Compositions and methods for treating canine parvovirus infection

Abstract

Provided herein are treatments that reduce the symptoms associated with canine parvovirus infection, shorten the time to recovery, and reduce the time that infected dogs spend in hospital isolation. TIFF2024507417000002.tif76128

Description

[Technical Field]

[0001] Reference to Related Application This application claims the benefit of priority from U.S. Provisional Application No. 63 / 153,230, filed on February 24, 2021, the entire content of which is incorporated herein by reference.

[0002] 1. Field The present invention relates generally to the field of veterinary medicine. More specifically, the invention relates to compositions and methods for treating Canine parvovirus. [Background Art]

[0003] 2. Description of Related Art Canine parvovirus is a particularly lethal disease among young puppies, with a mortality rate of about 80%, causing gastrointestinal disorders, dehydration, and even cardiac syndrome in young puppies. It is transmitted by contact with feces from infected dogs. Symptoms include lethargy, severe diarrhea, fever, vomiting, loss of appetite, and dehydration. Current treatments for canine parvovirus infection are non-specific, primarily including supportive care and prevention of secondary infections, and usually require prolonged hospitalization in an isolation room to prevent the spread of the highly contagious virus. New therapeutic approaches are needed to treat dogs with canine parvovirus infection. [Summary of the Invention]

[0004] Summary In one aspect, there is provided herein a pharmaceutical composition comprising an iron-containing compound, a pharmaceutical excipient, and optionally glucose. In some aspects, the iron-containing compound is a ferric-containing compound. In some aspects, the ferric-containing compound is ferric citrate. In some aspects, the glucose is D-glucose. In some aspects, the pharmaceutical composition is formulated for oral administration.

[0005] In some aspects, the pharmaceutical composition further comprises a salt solution. In some aspects, the salt solution comprises one or more of the following: sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, and sodium bicarbonate. In some aspects, the pharmaceutical composition comprises ferric citrate, sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, sodium bicarbonate, and D-glucose.

[0006] In one embodiment, this specification provides a method for treating a disease or disorder in a subject in need, the method comprising the step of administering a therapeutically effective amount of any one of the pharmaceutical compositions of this embodiment to the subject. In some aspects, the subject has a viral infection caused by a virus that uses the iron transport pathway. In some aspects, the subject has a Parvoviridae infection. In some aspects, the subject is a dog with a parvovirus infection. In some aspects, the dog is a young puppy and / or weighs less than 5 pounds. In some aspects, this method shortens the course of the disease. In some aspects, this method improves many of the symptoms of canine parvovirus infection, including vomiting, diarrhea, lethargy, and loss of appetite. In some aspects, the dog also has a hookworm infection. In some aspects, the administration is performed orally. In some aspects, the administration is performed twice daily. In some aspects, the subject is a cat with a Parvoviridae infection. In some cases, the subject is a human with an Arenaviridae infection. In some cases, the subject has a bacterial infection caused by pathogenic bacteria that use siderophores to sequester iron. In some cases, the subject has gastroenteritis or anemia.

[0007] [Invention 1001] A method for treating Parvoviridae infections in dogs, The step of administering a therapeutically effective dose of a pharmaceutical composition containing iron-containing compounds and pharmaceutical additives. Methods that include... [Invention 1002] The method of the present invention 1001, wherein the iron-containing compound is a ferric-containing compound. [Invention 1003] The method of the present invention 1002, wherein the ferric-containing compound is ferric citrate. [Invention 1004] The method of the present invention 1001, wherein the dog is a young puppy and / or weighs less than 5 pounds. [Invention 1005] A pharmaceutical composition comprising an iron-containing compound, a pharmaceutical additive, and optionally glucose. [Invention 1006] A pharmaceutical composition according to the present invention 1005, wherein the iron-containing compound is a ferric-containing compound. [Invention 1007] A pharmaceutical composition according to Invention 1006, wherein the ferric-containing compound is ferric citrate. [Invention 1008] A pharmaceutical composition according to any one of the present invention 1005 to 1007, wherein glucose is D-glucose. [Invention 1009] A pharmaceutical composition according to any one of the present invention 1005 to 1008, further comprising a salt solution. [Invention 1010] A pharmaceutical composition according to Invention 1009, wherein the salt solution comprises one or more of the following: sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, and sodium bicarbonate. [Invention 1011] A pharmaceutical composition according to any of the present invention 1005 to 1010, comprising ferric citrate, sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, sodium bicarbonate, and D-glucose. [Invention 1012] A pharmaceutical composition according to any of the present invention 1005 to 1011, formulated for oral administration. [Invention 1013] A method for treating a disease or disorder in a person who needs it, A step of administering a therapeutically effective amount of any of the pharmaceutical compositions described in items 1005 to 1012 of the present invention to the subject. Methods that include... [Invention 1014] The method of the present invention 1013, wherein the subject has a viral infection caused by a virus that uses iron transport routes. [Invention 1015] The method of the present invention 1013 or 1014, wherein the subject has a parvoviridae infection. [Invention 1016] The method according to any one of the present invention 1013 to 1015, wherein the subject is a dog having a parvovirus infection. [Invention 1017] The method of the present invention 1016, wherein the dog is a young puppy and / or weighs less than 5 pounds. [Invention 1018] A method according to the present invention 1016 for shortening the course of a disease. [Invention 1019] The method of the present invention 1016, which improves many of the symptoms of canine parvovirus infection, including vomiting, diarrhea, lethargy, and loss of appetite. [Invention 1020] The method according to any one of the present invention 1016 to 1019, wherein the dog further has a hookworm infection. [Invention 1021] Any method of the present invention 1013 to 1020, wherein the administration step is performed orally. [Invention 1022] The method of any of the present invention 1013 to 1015 and 1021, wherein the subject is a cat having a parvoviral infection. [Invention 1023] The method according to any one of 1013, 1014, and 1021 of the present invention, wherein the subject is a human having an Arenaviridae infection. [Invention 1024] The method of the present invention 1013, wherein the subject has a bacterial infection caused by pathogenic bacteria that use siderophores to enclose iron. [Invention 1025] The method of the present invention 1013, wherein the subject has gastroenteritis or anemia. Other objects, features, and advantages of the present invention will become apparent from the following detailed description. However, it should be understood that the detailed description and specific examples, while illustrating preferred embodiments of the present invention, are provided for illustrative purposes only; for various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. [Brief explanation of the drawing]

