Treatment or prevention of coronavirus infections

Abstract

1. A method for treating or preventing a coronavirus infection in a subject, comprising administering a therapeutically effective amount of a compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof, wherein the coronavirus comprises at least one selected from the group consisting of 2019-nCov virus, HCov 229E virus, SARS virus, and MERS virus. TIFF2022539640000018.tif6288(I)

Description

[Technical Field]

[0001] The present invention relates to the medical field, in particular to the treatment or prevention of coronavirus infections in a subject. [Background technology]

[0002] The World Health Organization recently declared severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) a public health emergency of international concern. As of June 1, 2020, there have been over 6 million confirmed cases and over 370,000 confirmed deaths worldwide.

[0003] Coronaviridae (also known as "coronaviruses") are a family of enveloped, single-stranded, positive-sense RNA viruses. Coronaviruses can cause disease in mammals and birds. In humans, the viruses usually cause mild respiratory infections, including the common cold, although rare forms, such as SARS (including the one that causes COVID-19) and MERS, can be fatal.

[0004] Therefore, there is an increasing demand for research and development of more effective methods for treating or preventing coronavirus infections. Summary of the Invention

[0005] The following is merely a summary of certain embodiments of the present invention and is not intended to limit the present invention. All citations herein are incorporated by reference in their entirety. Where the disclosure of the present invention differs from the citations, the disclosure of the present specification shall control.

[0006] During the research and development process, the present inventors have surprisingly found that in multiple experiments, WS-635 (the compound of formula (I)) exhibits significantly high efficacy and activity in treating or preventing coronavirus infections.

[0007] In one embodiment of the present disclosure, there is provided a method of treating or preventing a coronavirus infection in a subject, said method comprising administering to said subject in need thereof a therapeutically effective amount of a compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof; [ka] (I) The method is provided, wherein the coronavirus family includes at least one selected from the group consisting of 2019-nCov virus, HCov 229E virus, SARS virus, and MERS virus.

[0008] In another embodiment of the present disclosure, there is provided a pharmaceutical composition for treating or preventing a coronavirus infection, said pharmaceutical composition comprising a compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof: [ka] (I) The pharmaceutical composition is provided, wherein the coronavirus comprises at least one selected from the group consisting of 2019-nCov virus, HCov 229E virus, SARS virus, and MERS virus.

[0009] In another embodiment of the present disclosure, there is provided a compound of Formula I, or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof, for use in treating or preventing a coronavirus infection in a subject: [ka] (I) The coronavirus family includes at least one selected from the group consisting of 2019-nCov virus, HCov 229E virus, SARS virus, and MERS virus.

[0010] In another embodiment of the present disclosure, there is provided the use of a compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof in the manufacture of a medicament for treating or preventing a coronavirus infection, comprising: [ka] (I) The coronavirus family includes at least one selected from the group consisting of 2019-nCov virus, HCov 229E virus, SARS virus, and MERS virus.

[0011] The foregoing is merely a summary of particular embodiments disclosed herein and is not intended to be limiting in any way. These, other embodiments, and further aspects, features, and advantages of the present invention will be apparent from the following detailed description and from the practice of the invention. [Brief explanation of the drawings]

[0012] These and other embodiments and advantages of aspects of the present disclosure will become apparent and more readily understood from the following detailed description taken in conjunction with the accompanying schematics and figures. [Figure 1] Figure 1 shows the inhibitory effect of WS-635 on SARS-CoV-2 coronavirus at the cellular level. [Figure 2] Figure 2 shows the inhibitory effect of WS-635 on SARS-CoV-2 coronavirus at the cellular level. [Figure 3] Figure 3 shows the activity results of WS-635 against human coronavirus (HCoV) 229E.

[0013] definition Unless otherwise stated, the following terms and phrases used herein have the following meanings:

[0014] "Ester" means any ester of a compound, wherein the -COOH functional group of the ester molecule is a substituted -C(O)OR functional group, or the -OH functional group of the ester molecule is an -OC(O)R functional group, where R may form a stable ester moiety, which may be any carbon-containing group, including, but not limited to, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heterocyclyl, heterocycloalkyl, and substituted derivatives thereof.

