Azelastine as an antiviral treatment

Abstract

An azelastine compound in an antivirally effective amount for use as an antiviral agent in a pharmaceutical formulation for use in the prophylactic or therapeutic treatment of a subject in need of antiviral treatment.

Description

[Technical Field]

[0001] The present invention relates to novel uses of antihistamine compounds for treating viral infections, particularly methods and compounds for treating coronavirus infections (e.g., SARS virus or MERS virus), adenovirus infections, human respiratory multinuclear virus infections, and influenza infections. [Background technology]

[0002] Coronaviruses are single-stranded RNA viruses with a diameter of approximately 120 nanometers. Coronaviruses are prone to mutation and recombination and are therefore highly diverse. There are about 40 different variants, which primarily infect humans, non-human mammals, and birds. These variants are found in bats and wild birds and can spread to other animals, and thus to humans.

[0003] Based on their genome structure, there are four main genera (alpha, beta, gamma, and delta coronaviruses). Alpha- and beta coronaviruses infect only mammals, typically causing respiratory symptoms in humans and gastroenteritis in other animals. As of December 2019, only six different coronaviruses were known to infect humans. Four of these (HCoV-NL63, HCoV-229E, HCoV-OC43, and HKU1) typically cause mild, cold-like symptoms in immunocompetent humans, while the remaining two have caused pandemics over the past two decades. In 2002-2003, severe acute respiratory syndrome coronavirus (SARS-CoV) caused the SARS epidemic, resulting in a 10% mortality rate. Similarly, Middle East respiratory syndrome coronavirus (MERS-CoV) caused a pandemic in 2012 with a 37% mortality rate.

[0004] In the latter half of 2019 and the first half of 2020, a novel coronavirus, SARS-CoV-2, was discovered to be closely related to SARS-CoV and was the cause of a large-scale, rapidly spreading respiratory illness, including pneumonia. Since the recognition of the novel coronavirus, the disease it causes has been called coronavirus infection 2019 (COVID-19).

[0005] SARS-related coronaviruses are coated with a spike protein containing a variable receptor-binding domain (RBD). This RBD binds to angiotensin-converting enzyme-2 (ACE-2) receptors found in the heart, lungs, kidneys, and gastrointestinal tract, thereby facilitating viral entry into target cells.

[0006] The initial virus strain that spread from Wuhan was considered a "wild-type" virus, and this virus quickly gave rise to mutant strains, i.e., variants that evolved through natural selection based on higher infectivity. In early March 2020, a mutant strain with a D614G mutation in the spike protein (B.1 mutant) was identified in Europe, and this mutant strain quickly replaced the initial Wuhan strain globally. Subsequently, SARS-CoV-2 mutant strains B.1.1.7 (also known as 20I / 501Y.V1, VOC 202012 / 01) and B.1.351 (also known as 20H / 501Y.V2) were identified in the UK and South Africa, respectively, and have since spread to many countries. These mutant strains have diverse mutations in the gene encoding the spike protein. One of these mutations is located at position 501 of the receptor-binding domain (RBD that binds to human ACE2), where the amino acid asparagine (N) is replaced with tyrosine (Y) (mutation N501Y). The B.1.1.7 mutant also has several other mutations, including a 69 / 70 deletion that likely results in a conformational change of the spike protein, and P681H near the S1 / S2 furin cleavage site. The combination of these mutations results in higher receptor binding, more efficient diffusion, and higher pathogenicity compared to the initial version that appeared in Wuhan, China in 2019. The B.1.351 mutant also has E484K and K417N mutations in addition to N501Y, but lacks the 69 / 70 deletion. Several pieces of evidence suggest that this mutant emerged due to immunosuppression, i.e., as a means of escaping the human immune response resulting from natural infection or active immunity acquisition using passive immunization therapy or different vaccines. The E484K mutation is thought to be responsible for the reduced vaccine effectiveness. This mutation (along with 16 other mutations, N501Y and K417T) was also detected in the Brazilian variant, which was named P.1. The P.1 variant caused widespread infection in Brazil and was later detected worldwide. This variant is suspected to cause more severe illness and a higher mortality rate in relatively younger individuals and to evade the immune response induced by previous variants and some vaccines.

[0007] Mutant B.1.617 was first detected and spread in India. This mutant has 13 mutations, including three mutations of concern regarding immune evasion: E484Q, L452R, and P681R. Strain B.1.618 has become one of the dominant mutants in West Bengal and possesses two amino acid deletions (H146del and Y145del), as well as E484K and D614G mutations in the spike protein. This strain is suspected to be more infective and pose a threat that evades natural or vaccine-induced immunity.

[0008] Widespread vaccination is highly likely to induce the emergence of further mutant strains with different combinations of currently known mutations and new mutations. Accelerated global vaccination, along with the simultaneous and intense spread of the virus, is placing immense evolutionary stress on the emergence of evasive variants. Almost without exception, all vaccines rely on triggering an immune response against the spike protein. Furthermore, according to current views, a neutralizing immune response against the receptor-binding domain of the spike protein is considered a major component of defense. Based on these two facts, among naturally emerging viral variants, those expressing a mutated spike protein that offers a selective advantage (higher binding to the receptor and / or evasion of the neutralizing immune response) are selected. Subsequent variants may accumulate several consecutive mutations that have higher overall pathogenicity / infectivity and / or a higher likelihood of immune evasion. It is anticipated that further new viral variants will emerge in addition to the aforementioned variants, potentially necessitating a second-generation vaccine for optimal protection against such variants.

[0009] Therefore, antiviral compounds that are active against the now-dominant UK(B1.1.7) and SA(B.1.351) variants, as well as other emerging variants associated with significantly reduced vaccine-induced protection, are highly relevant today, both for prevention and treatment.

[0010] Adenoviruses are large, non-enveloped viruses with a double-stranded DNA genome. The Adenoviridae family contains six genera with broad host specificity. All seven species of human adenoviruses (A-G) belong to the genus Mastadenovirus. Currently, human viruses are classified into 88 different (sero) types, and these serotypes cause a variety of mucosal infections. Types within families B (HAdV-B) and C (HAdV-C) cause upper respiratory tract infections, while HAdV-F (mainly types 40 and 41) and HAdV-G (mainly type 52) types cause gastroenteritis. Conjunctivitis is associated with HAdV-B and HAdV-D. These mucosal infections are usually self-healing, but can be more severe in immunocompromised hosts. Adenovirus serotype 14 is a newly emerging pathogen that causes severe respiratory infections that can be fatal even to immunocompetent hosts.

[0011] Adenoviruses exhibit a classic icosahedral capsid composed of 240 hexons and 12 penton proteins. The penton base associates with protruding fibers that bind to host cell receptors: CD46 (family B) and coxsackie / adenovirus receptors (CAR, in all other families). Following this initial binding, the penton base viral structure interacts with αV integrin, which stimulates endocytosis of the viral particle. Inside the cell, the capsid is destabilized, the endosome is degraded, and the viral DNA enters the nucleus through the nuclear pore. After association with histone proteins, the host transcription machinery is used for viral gene expression without the viral genome being integrated into the host genome. Viral proteins are expressed in early (primarily regulatory proteins) and late (structural proteins) phases, separated by genome replication. Finally, the viral genome is packed into a protein shell and released from the host cell in a virus-induced lysis procedure.

[0012] Adenoviruses are relatively resistant to disinfectants and cleaning agents (unencapsulated) and can survive for long periods on surfaces and in water. Since there are no antiviral drugs proven to treat adenovirus infections, treatment is largely focused on managing symptoms. Currently, there are no adenovirus vaccines available to the general public, although vaccines for types 4 and 7 have been used by the U.S. military. More recently, adenoviruses engineered as viral vectors for heterologous vaccine antigen delivery have been successfully used in gene therapy (Ebola and Covid).

[0013] Human respiratory multinuclear virus (hRSV) is an enveloped virus with a negative-strand single-stranded RNA genome. The genome is linear and contains 10 genes encoding 11 proteins. RSV is divided into two antigenic subtypes, A and B, with 16 and 22 clades (or strains), respectively.

[0014] RSV is highly contagious and can cause outbreaks from both community and hospital sources. Approximately 30 million cases of acute respiratory illness and over 60,000 child deaths worldwide are caused by RSV each year. Transmission occurs through contaminated aerosol droplets encountering the mucous membranes of the nose, mouth, or eyes. Infection of the ciliated cells of the upper respiratory tract is followed by spread to the lower respiratory tract. Infection of the ciliated cells of the upper respiratory tract is one of the most common childhood infections, ranging in severity from mild upper respiratory tract infections to viral pneumonia via bronchiolitis, which in the most severe cases may require mechanical ventilation. Immunocompromised individuals (including premature infants) are at higher risk of more severe disease outcomes. Treatment options are typically limited to supportive care, although ribavirin is approved for RSV infections in children. However, a vaccine is not available (despite considerable development efforts), and passive immunization with monoclonal antibodies (palivizumab) has become available as a preventive option.

[0015] Influenza, commonly known as "the flu," is an infectious disease caused by the influenza virus. Each season (winter months in the Northern Hemisphere), 5–15% of the population contracts influenza, with approximately 3–5 million cases being severe. More than 500,000 deaths occur annually among high-risk groups, including infants, the elderly, and those with chronic health conditions. Following an incubation period of 1–4 days, the onset of symptoms is sudden, including fever, chills, headache, muscle aches or pains, loss of appetite, fatigue, and confusion. Pneumonia may be caused by the primary viral infection or by a secondary bacterial infection, such as Streptococcus pneumoniae or Staphylococcus aureus. Influenza is transmitted through aerosol droplets. The primary site of infection is the upper respiratory tract, followed by the lower respiratory tract, where invasive infection occurs.

[0016] There are four types (species) of influenza viruses: A, B, C, and D. Seasonal outbreaks (i.e., influenza seasons) are caused by human influenza A (IAV) and B (IBV) viruses, while C and D are rarely associated with symptomatic infection in humans. Influenza A viruses are divided into subtypes based on two proteins on the viral surface: hemagglutinin (H) and neuraminidase (N). Potentially, there are 198 different influenza A subtype combinations, but currently, H1N1 and H3N2 types circulate worldwide. Influenza B viruses are not divided into subtypes, but instead are further classified into two lineages: B / Yamagata and B / Victoria. Based on these lineages, seasonal influenza vaccines usually contain a combination of two A (H1N1 and H3N2) and one or two B strains. Influenza viruses have a segmented minus-stranded single-stranded RNA genome. Both IAV and IBV contain eight segments, which are thought to be able to combine with the genomes of other influenza viruses in co-infection of the same cells. This process (called reassortment) produces offspring with significantly altered viral antigenic compositions. Such reassorted viral mutants are novel to human populations and can cause pandemics (e.g., the 1918 Spanish flu, H1N1). Reassortment can also occur between viruses that are specific to different hosts. For example, the 2009 "swine flu" pandemic was caused by a triplet conjugation mutant virus (H1N1) possessing a combination of pig-, bird-, and human-specific viral sequences. Similarly, avian influenza strains, commonly found in wild waterfowl, occasionally infect humans ("avian or bird flu," H5N1). When such avian viruses reassort with human influenza viruses, they can produce mutants that enable human-to-human spread and thus have the potential to cause a global pandemic.

[0017] Treatment for individuals in lower-risk groups focuses primarily on symptomatic treatment (fever) and isolation. Patients with severe or progressive clinical illness associated with suspected or confirmed influenza virus infection are treated with antiviral drugs (e.g., oseltamivir or other neuraminidase inhibitors) and supportive care (e.g., anti-inflammatory drugs). High-risk groups (elderly, pregnant women, immunocompromised individuals, and those with associated chronic diseases) should receive seasonal influenza vaccinations annually as a preventive measure.

[0018] Azelastine, a phthalazine derivative, is an antihistamine available as an intranasal spray for the treatment of allergic and vasomotor rhinitis, and as eye drops for the treatment of allergic conjunctivitis. Although azelastine is a racemic mixture, there are no significant differences in pharmacological activity between the enantiomers, and it was first approved by the FDA in 1996.

[0019] Gysi et al. ("Network Medicine Framework for Identifying Drug Repurposing Opportunities for COVID-19," arXiv:2004.07229[q-bio.MN] dated April 15, 2020) disclose a network-based toolset for COVID-19 to arrive at certain drug candidates based on their potential efficacy in COVID-19 patients, which included azelastine, but does not show whether antihistamines directly affect the virus underlying the disease.

[0020] Hasanain Abdulhameed Odhar et al. (Bioinformation 2020 16(3):236-244) describe molecular docking and dynamic simulations of FDA-approved drugs with major proteases derived from the novel coronavirus 2019. The drugs were ranked according to their minimum binding activity to the major protease crystal of 2019-nCoV, with azelastine receiving a less favorable ranking than conivaptan.