[0008] The following drawings constitute part of this specification and are included to further illustrate specific aspects of the invention. The invention can be better understood by referring to one or more of these drawings in conjunction with the detailed description of the specific embodiments presented herein. [Figure 1] Figure 1 shows that iron availability improves cell survival in the presence of CPV infection. [Figure 2] Figure 2 shows that, in the absence of CPV virus infection, the addition of ferric citrate does not change cell viability. [Figure 3] Figure 3 shows that treating dogs with increasing concentrations of ferric citrate increases their survival rate from spontaneous infection with CPV. [Figure 4] Figure 4 shows that there was no significant difference in median body weight among puppies treated with each concentration of ferric citrate. [Modes for carrying out the invention]

[0009] Detailed explanation This specification provides a solution containing an iron-containing compound, such as ferric citrate, in a glucose-containing salt solution. This specification also provides a method for treating canine parvovirus using this solution. In vitro, this solution protects against cell death induced by canine parvovirus. When this solution is administered orally to dogs naturally infected with canine parvovirus, many of the symptoms of canine parvovirus infection, including vomiting, diarrhea, lethargy, and loss of appetite, are improved, the course of the disease is shortened, and complete recovery begins as quickly as 12 hours after the first treatment.

[0010] I. Canine parvovirus As will be further discussed below, current treatments for canine parvovirus infection are nonspecific and mainly consist of supportive care and prevention of secondary infections, and usually require long-term hospitalization in isolation to prevent the spread of the highly contagious virus. This specification provides a specific treatment that reduces the symptoms associated with canine parvovirus infection, shortens the time to recovery, and reduces the amount of time infected dogs spend in isolation.

[0011] Canine parvovirus (CPV, CPV2, or parvo) is a contagious DNA virus belonging to the Parvoviridae family that primarily infects dogs. However, CPV can also infect other mammals, such as foxes, wolves, cats, and skunks. CPV is highly contagious and spreads from dog to dog through direct or indirect contact with dog feces. While this infection can be prevented by vaccination, the mortality rate can reach 91% if left untreated. Treatment often requires hospitalization at a veterinary hospital.

[0012] There are two types of CPV, called canine minute virus (CPV1) and CPV2. CPV2 causes the most serious illness and infects domestic dogs and wild canids. In severe cases, dogs may die within 48 to 72 hours without fluid therapy. In the more common and less severe form, the mortality rate is about 10%. Factors such as age, breed, stressful environments, bacteria, parasites, and co-infection with canine coronaviruses increase a dog's risk of developing a severe infection.

[0013] Dogs that develop symptoms will show signs of illness within 3 to 7 days. These signs include lethargy, vomiting, fever, and diarrhea (usually with blood). Generally, the first sign of CPV is lethargy. Secondary signs include weight loss and loss of appetite, or diarrhea followed by vomiting. Diarrhea and vomiting can cause dehydration, disrupting electrolyte balance and potentially having serious consequences for the dog. Secondary infections occur as a result of a weakened immune system. The normal intestinal lining is also damaged, allowing blood and protein to leak into the intestines, leading to anemia and protein loss, and endotoxins to leak into the bloodstream, causing endotoxemia.

[0014] Diagnosis is made by detecting CPV in the stool using either ELISA or hemagglutination testing, or by electron microscopy. PCR is now available for the diagnosis of CPV and can be used in the later stages of the disease; in the later stages, the amount of virus shed in the stool may be less, and it may not be detectable by ELISA.

[0015] Survival rates vary depending on how early CPV is diagnosed, the dog's age, and how aggressive the treatment is. There is no approved treatment, and the current standard of care is supportive, requiring prolonged hospitalization due to the possibility of severe dehydration and damage to the intestines and bone marrow.

[0016] II. Therapeutic compositions This specification provides therapeutic compositions comprising an iron-containing compound, such as ferric citrate, in a salt solution, which optionally contains glucose.

