Use of betrixaban or its pharmaceutically acceptable salt in the preparation of antiviral drugs

Abstract

The present invention discloses the use of betrixaban or a pharmaceutically acceptable salt thereof for preparing an antiviral drug. The present invention has found, through drug screening and in vitro drug experiments, that betrixaban or a pharmaceutically acceptable salt thereof can inhibit viral replication and infectivity, and can block virus-induced syncytia, thereby reducing cell death caused by syncytia. In addition, betrixaban or a pharmaceutically acceptable salt thereof of the present invention has a broad spectrum of antiviral activity and is also safe and highly effective.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and in particular relates to the use of betrixaban or a pharmaceutically acceptable salt thereof in the preparation of antiviral drugs. Background Art

[0002] In recent years, due to changes in the natural environment and the expansion of human activities, viral diseases have become more frequent, posing a significant threat to human public health and safety. The spread of viral diseases is a major public health and safety issue that threatens human health. Therefore, there is an urgent need to develop safe, broad-spectrum, and highly effective antiviral drugs. Summary of the Invention

[0003] To address at least some of the technical problems in the prior art, the present invention provides the use of betrixaban or a pharmaceutically acceptable salt thereof in the preparation of an antiviral drug. Specifically, the present invention includes the following contents.

[0004] In a first aspect, the present invention provides use of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof in the preparation of an antiviral drug.

[0005] In certain embodiments, according to the use of the present invention, the virus comprises a DNA virus and / or an RNA virus.

[0006] In certain embodiments, according to the use of the present invention, the DNA virus comprises at least one of herpes simplex virus type 1, herpes simplex virus type 2, varicella-zoster virus, cytomegalovirus, smallpox virus, monkeypox virus, adenovirus, human papillomavirus, parvovirus, bocavirus and hepatitis B virus.

[0007] In certain embodiments, according to the use of the present invention, the RNA virus includes at least one of vesicular stomatitis virus, encephalomyocarditis virus, mouse hepatitis virus, influenza A virus, influenza B virus, influenza C virus, dengue virus, Zika virus, Ebola virus, Marburg virus, Nipah virus, coronavirus, hepatitis A virus, hepatitis C virus, rotavirus, measles virus, human immunodeficiency virus, respiratory syncytial virus and rabies virus.

[0008] In certain embodiments, according to the use of the present invention, the antiviral agent comprises at least one of the following:

[0009] (1) Inhibit virus invasion of host cells;

[0010] (2) inhibiting viral genome replication;

[0011] (3) inhibiting viral protein processing or assembly;

[0012] (4) inhibiting viral release;

[0013] (5) Reduce the amount or activity of the virus.

[0014] In certain embodiments, the use according to the present invention is achieved by administering a therapeutically effective amount of the drug to the subject.

[0015] In certain embodiments, according to the use of the present invention, the subject comprises a mammal.

[0016] In certain embodiments, the use according to the present invention, wherein the mammal comprises a human.

[0017] In certain embodiments, according to the use of the present invention, the therapeutically effective amount is 0.01-1000 mg / Kg.

[0018] The second aspect of the present invention provides the use of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof and other drugs in the preparation of a combined antiviral drug.

[0019] The present invention has found through drug screening and in vitro drug experiments that betrixaban or its pharmaceutically acceptable salts, solvates, hydrates, prodrugs, isomers, analogs, derivatives or metabolites can inhibit viral replication and infectivity, and can block virus-induced syncytia, thereby reducing cell death caused by syncytia. In addition, betrixaban or its pharmaceutically acceptable salts, solvates, hydrates, prodrugs, isomers, analogs, derivatives or metabolites of the present invention have broad-spectrum antiviral activity and are safe and highly effective. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Shown is the inhibitory effect of betrixaban on vesicular stomatitis virus (VSV) virus in human fibrosarcoma cells (2fTGH cells).

[0021] Figure 2 Inhibitory effects of betrixaban on other viral transcription and on syncytium formation are shown.

[0022] Figure 3 The inhibitory effect of betrixaban on VSV virus in other cells is shown.