[0021] Xia Xiao et al. (bioRxiv, July 6, 2020, DOI: 10.1101 / 2020.07.06.188953) describe the antiviral activity of a series of compounds against OC43, among which azelastine as an antihistamine.

[0022] Fu et al. (Cell Prolif., 2014, 47: 326 - 335) describe azelastine hydrochloride targeting the sodium taurocholate cotransporting polypeptide (NTCP) in hepatitis B virus (HBV) therapy. NTCP is a transmembrane protein highly expressed in human hepatocytes that mediates the transport of bile acids, and this transport plays an important role in the entry of HBV into hepatocytes.

[0023] M.W. Simon (Pediatric Asthma, Allergy & Immunology, 2003, Vol. 16: 275 - 282) evaluates azelastine as a potent and selective second - generation histamine H1 - receptor antagonist that down - regulates the expression of the intracellular adhesion molecule - 1 (ICAM - 1) receptor. The intracellular adhesion molecule - 1 (ICAM - 1) receptor plays an important role in the mucosal adhesion of human rhinovirus, coxsackievirus A, adenovirus type 5, human parainfluenza virus types 2 and 3, and respiratory syncytial virus and the recruitment of immune effector cells. Summary of the Invention Problems to be Solved by the Invention

[0024] In particular, it is an object of the present invention to provide new antiviral treatments, pharmaceutical products and pharmaceuticals that can be used to prevent viral infection and / or viral spread in subjects exposed to or infected with a virus or at risk of infection. The object is solved as described further in the subject matter of the claims and in this specification. Means for Solving the Problems

[0025] The present invention provides an azelastine compound in an antivirally effective amount for use as an antiviral substance in a pharmaceutical formulation for use in preventive or therapeutic treatment of subjects requiring antiviral treatment.

[0026] Specifically, azelastine compounds are azelastine, or pharmaceutically acceptable salts thereof, such as azelastine hydrochloride. In certain embodiments, a pharmaceutical preparation is a medical product or drug. Specifically, a pharmaceutical preparation comprises an azelastine compound and a pharmaceutically acceptable carrier.

[0027] Specifically, the target group requires antiviral treatment that targets respiratory viruses such as influenza virus, polynuclear respiratory viruses, adenoviruses, and coronaviruses, or upper respiratory tract viruses such as rhinoviruses.

[0028] In this specification, respiratory viruses are understood to be, in particular, viruses that cause respiratory illness. Some of the target viruses further described herein may affect other parts of the body in addition to causing respiratory illness, but such viruses are still understood in this specification to be “respiratory viruses.” Specifically, antiviral treatment involves targeting one or more human viruses, in particular human respiratory viruses, such as those selected from the Viridae, Coronaviridae, Adenoviridae, Paramyxoviridae, or Orthomyxoviridae families.

[0029] in particular, a) Preferably, a coronavirus selected from the group consisting of β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, HCoV-HKU1, α-coronaviruses such as HCoV-NL63, HCoV-229E, or PEDV, and naturally occurring variants or mutants of any of the aforementioned viruses, b) Adenoviruses, preferably human adenoviruses such as HAdVB, HAdVC, or HAdVD; c) Human respiratory multinuclear virus (RSV), such as RSV subtype A or B; or d) Influenza viruses such as human influenza viruses, preferably influenza virus A (IVA) such as H1N1, H3N3, or H5N1, or influenza virus B (IVB), or influenza virus C (IVC), or influenza virus D (IVD), A disease state caused by or associated with infection resulting from one or more of these conditions is treated.

[0030] Specifically, the treatment will address disease conditions caused by or associated with infection by one or more coronaviruses, particularly one or more different coronaviruses.

[0031] In a particular embodiment, the coronavirus family virus is preferably selected from the group consisting of β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, or HCoV-HKU1, or α-coronaviruses such as human coronavirus NL63 (HCoV-NL63, New Haven coronavirus), HCoV-229E, or porcine epidemic diarrhea virus (PEDV), and naturally occurring variants or mutants of any of the aforementioned viruses.

[0032] Specifically, the one or more different coronaviruses referred to herein are naturally occurring SARS-CoV-2 mutants or variants, in particular mutants or variants that include one or more mutations in the spike protein (SARS-CoV-2 S-protein), such as one or more of the following mutations: K417N, L452R, N501Y, D614G, P681H, P681R, E484K, E484Q, or 69 / 70 deletions.

[0033] Specifically, the spike protein contains or consists of the amino acid sequence identified as Sequence ID No. 4 (the sequence provided in Figure 5, NCBI accession number QII57161.1, SARS-CoV-2, S-protein).

[0034] Preferably, the one or more different coronaviruses are naturally occurring SARS-CoV-2 mutants or variants selected from the group consisting of the UK (B1.1.7) mutant, the South Africa (B.1.351) mutant, the Brazil (P.1) mutant, the India (B.1.617) mutant, and the Bengal (B.1.618) mutant.

[0035] The formulations, methods, and uses described herein include, in particular, an antiviral effective dose of an azelastine compound. This antiviral effective dose may target one specific virus or more different viruses, viral variants, or viral mutants, such as those further described herein.

[0036] The target viruses for antiviral effects are understood to be viruses that are (directly or indirectly) related to the indications for treatment of disease conditions or diseases caused by or otherwise associated with the treatments described herein.

[0037] Specifically, the targets of the formulations, methods and uses described herein, in particular the targets of the antiviral effects described herein, may be any one or more different virus species, virus variants or virus mutant strains, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 different virus species, variants or mutant strains.

[0038] Specifically, the targets could be any two or more viruses, for example, 2, 3, 4, 5, 6, 7, 8, 9, or 10 different virus species, virus variants, or virus mutant strains, and at least 2, 3, or 4 of those viruses belong to the family: a) Coronaviruses (β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, HCoV-HKU1; or α-coronaviruses such as HCoV-NL63, HCoV-229E, or PEDV, and naturally occurring variants or offshoots of any of the aforementioned viruses); b) Adenoviridae (adenoviruses or human adenoviruses, such as HAdVB, HAdVC, or HAdVD); c) Paramyxoviridae (RSV or human RSV, e.g., hRSV subtype A or B); or d) Orthomyxoviridae (influenza viruses or human influenza viruses, preferably influenza virus A (IVA) such as H1N1, H3N3, or H5N1, or influenza virus B (IVB), or influenza virus C (IVC), or influenza virus D (IVD), etc.) These are different virological families, such as those selected from the above.

[0039] Specifically, the viruses listed in a) through d) above are referred to in this specification as exemplary "respiratory viruses." In specific examples, the target can be at least one virus selected from a) and at least one virus selected from b), c), or d). Specifically, the target can be at least one β-coronavirus such as the SARS virus, particularly SARS-CoV-2, and at least one influenza virus.

[0040] Specifically, the targets could be at least one β-coronavirus, particularly SARS-CoV-2, and at least one adenovirus HAdVB, HAdVC, or HAdVD.

[0041] Specifically, the targets could be SARS viruses, particularly at least one β-coronavirus such as SARS-CoV-2, and at least one hRSV subtype A or B.

[0042] In a specific example, the target could be at least one virus selected from d) and at least one virus selected from a), b), or c). Specifically, the targets can be at least one influenza virus or human influenza virus, and at least one adenovirus HAdVB, HAdVC, or HAdVD.

[0043] Specifically, the targets can be at least one influenza virus or human influenza virus, and at least one hRSV subtype A or B.

[0044] In a specific example, the target could be at least one virus selected from b) and at least one virus selected from a), c), or d). Specifically, the targets can be at least one adenovirus HAdVB, HAdVC, or HAdVD, and at least one hRSV subtype A or B.

[0045] In specific examples, the target could be at least one virus selected from a) and at least one virus selected from b), and optionally at least one virus selected from c) or d). Specifically, the target could be at least one β-coronavirus such as the SARS virus, particularly SARS-CoV-2, and at least one influenza virus, and optionally at least one adenovirus HAdVB, HAdVC, or HAdVD, and / or at least one hRSV subtype A or B.

[0046] The present invention further provides azelastine compounds (as further described herein) in antiviral effective amounts for use as antiviral substances in pharmaceutical formulations for the prophylactic or therapeutic treatment of disease conditions caused by or associated with infection by one or more different viruses of the Coronaviridae family, which are preferably β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, or HCoV-HKU1, or α-coronaviruses such as HCoV-NL63, HCoV-229E, or PEDV, and naturally occurring variants or mutants of any of the aforementioned viruses, for example, preferably one or more naturally occurring SARS-CoV-2 variants or mutants referred to herein, selected from the group consisting of UK(B1.1.7) variant, SA(B.1.351) variant, Brazil(P.1) variant, India(B.1.617) variant, and Bengal(B.1.618) variant.

[0047] Specifically, disease conditions associated with or caused by target viruses, or respiratory viruses described herein, particularly viruses of the Coronaviridae, Adenoviridae, Paramyxoviridae, or Orthomyxoviridae families, include the common cold, nasal, sinusitis, pharyngeal and laryngeal infections, bronchiolitis, diarrhea, skin rashes, or pneumonia, acute respiratory distress syndrome (ARDS). Specifically, disease conditions can be symptoms associated with one or more of the aforementioned viruses. Specifically, symptoms treated can be any of the following: cough, pharyngitis, runny nose, sneezing, headache, and fever.

[0048] In certain embodiments, an effective antiviral dose is effective in preventing infection of susceptible cells by the virus and thereby treating the disease state. Specifically, susceptible cells are located on or within a biological surface or object.

[0049] In a particular embodiment, the effective antiviral dose is 0.1 to 500 μg / dose, preferably less than any one of 100, 90, 80, 70, 50, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, or 1 μg / dose. Specifically, the number of doses is 1 to 10 per day.

[0050] In certain embodiments, the effective antiviral dose is 15 μg to 150 μg per dose, preferably less than 100 μg or less than 50 μg. Specifically, the effective antiviral dose may be much lower, ranging from 15 μg to 150 μg, and less than one of the following percentages: 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1%. These relatively low doses have been particularly proven effective against coronaviruses (or viruses of the Coronaviridae family) and / or viruses other than coronaviruses (or viruses other than Coronaviridae), such as viruses of the Adenoviridae, Paramyxoviridae, or Orthomyxoviridae families.

[0051] For example, the antiviral effect targeting any of the target viruses described herein has been shown to be highly effective at 0.1% (2.39 mM) and 5× dilution (0.02%, or 478 microM) doses of commercially available azelastine nasal spray formulations, thereby reducing the viral load in humans.

[0052] For example, the anti-influenza virus effect has been found to be highly effective with 5× dilutions (0.02%, or 478 microM) and 10× dilutions (0.01%, or 239 microM) of commercially available azelastine preparations (0.1%, or 239 microM).

[0053] For example, the anti-RSV effect has been shown to be highly effective in vitro at concentrations ranging from 0.4 to 6.4 microM. According to certain embodiments, the pharmaceutical formulation is formulated for topical administration or topical mucosal administration, preferably for use in the upper and lower respiratory tracts, nasal cavity, lungs, oral cavity, eyeball, or skin, or preferably for systemic administration by intravenous, intramuscular, subcutaneous, intradermal, transdermal, or oral administration. Typically, for parenteral administration, intravenous or oral administration is preferred.

[0054] In a particular embodiment, the pharmaceutical preparation is administered to a target as a spray such as a nasal spray, a powder such as an instant powder or inhalation powder, or as a health medical device such as a surface or fabric impregnated with an inhalation azelastine compound, a gel, ointment, cream, bubbles, or liquid solution, lotion, mouthwash, aerosolized powder, aerosolized liquid, granules, capsules, drops, tablets, syrup, lozenges, eye drops, or a preparation for injection or injection.

[0055] Specifically, antiviral formulations and dosage forms for veterinary and human use are provided. Specifically, the formulations contain a predetermined amount of an azelastine compound as an active ingredient, for example, as a powder or granules; as a solution or suspension in an aqueous or non-aqueous liquid; or as an oil-in-water or water-in-oil liquid emulsifier.

[0056] In certain embodiments, the azelastine compound is used in an antiviral effective dose that gives a peak (or maximum) concentration in blood or plasma, the concentration of which is approximately 0.01–2 μg / mL, or up to one of 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 μg / mL.

[0057] According to a particular embodiment, the azelastine compound is used in the formulation at a concentration of 1 μM to 10 mM, preferably up to one of 1 mM, 100 μM, 90 μM, 80 μM, 70 μM, 60 μM, or 50 μM. According to a particular embodiment, the concentration is 3 to 50 μM.

[0058] Specifically, the solution or dispersion is used for nasal administration, such as by nasal drops or nasal spray, and preferably the antiviral effective dose is 1 to 1000 μg per nostril, preferably 1 to 500 μg per nostril, or up to one of 200, 190, 180, 170, 160, 150, 140, 130, 120, or 110 per nostril, and more preferably up to 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2, or 1 μg.