[0017] In some contexts, iron-containing compounds are ferric compounds, such as ferric citrate. Ferric citrate, or iron(III) citrate, represents one of several complexes formed by the bonding of a ferric ion to one of several conjugate bases derived from citric acid. These complexes contain two or more Fe(III) centers. In some contexts, iron-containing compounds are ferrous ascorbate, ferrous carbonate, ferrous citrate, ferrous fumarate, ferrous gluconate, ferrous lactate, ferrous succinate, ferrous sulfate, ferrous glycinate, ferric ammonium citrate, ferric chloride, ferric oxide, ferric phosphate, ferric polymaltose, ferric protein succinylate, ferric pyrophosphate, sodium ferric pyrophosphate, or ferric sulfate.

[0018] In some cases, the therapeutic composition further comprises a salt solution containing one or more of the following: sodium chloride (NaCl), potassium chloride (KCl), monobasic potassium phosphate (KH2PO4), dibasic sodium phosphate (Na2HPO4), magnesium sulfate (MgSO4), calcium chloride (CaCl2), and sodium bicarbonate (NaHCO3).

[0019] In some cases, the therapeutic composition further includes glucose, such as D-glucose.

[0020] In some cases, the therapeutic composition contains ferric citrate (C6H5FeO7), sodium chloride (NaCl), potassium chloride (KCl), monobasic potassium phosphate (KH2PO4), dibasic sodium phosphate, anhydrous (Na2HPO4), magnesium sulfate, anhydrous (MgSO4), calcium chloride, anhydrous (CaCl2·2H2O), sodium bicarbonate (NaHCO3), and D-glucose.

[0021] In some cases, the therapeutic composition contains 2-50 g / L of ferric citrate (C6H5FeO7) (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 25, 30, 35, 40, 45, or 50 g / L, or any value derived therefrom, e.g., 15 g / L) and 0-0.5 g / L of sodium chloride (e.g., 0.01, 0.1, 0.2, 0.3, 0.4, or 0.5 g / L, or any value derived therefrom, e.g., 0.185 g / L) NaCl, potassium chloride (KCl) in concentrations of 0-0.5 g / L (e.g., 0.01, 0.1, 0.2, 0.3, 0.4, or 0.5 g / L, or any value derived therefrom, e.g., 0.4 g / L), monobasic potassium phosphate (KH2PO4) in concentrations of 0-0.2 g / L (e.g., 0.01, 0.02, 0.05, 0.1, or 0.2 g / L, or any value derived therefrom, e.g., 0.06 g / L), 0-0.2 g / L (e.g., 0.01, 0.02, 0.05, Dibasic sodium phosphate, anhydrous (Na2HPO4) in 0.1 or 0.2 g / L, or any value derived therefrom, e.g., 0.048 g / L; magnesium sulfate, anhydrous (MgSO4) in 0-0.2 g / L (e.g., 0.01, 0.02, 0.05, 0.1, or 0.2 g / L, or any value derived therefrom, e.g., 0.098 g / L); 0-0.5 g / L (e.g., 0.01, 0.1, 0.2, 0.3, 0.4, or 0.5 g / L, or from there) It contains anhydrous calcium chloride (CaCl2·2H2O) in any derived value, e.g., 0.185 g / L, sodium bicarbonate (NaHCO3) in 0 to 0.5 g / L (e.g., 0.01, 0.1, 0.2, 0.3, 0.4, or 0.5 g / L, or any derived value therefrom, e.g., 0.35 g / L), and D-glucose in 0 to 5 g / L (e.g., 0.1, 0.5, 1, 2.5, or 5 g / L, or any derived value therefrom, e.g., 1 g / L).

[0022] In some cases, therapeutic compositions are formulated for oral administration. The therapeutic mixture is prepared as a liquid concentrate, which may be packaged in individual (i.e., single-use) unit doses or in multi-use dose forms. The concentrated liquid formulation may contain the therapeutic agent and a solvent (e.g., an organic solvent or an aqueous solvent). A liquid composition for oral administration can be obtained by mixing the concentrated liquid formulation with an aqueous medium.

[0023] In another aspect, when administered to patients requiring such treatment, the therapeutic composition (also called a pharmaceutical preparation, pharmaceutically appropriate composition, pharmaceutical preparation, pharmaceutical product, drug, medicine, agent, or pharmacopoeia) contains a therapeutically effective amount of the iron-containing compound disclosed herein, formulated with one or more pharmaceutical excipients and / or drug carriers suitable for the indicated route of administration. In some embodiments, the iron-containing compound disclosed herein is formulated in a manner suitable for the treatment of human and / or animal patients. In some embodiments, formulation involves mixing or combining one or more iron-containing compounds disclosed herein with one or more of the following pharmaceutical excipients: lactose, sucrose, starch powder, cellulose esters of alkanates, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric acid and sulfate, gelatin, acacia, sodium alginate, polyvinylpyrrolidone, and / or polyvinyl alcohol. In some embodiments, for example, for oral administration, the pharmaceutical preparation may be in tablet, encapsulated, or liquid form. In some embodiments, the compound may be dissolved or slurryed in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, and / or various buffers. In some embodiments, the pharmaceutical formulation may be subjected to pharmaceutical operations such as sterilization and / or may contain drug carriers and / or pharmaceutical additives, such as preservatives, stabilizers, wetting agents, emulsifiers, encapsulating agents such as lipids, dendrimers, and polymers, proteins such as albumin, nucleic acids, and buffers.

