Use of compounds and pharmaceutically acceptable salts thereof in the treatment of coronavirus

Abstract

Provided are uses of a compound represented by Formula I and pharmaceutically acceptable salts thereof in anti-coronavirus, especially in treating or preventing COVID-19. Also provided is a composition comprising the compound represented by Formula I.

Description

Technical Field

[0001] This invention relates to the field of prevention and / or treatment of coronavirus infection, and more particularly to the use of a compound and its pharmaceutically acceptable salt in the fight against coronaviruses. Background Technology

[0002] Coronaviruses are non-segmented, single-stranded, positive-sense RNA viruses belonging to the subfamily Orthocoronavirinae of the family Coronaviridae within the order Nidovirales. Based on serotype and genomic characteristics, the Orthocoronavirinae is divided into four genera: α, β, γ, and δ. They can infect many animal species, including humans, bats, dogs, pigs, mice, birds, cattle, whales, horses, goats, and monkeys. Six coronaviruses are known to infect humans, including α-coronaviruses 229E and NL63, β-coronaviruses OC43 and HKU1, Middle East Respiratory Syndrome-associated Coronavirus (MERSr-CoV), and Severe Acute Respiratory Syndrome-associated Coronavirus (SARSr-CoV).

[0003] The novel coronavirus (SARS-CoV-2) belongs to the β genus of coronaviruses. It is enveloped, and its particles are round or oval, often pleomorphic, with a diameter of 60–140 nm. Its genetic characteristics are significantly different from SARSr-CoV and MERSr-CoV. The development of effective antiviral small molecule drugs is urgently needed. Developing low-toxicity and highly effective antiviral small molecule drugs against SARS-CoV-2 coronavirus to meet the clinical needs of SARS-CoV-2 coronavirus-infected patients both domestically and internationally has significant social implications.

[0004] Therefore, there is an urgent need to develop inhibitors against the SARS-CoV-2 coronavirus for the treatment or prevention of coronavirus infection. Summary of the Invention

[0005] In order to solve one of the aforementioned technical problems in the prior art, the present invention has discovered a class of compounds that can effectively treat or prevent coronavirus infection.

[0006] According to a first aspect of this disclosure, the present invention provides the use of compounds of Formula I or pharmaceutically acceptable salts thereof in the preparation of medicaments for treating or preventing coronavirus infection.

[0007]

[0008] Z is selected from H, OH, COOH, CN, NH2 or halogen;

[0009] R1, R2, R3, R4, R5, R6, R7, R8, R9, R 10 and R11 Each is independently selected from H, OH, COOH, NH2, CN, halogen, optionally substituted straight or branched C1-C6 alkyl, optionally substituted straight or branched C2-C6 alkenyl, or optionally substituted straight or branched C2-C6 alkynyl; and

[0010] R7, R8, R9, R 10 and R 11 At least one of them is a halogen.

[0011] According to some embodiments, in the compounds represented by Formula I of this disclosure, Z is selected from H, OH, or COOH. According to some embodiments, Z is selected from OH.

[0012] According to some embodiments, the compound represented by Formula I of this disclosure, wherein R1, R2, R3, R4, R5, R6, R7, R8, R9, R 10 and R 11 Each is independently selected from H, OH, COOH, NH2, CN, halogen, optionally substituted straight or branched C1-C6 alkyl, optionally substituted straight or branched C2-C6 alkenyl, or optionally substituted straight or branched C2-C6 alkynyl. According to some embodiments, the compounds of Formula I disclosed herein, wherein R1, R2, R3, R4, R5, R6, R7, R8, R9, R... 10 and R 11 Each of the substituents is independently selected from H, OH, COOH, NH2, CN, halogen, optionally substituted straight-chain or branched C1-C3 alkyl, optionally substituted straight-chain or branched C2-C3 alkenyl, or optionally substituted straight-chain or branched C2-C3 alkynyl. According to some embodiments, the substituents may be selected from halogen, phenyl, C1-C3 alkyl, C2-C3 alkenyl, or C2-C3 alkynyl.