[0015] "Pharmaceutically acceptable" means suitable for use in pharmaceutical preparations, generally regarded as safe, approved for use by a national or local regulatory body, or listed in the Pharmacopeia or other generally recognized pharmacopeia for use in animals, particularly humans.

[0016] "Pharmaceutically acceptable carrier" means a pharmaceutically acceptable diluent, adjuvant, excipient, carrier, or other ingredient that is administered in combination with a compound of the invention.

[0017] "Pharmaceutically acceptable salts" refers to those salts in which the desired pharmacological activity is enhanced. These pharmaceutically acceptable salts with inorganic or organic acids include acid addition salts, metal salts, and amine salts. Acid addition salts may be formed with inorganic and organic acids.

[0018] "Prodrug" means that the active ingredient drug substance can be formed by a biological system upon administration as a result of spontaneous chemical reaction, enzyme-catalyzed chemical reaction, photolysis, and / or metabolism of a chemical process. Thus, the prodrug is a covalently modified analog or latent form of the therapeutically active compound. Non-limiting examples include prodrug esters, quaternary ammonium molecules, glycol molecules, etc.

[0019] "Therapeutically effective amount" means that the compound is administered for treating a disease in an amount sufficient to effect treatment for said disease.

[0020] "Treat" a therapeutically effective amount of a compound means and includes any of the following uses: (a) administering to a patient a therapeutically effective amount of a therapeutically effective compound; (1) preventing disease pathology or symptoms from being experienced or developed in disease-prone animals that may lead to disease; (2) inhibiting disease pathology or symptoms experienced or occurring in an animal (i.e., preventing further progression of pathology and / or symptoms); or (3) Ameliorating the pathology or symptoms of a disease experienced or occurring in an animal (i.e., alleviating the pathology or symptoms).

[0021] compound The present disclosure relates to compounds of Formula I or a stereoisomer, tautomer, N-oxide, solvate, metabolite, pharmaceutically acceptable salt, or prodrug thereof: [ka] (I)

[0022] When treating or preventing coronavirus infections in a subject in need thereof, The compound of formula I or stereoisomers, tautomers, N-oxides, solvates, metabolites, pharmaceutically acceptable salts, or Prodrug is administered In some examples, the coronavirus family can include at least one selected from the 2019-nCov virus, the HCov 229E virus, the SARS virus, and the MERS virus.

[0023] In some examples of the present disclosure, the compounds of Formula I can be used as pharmaceutically acceptable salts, including phosphate, hydrochloride, hydrobromide, hydroiodide, sulfate, nitrate, ethanesulfonate, toluenesulfonate and benzenesulfonate, acetate, trifluoroacetate, tartrate, maleate, succinate, citrate, benzoate, salicylate, and ascorbate, adipate, alginate, alginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogen phosphate, dinitrobenzoate, dodecyl sulfate, and fumarate. , galactarate, galacturonate, glucoheptonate, gluconate, glutamate, glycerophosphate, hemisuccinate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, isethionate, isobutyrate, lactate, lactobionate, malate, malonate, mandelate, metaphosphate, methanesulfonate, methyl benzoate, monohydrogen phosphate, 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, oleate, pamoate, pectinate, persulfate, phenyl acetate, 3-phenylpropionate, phosphate, phosphonate, and phthalate.

[0024] Pharmaceutical Preparations and Uses The compounds of the present disclosure may be formulated with conventional carriers and excipients, or carriers and excipients suitable for selected common procedures. Tablets may contain excipients, lubricants, fillers, binders, etc. Aqueous preparations, particularly those intended for oral administration as isotonic solutions, are formulated aseptically.

[0025] The active ingredient can be administered alone, but is preferably formulated into a pharmaceutical preparation. The formulation of the present invention, whether for human or animal use, comprises at least one active ingredient, one or more acceptable carriers, and any other therapeutic ingredients.