[0024] In one embodiment, the therapeutic composition is prepared by dissolving ferric citrate (C6H5FeO7) in a solution containing sodium chloride (NaCl), potassium chloride (KCl), monobasic potassium phosphate (KH2PO4), dibasic sodium phosphate, anhydrous (Na2HPO4), magnesium sulfate, anhydrous (MgSO4), calcium chloride, anhydrous (CaCl2·2H2O), sodium bicarbonate (NaHCO3), and D-glucose.

[0025] In some cases, liquid oral therapeutic compositions include at least one buffering agent. Examples of buffering agents include, but are not limited to, sodium bicarbonate, potassium bicarbonate, magnesium hydroxide, magnesium lactate, magnesium gluconate, other magnesium salts, aluminum hydroxide, aluminum hydroxide / sodium bicarbonate coprecipitate, mixtures of amino acids and buffers, mixtures of aluminum glycinate and buffers, mixtures of amino acid salts and buffers, and mixtures of amino acid alkali salts and buffers. Additional buffering agents include, but are not limited to, sodium citrate, sodium tartrate, sodium acetate, sodium carbonate, sodium carbonate, sodium polyphosphate, potassium polyphosphate, sodium pyrophosphate, potassium pyrophosphate, disodium hydrogen phosphate, dipotassium hydrogen phosphate, trisodium phosphate, tripotassium phosphate, sodium acetate, potassium metaphosphate, magnesium oxide, magnesium hydroxide, magnesium carbonate, magnesium silicate, calcium acetate, calcium glycerophosphate, calcium chloride, calcium hydroxide, calcium lactate, calcium carbonate, calcium bicarbonate, and other calcium salts.

[0026] In some aspects, the liquid oral therapeutic composition comprises at least one pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier of the liquid oral therapeutic composition may contain a bicarbonate of a Group IA metal as a buffer, which can be prepared by mixing a bicarbonate of a Group IA metal, preferably sodium bicarbonate, with water. The concentration of the bicarbonate of a Group IA metal in the composition is generally in the range of about 5.0% to about 60.0%.

[0027] Furthermore, various additives can be incorporated into the therapeutic composition to enhance its stability, sterility, and isotonicity. Antimicrobial preservatives, antioxidants, chelating agents, and additional buffers may be added. Various antimicrobial and antifungal agents, such as parabens, chlorobutanol, phenol, and sorbic acid, can enhance the prevention of microbial activity.

[0028] In some cases, it may be desirable to include isotonic agents, such as sugars or sodium chloride. Furthermore, thickeners such as methylcellulose may also be desirable.

[0029] In some cases, the therapeutic composition can be alternatively formulated as a powder, tablet, suspension tablet, chewable tablet, capsule, two-part tablet or capsule, effervescent powder, effervescent tablet, pellet, and granule.

[0030] Pharmaceutical formulations can be administered in various ways, for example, orally or by injection (e.g., subcutaneous, intravenous, or intraperitoneal injection). Depending on the route of administration, the iron-containing compounds disclosed herein may be coated with a substance to protect them from the action of acids and other natural conditions that may inactivate them. To administer the active compound by means other than parenteral administration, it may be necessary to coat the compound with a substance to prevent its inactivation or to co-administer the compound with that substance. In some embodiments, the active compound may be administered to the patient in a suitable carrier, such as a liposome or diluent. Pharmaceutically acceptable diluents include physiological saline and aqueous buffer solutions.

[0031] The iron-containing compounds disclosed herein may also be administered parenterally, intraperitoneally, intrathecally, or intracerebrally. Dispersions may be prepared in glycerol, liquid polyethylene glycol, and mixtures thereof, or in oil. Under normal storage and use conditions, these formulations may contain preservatives to prevent microbial growth.

[0032] Pharmaceutical compositions suitable for injection include sterile aqueous solutions (if water-soluble) or dispersions, and sterile powders for the immediate preparation of sterile injections or dispersions. The carrier may be a solvent or dispersion medium, for example, containing water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. Appropriate fluidity can be maintained, for example, by the use of coating agents such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants. Prevention of microbial activity can be achieved by various antimicrobial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, and thimerosal. Often, it is preferable to include isotonic agents in the composition, such as sugars, sodium chloride, or polyhydric alcohols such as mannitol or sorbitol. Sustained absorption of the injection composition can be achieved by including substances that slow absorption, such as aluminum monostearate or gelatin, in the composition.