[0059] Specifically, doses of approximately 2 to 2000 μg per dose, preferably 2 to 1000 μg per dose, or up to one of 400, 380, 360, 340, 320, 300, 280, 260, 240, or 220, and more preferably up to 200, 180, 160, 140, 120, 100, 80, 60, 40, 20, 10, 5, 4, 3, or 2 μg per dose, can be administered intranasally.

[0060] The formulation is preferably applied as an aerosol, such as a nasal spray, nasal drop, aerosolized liquid or powder, for example, as a throat spray or for intrapulmonary administration, or as eye drops.

[0061] Exemplary formulations may contain an azelastine compound as an active ingredient in amounts ranging from 0.001% to 2% (w / w), such as approximately 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.05%, 0.1%, 0.15%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, or 2.0% (w / w).

[0062] Specifically, a volume of 100-1000 μL per dose can be applied to a sprayable formulation, for example, up to a spray volume of 500 μL. For example, a nasal spray can deliver a volume of approximately 100-150 μL per spray. Typically, it is applied once or twice a day per nostril.

[0063] When using a spray, it is preferable to use a measuring spray to dispense a specific amount or dose of spray solution per puff. Formulations suitable for intrapulmonary administration may have particle sizes ranging from 0.1 to 500 microns, and these formulations can be administered by nasal inhalation or oral inhalation to reach the alveolar sacs. Suitable formulations include aqueous or oily solutions of azelastine compounds. Formulations suitable for aerosol or dry powder administration may be prepared according to conventional methods and may be delivered together with other therapeutic agents, such as compounds used in the treatment or prevention of pneumonia or lung disease.

[0064] Specifically, the solution or dispersion is used for parenteral administration, such as by infusion or injection, and preferably provides an antiviral effective dose of about 1 to 500 mg. Specifically, a single loading dose of approximately 1 to 500 mg may be administered parenterally, followed by a maintenance dose of approximately 10 to 200 mg, or approximately 100 mg, or approximately 200 mg, administered daily, for example, for 1 to 10 days, or until a specific clinical response is reached.

[0065] Specifically, tablets, gels, or lozenges are used for oral administration, and preferably the antiviral effective dose is 1 μg to 12 mg per dose, preferably up to one of 5, 4, 3, 2, or 1 mg, or up to 100 μg per dose.

[0066] Specifically, tablets containing an azelastine compound that can be administered once to three times a day may be used. Formulations suitable for topical oral administration include lozenges containing the active ingredient in a flavor base such as sacrose, gum arabic, or tragacanth; lozenges containing an azelastine compound in an inert base such as gelatin and glycerin, sacrose, or gum arabic; or mouthwashes containing an azelastine compound in a suitable liquid carrier.

[0067] When formulated as a topically applied gel or ointment, the active ingredient may be used with a paraffin or water-miscible ointment base. Alternatively, the active ingredient may be formulated as a cream with an oil-in-water cream base.

[0068] Formulations suitable for topical administration to the eye may include eye drops, gels, or creams in which the azelastine compound is dissolved or suspended in a suitable carrier, particularly an aqueous solution or oil / water emulsion.

[0069] For example, the formulation for topical administration in the oral cavity, or a formulation applied topically such as a gel or ointment, contains the azelastine compound in concentrations of approximately 0.001%, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.05%, 0.1%, 0. It may be included in concentrations of 0.001 to 20% (w / w), such as 0.15%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 9%, or 20% (w / w).

[0070] In certain embodiments, treatment with an azelastine compound can be combined with further treatment involving the administration of an azelastine compound (which may be the same or a different compound) in a different dosage form. For example, treatment with an intranasal or throat spray containing azelastine hydrochloride can be combined with a tablet containing an azelastine compound (which may be azelastine hydrochloride or a different compound).

[0071] In certain embodiments, the azelastine compound is administered as the sole antiviral agent, or the treatment is combined with further treatments such as additional antiviral, anti-inflammatory, and / or antibiotic treatments, including, for example, the administration of one or more antiviral agents or drugs, and / or one or more anti-inflammatory and / or antibiotic or drugs, by one or more different formulations and / or one or more different routes of administration.

[0072] Specifically, azelastine compounds can be combined with one or more additional active therapeutic agents in an integrated dosage form for simultaneous, concurrent, or sequential administration to the target. Combination therapy may be administered, for example, as a simultaneous, parallel, or sequential regimen. When administered sequentially, the combination may be administered in two or more doses.

[0073] In certain embodiments, the subjects being treated are those infected with or at risk of being infected with the virus, preferably humans or non-human mammals such as dogs, cats, horses, camels, cattle, or pigs.

[0074] Specifically, the subjects are those who are exposed to the virus, have been exposed to it, or are at risk of becoming infected with the virus through other means. Specifically, the target group has a weakened immune system and is at higher risk of developing a viral disease or experiencing increased severity of a viral disease.

[0075] Specifically, the subjects are those who have been determined to be infected with the virus or have been diagnosed with it. In certain embodiments, subjects who are diseased or patients suffering from a disease caused by a coronavirus, such as a disease caused by the SARS virus, or COVID-19-related pneumonia, are treated by contact with a pathogen such as COVID-19.

[0076] In a further specific embodiment, a disease caused by the influenza virus, for example, a diseased subject or patient suffering from influenza, is treated. The present invention further provides azelastine compounds described herein for use as antiviral substances in medical products to treat biological surfaces to prevent viral infection and / or viral spread.

[0077] According to a particular embodiment, the medical product is formulated for topical use, preferably for application to the upper and lower respiratory tracts, nasal cavity, lungs, mouth, eyeball, or skin. Topical application typically refers to the surface of the skin, wounds, and / or mucosal cells or tissues (e.g., alveoli, buccal, tongue, masticatory, or nasal mucosa, etc.).

[0078] According to a particular embodiment, the medical product is used in formulations suitable for topical administration, particularly for mucosal administration, such as sprays, solutions, dispersions, dry powders, or aerosolized liquids or powders.

[0079] According to a particular embodiment, the azelastine compound is applied to the biological surface in an antiviral effective amount, preferably in an amount of 1 cm². 2 1 ng to 1000 ng per unit, preferably 10 to 800 ng / cm³ 2 , or 800, 700, 600, 500, 400, 300, 200, 100, 90, 80, 70, 60, 50 ng / cm 2 You may choose any one of the following.

[0080] Any medical device used appropriately for topical treatment as described herein can be used to treat biological surfaces. According to a particular embodiment, the biological surface includes or comprises a mucous membrane surface that is infected with or at risk of becoming infected with the virus.

[0081] The present invention further provides the use of azelastine compounds described herein as antiviral agents, particularly suitable for treating living surfaces or non-living surfaces such as sanitary devices, face masks, etc. Specifically, living or non-living surfaces can be treated with antiseptic agents.

[0082] Specifically, antiviral disinfectants are antiviral preparations such as medical supplies. In certain embodiments, the present invention provides azelastine compounds described herein for use in preventing or treating coronavirus infections, particularly SARS virus infections, in humans or non-human mammals.

[0083] In certain embodiments, the present invention provides azelastine compounds described herein for use in preventing or treating coronavirus infections, particularly SARS virus infections, in humans or non-human mammals.

[0084] In certain embodiments, the present invention provides azelastine compounds described herein for use in preventing or treating adenoviral virus infections, particularly HAdVB, HAdVC, or HAdVD virus infections in humans.

[0085] In a particular embodiment, the present invention provides azelastine compounds described herein for use in preventing or treating paramyxoviridae virus infections, particularly human RSV virus infections, in humans.

[0086] In certain embodiments, the present invention provides azelastine compounds described herein for use in preventing or treating orthomyxoviridae virus infections, particularly influenza virus infections, in humans or non-human mammals.

[0087] In certain embodiments, a kit is provided comprising one or more individual dose units of azelastine compounds further described herein, and instructions for their use in treating coronavirus infection or disease caused by coronaviruses in human or non-human mammals.

[0088] In another specific embodiment, a kit is provided comprising one or more individual dosing units of the azelastine compounds further described herein, and instructions for their use in treating adenoviral virus infections, particularly HAdVB, HAdVC, or HAdVD virus infections, in human or non-human mammals.

[0089] In another specific embodiment, a kit is provided comprising one or more individual dosing units of azelastine compounds further described herein, and instructions for their use in treating paramyxoviridae virus infections in humans, particularly human RSV virus infection.

[0090] In another specific embodiment, a kit is provided comprising one or more individual dosing units of azelastine compounds further described herein, and instructions for their use in treating orthomyxoviridae virus infections, particularly influenza virus infections, in human or non-human mammals.

[0091] In certain embodiments, the present invention provides an antiviral pharmaceutical formulation comprising an azelastine compound and a pharmaceutically acceptable carrier, as further described herein. Specifically, the pharmaceutical formulations are provided for medical use, particularly for the preventive or therapeutic treatment of disease conditions caused by coronaviruses such as COVID-19.

[0092] Specifically, the pharmaceutical formulations are provided for medical use, in particular for the prophylactic or therapeutic treatment of disease conditions caused by coronaviruses and / or adenoviruses and / or paramyxoviridae and / or orthomyxoviridae viruses.

[0093] In a particular embodiment, the present invention further provides a method for treating a subject who is infected with or at risk of being infected with one or more different viruses, such as one or more coronaviruses and / or adenoviruses and / or paramyxoviruses and / or orthomyxoviruses, comprising the step of administering an antiviral effective amount of an azelastine compound and each of the respective medical products or pharmaceutical formulations further described herein.

[0094] In a further specific embodiment, the present invention provides antiviral formulations (such as medical products, pharmaceutical formulations or disinfectants) of azelastine compounds described herein, and a method for producing such antiviral formulations, comprising the step of formulating an antiviral effective amount of azelastine compound together with a pharmaceutically acceptable carrier to produce an antiviral formulation, in particular a medical product or pharmaceutical formulation.

[0095] Topical administration of any of the antiviral agents described herein (such as medical products, pharmaceutical formulations, or disinfectants) to a biological surface is preferably such that, for example, 10 minutes to 24 hours and / or a specific contact time of up to 24, 18, 12, 6, 5, 4, 3, 2, or 1 hour, the contact reduces the virus on the surface by at least 0.5 times (half), or by 1-log, 2-log, 3-log, 4-log, or 5-log. [Brief explanation of the drawing]

[0096] [Figure 1] Figure 1 illustrates the Shannon entropy approach for identifying drug homologs with matching pathway profiles (A) and the SARS-CoV-2 related pathway (B). Pathway profiles were calculated for hydroxychloroquine and SARS inhibitors using clearly defined mechanisms and modes of action: SSAA09E2, a small molecule ACE2 inhibitor, and SSAA09E3, a general inhibitor of viral-host membrane fusion. [Figure 2] Figure 2 shows the prevention of the cytopathic effects of SARS-CoV-2 on Vero E6 cells based on phase-contrast microscopy images. A: Uninfected (control) culture; B: Cells infected with SARS-CoV-2; C-F: Cells infected with SARS-CoV-2 in the presence of control buffer (containing 0.5% DMSO); D-G: Cells infected with SARS-CoV-2 in the presence of gradually increasing concentrations of azelastin-HCl: 3.125, 6.25, 12.5, and 25 μM. Cells were infected with MOI 0.1 SARS-CoV-2 virus for 30 minutes, then the culture medium was removed and replaced with fresh culture medium free of virus (but containing azelastin), and microscopic images were taken 48 hours after infection. [Figure 3] Figure 3 shows the reduced cytopathic effect of SARS-CoV-2 on reconstituted human nasal tissue (MucilAir). Human nasal tissue was infected with SARS-CoV-2 and subsequently treated with a 5-fold diluted 0.1% azelastine-HCl nasal spray for 20 minutes every 24 hours for 3 days. The figure shows low-resolution microscopic images of human nasal tissue 48 and 72 hours post-infection. Mucin production (as an indicator of intact tissue function) is shown as the presence of dark areas on the tissue. [Figure 4]Figure 4 (in vitro inhibition of SARS-CoV-2 B.1.351 and B.1.1.7 infection by azelastin) shows the effect of azelastin on inhibiting infection of Vero-TMPRSS2 / ACE2 cells with SARS-CoV-2 B.1.351 (A) or B.1.1.7 (B). Cells were infected with MOI 0.01 SARS-CoV-2 virus for 30 minutes, and azelastin was added to infected cells immediately before infection (preventive setting) or 30 minutes after infection (therapeutic setting). Viral copy number 48 hours after infection was determined by quantitative PCR. Inhibition of infection is expressed as viral copy number compared to wells with only virus (no azelastin treatment). The graph shows the mean and standard deviation calculated from the individual values ​​of 9 (A) or 6 (B). The azelastine concentration (EC50), which inhibits 50% of infections, was calculated using nonlinear regression (log(agonist) vs. normalized response - variable slope) with GraphPad Prism 8.4.3. [Figure 5] Figure 5 shows Sequence ID No. 4: (NCBI accession number QII57161.1, SARS-CoV-2, S-protein). [Figure 6] Figure 6 shows the antiviral effects of azelastine at two different concentrations, as determined in Calu-3 human cells infected with SARS-CoV-2 at an MOI of 0.01, as well as the cytotoxicity of the same concentration of the compound measured in the same cells without viral infection. The graph shows the average of three results from one representative experiment. [Figure 7] Figure 7 shows the viability of uninfected Hep-2 cells treated for 48 hours with azelastin-HCl and the corresponding diluent, DMSO, within the indicated final concentration range. Mean and SEM values ​​were calculated from three independent experiments. The threshold for statistical significance is P<0.05. ****P<0.0001 [Figure 8A]Figure 8 shows the antiviral effect of azelastine-HCl on RSV infection of Hep-2 cells within the indicated final concentration range. Azelastine-HCl was applied prior to (A), simultaneously with (B), or after viral infection, and replication was assessed by visualizing RSV-infected cells as described in the text. DMSO was the solvent for the azelastine-HCl stock and was diluted accordingly to serve as a control. The experiment was repeated twice using pairs. Bars represent the mean using the mean standard error as error bars. [Figure 8B] Same as above. [Figure 8C] Same as above. [Figure 9] Figure 9 shows the effects of two different concentrations of azelastine-HCl and oseltamivir on viral copy numbers obtained from tip lavage of reconstituted nasal tissue 24 hours after infection with influenza H1N1 virus. Statistical significance is indicated by asterisks. The graph shows three mean results from one representative experiment. *P<0.05; **P<0.01 [Figure 10A] Figure 10 shows the effects of two different concentrations of azelastine and oseltamivir on the levels of IL-8 (Panel A) and RANTES (Panel B) secreted by reconstituted nasal tissue infected with influenza H1N1 virus. Statistical significance is indicated by asterisks. The graph shows the mean values ​​of three different results from one representative experiment. **P<0.01;***P<0.001;****P<0.0001 [Figure 10B] Same as above. [Modes for carrying out the invention]