[0013] According to some embodiments, in the compounds represented by Formula I of this disclosure, R1, R2, R3, R4, R5 and R6 are each independently selected from H.

[0014] According to some embodiments, in the compound represented by Formula I of this disclosure, R7, R8, R9, R 10 and R 11 At least one of them is a halogen. According to some embodiments, the halogen is selected from F, Cl, Br or I atoms. According to some embodiments, the halogen is Cl.

[0015] According to specific embodiments, in the compound represented by Formula I of this disclosure, R7 is a halogen, and R8, R9, R 10 and R 11 All are H. According to specific embodiments, in the compound represented by Formula I of this disclosure, R8 is a halogen, and R7, R9, and R...10 and R 11 All are H. According to specific embodiments, in the compound represented by Formula I of this disclosure, R9 is a halogen, and R7, R8, and R... 10 and R 11 All are H. According to specific embodiments, in the compounds represented by Formula I of this disclosure, R... 10 It is a halogen, and R7, R8, R9 and R 11 All are H. According to specific embodiments, in the compounds represented by Formula I of this disclosure, R... 11 It is a halogen, and R7, R8, R9 and R 10 All are H. According to a specific implementation, the halogen is a Cl atom.

[0016] According to some embodiments, the compounds represented by Formula I of this disclosure can be in the form of their pharmaceutically acceptable salts. When the compounds represented by Formula I contain an acidic center, such as a carboxyl group, their pharmaceutically acceptable salts can be reacted with a suitable base to form the corresponding base addition salts. Such bases include, but are not limited to: alkali metal hydroxides, such as potassium hydroxide, sodium hydroxide, and lithium hydroxide; alkaline earth metal hydroxides, such as barium hydroxide and calcium hydroxide; alkali metal alkoxides, such as sodium ethoxide, potassium ethoxide, sodium propoxide, or potassium propoxide; alkali metal hydrides, such as sodium hydride or potassium hydride; and organic bases, such as piperidine, diethanolamine, N-methylglutamine, choline, diethanolamine, ethylenediamine, phenethylbenzylamine, diethylamine, piperazine, and tromethamine. When the compounds represented by Formula I contain a basic center, their pharmaceutically acceptable salts can be reacted with suitable organic and inorganic acids to form the corresponding acid addition salts. For example, pharmaceutically acceptable acid addition salts of compounds represented by Formula I may include, but are not limited to: sulfates, acetates, adipates, alginates, arginine salts, aspartate salts, benzoates, benzenesulfonates, hydrogen sulfates, bisulfites, bromides, butyrates, camphorates, camphorsulfonates, octanoates, chlorides, chlorobenzoates, citrates, cyclopentanepropionate, gluconate, dihydrogen phosphates, dinitrobenzoates, dodecyl sulfates, ethanesulfonates, etc. Horse acid salt, galacturonate, hemisuccinate, heptanate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, iodide, hydroxyethyl sulfonate, isobutyrate, lactate, sodium lactobionate, malate, maleate, malonate, mandelate, metaphosphate, methanesulfonate, methylbenzoate, hydrogen phosphate, 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, phenylacetate, phenylpropionate, phosphate, phosphonate, phthalate.

[0017] According to some embodiments, the pharmaceutically acceptable salt of the compound represented by Formula I of this disclosure can be at least one selected from phosphate, hydrochloride, sulfate, and maleate. According to specific embodiments, the compound represented by Formula I of this disclosure can be in the form of its phosphate salt.

[0018] According to some embodiments, the compound of Formula I can prevent or treat coronavirus infection. According to some embodiments, the compound of Formula I can prevent or treat infection caused by coronavirus.

[0019] According to some implementations, the coronavirus may include, but is not limited to, severe acute respiratory syndrome. crown styrovirus SARS-CoV, 2019 novel coronavirus (2019-nCoV or SARS-CoV-2) and its variants, Middle East Respiratory Syndrome Coronavirus (MERS-CoV), human coronavirus HCoV-OC43, human coronavirus HCoV-229E, human coronavirus HCoV-NL63, and human coronavirus HCoV-HKU1. According to some embodiments, the coronavirus may be one or more selected from SARS-CoV-2 coronavirus and its variants. According to some embodiments, the coronavirus may be one or more selected from SARS-CoV-2 coronavirus and its Alpha, Beta, Gamma, Delta, Lambda, and Omicron novel coronavirus variants.