[0033] The iron-containing compounds disclosed herein can be administered orally, for example, with an inert diluent or an absorbable food carrier. The compounds and other components can also be encapsulated in hard or soft-shell gelatin capsules, compressed into tablets, or mixed directly into the patient's diet. For oral therapeutic administration, the compounds disclosed herein can be used in combination with pharmaceutical excipients in the form of ingestible tablets, buccal tablets, lozenges, capsules, elixirs, suspensions, syrups, wafers, and the like. The proportion of the therapeutic compound in the composition and formulation can, of course, vary. The amount of the therapeutic compound in such a pharmaceutical formulation is such that an appropriate dose is obtained.

[0034] Iron-containing compounds can also be administered topically to the skin, eyes, ears, or mucous membranes. Topical administration of therapeutic compounds may include formulations of the compound as topical solutions, lotions, creams, ointments, gels, foams, transdermal patches, or tinctures. When a therapeutic compound is formulated for topical administration, it may be combined with one or more substances that increase the permeability of the compound from the tissue to which it is administered. In other embodiments, topical administration may be to the eyes. Such administration may be applied to the surface of the cornea, conjunctiva, or sclera. While not bound by any theory, administration to the surface of the eye is thought to allow the therapeutic compound to reach the posterior part of the eye. Topical administration to the eye may be formulated as a solution, suspension, ointment, gel, or emulsion. Finally, topical administration may also include administration to mucous membranes, such as in the mouth. Such administration may be done directly to specific locations within the mucous membrane, such as teeth, painful areas, or ulcers. Alternatively, if local delivery to the lungs is desired, the therapeutic compound can be administered by inhalation in the form of a dry powder or aerosol.

[0035] III. Treatment method This specification provides a method for treating certain viral infections, bacterial infections, and / or anemia using a solution containing an iron-containing compound, such as ferric citrate, in a glucose-containing salt solution. In a preferred embodiment, this method treats canine parvovirus. In other embodiments, this method also treats other viruses that utilize the iron transport pathway (for example, by binding to transferrin receptors), such as other members of the Parvoviridae family, such as feline panleukopenia virus, and members of the Arenaviridae family, which cause acute viral hemorrhagic fever in humans.

[0036] In some cases, the aforementioned method treats certain pathogenic bacterial infections because many pathogenic bacteria utilize siderophores to encapsulate iron from the host, potentially leading to local (near infection) or systemic iron deficiency in the host. Local iron availability is crucial, for example, for macrophage function (e.g., by the production of reactive oxygen species) and CD4+ T cell activation and proliferation. In the case of bacterial infections, this treatment can be applied orally, as in the treatment of canine parvovirus infection, to treat intestinal-specific or systemic infections, or applied topically in combination with antibiotic therapy.

[0037] In some cases, the method treats gastroenteritis of unknown cause (negative for canine parvovirus). In other cases, the method treats dogs that are anemic due to hookworm infection, which increases their risk of serious complications from canine parvovirus infection; this is because hookworms attach directly to the intestinal wall and draw blood from the host (dog), causing considerable intestinal pain and anemia.

[0038] In a preferred embodiment, the therapeutic composition provided herein is used to treat canine parvovirus infection, including in very young / small puppies (<5 pounds) that would otherwise die immediately from parvovirus infection despite hospitalization. In this preferred embodiment, the therapeutic composition is provided as an easily dispensable solution for oral administration. In some aspects, the method increases the patient's chances of survival.

[0039] Such treatments can also be used in combination with other treatment regimens, such as supportive care. Supportive care may consist of crystalloid IV infusions and / or colloids (e.g., hetastarch), antiemetic injections (e.g., maropitant, metoclopramide, drasetron, ondansetron, and prochlorperazine), and broad-spectrum antibiotic injections (e.g., cefazolin / enrofloxacin, ampicillin / enrofloxacin, metronidazole, thimentine, or enrofloxacin). IV infusions may be administered, and antiemetic injections and antibiotic injections may be administered subcutaneously, intramuscularly, or intravenously. The infusion may be a mixture of sterile balanced electrolyte solution and appropriate amounts of B vitamins, glucose, and potassium chloride. Analgesics may also be used to relieve the intestinal discomfort caused by frequent bouts of diarrhea.

[0040] Once the dog is able to retain fluids in its stomach, intravenous fluids can be gradually discontinued, and non-irritating foods can be slowly introduced. Depending on the white blood cell count and the patient's ability to prevent secondary infections, oral antibiotics may be administered for several days.

[0041] The therapeutic composition is intended for single or multiple doses. The preferred time interval for multiple doses can be determined by those skilled in the art simply by using routine tests. For example, the patient may be administered twice daily, approximately 12 hours apart. In some embodiments, the drug is administered once daily.

[0042] In various situations, preferred doses are ferric citrate of 0.1–10 mg / kg, 0.22–10 mg / kg, 0.1–5.5 mg / kg, or 0.22–5.5 mg / kg. Dosages may be ferric citrate of 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, or 10 mg / kg, or any value derived therefrom, such as 0.22, 1.1, 2.2 mg / kg. Dosages may be administered twice daily for 1, 2, 3, 4, 5 days, or longer.