[0026] The formulations of the present invention suitable for oral administration may be presented as discrete units, each containing a particular amount of the active ingredient, such as capsules, cachets, or tablets; powders or granules; in aqueous or non-aqueous solutions or suspensions; or as oil-in-water or water-in-oil emulsions. The active ingredient may also be in the form of a bolus, electuary, or paste.

[0027] The effective amount of the active ingredient will depend, at least, on the characteristics of the disease being treated, toxicity, the compound used for prophylaxis (low doses) or for active viral infection, the pharmaceutical formulation, and the method of administration, and will be determined by conventional dose escalation studies by the clinician.

[0028] In a preferred embodiment of the present disclosure, the present disclosure relates to a compound of formula I. In multiple experiments, the present inventors have surprisingly discovered that the compound of formula I exhibits significantly high efficacy and activity in treating or preventing coronavirus infections.

[0029] The compound of formula I or a pharmaceutically acceptable salt, ester, or prodrug thereof may be administered to a subject in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, a granule, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, a microparticle, or a pill. In some examples, the compound of formula I or a pharmaceutically acceptable salt, ester, or prodrug thereof may be administered in a daily amount of less than 1000 mg, for example, 100 mg to 1000 mg, or 100 mg to 500 mg, of the compound of formula I.

[0030] Furthermore, those skilled in the art will understand that combination therapy may also be used. The compound of Formula I or a pharmaceutically acceptable salt, ester, or prodrug of the compound of Formula I may be combined with one or more active ingredients. Suitable combination active ingredients include, but are not limited to, one or more selected from the group consisting of corticosteroids, anti-inflammatory signaling modulators, β2-adrenergic receptor agonist bronchodilators, anticholinergic agents, mucolytic agents, hypertonic saline, cytokine storm suppressants, other drugs for treating COVID-19 virus infection, or mixtures thereof. In some examples, a method for treating or preventing COVID-19 virus infection in a subject is provided, comprising administering a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof to the subject in need thereof. [ka] (I)

[0031] In particular embodiments, the method further comprises administering a therapeutically effective amount of at least one other therapeutic agent or composition selected from the group including corticosteroids, anti-inflammatory signaling modulators, beta-2 adrenergic receptor agonist bronchodilators, anticholinergic agents, mucolytic agents, hypertonic saline, cytokine storm suppressants, other drugs for treating COVID-19 virus infection, or mixtures thereof.

[0032] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered to a subject in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, granules, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, microparticles, or a pill.

[0033] In a specific embodiment, the compound of Formula I, or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof, is administered in a daily amount of less than 1000 mg of the compound of Formula I.

[0034] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered in a daily dose of 100 mg to 1000 mg of the compound of formula I.

[0035] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered in a daily dose of 100 mg to 500 mg of the compound of formula I.

[0036] In some other examples, there is provided a pharmaceutical composition for treating or preventing a covid-19 virus infection in a subject in need thereof, the pharmaceutical composition comprising a compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof. [ka] (I)

[0037] In specific examples, the composition further comprises at least one other therapeutic agent or composition selected from the group comprising corticosteroids, anti-inflammatory signaling modulators, beta-2-adrenergic receptor agonist bronchodilators, anticholinergic agents, mucolytic agents, hypertonic saline, cytokine storm suppressants, other drugs for treating COVID-19 viral infection, or mixtures thereof.

[0038] In specific examples, the pharmaceutically acceptable salts include phosphate, hydrochloride, hydrobromide, hydroiodide, sulfate, nitrate, ethanesulfonate, toluenesulfonate, and benzenesulfonate, acetate, trifluoroacetate, tartrate, maleate, succinate, citrate, benzoate, salicylate, and ascorbate, adipate, alginate, alginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogen phosphate, dinitrobenzoate, dodecyl sulfate, fumarate, galactarate, and the like. e) at least one selected from the group comprising galacturonate, glucoheptonate, gluconate, glutamate, glycerophosphate, hemisuccinate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, isethionate, isobutyrate, lactate, lactobionate, malate, malonate, mandelate, metaphosphate, methanesulfonate, methyl benzoate, monohydrogen phosphate, 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, oleate, pamoate, pectinate, persulfate, phenyl acetate, 3-phenylpropionate, phosphate, phosphonate, and phthalate.