[0023] Figure 4 The results of a study on the antiviral mechanism of betrixaban are shown. DETAILED DESCRIPTION

[0024] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0025] It should be understood that the terms described in the present invention are only for describing particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges in the present invention, it should be understood that the upper and lower limits of the ranges and each intermediate value therebetween are specifically disclosed. Each smaller range between any stated value or intermediate value within a stated range and any other stated value or intermediate value within the stated range is also included in the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0026] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention belongs. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In the event of any conflict with any incorporated document, the content of this specification shall prevail.

[0027] use

[0028] One aspect of the present invention provides use of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof in the preparation of an antiviral drug.

[0029] As used herein, the term "pharmaceutically acceptable salt" refers to organic and inorganic salts of the compound betrixaban of the present invention. Pharmaceutically acceptable salts are well known in the art, and examples include, but are not limited to, inorganic acid salts such as hydrochlorides, hydrobromides, phosphates, sulfates, and perchlorates; organic acid salts such as acetates, oxalates, maleates, tartrates, citrates, succinates, and malonates; or salts obtained by other methods described in the literature, such as ion exchange methods. Other pharmaceutically acceptable salts include adipate, malate, 2-hydroxypropionate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, cyclopentylpropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, stearate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, and the like.

[0030] As used herein, the term "solvate" refers to an association or complex formed between one or more solvent molecules and a compound of the present invention. Solvents that form solvates include, but are not limited to, water, isopropanol, ethanol, methanol, dimethyl sulfoxide, ethyl acetate, acetic acid, aminoethanol, and the like.

[0031] The term "hydrate" as used herein refers to an association or complex formed when the solvent molecule is water and the compound of the present invention.

[0032] As used herein, the term "isomer" includes enantiomeric, diastereomeric, and geometric (or conformational) isomeric forms of a given structure. For example, the present invention includes R and S configurations for each asymmetric center, Z and E double bond isomers, Z and E conformational isomers, single stereochemical isomers, and enantiomeric, diastereomeric, and geometric (or conformational) isomer mixtures. Unless otherwise indicated, the present invention includes all tautomeric forms of the disclosed structures of betrixaban.

[0033] The term "prodrug" as used herein refers to a compound that is converted into a compound represented by the following formula (I) or formula (II) in vivo:

[0034] Formula (I);

[0035] Formula (II).

[0036] The term "prodrug" refers to a pharmacologically inactive derivative of a parent drug molecule that requires spontaneous or enzymatic biotransformation in vivo to release an active drug. Prodrugs are variants or derivatives of compounds of the present invention that have a cleavable group under metabolic conditions. When prodrugs undergo solvent degradation or undergo enzymatic degradation under physiological conditions, they become compounds of the present invention that are pharmaceutically active in vivo. Prodrug forms typically provide advantages of solubility, tissue compatibility, or delayed release in mammalian organisms (see Bundgard, Design of Prodrugs (prodrug design), pp.7-9, 21-24, Elsevier, Amsterdam 1985 and Silverman, The Organic Chemistry of Drug Design and Drug Action (organic chemistry and drug action of drug design), pp.352-401, Academic Press, San Diego, CA, 1992). Prodrugs known in the art include acid derivatives well known to those skilled in the art, such as esters prepared by reacting the parent acid with a suitable alcohol, or amides prepared by reacting the parent acid compound with an amine, or with a basic group which reacts to form an acylated base derivative.

[0037] As used herein, the term "metabolite" refers to a product obtained by metabolism of a specific compound or its salt in vivo. The metabolites of a compound can be identified by techniques known in the art, and their activity can be characterized by methods known in the art. Such products can be obtained by administering the compound through oxidation, reduction, hydrolysis, amidation, deamidation, esterification, defatting, enzymatic cleavage, and the like. Accordingly, the present invention includes metabolites of compounds of formula (I) or (II), including metabolites produced by contacting a compound of formula (I) or (II) of the present invention with a mammal or a cell line derived therefrom for a sufficient period of time.

[0038] In the present invention, the term "derivative" refers to a product derived from the substitution of atoms or functional groups in the compound of formula (I) or (II) by other atoms or functional groups. The derivative is not specifically limited and includes but is not limited to esters, acylates or metal complexes of the compound of formula (I) or (II).