[0043] Therapeutic compositions may be administered on a routine schedule. As used herein, a routine schedule refers to a predetermined period of time. A routine schedule may encompass periods of exactly the same length or different lengths, as long as the schedule is predetermined. For example, a routine schedule may include administration twice a day, daily, every two days, every three days, every four days, every five days, every six days, weekly, monthly, or any set number of days or weeks in between. Alternatively, a predetermined routine schedule may include administration twice a day for the first week, and once a day for the following months. In other embodiments, the present invention provides that therapeutic compositions may be taken orally, and that the timing of such administration may or may not depend on food intake.

[0044] The exact amount of therapeutic composition administered is determined at the physician's discretion and is specific to each patient. Various factors that influence the dosage include the patient's physical and clinical condition, the route of administration, the intended therapeutic goal, and the potency, stability, and toxicity of the particular therapeutic formulation.

[0045] The actual dose of the therapeutic composition of this disclosure administered to a patient will be determined by physical and physiological factors, such as the species, age, sex, weight, severity of symptoms, type of disease being treated, previous or simultaneous therapeutic interventions, the patient's idiopathic disease, and route of administration. These factors can be determined by those skilled in the art. Typically, the physician responsible for administration will determine the concentration of the active ingredient(s) in the composition and the appropriate dose(s) for the individual patient. If any complications occur, the individual physician may adjust the dose.

[0046] IV. Definition The terms “comprise,” “have,” and “include” are open-ended linking verbs. Any one or more forms or tenses of these verbs, such as “comprises,” “comprising,” “has,” “having,” “includes,” and “including,” are also open-ended. For example, the way in which one or more steps “comprises,” “has,” or “includes” is not limited to possessing only those one or more steps, but also covers other steps that are not listed.

[0047] Where the term “effective” is used herein and / or in the claims, it means sufficient to achieve the desired, expected, or intended result. Where used in the context of treating a patient or subject with an active ingredient, “effective dose,” “therapeutic dose,” or “pharmaceutical dose” means the amount of the active ingredient that, when administered to the patient or subject, is sufficient to achieve the treatment or prevention of a disease (these terms are defined below).

[0048] A "pharmaceutical excipient" is a pharmaceutically acceptable substance that is formulated together with the active ingredient(s) of a pharmaceutical, pharmaceutical composition, formulation, or drug delivery system. Pharmaceutical excipients may be used, for example, to stabilize a composition, to increase its volume (and therefore often called a "bulker," "filler," or "diluent" when used for this purpose), or to impart therapeutic enhancement effects to the active ingredient in the final dosage form, such as promoting drug absorption, reducing viscosity, or increasing solubility. Pharmaceutical excipients include pharmaceutically acceptable types of anti-sticking agents, binders, coatings, colorants, disintegrants, flavoring agents, flow enhancers, lubricants, preservatives, adsorbents, sweeteners, and vehicles. The main pharmaceutical excipient that acts as a medium for carrying the active ingredient is usually called a vehicle. Pharmaceutical excipients may also be used in the manufacturing process to facilitate the handling of active ingredients, for example, by promoting the fluidity or non-stickiness of powders, in addition to assisting in vitro stability, such as preventing denaturation or aggregation during the expected storage period. The suitability of pharmaceutical excipients will typically depend not only on the route of administration, dosage form, and active ingredient, but also on other factors.

[0049] As used herein, “pharmaceutically acceptable” means a compound, material, composition, and / or dosage form that, within the bounds of sound medical judgment, is suitable for use in contact with human and animal tissues, organs, and / or bodily fluids, without excessive toxicity, irritation, allergic reactions, or other problems or complications, in proportion to a reasonable benefit-to-risk ratio. Furthermore, in the case of administration to animals, it will be understood that the formulation must meet the sterility, pyrogenicity, general safety, and purity standards required by the FDA Office of Biological Standards.

[0050] As used herein, “pharmaceutically acceptable carriers” include all aqueous solvents (e.g., water, alcoholic / aqueous solutions, saline, parenteral vehicles, e.g., sodium chloride, Ringer's solution, glucose), non-aqueous solvents (e.g., propylene glycol, polyethylene glycol, vegetable oils, and organic esters for injection, e.g., ethyl oleate), dispersion media, coatings, surfactants, antioxidants, preservatives (e.g., antimicrobial or antifungal agents, antioxidants, chelating agents, and inert gases), isotonic agents, absorption retarders, salts, drugs, drug stabilizers, gels, binders, excipients, disintegrants, lubricants, sweeteners, flavorings, pigments, water and nutritional supplements, similar materials and combinations thereof as known to those skilled in the art. The pH and precise concentrations of various components in a pharmaceutical composition are adjusted according to well-known parameters.

[0051] A "therapeutic composition" (also called a pharmaceutical, pharmaceutical preparation, pharmaceutical composition, compound, pharmaceutical preparation, pharmaceutical product, drug, medication, medicament, or simply drug, drug, or preparation) is a composition used for the diagnosis, cure, treatment, or prevention of a disease, and contains an active pharmaceutical ingredient (API) and optionally one or more inactive components, also called pharmaceutical excipients.