[0097] As used herein, the terms “comprise,” “contain,” “have,” and “include” may be used as synonyms, are understood as open definitions, and allow for further members, parts, or elements. “Consisting” is considered the most exclusive definition and has no further elements that characterize the “consisting” definition. Thus, “compriseing” is broader and includes the “consisting” definition.

[0098] As used herein, the term "about" refers to a value that is the same as a given value or a value that is + / - 10% or + / - 5% different from that value. As used herein, the term "azelastine compound" refers to azelastine or a salt thereof.

[0099] The molecular formula for azelastine is C 22 H 24 It is ClN3O and has the following chemical structure.

[0100] [ka]

[0101] IUPAC name: 4-[(4-chlorophenyl)methyl]-2-(1-methylazepan-4-yl)phthalazine-1-one CAS number: 58581-89-8 The azelastine salt is preferably a pharmaceutically acceptable salt, such as a hydrochloride salt.

[0102] Azelastine hydrochloride is the hydrochloride form of azelastine and is used as an antihistamine compound formulated as a metered spray solution for intranasal administration. Commercial products containing azelastine hydrochloride are provided as nasal sprays containing 0.1% or 0.15% azelastine hydrochloride USP in aqueous solution at pH 6.8 ± 0.3.

[0103] Azelastine hydrochloride exists as a white or nearly white crystalline powder. It is slightly soluble in water and soluble in ethanol and dichloromethane. Molecular formula:C 22 H 25 Cl2N3O IUPAC name: 4-[(4-chlorophenyl)methyl]-2-(1-methylazepan-4-yl)phthalazine-1-one; hydrochloride CAS numbers: 58581-89-8; 37932-96-0; 79307-93-0 The choice of azelastine salt is primarily determined by how acidic or basic the chemical is (pH), the safety of its ionization form, the intended use of the drug, how the drug is administered (e.g., orally, by injection, or on the skin), and the type of dosage form (such as tablets, capsules, or liquid).

[0104] The term "pharmacologically acceptable," also known as "pharmacologically acceptable," means suitable for use in animals, particularly humans. The term "pharmacologically acceptable salts" includes pharmacologically acceptable acid addition salts and pharmacologically acceptable base addition salts.

[0105] As used herein, the term “pharmacologically acceptable acid addition salt” means any non-toxic organic or inorganic salt of any of the main components of this disclosure, or any intermediate thereof. Main components of this disclosure that may form acid addition salts include, for example, compounds containing a basic nitrogen atom. Examples of inorganic acids that form suitable salts include hydrochloric acid, hydrobromic acid, sulfuric acid and phosphoric acid, as well as metal salts such as sodium monohydrogen orthophosphate and potassium bisulfate. Examples of organic acids that form suitable salts include mono-, di- and tricarboxylic acids such as glycolic acid, lactic acid, pyruvic acid, malonic acid, succinic acid, glutaric acid, fumaric acid, malic acid, tartaric acid, citric acid, ascorbic acid, maleic acid, benzoic acid, phenylacetic acid, cinnamic acid and salicylic acid, as well as sulfonic acids such as p-toluenesulfonic acid and methanesulfonic acid derivatives. One, two, or three salts may be formed, and such salts may exist in hydrated, solvated, or substantially anhydrous forms. In general, the acid addition salts of the compounds disclosed herein are generally more soluble in water and various hydrophilic organic solvents and exhibit higher melting points than their free base forms. The selection of a suitable salt will be known to those skilled in the art. Other non-pharmacologically acceptable acid addition salts, such as oxalates, may be used in the isolation of the compounds disclosed herein for laboratory use or for subsequent conversion to pharmacologically acceptable acid addition salts.

[0106] As used herein, the term “pharmacologically acceptable basic salt” means any non-toxic organic or inorganic base addition salt, or intermediate thereof, of any acid compound of the present invention, which is suitable or compatible for treatment of animals, in particular humans. Acid compounds of the present invention that may form base addition salts include, for example, compounds containing carboxylic acids, sulfonic acids, sulfinic acids, sulfonamides, N-unsubstituted tetrazoles, phosphate esters, or sulfate esters. Examples of inorganic bases that form suitable salts include lithium hydroxide, sodium hydroxide, potassium hydroxide, calcium hydroxide, magnesium hydroxide, or barium hydroxide. Examples of organic bases that form suitable salts include aliphatic, alicyclic, or aromatic organic amines such as methylamine, trimethylamine, and picoline, or ammonia. The selection of suitable salts will be known to those skilled in the art. Other non-pharmacologically acceptable base addition salts may be used in the isolation of the compounds of this disclosure for laboratory use or for subsequent conversion to pharmacologically acceptable base addition salts. The formation of the desired compound salt is achieved using standard techniques. For example, a neutral compound is treated with a base in a suitable solvent, and the resulting salt is isolated by filtration, extraction, or any other suitable method.

[0107] As used herein, the term “antiviral” means any substance, drug, or formulation that brings about a reduction in viral load or infectivity by altering the biology of a virus, attenuating or inhibiting viral attachment, entry, replication, shedding, latency, or any combination thereof. The terms “attenuating,” “inhibiting,” “reducing,” or “prevention,” or any variation thereof, as used in the claims and / or specification, include any measurable reduction or complete inhibition to achieve the desired result, e.g., a reduction in the risk of viral infection (pre-exposure) or a reduction in post-exposure viral survival, load, or replication.

[0108] The exemplary antiviral formulations described herein are medical products, drugs, and disinfectants for in vivo, ex vivo, or in vitro use. As used herein, the term "biological surface" refers to a surface containing living cells, such as mammalian (human or non-human animal) cells, including, for example, the surface of a biological tissue such as epithelial or dermal tissue (e.g., skin), mucous membrane tissue, or membrane tissue.

[0109] As used herein, the term “effective dose” in relation to antiviral effects refers to the amount (in particular, a specified amount) that has a proven antiviral effect. This amount is typically sufficient to produce the desired outcome, including antiviral or clinical results, when applied to a surface or administered to a target; therefore, the effective dose or its synonyms depend on the setting in which the amount is applied.

[0110] The effective dose of a drug or medicine is intended to mean the amount of compound sufficient to treat, prevent or inhibit a disease, disease condition, or disorder. In particular, the effective dose is the amount of compound sufficient to result in the cure, prevention, or improvement of a condition relating to the disease or disorder described herein.

[0111] In the context of disease, the effective doses (in particular, prophylactic or therapeutic effective doses) of the azelastine compounds described herein are used specifically to treat, regulate, reduce, reverse, or influence a disease or condition that benefits from its antiviral effect. The amount of compound corresponding to such effective dose will vary depending on various factors such as a given drug or compound, formulation, route of administration, type of disease or disorder, identity of the subject or host being treated, assessment of the medical condition, and other relevant factors, but can nevertheless be routinely determined by a person skilled in the art.

[0112] The treatment or prophylactic regimens described herein using an effective amount of the azelastine compound may consist of a single application or administration, or they may instead comprise a series of applications and administrations. For example, the azelastine compound may be used at least once a month, or at least once a week, or at least once a day. However, in certain acute cases, for example, when exposure to the virus is suspected or confirmed, or after a viral infection has been confirmed, the azelastine compound may be used more frequently, for example, 1 to 10 times a day.

[0113] Specifically, the following are provided: treatment using the formulations described herein, as well as combination therapies including standard therapies for diseases caused by coronaviruses and / or other target viruses.

[0114] The dosage may be administered in combination with other active agents, such as antivirals, anti-inflammatory agents, or antibiotics, to prevent pathogen-associated reactions, for example, when there is a risk of the target virus spreading.

[0115] The treatment can be combined with antiviral, anti-inflammatory, or antibiotic treatments, and preferably the drug is administered before, during (e.g., by co-administration or in parallel with) or after the antiviral, anti-inflammatory, or antibiotic treatment.

[0116] Specifically, the azelastine compounds described herein can be combined with additional antiviral agents, which may be the same or different azelastine compounds. Specific embodiments refer to further antiviral agents selected from ACE2 inhibitors, viral protein M2 ion channel inhibitors, neuraminidase inhibitors, RNA replication and translation inhibitors, and polymerase inhibitors. The antiviral agent may be amantadine or rimantadine. Specifically, the antiviral agent may be oseltamivir, zanamivir, peramivir, ribavirin, lopinavir, or ritonavir. Specific examples of further antiviral agents include antiviral agents appropriately used for biological surface treatment, such as carrageenan, or antiviral agents currently under investigation for the treatment of SARS-CoV-2 infection, such as hydroxychloroquine or remdesivir.

[0117] Specifically, azelastine compounds are combined with anti-inflammatory agents such as standard steroidal anti-inflammatory drugs, glucocorticoids, and non-steroidal anti-inflammatory drugs (NSAIDs). Suitable NSAIDs include, but are not limited to, ibuprofen, naproxen, fenoprofen, ketoprofen, flurbiprofen, oxaprozin, indomethacin, sulindac, etodolac, ketorolac, diclofenac, nabumetone, piroxicam, meloxicam, tenoxicam, droxicam, lornoxicam, isoxicam, mefenamic acid, meclofenamic acid, flufenamic acid, tolfenamic acid, and celecoxib. Suitable steroidal anti-inflammatory agents include, but are not limited to, corticosteroids such as synthetic glucocorticoids. Specific examples include immunosuppressants such as fluticasone, COX-2 inhibitors, ibuprofen, hydroxychloroquine, heparin, LMW heparin, hirudin, or azathioprine, cyclosporine A, or cyclophosphamide.

[0118] Specifically, azelastine compounds include β-lactam antibiotics, aminoglycoside antibiotics, ansamycin, carbasephalm, carbapenem, cephalosporins, glycopeptides, lincosamides, lipopeptides, macrolides, monobactams, nitrofurans, oxazolidinones and other antibiotics, polypeptides, sulfonamides, clofazimine, dapsone, capreomycin, cycloserine, ethambutol, ethionamide, isoniazid, pyrazinamide, rifampicin, rifabutin, rifapentin, streptomycin, arsphenamine, chloramphenicol, fosfomycin, mupirocin, platensimycin, quinupristin / dalfopristin, thianphenicol, tigecycline, tinidazole, It can be combined with trimethoprim, teixobactin, malacidin, halicin, clindamycin, vancomycin, metronidazole, fusidic acid, thiopeptides, fidaxomycin, quinolones, tetracyclines, omadacyclines, rifamycin, kibdelomycin, oxazolidinone, ketolides, thiazolides, amixicile, teicoplanin, lamopranin, oritabancin, lanchiobiotics, capramycin, slotomycin, turicin, endolysin, avidocin CD, cadazol, lamisobactin, defensin, lysinirazole, medium-chain fatty acids, phages, berberine, and lactoferrin.

