner CELLSTAR® black, Sigma-Aldrich® M0562) in 200 μl / well of DMEM medium containing 5% FBS. After 3 days of culture, the monolayer was washed and 100 μl / well of 5000 TCID 50 / mL of strain A / swine / MN / A01483170 / 2014 (H1N1pdm) or 2500 TCID 50 / mL of strain A / swine / Belgium / 113 / 2013 (H3N2) was added and the cells were infected with swine influenza virus (SIV). One hour after infection, NHC was added at 1 / 3 serial dilutions to obtain a test concentration range of 270 μM to 123 nM, and the cells were cultured at 37 °C under 5% CO2 for 1 day. Inhibition of SIV strain replication by NHC was quantified by immunofluorescence assay using a mouse anti-SIV virus-specific monoclonal antibody (MSD AH, #HB-65), followed by Alexa Fluor488 fluorochrome-conjugated goat anti-mouse antibody (Invitrogen) and DAPI (Sigma Aldrich) nuclear staining. The degree of immunofluorescence was imaged and quantified as relative fluorescence units (RFU) at 488 nm excitation and 525 nm emission using a Cytation5 cell imaging multimode reader and Gen5 software version 3.10 (Agilent BioTek). The effective concentration (EC 50 ) of NHC that reduces SIV replication by 50% was calculated in a four-parameter non-linear regression model using GraphPad Prism8. The effective concentration (EC 50 ) of NHC at which SIV replication is halved was 1.3 μM for the H3N2 strain (Figure 4a) and 5.2 μM for the H1N1pdm strain (Figure 4b).

Claims

1. 1. A method for preventing or treating a viral disease in a non-human animal, comprising administering to the animal a pharmaceutical composition comprising the antiviral nucleoside β-D-N(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier; And wherein said viral disease is caused by swine influenza virus, porcine rotavirus, feline infectious peritonitis virus (FIPV) or feline calicivirus.

2. 10. The method of claim 1, wherein said administration is by oral, topical, or injection route, preferably oral.

3. 10. The method of claim 1, wherein the animal is a pig and the disease is caused by a swine influenza virus or a porcine rotavirus.

4. 2. The method of claim 1, wherein the animal is a cat and the disease is caused by feline infectious peritonitis virus or feline calicivirus.

5. The method of any one of claims 1 to 4, wherein the pharmaceutical composition is administered in a single dose or in multiple doses administered in multiple doses.

6. 1. A method for preventing or treating a viral disease in a non-human animal, comprising administering to the animal a pharmaceutical composition comprising β-DN(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier; and wherein said viral disease is caused by SARS-CoV-2 infection in said animal, and said animal is a pig, cow, horse, dog, or cat.

7. 1. A pharmaceutical composition for use in the prevention or treatment of a viral disease in an animal, comprising the antiviral nucleoside β-DN(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier, wherein the animal is a pig or a cat, and the animal is suffering from or susceptible to a viral disease caused by swine influenza virus, porcine rotavirus, feline infectious peritonitis virus or feline calicivirus.

8. 1. A pharmaceutical composition for inducing an antiviral response in an animal, comprising the antiviral nucleoside β-DN(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier, wherein the animal is a pig or a cat, the animal is suffering from or susceptible to a viral disease, and the viral disease is caused by swine influenza virus, porcine rotavirus, feline infectious peritonitis virus, or feline calicivirus; Pharmaceutical compositions.

9. 9. The pharmaceutical composition of claim 8, wherein the antiviral response results in the viral load in the animal remaining substantially the same.

10. 9. The pharmaceutical composition of claim 8, wherein the antiviral response is a reduction in the amount of virus in the animal.

11. 9. The pharmaceutical composition of claim 8, wherein the antiviral response is a substantial elimination of the viral load in the animal.

12. 1. A method for treating a viral infection, comprising administering to a non-human animal a pharmaceutical composition comprising the antiviral nucleoside β-DN(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier; wherein the viral infection is selected from the group consisting of porcine rotavirus, swine influenza virus, feline infectious peritonitis virus (FIPV) or feline calicivirus. method.

13. 1. A method for providing antiviral prophylaxis, comprising administering to a non-human animal a pharmaceutical composition comprising the antiviral nucleoside β-DN(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier; wherein the viral infection is selected from the group consisting of porcine rotavirus, swine influenza virus, feline infectious peritonitis virus (FIPV) or feline calicivirus. method.

14. 1. A method for preventing or treating a viral disease in a non-human animal, comprising administering to the animal a pharmaceutical composition comprising the antiviral nucleoside β-D-N(4)-hydroxycytidine, or a prodrug or salt thereof, and a pharmaceutically acceptable carrier; And wherein the viral disease is caused by a coronavirus.

15. 15. The method of claim 14, wherein the animal is a cat or a pig.

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