[0039] In the present invention, the viruses include DNA viruses and / or RNA viruses. Examples of DNA viruses include, but are not limited to, herpes simplex virus type 1, herpes simplex virus type 2, varicella-zoster virus, cytomegalovirus, smallpox virus, monkeypox virus, adenovirus, human papillomavirus, parvovirus, bocavirus, hepatitis B virus, etc.; examples of RNA viruses include, but are not limited to, vesicular stomatitis virus, encephalomyocarditis virus, mouse hepatitis virus, influenza A virus, influenza B virus, influenza C virus, dengue virus, Zika virus, Ebola virus, Marburg virus, Nipah virus, coronavirus, hepatitis A virus, hepatitis C virus, rotavirus, measles virus, human immunodeficiency virus, respiratory syncytial virus, rabies virus, etc. In a preferred embodiment, the DNA virus is herpes simplex virus type 1. In another preferred embodiment, the RNA virus is vesicular stomatitis virus. In another preferred embodiment, the RNA virus is encephalomyocarditis virus. In another preferred embodiment, the RNA virus is mouse hepatitis virus. In another preferred embodiment, the RNA virus is influenza A virus.

[0040] As used herein, "antiviral" refers to improving a condition before or after the onset of a disease or disorder. This relief or prevention is at least 5%, 10%, 20%, 40%, 50%, 60%, 80%, 90%, 95% or 100% as measured by any standard technique compared to an untreated control group under the same conditions. Beneficial or desired clinical results include, but are not limited to, the following, whether detectable or undetectable, including relief of symptoms, reduction in disease severity, stabilization of the disease state (i.e., no worsening), delay or slowing of disease progression, improvement or alleviation of the disease state, and reduction (whether partial or complete). Antiviral conditions include, but are not limited to (1) inhibition of viral invasion of host cells; (2) inhibition of viral genome replication; (3) inhibition of viral protein processing or assembly; (4) inhibition of viral release; and (5) reduction of viral amount or activity.

[0041] In the present invention, antiviral treatment is achieved by administering a therapeutically effective amount of the drug to a subject, wherein the subject includes but is not limited to mammals, and examples of mammals include but are not limited to humans, mice, rabbits, cats, dogs, cattle, sheep, pigs, etc.

[0042] In the present invention, the medicament contains a therapeutically effective amount of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof, particularly betrixaban maleate, and optionally, a pharmaceutically acceptable excipient or carrier. As used herein, the term "pharmaceutically acceptable excipient or carrier" refers to a pharmaceutically acceptable material, composition or medium, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, that participates in the transport or delivery of the medicament from one organ or part of the body to another organ or part of the body. Each excipient or carrier is "acceptable" in that it is compatible with the other ingredients of the formulation and does not harm the patient.

[0043] In the present invention, the mode of administration of the drug is not particularly limited. Representative modes of administration include, but are not limited to, oral, rectal, parenteral (intravenous, intramuscular, or subcutaneous), and topical administration. Accordingly, the drug of the present invention can be prepared into various clinically acceptable dosage forms, including oral dosage forms, injection dosage forms, topical dosage forms, or external dosage forms.

[0044] The therapeutically effective amount of the present invention refers to the pharmaceutically effective dosage, that is, the amount of the active compound (i.e., betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof, in particular betrixaban maleate) sufficient to significantly improve the condition without causing serious side effects. The daily dosage of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof is generally 0.01-1000 mg / Kg, preferably 0.01-500 mg / Kg, or 0.01-400 mg / Kg, or 0.01-300 mg / Kg, or 0.01-200 mg / Kg, or 0.01-150 mg / Kg, or 0.01-100 mg / Kg, or 0.01-50 mg / Kg, or 0.01-40 mg / Kg, or 0.01-30 mg / Kg, most preferably 0.01-20 mg / Kg. Exemplary effective dosages include, for example, 0.01 mg / Kg, 0.05 mg / Kg, 0.1 mg / Kg, 0.2 mg / Kg, 0.3 mg / Kg, 0.4 mg / Kg, 0.5 mg / Kg, 0.75 mg / Kg, 0.95 mg / Kg, 1 mg / Kg, 1.25 mg / Kg, 1.5 mg / Kg, 1.75 mg / Kg, 2 mg / Kg, 2.5 mg / Kg, 2.75 mg / Kg, 3 mg / Kg, 3.25 mg / Kg, 3.5 mg / Kg, 3.75 mg / Kg, 4 mg / Kg, 4.25 mg / Kg, 4.5 mg / Kg, 4.75 mg / Kg, 5 mg / Kg, 5.25 mg / Kg, 5.5 mg / Kg, 5.75 mg / Kg, 6 18 mg / Kg, 19 mg / Kg, 20 mg / Kg. Preferably, the above daily dosage is based on betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof. It can be administered as a single dose once daily, divided into multiple doses per day, or administered at intervals.