[0119] Specifically, treatments using the azelastine compounds described herein can be combined with treatments administering at least one other therapeutic agent selected from the group consisting of corticosteroids, anti-inflammatory signaling modulators, 2-adrenergic receptor agonist bronchodilators, anticholinergics, mucolytics, hypertonic saline, and other drugs for treating coronavirus infections and / or infections with any of the other target viruses described herein; or mixtures thereof. Certain pharmaceutical compositions may particularly include one or more anti-inflammatory agents and / or analgesics, PPAR-γ agonists, and immune response modulators.

[0120] The duration of treatment depends on various factors, including the severity of the disease, whether it is acute or chronic, the patient's age, and the concentration of the azelastine compound. It is also recognized that the effective dose used for treatment or prevention may increase or decrease during a particular treatment or prophylactic regimen. Changes in dosage may occur and be revealed by standard diagnostic assays known in the art.

[0121] In certain embodiments, the medicinal products or pharmaceutical compositions described herein contain an effective amount of the azelastine compound as defined herein. The formulations described herein may be provided for single or multiple dose use.

[0122] Single-dose or multi-dose containers, such as sealed ampoules and vials, or multi-use sprays may be used, and the formulation may contain a liquid or dry phase and be stored in a freeze-dried state, for example, by simply adding a sterile liquid carrier, such as sterile water for injection, immediately before use. Preferred unit-dose formulations are those containing a daily dose or a unit daily subdose of the azelastine compound, or multiple doses.

[0123] As used herein, the term “single dose” is understood as follows: A single dose, or as-needed dose, is a quantity intended for use in a single subject, such as a patient, human, or animal, for a single case / procedure / administration. Packages containing a single dose are typically, and therefore, labeled by the manufacturer. A single dose is understood as a daily dose, particularly for an individual such as a child or an adult, in order to provide an effective amount.

[0124] The medical products or pharmaceutical compositions described herein are provided, in particular, as human or veterinary medical products or pharmaceutical compositions. A medical product is understood to be a substance used to treat a disease, alleviate a complaint, or, primarily, prevent such disease or complaint. This definition applies whether the medical product is administered to humans or animals. The substance may act both internally or externally within the body.

[0125] The medicinal products or pharmaceutical compositions described herein are preferably dosage forms that contain one or more pharmaceutically acceptable adjuvants, enabling the active pharmaceutical compound to be administered with high bioavailability. Suitable adjuvants may be, for example, based on cyclodextrins. Suitable formulations may also incorporate synthetic polymer nanoparticles formed from polymers selected from the group consisting of acrylates, methacrylates, cyanoacrylates, acrylamides, polylactates, polyglycolates, polyanhydrides, polyorthoesters, gelatin, albumin, polystyrene, polyvinyl, polyacrolein, polyglutaraldehyde and their derivatives, copolymers, and mixtures.

[0126] Certain medicinal products or pharmaceutical compositions described herein include azelastine compounds and pharmaceutically acceptable carriers or excipients. “Pharmaceutically acceptable carriers” refers to components in a formulation intended for medicinal or medical use other than the active ingredient, which are non-toxic to the subject. pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives, particularly physiological saline, phosphate-buffered saline, glucose, glycerol, ethanol, and similar substances.

[0127] The azelastine compounds used herein can be formulated together with conventional carriers and excipients, which are selected according to common practice. Pharmacopoecitable carriers generally include any suitable solvents, dispersions, coatings, antiviral agents, antimicrobial agents, and antifungal agents, isotonic and absorption retardants, and similar substances that are physiologically compatible with the antiviral small molecule compounds or related compositions or combination formulations described herein.

[0128] In certain embodiments, azelastine compounds can be combined with one or more carriers suitable for a desired route of administration. Azelastine compounds may be mixed with, for example, lactose, sacrose, starch, cellulose esters of alkanates, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric acid and sulfate, acacia, gelatin, sodium alginate, polyvinylpyrrolidone, or polyvinyl alcohol, and optionally further tableted or encapsulated for conventional administration. Alternatively, azelastine compounds may be dispersed or dissolved in physiological saline, water, polyethylene glycol, propylene glycol, carboxymethylcellulose colloidal solution, ethanol, corn oil, peanut oil, cottonseed oil, sesame oil, tragacanth gum, and / or various buffers. Other carriers, adjuvants, and modes of administration are well known in the pharmaceutical field. The carriers may include sustained-release or time-delayed materials such as glyceryl monostearate or glyceryl distearate alone or with wax, or other materials well known in the art.

[0129] The compounds described herein may be provided as sustained-release formulations ("sustained-release formulations") in which the release of the azelastine compound is controlled and regulated to enable fewer doses or to improve the pharmacokinetic or toxicity profile of a given active ingredient.

[0130] The pharmaceutical composition may be coated, typically with a pH or time-dependent coating, by conventional methods, so that the subject drug is released near the desired topical application to the gastrointestinal tract or at various times to extend the desired effect. Such dosage forms typically include, but are not limited to, cellulose phthalate acetate, polyvinyl acetate phthalate, hydroxypropyl methylcellulose phthalate, ethylcellulose, wax, and shellac.

[0131] Additional pharmaceutically acceptable carriers are known in the art and are described, for example, in Remington: The Science and Practice of Pharmacy, 22nd revised edition (Allen Jr., LV, ed., Pharmaceutical Press, 2012). Liquid formulations may be solutions, emulsions, or suspensions and may contain excipients such as suspending agents, solubilizers, surfactants, preservatives, and chelating agents.

[0132] A preferred formulation is a ready-to-use, storage-stable form with a shelf life of at least one or two years. As used herein, the term “formulation” refers to a formulation that is ready for use in a particular manner. Specifically, the compositions described herein include azelastine compounds and pharmaceutically acceptable diluents, carriers, or excipients.

[0133] In certain embodiments, formulations are provided that include a pharmaceutically acceptable medium for nasal, intrapulmonary, oral, topical, mucosal, or parenteral administration. Administration may also be intradermal or transdermal. Furthermore, the disclosure includes compounds that are lyophilized and can be reconstituted to form pharmaceutically acceptable formulations for administration.

[0134] Certain medical products or pharmaceutical compositions described herein are formulated for intranasal administration or by other local routes onto biological surfaces, such as mucous membranes or skin. Suitable pharmaceutical carriers for facilitating such administration are well known in the art.

[0135] Specifically, a nasal spray may be used that contains 0.001% or 0.15% (w / w) of an azelastine compound in an aqueous solution at pH 6.8 ± 0.3, and optionally further contains one or more preservatives such as citric acid monohydrate, disodium hydrogen phosphate dodecahydrate, disodium edetate, hypromellose, purified water, sodium chloride, and benzalkonium chloride.

[0136] To administer azelastine compounds by any route other than parenteral administration, it may be necessary to coat the activator with a material that prevents its deactivation, or to administer the activator simultaneously with this material. For example, a suitable carrier, such as liposomes, or a diluent may be used. Pharmacochemically acceptable diluents include physiological saline and aqueous buffers.

[0137] Azelastine compounds can be administered orally, for example, with an inert diluent or an assimilated or edible carrier. For example, formulations may be encapsulated in hard or soft-shelled gelatin capsules or compressed into tablets. In oral therapeutic administration, azelastine compounds can be incorporated with excipients and used in the form of ingestible tablets, oral tablets, lozenges, capsules, elixirs, suspensions, syrups, wafers, and the like. The percentage of compounds in compositions and formulations can, of course, vary. The amount of azelastine compound in the therapeutically useful composition is such that an appropriate dosage can be obtained.

[0138] Tablets contain excipients, lubricants, fillers, binders, disintegrants, lubricants, fragrances, and the like. Granules may be manufactured using isomaltose. It is even more preferable to provide formulations that act locally, for example, on mucosal sites (such as the nose, mouth, eyes, esophagus, throat, and lungs), without systemic effects. Aqueous formulations are prepared in a sterile manner and are generally isotonic if intended for delivery by means other than oral administration.

[0139] In relation to the administration or application of a formulation for treating a subject, or other mucosal use, or with respect to each formulation, the term "mucosal" means administration via a mucosal route, including systemic or topical administration, where the active ingredient is taken up by contact with the mucosal surface. This includes nasal, pulmonary, oral, or oral administration and formulations, such as liquids, syrups, lozenges, eye drops, tablets, sprays, powders, instant powders, granules, capsules, creams, gels, drops, suspensions, or emulsions.

[0140] Oral formulations may include liquid solutions, emulsions, suspensions, and similar substances. Pharmaceutically acceptable solvents suitable for the preparation of such compositions are well known in the art. Typical components of carriers for syrups, elixirs, emulsions, and suspensions include ethanol, glycerol, propylene glycol, polyethylene glycol, liquid sacrose, sorbitol, and water. In suspensions, typical suspending agents include methylcellulose, sodium carboxymethylcellulose, tragacanth, and sodium alginate; typical wetting agents include lecithin and polysorbate 80; and typical preservatives include methylparaben and sodium benzoate. Oral liquid compositions may also contain one or more components such as sweeteners, flavorings, and colorants disclosed above.

[0141] Other compositions useful for achieving systemic delivery of azelastine compounds or their respective formulations include sublingual, buccal, and nasal formulations. These compositions typically include soluble fillers such as sacrose, sorbitol, and mannitol; binders such as acacia, crystalline cellulose, carboxymethylcellulose, and hydroxypropylmethylcellulose; or one or more of lubricants, sweeteners, colorants, antioxidants, and flavorings.

[0142] Azelastine compounds or their respective formulations can also be administered topically to a subject, for example, by placing or spreading a composition containing the same directly onto the epidermis or epithelial tissue of the subject, or transdermally via a “patch.” Such compositions include, for example, lotions, creams, solutions, gels, and solids. These topical compositions may contain an effective amount of the azelastine compound, typically at least about 0.001 wt%, or about 0.1 wt% to 5 wt%, or 1 wt% to about 5 wt%. Suitable carriers for topical administration typically remain in place on the skin as a continuous covering and resist removal by sweating or immersion in water. Generally, the carrier is substantially organic, in which the therapeutic agent may be dispersed or dissolved. The carrier may contain pharmaceutically acceptable emollients, emulsifiers, thickeners, solvents, and the like.

[0143] Pharmaceutical compositions suitable for injection include sterile aqueous solutions (especially when the compound or pharmaceutically acceptable salt is water-soluble) or dispersions, and sterile powders for the immediate preparation of sterile injection solutions or dispersions. In particular, the compositions are particularly sterile and fluid to the extent that easy needle passage is present, and the compositions are stable under manufacturing and storage conditions and preserved against contamination by microorganisms such as bacteria and fungi.

[0144] Suitable pharmaceutically acceptable solvents include, but are not limited to, any non-immunogenic pharmaceutical adjuvant suitable for oral, parenteral, nasal, mucosal, transdermal, intravascular (IV), intra-arterial (IA), intramuscular (IM), and subcutaneous (SC) administration routes, such as phosphate-buffered saline (PBS).

[0145] As used herein, the term “subject” means warm-blooded mammals, in particular humans or non-human animals, including, for example, dogs, cats, rabbits, horses, cattle, and pigs. In particular, the treatments and medical uses described herein apply to subjects who require the prevention or treatment of disease conditions associated with infection with coronaviruses and / or any of the other target viruses described herein. Specifically, the treatments may involve preventing the pathogenesis of disease conditions in which coronaviruses and / or adenoviruses and / or paramyxoviruses or orthomyxoviralized viruses are the etiologists of the condition. Subjects may be patients at risk of or suffering from such disease conditions. Specifically, subjects may have a weakened immune system or pre-existing respiratory or cardiac conditions that are more likely than other conditions to increase the risk of disease or the severity of disease.

[0146] The term "at risk" for a particular disease state refers to an individual who is potentially at risk of developing the disease state due to a specific predisposition, exposure to a virus, or to an infected individual, or, in particular, an individual who is already suffering from the disease state at various stages, especially in relation to other conditions or complications that follow as a result of other causative disease states or viral infections. Risk assessment is especially important for individuals whose disease has not yet been diagnosed. Therefore, this risk assessment includes early diagnosis to enable preventive treatment. Specifically, azelastine compounds are used in high-risk individuals, for example, those with a high probability of developing the disease.

[0147] The term “patient” includes human and other mammalian subjects receiving preventive or therapeutic treatment. As used herein, the term “patient” always refers to healthy subjects. Therefore, the term “treatment” is intended to include both preventive and therapeutic treatments.

[0148] Specifically, the term "prevention" refers to preventive measures intended to include the prevention of the onset of disease development or to preventive measures aimed at reducing the risk of disease development. As used herein with respect to the treatment of a subject, “treatment” means the medical management of a subject for the purpose of curing, improving, stabilizing, reducing the incidence of, or preventing a disease, pathological condition, or disorder, which is understood individually or collectively as a “disease condition.” This term includes dynamic treatment directed in particular to the improvement of a disease condition, preventive treatment directed in particular to the prevention of a disease condition, and causal treatment directed to the removal of the causes of the associated disease condition. Furthermore, this term includes symptomatic treatment aimed at alleviating symptoms rather than curing a disease condition, directed to curing the disease condition and minimizing or partially or completely inhibiting the progression of the further associated disease condition, as well as supportive treatment used to complement other specific therapies directed to the improvement of the associated disease condition.