[0052] "Prevention" or "prevention" includes: (1) suppressing the onset of a disease in subjects or patients who may be at risk and / or predisposed to the disease but have not yet experienced or manifested any or all of the pathology or symptoms of the disease; and / or (2) delaying the onset of the pathology or symptoms of the disease in subjects or patients who may be at risk and / or predisposed to the disease but have not yet experienced or manifested any or all of the pathology or symptoms of the disease.

[0053] "Treatment" or "to treat" includes: (1) suppressing a disease in a subject or patient experiencing or exhibiting its pathology or signs (e.g., preventing further progression of the pathology and / or signs); (2) improving a disease in a subject or patient experiencing or exhibiting its pathology or signs (e.g., reversing the pathology and / or signs); and / or (3) resulting in a measurable reduction of the disease or its symptoms in a subject or patient experiencing or exhibiting its pathology or signs.

[0054] As used throughout this application, the terms “therapeutic benefit” or “therapeutic effective” refer to anything that promotes or enhances the well-being of the subject in relation to the medical treatment of the disease. This includes, but is not limited to, a reduction in the frequency or severity of the signs or symptoms of the disease.

[0055] Patient responsiveness to treatment refers to the clinical or therapeutic benefit provided to a patient who is at risk of or has a disease or disability. Such benefits include cellular or biological responses, complete responses, partial responses, disease stability (without progression or relapse), or responses that later relapse.

[0056] The term "unit dose" refers to a formulation of a compound or composition that is prepared in such a manner that it is sufficient to deliver a single therapeutically effective amount of the active ingredient to a patient in a single administration. Such unit dose formulations that can be used include, but are not limited to, a single tablet, capsule, or other oral formulation, or a single vial containing a syringe-injectable liquid or other injectable formulation.

[0057] As used herein, "Canine" includes what is commonly called a dog, but also other members of the Canidae family.

[0058] As used herein, the terms “patient” or “subject” refer to living mammalian organisms, such as humans, monkeys, cattle, sheep, goats, dogs, cats, foxes, wolves, skunks, mice, rats, guinea pigs, or their transgenic species. In certain embodiments, the patient or subject is a dog.

[0059] As used herein, “oral” administration refers to the introduction of a substance, such as a therapeutic composition, into the body of a subject through or via the mouth, and includes swallowing, passage through the oral mucosa (e.g., sublingual absorption or buccal absorption), or both. Intratracheal administration is also a means of oral administration.

[0060] As used herein with respect to a particular component, “essentially absent” means that the particular component is intentionally and completely absent from the composition, and / or present only as a contaminant or in trace amounts. Therefore, the total amount of a particular component resulting from unintentional contamination of the composition is well below 0.05%, preferably less than 0.01%. Most preferably, the amount of the particular component is undetectable by standard analytical methods.

[0061] As used herein, "a" or "an" may mean one or more. As used in the claims, "a" or "an," when used in conjunction with the word "including," may mean one or more.

[0062] The use of the term “or” in the claims is used to mean “and / or” unless it is expressly indicated that it refers only to substitutes, or that the substitutes are not mutually exclusive; however, in this disclosure, the definition of “and / or” referring to only one substitute is supported. As used herein, “another” may mean at least a second or subsequent one.

[0063] Throughout this application, the term “approximately” is used to indicate that the value includes variations in error inherent to the device, variations inherent to the method used to measure the value, variations present between the subjects of study, or values ​​within 10 percent of the stated value.

[0064] The above definitions take precedence over any conflicting definitions in the references incorporated herein by reference. However, the fact that certain terms are defined should not be interpreted as indicating that any undefined terms are indefinite. Rather, all terms used are intended to describe the invention in a manner that enables those skilled in the art to understand its scope and implement it. [Examples]

[0065] V. Examples The following embodiments are included to illustrate preferred embodiments of the present invention. As those skilled in the art will understand, the techniques disclosed in the following embodiments represent techniques found by the inventors to work well in carrying out the present invention and can therefore be considered to constitute preferred forms for carrying out the present invention. However, those skilled in the art should understand that, in light of this disclosure, many modifications can be made to the specific embodiments disclosed without departing from the spirit and scope of the present invention, and similar or comparable results can still be obtained.

[0066] Example 1 - In vitro test Methods: Canine fibroblasts were cultured at 38°C in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 5% fetal bovine serum, 2 mM L-glutamine, 0.1 mM MEM non-essential amino acids, 1 mM sodium pyruvate, 0.05 mM 2-mercaptoethanol, 10 mM HEPES, 100 units / mL penicillin, and 100 mcg / mL streptomycin, under 5% CO2. Canine parvovirus type 2 (CPV2) was isolated from fecal samples collected from naturally infected dogs. The virus was grown in canine fibroblasts and 10 13.37 The titer of tissue culture infectious dose (TCID50) / mL was used in the viability determination assay. In the viability determination assay, cells were periodically passaged and 5 × 10¹⁶ cells were placed in 200 μL of complete DMEM medium in a 96-well plate. 4 Cells were seeded at 1 / mL and incubated overnight at 38°C and 5% CO2. The medium was then removed, and the cells were infected with CPV2 at 38°C and 5% CO2 for 1 hour. After 1 hour, ferric citrate (1 μM or 10 μM), deferoxamine (1 μM), or fresh medium was added to the wells in sets of three, and the cells were cultured for 72 hours. Cell viability was then analyzed using trypan blue exclusion. At least three independent experiments were performed for each assay.