[0045] Joint use

[0046] In one aspect of the present invention, there is provided the use of betrixaban or a pharmaceutically acceptable salt, solvate, hydrate, prodrug, isomer, analog, derivative or metabolite thereof and other drugs in the preparation of a combined antiviral drug.

[0047] In the present invention, other drugs include but are not limited to remdesivir, ribavirin, favipiravir, acyclovir, valacyclovir, famciclovir, ganciclovir, brivudine, interferon, oseltamivir phosphate, peramivir, zanamivir, mabaloxavir, favipiravir, arbidol, etc.

[0048] Example

[0049] The inhibitory effect of betrixaban on viruses is exemplified below.

[0050] 1. Inhibitory effect of betrixaban on VSV in 2fTGH cells

[0051] 1.1 Experimental Methods

[0052] 2fTGH cells were treated with betrixaban (DMSO) and VSV at concentrations of 10, 40, 60, and 80 μM, respectively. DMSO+VSV served as the control group. After 12 h and 24 h of treatment, the expression levels of VSV gene and protein were detected by qPCR and Western Blot, respectively. The content of viral particles in the cell supernatant was detected by plaque assay.

[0053] 1.2 Experimental Results

[0054] The results are as follows Figure 1 Specifically, the qPCR results showed that as the concentration of betrixaban increased, the relative amount of VSV viral gene expression decreased, and the effect of the drug on inhibiting viral transcription was not enhanced by the duration of drug action ( Figure 1 Western Blot analysis showed that the expression of VSV virus protein decreased with the increase of betrixaban concentration compared with the internal reference protein ( Figure 1 B), Figure 1 The concentrations of betrixaban in B are 20, 40, and 80 μM from left to right. The results of the viral plaque assay showed that the viral titer decreased significantly with the increase of betrixaban concentration. Compared with the DMSO group, the viral load was almost 0 when the concentration of betrixaban was 60 μM, indicating that the replication and infection ability of the virus was negatively correlated with the concentration of betrixaban ( Figure 1 C).

[0055] In summary, betrixaban can reduce VSV viral transcription and expression, and inhibit viral replication and infection ability.

[0056] 2. Inhibitory effects of betrixaban on other viruses

[0057] 2.1 Experimental Methods

[0058] 2fTGH cells were treated with betrixaban and herpes simplex virus type 1 (HSV-1) at concentrations of 20, 40, and 80 μM, respectively, and DMSO+HSV-1 served as the control group for 12 h. 2fTGH cells were treated with betrixaban and encephalomyocarditis virus (EMCV) at concentrations of 20, 40, and 80 μM, respectively, and DMSO+EMCV served as the control group for 12 h. 2fTGH cells were treated with betrixaban and mouse hepatitis virus (MHV) at concentrations of 20, 40, and 80 μM, respectively, and DMSO+MHV served as the control group for 12 h. 2fTGH cells were treated with betrixaban and influenza A virus (IAV) at concentrations of 20, 40, and 80 μM, respectively, and DMSO+IAV served as the control group for 12 h. The expression changes of viral genes were quantitatively detected by qPCR. In addition, the virus group and DMSO group were used as controls to observe the effect of betrixaban on the syncytium formation induced by HSV-1 and MHV in 2fTGH and J774.1 cells.