[0039] In specific examples, the compound of Formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, granules, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, microparticles, or a pill.

[0040] In specific examples, the composition comprises the compound of formula I, or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof, in a dosage of less than 1000 mg, less than 800 mg, less than 600 mg, less than 500 mg, or less than 200 mg.

[0041] The present disclosure provides a compound of formula I, or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof, for use in treating or preventing a covid-19 virus infection in a subject. [ka] Formula (I)

[0042] In specific examples, the pharmaceutically acceptable salts include phosphate, hydrochloride, hydrobromide, hydroiodide, sulfate, nitrate, ethanesulfonate, toluenesulfonate, and benzenesulfonate, acetate, trifluoroacetate, tartrate, maleate, succinate, citrate, benzoate, salicylate, and ascorbate, adipate, alginate, alginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogen phosphate, dinitrobenzoate, dodecyl sulfate, fumarate, galactarate, and the like. e) at least one selected from the group comprising galacturonate, glucoheptonate, gluconate, glutamate, glycerophosphate, hemisuccinate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, isethionate, isobutyrate, lactate, lactobionate, malate, malonate, mandelate, metaphosphate, methanesulfonate, methyl benzoate, monohydrogen phosphate, 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, oleate, pamoate, pectinate, persulfate, phenyl acetate, 3-phenylpropionate, phosphate, phosphonate, and phthalate.

[0043] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered to the subject in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, granules, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, microparticles, or a pill.

[0044] In a specific embodiment, the compound of Formula I, or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof, is administered in a daily amount of less than 1000 mg of the compound of Formula I.

[0045] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered in a daily dose of 100 mg to 1000 mg of the compound of formula I.

[0046] In a specific example, the compound of formula I or a pharmaceutically acceptable salt, ester, hydrate, or prodrug thereof is administered in a daily dose of 100 mg to 500 mg of the compound of formula I. [Example]

[0047] Example 1. Evaluation of the inhibitory effect of WS-635 on SARS-CoV-2 coronavirus at the cellular level WS-635 as a powder provided by Waterstone Pharmaceuticals Inc. Viral cells were preserved by the Virology Laboratory at the Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences. The SARS-CoV-2 coronavirus isolate (deposit code: 2019-nCoV BetaCoV / Beijing / AMMS01 / 2020) was deposited by the Academy of Military Medical Sciences, Institute of Microbiology and Epidemiology. EC of WS-635 by viral nucleic acid quantification 50 Identifying: Human Vero cells were purchased from ATCC (US), and SARS-Cov-2 virus was isolated by Beijing Military Medical Research Institute under the deposit code: 2019-nCoV BetaCoV / Beijing / AMMS01 / 2020.