[0067] To investigate the role of iron availability during CPV infection, ferric citrate or deferoxamine was added to CPV-inoculated cell cultures. Cell viability was assayed 72 hours after treatment. Cells infected with CPV alone had a 28% lower viability compared to untreated cells (Figure 1). Increased iron availability due to the addition of 1 μM or 10 μM ferric citrate significantly increased cell viability after CPV infection; conversely, decreased iron availability due to the addition of 1 μM deferoxamine resulted in a 45% decrease in viability compared to untreated cells and a 23% decrease compared to CPV-infected cells. This suggests that iron availability is crucial for cell survival during CPV infection. The data presented are mean ± SEM and pooled from three independent experiments.

[0068] To investigate whether additional ferric citrate affects cell survival in the absence of CPV infection, cells were treated with 1 μM or 10 μM ferric citrate, or 1 μM deferoxamine, for 72 hours, and then cell survival was assayed. In all cases, there was no significant difference in survival between untreated and treated cells (Figure 2). The data shown are mean ± SEM and pooled from three independent experiments.

[0069] Example 2 - In vivo study When this solution is administered orally to dogs naturally infected with canine parvovirus, many of the symptoms of canine parvovirus infection, such as vomiting, diarrhea, lethargy, and loss of appetite, improve, the course of the disease is shortened, and complete recovery begins as quickly as 12 hours after the first treatment.

[0070] Methods: Thirteen puppies with no prior vaccination history against CPV were induced to develop CPV infection via natural routes and treated according to a standard veterinary protocol, which included oral administration of 0.22, 1.1, or 2.2 mg / kg ferric citrate solution twice daily for up to 5 days, followed by monitoring for 14 days.

[0071] Puppies exhibiting symptoms of CPV infection, such as vomiting, diarrhea, bloody stools, lethargy, and loss of energy, were tested for CPV infection using the IDEXX Laboratories SNAP® Parvo Test according to the manufacturer's protocol. If a CPV positive result was confirmed, treatment of the puppies was initiated according to a standard veterinary protocol for CPV infection, including intravenous or subcutaneous fluid administration with the addition of 0.22, 1.1, or 2.2 mg / kg of ferric citrate solution, antiemetics, and antibiotic injections.

[0072] The survival rate for puppies supplemented with the highest concentration of ferric citrate was 100% (5 / 5), while the survival rates for lower concentrations were 40% (2 / 5) and 33.33% (1 / 3), respectively (Figure 3).

[0073] Young puppies with low body weight (less than 5 pounds) are at high risk of death if infected with CPV. The lowest concentration of ferric citrate tested, 0.22 mg / kg, did not prevent death in the smallest puppies (less than 5 pounds), but the highest concentration tested, 2.2 mg / kg, resulted in a 100% survival rate for all puppies; the smallest puppy weighed only 1.6 pounds at the start of treatment (Figure 4).

[0074] All methods disclosed and claimed herein can be carried out and achieved in light of this disclosure without undue experimentation. While the compositions and methods of the present invention are described in terms of preferred embodiments, it will be apparent to those skilled in the art that modifications can be applied to the methods, steps of the method, or order of steps described herein without departing from the concept, spirit, and scope of the invention. More specifically, it will be apparent that certain chemically and physiologically related agents can be used in place of the agents described herein, and the same or similar results can still be obtained. All such similar substitutes and modifications, which are apparent to those skilled in the art, are considered to be within the spirit, scope, and concept of the invention as defined by the appended claims.

Claims

1. A method for treating Parvoviridae infections in dogs, The step of administering a therapeutically effective dose of a pharmaceutical composition formulated for oral administration, which includes an iron-containing compound, ferric citrate, and a pharmaceutical additive. Methods that include...

2. The method according to claim 1, wherein the dog is a young puppy and / or weighs less than 2.268 kg (5 pounds).

3. The method according to claim 1, wherein the pharmaceutical composition further comprises glucose.

4. The method according to claim 3, wherein glucose is D-glucose.

5. The method according to any one of claims 1 to 4, wherein the pharmaceutical composition further comprises a salt solution.

6. The method according to claim 5, wherein the salt solution comprises one or more of sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, and sodium bicarbonate.

7. The method according to any one of claims 1 to 6, comprising ferric citrate, sodium chloride, potassium chloride, monobasic potassium phosphate, dibasic sodium phosphate, magnesium sulfate, calcium chloride, sodium bicarbonate, and D-glucose.

8. The method according to any one of claims 1 to 7, for shortening the course of a disease.

9. The method according to any one of claims 1 to 7, which improves many of the symptoms of canine parvovirus infection, including vomiting, diarrhea, lethargy, and loss of appetite.

10. The method according to any one of claims 1 to 9, wherein the administration step is performed orally.

11. The method according to any one of claims 1 to 10, wherein ferric citrate is administered at a dose of 2.2 mg / kg body weight.

12. The method according to any one of claims 1 to 11, wherein ferric citrate is administered twice a day.

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