[0149] The foregoing description will be understood more fully by referring to the following embodiments. However, these embodiments are representative of ways of carrying out one or more embodiments of the present invention and should not be read as limiting the scope of the present invention. [Examples]

[0150] Example 1 Antiviral drug identification The development of mathematical representations of small molecules (drugs) is a research area of ​​immeasurable value for modern drug research, and therefore, numerous molecular descriptors have been developed to effectively utilize the 2D and / or 3D features of chemical structures. These descriptors have been extremely valuable in evaluating quantitative structure-activity relationships. Specifically, atomic center feature pairs have proven to be highly relevant to drug discovery programs because they provide a cost-effective approach to high-throughput structure-activity relationship analysis, compound selection, hypothetical chemical screening, and pharmacological profiling. One commonly used chemical descriptor is called SHED, which stands for Shannon entropy descriptor (1). In this approach, the phase distribution of atomic center feature pairs is quantified based on information theory (Shannon entropy). A particular benefit is that chemically distinct but topologically related chemical skeletons can be identified using the SHED approach.

[0151] To identify novel antiviral drugs, a drug identification strategy based on a biochemical pathway-based intervention strategy was used. Clinically approved drugs that fit a predefined mechanism profile (mode of action) were identified as candidate antiviral drugs using a Shannon entropy-based description of the intrinsic chemical characteristics of the small molecules (drugs). The rationale behind this approach is that ligands with similar Shannon entropy vectors bind to similar protein targets.

[0152] Here, we used the DRUGBANK database, a repository of approved drugs and their experimentally validated protein targets. Drug similarity was assessed by calculating the Euclidean distance (0.25 was considered the cutoff value).

[0153] As a starting point for the analysis, available information regarding the mechanisms of SARS-CoV-2 infection was extracted from recent bioinformatics analyses (3) along with the analysis of certain query antiviral compounds. The pathway profiles of query antiviral compounds with known SARS-CoV-2 activity (hydroxychloroquine (RS)-2-[{4-[(7-chlor-4-sinnolinyl)amino]phenyl}(ethyl)amino]ethanol; SSAA09E2{N-[[4-(4-methylpiperazine-1-yl)phenyl]methyl]-1,2-oxazole-5-carboxamide}; and SSAA09E3[N-(9,10-dioxo-9,10-dihydroanthracene-2-yl)benzamide]) were predicted using the Shannon entropy approach and, interestingly, showed significant overlapping portions.

[0154] Secondly, pathway profiles for SARS inhibitors were predicted using well-defined mechanisms and modes of action: SSAA09E2, a small molecule ACE2 inhibitor, and SSAA09E3, a general inhibitor of viral-host membrane fusion (4). Both inhibitors were found to share a significant number of pathways with hydroxychloroquine. In summary, the analysis revealed that similar pathways are involved in SARS-CoV-2 infection, targeted by hydroxychloroquine, and countered by inhibitors through well-defined mechanisms. Thus, it was concluded that these pathways are highly relevant to antiviral activity and could provide a basis for novel drug reuse for antiviral activity by searching for clinically approved drugs that were not previously known to have antiviral activity and possess a matching pathway profile.

[0155] Clinically approved drugs were investigated for their fit to individual pathway profiles. The SELLECKCHEM database of approved (and commercially available) drugs was screened using predicted pathway profiles obtained for different query compounds (hydroxychloroquine, SSAA09E2, and SSAA09E3). The rationale for candidate selection was based on the co-occurrence of approved drugs in different (predicted) datasets. Hydroxychloroquine and SSAA09E2 (ACE2 inhibitor) showed significant overlap with each other and with drugs obtained using SARS-CoV-2 pathway profiles. After eliminating drugs based on chemical composition, two approved drugs were selected as candidate antiviral agents for further testing in in vitro SARS-CoV-2 infection models: azelastine and maraviroc.

[0156] References (1) Gregori-Puigjane, E. and Mestres, J. SHED: Shannon Entropy Descriptors from Topological Feature Distributions. J.Chem.Inf.Model. 46, 1615-1622 (2006) (2) Wishart, DS., Knox, C., Guo, AC:, Cheng, D., Shrivastava, S., Tzur, D., Gautam, B. and Hassanali, M. DrugBank: a knowledgebase for drugs, drug action and drug targets. Nucleic Acids Res. D901-D906, (2008) (3) Zhou, Y, Hou, Y., Shen, J, Huang, Y., Martin, W. and Cheng, F. Network-based drug repurposing for novel coronavirus 2019-nCoV / SARS-CoV-2. Cell Discovery 6,14-32, (2020) (4) Adedeji, AO., Severson, W., Jonsson, C., Singh, K., Weiss SR. and Sarafanos, SG. Novel Inhibitors of Severe Acute Respiratory Syndrome Coronavirus Entry That Act by Three Distinct Mechanisms. J Virol. 87, 8017-8028, (2013)

[0157] Example 2 Preventing SARS-CoV-2 viral infection of Vero E6 cells. To detect the effect of azelastin-HCl on SARS-CoV-2 infection, ACE2-expressing Vero E6 (monkey kidney) cells were infected with SARS-CoV-2 in the absence or presence of azelastin-HCl, and the cytopathic effect was evaluated by microscopic examination.

[0158] Experimental procedure: Vero E6 cells (ATCC CRL-1586) were seeded on 96-well plates. After 2 days, the cell culture reached culture density and formed a homogeneous monolayer. The cells were given fresh cell culture medium (DMEM+FBS?). Azelastine-HCl (Seleckchem catalog no. S2552, 10 mM preservative solution dissolved in DMSO) and maraviroc (Seleckchem catalog no. S2003, 10 mM preservative solution dissolved in DMSO), anti-HIV antiviral agents, were added to the cell culture medium at final concentrations of 50, 25, 12.5, 6.25, and 3.125 μM (dilutions were prepared in the culture medium). In viral infection, SARS-CoV-2 virus (hCoV-19 / Hungary / SRC_isolate_2 / 2020, Accession ID: EPI_ISL_483637) was added to the supernatant immediately after (essentially simultaneously with) the change of culture medium at an MOI of 0.1 (infection multiplicity: 1 virus particle per 10 cells). Virus stocks were prepared by proliferation in Vero E6 cells, and infectivity titers were determined. After 30 minutes of incubation with the virus, the culture medium was removed and replaced with fresh culture medium containing azelastine-HCl or maraviroc at the above concentrations (simultaneous administration: simulating prevention). The experiment was conducted, in a sense, using azelastine only when the drug was administered after 30 minutes of incubation with the virus, but not during this period (post-infection administration: post-exposure / treatment setting). 48 hours after infection, cells were evaluated by microscopic observation, and the supernatant was collected and stored at -80°C for quantitative PCR analysis. Viral RNA was extracted from the culture supernatant sample using the Monarch Total RNA Miniprep kit (New England BioLabs, catalog number T2010S) according to the manufacturer's instructions. Briefly, 300 μl of lysis buffer was mixed with 100 μl of culture supernatant, gDNA contamination was removed using a dedicated column (DNA was retained), and the pass-through fraction containing RNA was poured into an RNA-binding column. After washing the column, the RNA was eluted with water, and the sample was stored at -80°C until analysis. After reverse transcription, the DNA was amplified with F, R, and P2 primers derived from the RdRp gene. The primers and probes used were specific to the SARS-CoV-2 RdRp gene: Reverse primer: CARATGTTAAASACACTATTAGCATA (SEQ ID NO: 1) Forward primer: GTGARATGGTCATGTGTGGCGG (SEQ ID NO: 2) Probe: FAM-CAGGTGGAACCTCATCAGGAGATGC-BBQ (Sequence ID 3) A droplet PCR kit was used (BioRad ddPCR®, Bio-Rad Laboratories GmbH, Germany). The results of the RT-PCR reaction were quantified and calculated as virus particles / μl.

[0159] result: The dense, homogeneous layer of cells (uninfected, Figure 2A) is disrupted, and "pores" indicating virus-induced cell death appear (Figure 2B). In the presence of azelastine-HCl at all tested concentrations, SARS-CoV-2 infected cells were significantly protected from death, providing evidence of a direct antiviral effect (Figures 2C-F). Maraviroc was completely ineffective at lower concentrations and only offered minimal protection at higher concentrations (12.5-50 μM), resulting in a high cytopathic score (Table 1). Surprisingly, azelastine proved to be an effective antiviral agent, at least as effective as hydroxychloroquine.

[0160] These data suggest that azelastin compounds were able to immediately halt SARS-CoV-2 infection as soon as they were applied to cells. Since viruses need to enter cells in order to replicate and spread throughout the body, azelastin is expected to prevent COVID-19 on the mucous membrane surface of the respiratory tract, precisely where the virus would normally infect the human body.

[0161] [Table 1]

[0162] Scoring: 0: No cytopathic effect (CPE); cells appeared identical to those of the uninfected control. 1: Only a very small area showed low levels of CPE. 2: CPE was observed in a small area of ​​the cell culture. 3: This is an even stronger CPE, but not as strong as that of an infection control group. 4: CPE is as strong as infection control. Quantitative PCR analysis revealed that azelastine is highly effective in reducing viral particle counts by up to 99% in co-administration (simulating prevention) and up to 97% in post-infection (simulating post-exposure or treatment) settings (Table 2), suggesting that azelastine can be used in both preventive and therapeutic settings. As expected, co-administration was more effective, demonstrating that low viral counts at 25 μm azelastine concentrations can not only prevent but also halt ongoing infection.

[0163] [Table 2]

[0164] Example 3 Effectiveness against viral infection of reconstituted human nasal tissue caused by SARS-CoV-2. To confirm the efficacy of azelastine-HCl against SARS-CoV-2 in human cells, reconstituted human nasal 3D tissue (MucilAir, 3D human airway epithelium reconstituted in vitro) was infected with SARS-CoV-2 and treated with a commercially available nasal spray containing azelastine. The cytopathic effect was evaluated by microscopic observation of the tissue, and the number of viral particles was determined by droplet PCR.

[0165] Experimental procedure: MucilAir human nasal tissue (Epithelix Sarl (Geneva, Switzerland), catalog number: EP02MP) prepared from a healthy donor was infected apical with SARS-CoV-2 (SARS-CoV-2 virus, hCoV-19 / Hungary / SRC_isolate_2 / 2020, Accession ID: EPI_ISL_483637) at a multiple of infection (MOI) of 0.01. After incubation at 37°C in 5% CO2 for 20 minutes, the virus-containing medium was completely removed. Next, a 5-fold dilution of Allergodil nasal spray (0.1% azelastine-HCl, Mylan) (in MucilAir culture medium) was added to the apical side for 20 minutes (in a volume of 200 μl). Following the treatment, the diluted nasal spray was completely removed from the cell surface to provide a liquid-air interface and incubated for 24 hours. Treatment with diluted allergodil for 20 minutes was repeated at 24 and 48 hours post-infection (hpi). After 24, 48, and 72 hpi, the apical side of the cells was washed with MucilAir culture medium for 15 minutes, and the solution was collected for quantification of infectious virus particles. The cells were examined in detail under an inverted microscope at 48 and 72 hpi.

[0166] Total RNA was extracted from apical wash (100 μl) using the Monarch Total RNA Miniprep kit (Promega, catalog number: T2010S) according to the manufacturer's instructions. As described in Example 2, droplet digital PCR technology was applied to quantify the viral copy number (Bio-Rad Laboratories Inc. QX200 Droplet Digital PCR System).

[0167] result: Microscopic analysis of tissues at 48 and 72 hours revealed a decrease in mucin production in infected cells, which appeared as a complete absence of black spots in microscopic images compared to control cells (untreated with virus or drug, showing a large amount of black substance on the cells) (Figure 3). Importantly, mucin production, indicated by the presence of black substance in microscopic images, was clearly demonstrated in the presence of a 5-fold dilution of Allergodil treatment, comparable to the black substance seen in negative controls (untreated with virus or drug). In terms of tissue morphology, no differences were detected between control and azelastin-treated cells (uninfected with virus), and ciliary movement was detected in all tissues.

[0168] Droplet digital PCR analysis confirmed effective SARS-CoV-2 infection and rapid viral replication, reaching several thousand copies per microliter in the apical compartment of tissue inserts by 72 hours post-infection. A 5-fold diluted nasal spray (0.02% azelastine-HCl) used for 20 minutes daily drastically reduced viral particle counts by 48 and 72 hours post-infection (>99.9% inhibition) (Table 3).