[0048] 1) CC of drugs 50 The activity of Vero cells was determined by the MTS (Promega) method. Details are as follows: Vero cells were seeded into a 96-well cell culture plate at a concentration of 10,000 / well and cultured overnight at 37°C in 5% CO2. When the cells grew into a complete monolayer, 100 μL / well of 2% FBS-DMEM maintenance solution containing the drug at different concentrations (initial concentration: 400 μM) was added, and four wells were measured for each concentration. The culture was continued, and the state of the cells was observed daily under a microscope. Until the fourth day of administration, 20 μL of MTS solution was added, and the cells were cultured at 37°C for 1 hour, and the OD490 value was measured. 2) EC of the drug 50 The virus was identified by nucleic acid quantification as follows: One day before, Vero cells were seeded into a 96-well cell culture plate at a concentration of 10,000 / well. The 2019-nCoV virus was diluted with 2% cell maintenance medium and added to the 96-well plate, and the viral load in each well was 100 TCID. 50 WS-635 dissolved in DMSO was prepared using 2% FBS-DMEM maintenance solution at concentrations of 400, 200, 100, 50, 25, and 12.5 μM. The cell culture supernatant was discarded, and T705 (100 μL / well) at different concentrations was added. Four wells were used for each drug, and 100 TCID50 A virus solution was added to each droplet. A drug positive control (10 μM Remdesivir (RDV)) group, a virus control group, and a normal cell control group were prepared and cultured in an incubator at 37°C and 5% CO2. Two days after infection, 50 μL of cell supernatant was removed from each well and nucleic acid was extracted. The amount of virus was detected by quantitative RT-PCR, and EC 50 was calculated by fitting dose-response curves in GraphPad Prism 7. 3).Selective Index SI=CC 50 / EC 50

[0049] The inhibitory effect of WS-635 on SARS-CoV-2 novel coronavirus at the cellular level was examined using nucleic acid assay, and the EC 50 was calculated. Figure 1 shows that WS-635 has an inhibitory effect on the 2019-nCoV novel coronavirus at the cellular level, and its EC 50 is 14.28μM (Figure). CC 50 >100 μM, SI>7.

[0050] Example 2. Ex vivo assay of WS-635 for SARS-CoV-2 replication Experiments were carried out in the P3 laboratory at the Chinese Institute of Virology. Human Vero cells (ATCC) were infected with a virus concentration of 100 TCID 50 The cells were infected with SARS-CoV-2 (2019-nCOV BetaCoV / Beijing / AMMS01 / 2020) virus so that the RNA viral load was measured by incubating the cells with different compound concentrations.

[0051] One day before, Vero cells were seeded in a 96-well plate at a concentration of 10,000 / well. The 2019-nCov virus was diluted with 2% cell maintenance medium and added to the 96-well plate, and the viral load in each well was 100 TCID 50WS-635 dissolved in DMSO was prepared using 2% FBS-DMEM maintenance solution at concentrations of 400, 200, 100, 50, 25, and 12.5 μM. The cell culture supernatant was discarded, and T705 (100 μL / well) at different concentrations was added. Four wells were used for each drug, and 100 TCID 50 A virus solution was added to each droplet. A drug positive control (10 μM Remdesivir (RDV)) group, a virus control group, and a normal cell control group were prepared and cultured in an incubator at 37°C and 5% CO2. Two days after infection, 50 μL of cell supernatant was removed from each well and nucleic acid was extracted. The amount of virus was detected by quantitative RT-PCR, and EC 50 was calculated by fitting dose-response curves in GraphPad Prism 7.

[0052] The inhibition and cytotoxicity curves are shown in Figure 2 and Table 1. The figure shows that WS-635 inhibits SARS-CoV-2 (2019-nCOV BetaCoV / Beijing / AMMS01 / 2020) in a dose-dependent manner.

[0053] [Table 1]

[0054] Example 3. Evaluation of the antiviral activity of test compound WS-635 against human coronavirus (HCoV) 229E in a cytopathic effect (CPE) assay 3.1. Test Compounds and Control Compounds The test compound WS-635 was provided by the supplier as a dry powder and prepared as a 60 mM stock solution in 100% DMSO solution. The control compound Remdesivir was provided by WuXi AppTec. The 50% effective concentration (EC 50 ), and 50% cytotoxic concentration (CC 50 To identify the activity of DMSO, compounds were tested in duplicate at eight concentrations, 3-fold dilutions. The highest concentration tested is shown in Table 2. The final concentration of DMSO in the cell culture medium was 0.5%.