[0059] 2.2 Experimental Results

[0060] The results are as follows Figure 2 As shown in Figure 2, specifically, the qPCR relative quantitative results showed that betrixaban inhibited the transcription of HSV-1, EMCV, MHV and IAV viral genes, and as the drug concentration increased, the relative expression of viral genes decreased ( Figure 2 A). The cell syncytium experiment showed that the cell syncytium formation in the betrixaban-treated group was significantly reduced compared with the virus group and DMSO group, indicating that betrixaban can effectively inhibit the cell syncytium formation caused by MHV and HSV infection ( Figure 2 B).

[0061] These results indicate that betrixaban has a broad-spectrum antiviral activity and can block virus-induced syncytia, thereby reducing cell death caused by syncytia.

[0062] 3. Inhibitory effect of betrixaban on VSV in other cells

[0063] 3.1 Experimental Methods

[0064] A549, HeLa, HT29, Pan02, and Raw264.7 cells were co-treated with betrixaban (80 μM) and VSV, and viral gene expression was quantitatively analyzed by qPCR. In addition, HeLa cells were treated with 30 and 50 μM betrixaban and VSV-GFP (green fluorescent protein), respectively, using mock and DMSO groups as controls, and fluorescence intensity was observed using a fluorescence microscope.

[0065] 3.2 Experimental Results

[0066] The results are as follows Figure 3 Specifically, the qPCR results showed that the VSV virus mRNA expression level in the experimental group (betrixaban + VSV) was significantly lower than that in the control group (DMSO + VSV). This shows that betrixaban can significantly inhibit the transcriptional expression of VSV in these cell lines ( Figure 3 A). Microscopic results showed that compared with the control group, the number of green fluorescent cells decreased significantly with the increase of betrixaban concentration, indicating that betrixaban can inhibit the expression of VSV-GFP ( Figure 3 In summary, betrixaban has significant antiviral activity and can inhibit viral replication in multiple cell lines.

[0067] 4. Study on the antiviral mechanism of betrixaban

[0068] 4.1 Experimental Methods

[0069] cGAS knockout and wild-type 2fTGH cells were treated simultaneously with 20, 40, and 80 μM betrixaban and VSV, respectively. The expression levels of the antiviral genes MX1 and MX2 and the inhibition of VSV viral transcription were then detected by qPCR.

[0070] 4.2 Experimental Results

[0071] The results are as follows Figure 4 As shown by Figure 4 As shown in A, compared with the betrixaban control group, the expression of MX1 and MX2 genes in the cGAS knockout group was significantly reduced, indicating that the cGAS pathway was inhibited. Figure 4 As shown in Figure 2, the VSV virus mRNA gene detection results showed that in the wild-type 2fTGH model, compared with the DMSO control group, the expression of VSV mRNA in the betrixaban group was significantly decreased, and the expression level decreased with the increase of drug concentration; similarly, in the cGAS knockout model, the expression level of VSV mRNA in the betrixaban group decreased with the increase of drug concentration. This shows that betrixaban not only produces antiviral effects through the cGAS pathway, that is, cGAS is an important pathway among multiple antiviral pathways.

[0072] Finally, it should be noted that the above embodiments are intended only to illustrate the technical solutions of the present invention, and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that the technical solutions described in the above embodiments may be modified or some of the technical features thereof may be replaced with equivalents. Such modifications or replacements do not deviate from the spirit and scope of the technical solutions of the various embodiments of the present invention.

Claims

1. The use of betrixaban in the preparation of antiviral drugs, characterized in that: The virus is vesicular stomatitis virus.

2. The use according to claim 1, characterized in that The anti-virus includes at least one of the following situations: (1) Inhibit virus invasion of host cells; (2) inhibiting viral genome replication; (3) inhibiting viral protein processing or assembly; (4) inhibiting viral release; (5) Reduce the amount or activity of the virus.

3. The use according to claim 1, characterized in that The antiviral effect is achieved by administering a therapeutically effective amount of the drug to the subject.

4. The use according to claim 3, characterized in that The subject includes a mammal.

5. The use according to claim 4, characterized in that The mammals include humans.

6. The use according to claim 3, characterized in that The therapeutically effective amount is 0.01-1000 mg / Kg.

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