[0169] [Table 3]

[0170] Example 4 Demonstration of the in vitro efficacy of azelastine against SARS-CoV-2 mutant strains B.1.1.7 and B.1.351 of concern. To detect the antiviral effect of azelastin-HCl against SARS-CoV-2 mutant viruses, ACE2 and TMPRSS2-expressing Vero (monkey kidney) cell lines were infected with the B.1.1.7 and B.1.351 mutants of concern, either in the absence or in the presence of azelastin-HCl, and the effect on viral replication was evaluated by quantitative PCR.

[0171] Experimental procedure: SARS-CoV-2 infection assay using Vero-TMPRSS2 / ACE2 Vero cells stably overexpressing the human serine protease TMPRSS2 and the ACE2 receptor (Riepler et al., Comparison of Four SARS-CoV-2 Neutralization Assays. Vaccines (Basel), 2020; 9(1):13) were placed in a 96-well plate the day before infection. 4Cells were seeded in one well. Azelastine hydrochloride (Sigma-Aldrich, PHR1636-1G, lot number LRAC4832), dissolved in DMSO to a concentration of 10 mM, was diluted in Dulbecco's modified Eagle medium (Merck, Darmstadt, Germany) containing 2% FBS to final concentrations ranging from 50 μM to 0.4 μM (2-fold serial dilution). Prior to cell infection, the cell culture supernatant was aspirated and replaced with 50 μl of azelastine-HCl dilution in the prophylactic (concurrent administration) setting, and with 50 μl of medium in the post-infection setting. Three measurements were performed for each azelastine concentration. Subsequently, cells were infected with SARS-CoV-2 isolates belonging to type B.1.1.7 or B.1.351 at 37°C for 30 minutes at an MOI of 0.01. In both experimental setups, the supernatant was then aspirated and replaced with 50 μl of fresh medium and 50 μl of the same previously used azelastin concentration, resulting in azelastin concentrations ranging from 25 μM to 0.2 μM. 48 hours after infection, the cytopathic effect was evaluated, and the supernatant was mixed in a 1:1 ratio with DLR buffer (0.5% IGEPAL, 25 mM NaCl buffer in 10 mM Tris-HCl buffer, and 15 μl of RiboLock RNase Inhibitor (ThermoScientific, 40 U / μl, EO0381 per 1 ml of DLR buffer) to isolate viral RNA. SARS-CoV-2 genome copies were quantified by qPCR using E gene-specific primers (5'ACA GGT ACG TTA ATA GTT AAT AGC GT3' (SEQ ID NO: 5) and 5'ATA TTG CAG CAG TAC GCA CAC A3' (SEQ ID NO: 6)), a FAM-labeled probe (FAM-ACA CTA GCC ATC CTT ACT GCG CTT CG-BHQ1 (SEQ ID NO: 7)), and an iTaq Universal Probe One-Step Kit (BioRad, catalog no. 1725141). qPCR results were quantified using a vitro transcription RNA standard (SARS-CoV-2 E gene). Wells containing only virus without azelastine treatment were set up to 100%, and percentage inhibition was calculated for each sample compared to the virus-only wells.

[0172] result: Quantitative PCR analysis revealed that azelastine is highly effective in reducing viral particle counts in both prophylactic (concurrent administration) and post-infection (simulating post-exposure or treatment) administration settings (Tables 4 and 5 and Figure 4). Similar to results with wild-type virus, concurrent administration was slightly more effective, but the low viral counts at 12.5 μm azelastine concentrations (over 90% inhibition) indicate that it can not only prevent but also halt ongoing infection. The effective azelastine concentrations required to inhibit 50% of infection were 4–6.5 μM, depending on the setting and viral variant, which is within the EC50 range (~6 μM) observed for wild-type virus. This indicates that the effectiveness of azelastine against SARS-CoV-2 is independent of high-risk mutations.

[0173] [Table 4]

[0174] [Table 5]

[0175] Conclusion: These data suggest that azelastin compounds were able to immediately halt infection by SARS-CoV-2 variants B.1.351 and B.1.1.7 as soon as they were applied to cells. Since viruses need to enter cells in order to replicate and spread throughout the body, azelastin is expected to prevent COVID-19 at the very spot where the virus infects the mucous membrane surface of the human respiratory tract.

[0176] Example 5 To prevent SARS-CoV-2 viral infection of Calu-3 cells. To confirm the antiviral effect of azelastin-HCl on cells expressing the surface protease TMPRSS2, human lung adenocarcinoma cell line, Calu-3, was infected with SARS-CoV-2 in the absence or presence of azelastin-HCl, and the number of viral particles was measured after 48 hours.

[0177] Experimental procedure: Calu-3 (ATCC® HTB-55®) was seeded on 96-well plates. After 2 days, when the cells reached culture density, the culture was given fresh cell culture medium (EMEM + 10% FBS + 1% P / S, 1% L-glutamine, 1% non-essential amino acids). Azelastine-HCl (Seleckchem catalog no. S2552, 10 mM stock solution dissolved in DMSO) and hydroxychloroquine sulfate (TCI catalog no. H1306, 10 mM stock solution dissolved in H2O and aseptically filtered) were added to the cell culture medium at final concentrations of 50 and 25 μM, respectively (dilutions were prepared in the culture medium). Hydroxychloroquine sulfate has a strong anti-SARS-CoV-2 effect against Vero E6 cells (which do not express TMPRSS2), but its efficacy is significantly reduced when tested in cells that express the surface protease TMPRSS2, including Calu-3 cells (see Hoffmann et al., Nature 2020, pp. 585:588-590).

[0178] Virus stocks were prepared by proliferation in Vero E6 cells and infectivity titers were determined. For viral infection, SARS-CoV-2 virus was added to the supernatant immediately after (essentially simultaneously with) a change in culture medium at an MOI of 0.01 (infection multiplicity: 1 virus particle per 100 cells). After incubation with the virus for 30 minutes, the culture medium was removed and replaced with fresh culture medium containing azelastine-HCl or hydroxychloroquine sulfate at the above concentrations. 48 hours after infection, the supernatant was collected and stored at -80°C for quantitative PCR analysis. Viral RNA was extracted from the culture supernatant sample using the Monarch Total RNA Miniprep kit (New England BioLabs, catalog number T2010S) according to the manufacturer's instructions. In short, 300 μl of lysis buffer was mixed with 100 μl of culture supernatant, gDNA contamination was removed using a dedicated column (while retaining the DNA), and the RNA-containing fraction was poured into an RNA-binding column. After washing the column, the RNA was eluted with H2O, and the samples were stored at -80°C until analysis. Following reverse transcription, the DNA was amplified using the 1-Step RT-ddPCR Advanced Kit for Probes (BioRad ddPCR™, catalog numbers 1864021 and 1864022, https: / / www.bio-rad.com / de-at / product / 1-step-rt-ddpcr-advanced-kit-for-probes?ID=NTGCRI15) with F, R, and P2 primers derived from the RdRp gene.

[0179] [Table 6]

[0180] The results of the RT-PCR reaction were quantified and calculated as viral copy number / μl, and compared to the copy number measured in infected mock-treated (buffer) cells (positive control). Simultaneously, cell viability was measured on uninfected Calu-3 cells treated with azelastin-HCl or hydroxychloroquine sulfate for 48 hours using the CellTiter-Glo® 2.0 cell viability assay (Promega® catalog number G9241) according to the manufacturer's instructions. Briefly, the CellTiter-Glo® 2.0 reagent was added to the cells in a volume equal to the volume of the culture medium and incubated at room temperature for 10 minutes. The luminescence signal was measured and compared to the signal measured in untreated cells (negative control).

[0181] result: Quantitative PCR analysis revealed that azelastine-HCl is highly effective in reducing the number of viral particles in human lung adenocarcinoma cells by >99% at a 50 μM concentration and by more than 90% at a 25 μM concentration (Figure 6), confirming that azelastine exerts its anti-SARS-CoV-2 effect against human respiratory epithelial cells, which express the surface protease TMPRSS2 and are best suited for SARS-CoV-2 infection.

[0182] Example 6 Prevent and treat HEp-2 cell infection caused by respiratory multinuclear virus (RSV). To confirm the broad antiviral effects of azelastine-HCl, HEp-2 cells (ATCC CCL-23) were infected with RSV in the absence or presence of azelastine-HCl. Azelastine-HCl was tested in three settings: prophylaxis (azelastine-HCl treatment followed by infection), co-administration (cells were infected simultaneously and treated with azelastine-HCl), and therapeutic settings (infected cells were treated with azelastine-HCl 1 hour after infection). Spot count and viral genome copy number were determined and compared with cells treated with buffer.

[0183] Experimental procedure: Azelastine-HCl (Sigma catalog number PHR1636-1G) was tested for anti-RSV activity in the 0.4–25 μM concentration range using a 10 mM stock prepared in DMSO.

[0184] 1×10 4 HEp-2 cells (ATCC CCL-23) were seeded in a 96-well plate and cultured at 37 °C, 5% CO2 and 100% humidity for 3 hours in 50 μl / well of DMEM medium supplemented with 10% FCS and 2 mM L-glutamine. After 3 hours, the cells had reached adhesion and were infected with the RSV Long strain (ATCC VR-26, donated by T. Grunwald, Fraunhofer Institute for Cell Therapy and Immunology, Leipzig, Germany) at an MOI of 0.01. Azelastine-HCl was added to the cells 1 hour before infection, simultaneously with infection or 1 hour after infection. The azelastine-HCl stock was serially diluted with culture medium (DMEM / 10% FCS / 2 mM L-glutamine) to the desired final concentration. DMSO was diluted in the same way and added to the cells as a solvent control. The cells infected and treated at a total volume of 200 μl were incubated at 37 °C, 5% CO2 and 100% humidity for 48 hours. RSV infection plaques were determined by immunocytochemical staining using a polyclonal goat antibody against RSV (anti-RSV Gt X (IgG Frac), Merck) and an HRP-conjugated rabbit polyclonal anti-goat IgG (Novusbio). 3-Amino-9-ethylcarbazole (AEC, Sigma) was used as a chromogen in immunohistochemistry to visualize RSV-infected cells. The plaques were counted manually. The viral genome copy number was determined by RT-PCR using the primers RSV-1 (5’-AGA TCA ACT TCT GTC ATC CAG CAA-3’, SEQ ID NO: 18) and RSV-2 (5’-GCA CAT CAT AAT TAG GAG TAT CAA T-3’, SEQ ID NO: 19) previously described by Wilmschen et al. (Vaccines, 2019, 7:59). The Ct values obtained after azelastine-HCl treatment were compared only with those of the solvent-treated cells. To calculate the viral genome copy number in the treated samples compared to the solvent, the inventors used the following equation: 1 Ct increase is equal to 50% fewer copies.

[0185] The cytotoxicity of azelastin-HCl against HEp-2 cells was determined using a flow cytometry-based assay with the viral dye propidium iodide. The experiment was conducted independently 3 to 4 times.

[0186] result: Azelastin-HCl exhibited cytotoxicity against HEp-2 cells at concentrations of 12.8 and 25.6 μM (Figure 7). Therefore, the effect of azelastin-HCl was evaluated at 6.4 μM and lower concentrations. RSV infection of cells was determined by the number of spots, and in all three settings, it was reduced by approximately 50% at concentrations of 3.2 and 6.4 μM compared to the solvent (DMSO) treated group (Figure 8), suggesting a potent prophylactic and therapeutic effect of azelastin-HCl against RSV infection of HEp-2 cells. This effect at a 6.4 μM azelastin-HCl concentration was confirmed by viral genome copy number determination (Table 6).

[0187] [Table 7]

[0188] Example 7 Prevents infection with MucilAir (trademark) caused by influenza A virus H1N1. The efficacy of repeated-dose azelastine-HCl nasal spray (Pollival, UrsaPharm Arzneimittel) against influenza H1N1 infection was tested in fully differentiated human nasal epithelial cells (MucilAir® Pool, Epithelix Sarl - primary cells derived from a pool of 14 different normal nasal donors) cultured at the gas-liquid interface.

[0189] Experimental procedure: The antiviral effect of azelastine-HCl against influenza H1N1 was tested as described by Boda et al. (Antiviral Research 156 (2018), pp. 72-79). Briefly, following a apical wash of a MucilAir® Pool in MucilAir® culture medium (200 μl for 10 minutes), 10 μl of azelastine-HCl at a concentration of 0.02 or 0.01%, diluted in the proprietary diluent from Polival® nasal spray (Ursapharm Arzneimittel GmbH), was poured onto the apical side of the MucilAir® pool for 10 minutes. Subsequently, 100 μl of influenza H1N1 (ATCC VR-95) was added, with 10% azelastine-HCl per ml. 6 The genomic copy number (from stock) was applied to the apical end and incubated for 3 hours. The inoculum was washed away by washing the apical end of the cells three times with 200 μl of MucilAir culture medium (20-minute incubation). After the third wash, 10 μl of the same concentration of azelastin-HCl was added to the apical end of the cells and incubated for 21 hours. At 24, 48, and 72 hours, the MucilAir® cell wash was repeated, and then azelastin was replaced on the apical end as before. The viral copy number was determined from the apical wash at each time point. The basal medium was also removed and replaced daily with 500 μl of fresh culture medium. LDH release was measured from the nasal lavage (to assess cell death at 96 hours), and cytokine levels (IL-8 and RANTES) were determined at 48 and 96 hours. All incubation steps were performed at 34°C, 5% CO2, and 100% humidity. The viral copy number was determined from the washed apical end; therefore, RNA was extracted using the QIAamp® viral RNA kit (Qiagen), and viral RNA was quantified by RT-PCR (QuantyTect Probe RT-PCR, Qiagen) with the qTOWER3 detection system. Ct data were reported aligned to a standard curve and expressed as genome copy number / ml.