[0055] [Table 2]

[0056] 3.2. Cell lines and virus strains HCoV 229E (ATCC VR-740) and MRC-5 cells (ATCC CCL-171) were obtained from ATCC. MRC-5 cells were maintained in minimum essential medium (Sigma) supplemented with 10% FBS (Hyclone), 1% L-glutamine (Gibco), 1% NEAA (Gibco), and 1% penicillin-streptomycin (Hyclone). Minimum essential medium supplemented with 5% FBS, 1% L-glutamine, 1% NEAA, and 1% penicillin-streptomycin was used as the assay medium.

[0057] 3.3. Reagents and Equipment The assay used the CellTiter Glo (Promega) luminescent cell viability assay kit as the main reagent and the Synergy2 Microplate Reader (BioTek) as the main instrument.

[0058] 3.4. Method The antiviral assay results are summarized in Table 3.

[0059] [Table 3]

[0060] HCoV 229E CPE Assay MRC-5 cells were added to a 96-well plate at 20,000 cells / well and cultured overnight at 37°C and 5% CO. The next day, serially diluted compounds and virus-containing medium (200 TCID 501000mg / well) was added. The resulting culture medium was maintained at 35°C and 5% CO2 for an additional 3 days until the virus infection in the virus control (cells infected with virus without compound treatment) showed significant CPE. Cell viability was measured using CellTiter Glo according to the manufacturer's manual. Luminescence signals were measured using a Microplate Reader Synergy2 (Molecular Devices). The antiviral activity of each compound was calculated based on the inhibition of CPE at each concentration normalized by the virus control.

[0061] 3.4.2. Cytotoxicity Assay In parallel, the cytotoxicity of the compounds was evaluated under identical conditions but without viral infection. Cell viability was measured with CellTiter Glo according to the manufacturer's manual. 50 Values ​​were calculated based on cytotoxicity at the test concentrations normalized to the medium control (medium only).

[0062] Data Analysis The antiviral activity and cytotoxicity of the compounds are expressed as % inhibition and % cell viability, respectively, and are calculated by the following formulas: Inhibition (%) = (raw data CPD -average VC ) / (average CC -average VC ) x 100 Cell viability (%) = (raw data CPD -average MC ) / (average CC -average MC ) x 100 The raw data CPD indicates the number of compound-treated wells, and the mean VC ,average CC , and the average MC indicates the mean values ​​of the virus control, cell control (cells without virus infection or compound treatment), and medium control, respectively. EC 50 and CC 50Values ​​were calculated using the logarithmic equation for variable slope responses using GraphPad Prism software (Version 5), and SI (CC 50 / EC 50 ) was calculated.

[0063] 3.5. Results and Discussion The antiviral experiments were quality controlled with reference compounds. Remdesivir showed the expected antiviral activity and had an effect on cell viability, demonstrating the reliability of the experiments. The antiviral and cytotoxicity results of the test compounds are summarized in Table 3. WS-635 has an EC of 5.16 μM against HCoV 229E. 50 WS-635 showed inhibitory activity in CC 50 The SI values ​​were 46.79 μM and 9.07, respectively, showing obvious cytotoxicity in MRC-5 cells. The inhibition and cytotoxicity curves are shown in Figure 3 and Table 4, which show that WS-635 has significant cytotoxicity in MRC-5 cells infected with HCoV229E, indicating that it functions by blocking the spread of the virus.

[0064] [Table 4]

[0065] Throughout this specification, reference to "an embodiment," "an embodiment," "one embodiment," "another example," "example," "embodiment," or "some examples" means that a particular feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the disclosure. Thus, the appearances of phrases such as "in one embodiment," "in one embodiment," "in another example," "in an example," "in an embodiment," or "some examples" in various places throughout this specification are not necessarily all referring to the same embodiment or example of the disclosure. Furthermore, particular features, structures, materials, or characteristics may be combined in any suitable manner in one or more embodiments or examples.

[0066] While exemplary embodiments have been shown and described, those skilled in the art will recognize that these embodiments are not to be construed as limiting the present disclosure, and that changes, substitutions, and improvements may be made in the embodiments without departing from the spirit, principles, and scope of the present disclosure.