[0190] Uninfected cells were included as a negative control. Cells infected with influenza H1N1 but treated only with buffer were used as a positive control. The antiviral effect of azelastine-HCl was compared to that of oseltamivir carboxylate (oseltamivir, 10 μM, Compton, UK, Carbosynth) added to the basal outer compartment of infected cells. Three different compounds were tested.

[0191] result: Treatment of MucilAir® cells with azelastin-HCl at both concentrations resulted in a statistically significant reduction in viral genome copies at 24 hours post-infection compared to buffer-treated cells. The reduction in viral copies was approximately 1.9 log (98.74% reduction) with 0.02% azelastin-HCl and approximately 0.9 log (87.4% reduction) with 0.01% azelastin-HCl (Figure 9).

[0192] Furthermore, azelastine-HCl demonstrated anti-inflammatory effects during MucilAir™ H1N1 infection; both IL-8 (Figure 10 / A) and RANTES (Figure 10 / B) levels were significantly reduced compared to the buffered control on post-infection days 2 and 4.

[0193] A 0.02% concentration of azelastin-HCl showed slight cytotoxicity (6.5%) in uninfected cells, but showed no cytotoxicity in infected cells or at lower concentrations (0.01%).

[0194] Example 8 Prevent and treat A549 cell infection caused by adenovirus. To confirm the broad antiviral effects of azelastine-HCl, human lung adenocarcinoma cells (A549) were infected with adenovirus in the absence or presence of azelastine-HCl. Azelastine-HCl was tested in both co-administration and therapeutic settings.

[0195] Experimental procedure: Azelastine-HCl (Sigma catalog number PHR1636-1G) was tested for anti-adenovirus activity in the concentration range of 0.78–50 μM using a 10 mM stock prepared in DMSO.

[0196] 3 x 10 4 A549 cells (ATCC CCL-185®) were seeded in 100 μl of complete growth medium (DMEM, high glucose, GlutaMAX® supplement, 10% FBS, QUALIFIED, HI 500ML, catalog number: 10500064, lot number: 08Q6291K) in a 96-well plate and incubated at 37°C, 5% CO2, and 100% humidity. After 24 hours, infection was initiated by adding 25 μl of fresh medium to the cells along with adenovirus hAdv5 (ATCC VR-5) MOI 0.01 TCID50. In the co-administration setting, 25 μl of azelastin-HCl diluted in infection growth medium was added to the cells simultaneously with infection. In the therapeutic setting, azelastin-HCl was added to the cells 6 hours after infection. Cells were incubated at 37°C, 5% CO2, and 100% humidity 48 hours after infection. Subsequently, the plates were frozen at -80°C, and the virus was released from the cells through three freeze-thaw cycles. Viral titers were determined according to the manufacturer's instructions using the Adeno-X® Rapid Titer kit (3 repeats per well, Clontech, Takara Bio, catalog number PT3651-2).

[0197] The cytotoxicity of azelastin against uninfected A549 cells was determined using the CellTiter Glo assay (Promega, USA) with the same protocol described above, except that the cells were not infected with adenovirus. In some embodiments, the present invention may be described as follows. [Aspect 1] An azelastine compound in an antivirally effective amount for use as an antiviral substance in a pharmaceutical formulation for use in preventive or therapeutic treatment of subjects requiring antiviral treatment. [Aspect 2] a) Coronavirus family (β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, HCoV-HKU1; or α-coronaviruses such as HCoV-NL63, HCoV-229E, or PEDV, and naturally occurring variants or mutants of any of the aforementioned viruses); b) Adenoviridae (adenoviruses or human adenoviruses, such as HAdVB, HAdVC, or HAdVD); c) Paramyxoviridae (RSV or human RSV, e.g., hRSV subtype A or B); or d) Orthomyxoviridae (influenza viruses or human influenza viruses, preferably influenza virus A (IVA) such as H1N1, H3N3, or H5N1, or influenza virus B (IVB), or influenza virus C (IVC), or influenza virus D (IVD), etc.) An azelastine compound for use according to embodiment 1, wherein a disease condition caused by or associated with infection by one or more viruses selected from among is treated. [Aspect 3] The azelastine compound for use according to Aspect 2, wherein one or more different coronaviruses are naturally occurring SARS-CoV-2 mutants or variants such as SARS-CoV-2 mutants or variants containing one or more mutations in the SARS-CoV-2 S-protein, preferably K417N, L452R, N501Y, D614G, P681H, P681R, E484K, E484Q, or 69 / 70 deletion in SEQ ID NO: 4, and preferably the SARS-CoV-2 mutant or variant is selected from the group consisting of B.1.1.7 (UK mutant), B.1.351 (South Africa), P.1 (Brazil), B.1.617 (India), and B.1.618 (Bengal) variants. [Aspect 4] An azelastine compound for use according to any one of aspects 1 to 3, wherein the pharmaceutical preparation is a medical product or drug comprising an azelastine compound and a pharmaceutically acceptable carrier. [Aspect 5] An azelastine compound for use according to any one of aspects 1 to 4, wherein the disease condition is a common cold, nasal or sinusitis, pharyngeal and laryngeal infection, bronchiolitis, diarrhea, skin rash, or pneumonia, acute respiratory distress syndrome (ARDS). [Aspect 6] An azelastine compound for use according to any one of aspects 1 to 5, wherein the antiviral effective amount is effective in preventing infection of susceptible cells by the virus and thereby treating the disease state. [Aspect 7] An azelastine compound for use according to any one of aspects 1 to 5, wherein the antiviral effective dose is 0.1 to 500 μg / dose. [Aspect 8] An azelastine compound for use according to any one of aspects 1 to 7, wherein the pharmaceutical formulation is formulated for local administration, preferably for use in the upper and lower respiratory tract, nasal cavity, lungs, oral cavity, eyeball, or skin, or for systemic administration by intravenous, intramuscular, subcutaneous, intradermal, transdermal, or oral administration. [Aspect 9] An azelastine compound for use according to any one of aspects 1 to 8, wherein the pharmaceutical preparation is administered to a subject as a spray, powder, gel, ointment, cream, foam, or liquid solution, lotion, mouthwash, aerosolized powder, aerosolized liquid, granules, capsule, drop, tablet, syrup, lozenge, eye drop, or preparation for injection or injection. [Aspect 10] An azelastine compound for use according to any one of aspects 1 to 9, wherein the azelastine compound is applied to the target nose in an antiviral effective amount of 1 to 1000 μg per nostril. [Aspect 11] An azelastine compound for use according to any one of aspects 1 to 10, wherein the azelastine compound is administered as the sole antiviral substance, or the treatment is combined with a further treatment using one or more active substances selected from the group consisting of antiviral substances, anti-inflammatory substances, and antibiotics. [Aspect 12] An azelastine compound for use according to any one of aspects 1 to 11, wherein the subject being treated is infected with or at risk of becoming infected with a virus, preferably a human, dog, cat, horse, camel, cattle, or pig. [Aspect 13] An azelastine compound for use as an antiviral substance in a medical product for treating a biological surface to prevent viral infection and / or viral spread. [Aspect 14] The azelastine compound for use according to aspect 13, wherein the biological surface is a mucous membrane surface that is infected with or at risk of being infected with one or more different viruses. [Aspect 15] The azelastine compound for use according to aspect 14, wherein one or more different viruses are preferably selected from the group consisting of β-coronaviruses such as SARS-CoV-2, MERS-CoV, SARS-CoV-1, HCoV-OC43, or HCoV-HKU1; or α-coronaviruses such as HCoV-NL63, HCoV-229E, or PEDV; and naturally occurring variants or mutants of any of the aforementioned viruses, and are coronaviruses. [Aspect 16] The azelastine compound for use according to Aspect 15, wherein one or more different coronaviruses are naturally occurring SARS-CoV-2 mutants or variants, such as SARS-CoV-2 mutants or variants containing one or more mutations in the SARS-CoV-2 S-protein, preferably K417N, L452R, N501Y, D614G, P681H, P681R, E484K, E484Q, or 69 / 70 deletion in SEQ ID NO: 4, and preferably the SARS-CoV-2 mutant or variant is selected from the group consisting of B.1.1.7 (UK mutant), B.1.351 (South Africa), P.1 (Brazil), B.1.617 (India), and B.1.618 (Bengal) variants. [Aspect 17] An azelastine compound for use according to any one of aspects 13 to 16, wherein the medical product is formulated for topical use, preferably for application to the upper and lower respiratory tract, nasal cavity, lungs, oral cavity, eyeball, or skin. [Aspect 18] An azelastine compound for use according to any one of aspects 13 to 17, wherein the medical product is used as a solution, dispersion, dry powder, or aerosolized liquid or powder. [Aspect 19] The azelastine compound is preferably present in an amount of 1 ng to 1000 ng / cm³.2 An azelastine compound for use according to any one of embodiments 13 to 18, which is applied in an antiviral effective amount. [Aspect 20] Use of azelastine compounds as antiviral agents.

Claims

1. A pharmaceutical composition comprising an azelastine compound, for use in the prevention or treatment of viral diseases in human subjects requiring antiviral treatment, The pharmaceutical composition is used for topical mucosal administration. The azelastine compound is administered at a dose of 0.1 to 500 μg / dose. This viral disease, a) Beta coronavirus or alpha coronavirus; b) RSV; or c) Influenza virus; Caused by one or more viruses selected from, The above-mentioned pharmaceutical composition.

2. The pharmaceutical composition according to Claim 1, wherein the β-coronavirus is a naturally occurring SARS-CoV-2 mutant or variant, such as a SARS-CoV-2 mutant or variant containing one or more mutations of the SARS-CoV-2 S-protein, preferably K417N, L452R, N501Y, D614G, P681H, P681R, E484K, E484Q, or 69 / 70 deletion in SEQ ID NO: 4, and preferably the SARS-CoV-2 mutant or variant is selected from the group consisting of B. 1.1.7 (UK mutant), B. 1.351 (South Africa), P. 1 (Brazil), B. 1.617 (India), and B. 1.618 (Bengal) variants.

3. The pharmaceutical composition according to claim 1 or 2, further comprising a pharmaceutically acceptable carrier.

4. The pharmaceutical composition according to any one of claims 1 to 3, wherein the viral disease is a common cold, nasal, sinusitis, pharyngeal and laryngeal infection, bronchiolitis, diarrhea, skin rash, pneumonia, or acute respiratory spurt syndrome (ARDS).

5. The pharmaceutical composition according to any one of claims 1 to 4, wherein the amount is effective in preventing infection of susceptible cells by the virus and thereby treating a viral disease.

6. A pharmaceutical composition according to any one of claims 1 to 5, wherein the number of doses per day is up to 10.

7. The pharmaceutical composition according to any one of claims 1 to 6, wherein the pharmaceutical composition is formulated for use in the upper and lower respiratory tract, nasal cavity, lungs, oral cavity, eyeball, or skin.

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein the pharmaceutical composition is administered to a subject as a spray, powder, gel, ointment, cream, foam, liquid solution, lotion, mouthwash, aerosolized powder, aerosolized liquid, granules, capsule, drop, tablet, syrup, lozenge, or eye drops.

9. The pharmaceutical composition according to claim 8, wherein the azelastine compound is applied by nasal spray.

10. The pharmaceutical composition according to claim 9, wherein the nasal spray contains 0.001% or 0.15% (w / w) of an azelastine compound or a pharmaceutically acceptable salt thereof in an aqueous solution at pH 6.8 ± 0.

3.

11. The pharmaceutical composition according to any one of claims 1 to 10, wherein the azelastine compound is applied to the nose of a target in an amount of 1 to 1000 μg per nostril.

12. The pharmaceutical composition according to any one of claims 1 to 11, wherein the azelastine compound is administered as the sole antiviral substance, or the treatment is preferably combined with a further treatment using one or more active substances selected from the group consisting of antiviral substances, anti-inflammatory substances, and antibiotics.

13. A pharmaceutical composition according to any one of claims 1 to 12, wherein the human subject is infected with or at risk of becoming infected with a virus.

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