Claims

1. A pharmaceutical composition for use in the treatment or prevention of coronavirus infections, comprising: including compounds of formula I or a stereoisomer, tautomer, N-oxide, solvate, pharmaceutically acceptable salt, or hydrate thereof, 【Chemical 1】 (I) The pharmaceutical composition, wherein the coronavirus is the 2019-nCov virus.

2. 10. The pharmaceutical composition of claim 1, further comprising at least one other therapeutic agent or composition selected from the group comprising corticosteroids, anti-inflammatory signaling modulators, beta2-adrenergic receptor agonist bronchodilators, anticholinergic agents, mucolytic agents, hypertonic saline, cytokine storm suppressants, other drugs for the treatment of COVID-19 viral infection, or mixtures thereof.

3. 10. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition is in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, a granule, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, microparticles, or a pill.

4. The pharmaceutically acceptable salt may be phosphate, hydrochloride, hydrobromide, hydroiodide, sulfate, nitrate, ethanesulfonate, toluenesulfonate, benzenesulfonate, acetate, trifluoroacetate, tartrate, maleate, succinate, citrate, benzoate, salicylate, ascorbate, adipate, alginate, alginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogen phosphate, dinitrobenzoate, dodecyl sulfate, fumarate, galactarate, te), galacturonate, glucoheptonate, gluconate, glutamate, glycerophosphate, hemisuccinate, hemisulfate, heptanoate, hexanoate, hippurate, 2-hydroxyethanesulfonate, iodide, isethionate, isobutyrate, lactate, lactobionate, malate, malonate, mandelate, metaphosphate, methanesulfonate, methyl benzoate, monohydrogen phosphate, 2-naphthalenesulfonate, nicotinate, oxalate, oleate, pamoate, pectinate, persulfate, phenyl acetate, 3-phenylpropionate, phosphonate, and phthalate.

5. 10. The pharmaceutical composition of claim 1, further comprising one or more pharmaceutically acceptable carriers, diluents, or excipients.

6. 10. The pharmaceutical composition of claim 1, comprising the compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, or pharmaceutically acceptable salt thereof in a dose of less than 1000 mg, less than 800 mg, less than 600 mg, or less than 500 mg.

7. 1. Use of a compound of formula I or a stereoisomer, tautomer, N-oxide, solvate, pharmaceutically acceptable salt, or hydrate thereof in the manufacture of a medicament for use in the treatment or prevention of a coronavirus infection, comprising: 【Chemistry 2】 (I) The use, characterized in that the coronavirus family is the 2019-nCov virus.

8. The pharmaceutically acceptable salt may be selected from the group consisting of phosphate, hydrochloride, hydrobromide, hydroiodide, sulfate, nitrate, ethanesulfonate, toluenesulfonate, benzenesulfonate, acetate, trifluoroacetate, tartrate, maleate, succinate, citrate, benzoate, salicylate, ascorbate, adipate, alginate, alginate, aspartate, bisulfate, bisulfite, bromide, butyrate, camphorate, camphorsulfonate, caprylate, chloride, chlorobenzoate, cyclopentanepropionate, digluconate, dihydrogen phosphate, dinitrobenzoate, dodecyl sulfate, fumarate, galactarate, 8. The use according to claim 7, wherein the hydroxybenzoate is at least one selected from the group consisting of hydroxybenzoates, ...

9. 8. The use according to claim 7, wherein the medicament is in the form of a sol, a perfume, an aerosol, a topical inhalant, a powder, a granule, a tablet, an ointment, a paste, an emulsion, a suspension, a gas dispersion, a solid dispersion, microparticles, or a pill.

10. 8. The use of claim 7, wherein the medicament comprises the compound of Formula I or a stereoisomer, tautomer, N-oxide, solvate, or pharmaceutically acceptable salt thereof in a dose of less than 1000 mg, less than 800 mg, less than 600 mg, or less than 500 mg.

11. 8. The use according to claim 7, wherein the medicament further comprises one or more pharmaceutically acceptable carriers, diluents, or excipients.

Images