[0020] According to some embodiments, the compound of Formula I can prevent or treat novel coronavirus infection.

[0021] According to some embodiments, the compound of Formula I can inhibit the transcriptional activity of cyclic AMP-response element-binding protein (CREB), for example by inhibiting the binding of CREB to CREB-binding protein (CBP).

[0022] According to some embodiments, the concentration of the compound of Formula I is about 0.04 μM to 10 μM.

[0023] According to some embodiments, the concentration of the compound of Formula I is about 0.12 μM to 5 μM.

[0024] According to specific embodiments, the compound represented by Formula I of this disclosure can be 2-hydroxy-3-naphthoic acid-2-chloroaniline, also known as naphthol AS-E, and its structural formula is shown below.

[0025]

[0026] According to specific embodiments, the compound represented by Formula I of this disclosure can be 2-hydroxy-3-naphthoic acid-2-chloroaniline phosphate, also known as naphthol phosphate AS-E, and its structural formula is shown below:

[0027]

[0028] According to some embodiments, the dosage form of the drug may be selected from at least one of tablets, capsules, injections, granules, suspensions, solutions, and inhalants.

[0029] According to another aspect, a composition is provided comprising a compound represented by Formula I.

[0030]

[0031] Among them, Z, R1, R2, R3, R4, R5, R6, R7, R8, R9, R 10 and R 11 As defined above.

[0032] The compound shown in Formula I of this disclosure can effectively inhibit infection by coronaviruses, such as SARS-CoV-2, thereby effectively treating or preventing coronavirus-induced infections. Attached Figure Description

[0033] Figure 1 The effects of the compound represented by Formula I according to this disclosure against the original SARS-CoV-2 strain are shown.

[0034] Figure 2 The effects of the compound represented by Formula I according to this disclosure on the original SARS-CoV-2 strain and the VOC strain Delta are shown.

[0035] Figure 3 This study demonstrates that SARS-CoV-2 coronavirus infection enhances the transcriptional activity of the naphthol AS-E target gene CREB. Detailed Implementation

[0036] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. The specific embodiments described herein are for illustrative purposes only and are not intended to limit the invention in any way. Furthermore, descriptions of well-known structures and techniques are omitted in the following description to avoid unnecessarily obscuring the concepts of this disclosure. Such structures and techniques have also been described in many publications.

[0037] definition

[0038] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly used in the field to which this invention pertains. For the purposes of interpreting this specification, the following definitions will apply, and where appropriate, terms used in the singular will also include the plural forms, and vice versa.

[0039] Unless the context clearly indicates otherwise, the terms “a” and “an” as used herein include plural references. For example, reference to “a cell” includes multiple such cells and equivalents known to those skilled in the art, etc.

[0040] As used herein, the term "about" indicates a range of ±20% of the following value. In some embodiments, the term "about" indicates a range of ±10% of the following value. In some embodiments, the term "about" indicates a range of ±5% of the following value.

[0041] As used herein, the term "alkyl" refers to a saturated hydrocarbon group, which is a hydrocarbon group formed by removing one hydrogen atom from an alkane molecule. "C1-C3 alkyl" or "C1-C6 alkyl" refers to alkyl groups containing 1 to 3 carbon atoms or 1 to 6 carbon atoms, respectively. Examples of C1-C6 alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, n-butyl, tert-butyl, neopentyl, n-hexyl, etc.

[0042] As used herein, the term "alkenyl" refers to a hydrocarbon group formed by removing one or more hydrogen atoms from an alkane molecule, wherein the alkane has at least one carbon-carbon double bond. "C2-C3 alkenyl" or "C2-C6 alkenyl" refer to alkenyl groups containing 2 to 3 carbon atoms, or 2 to 6 carbon atoms, respectively. Representative examples of alkenyl groups include, but are not limited to, vinyl, propenyl, butenyl, 1-methyl-2-buten-1-yl, heptenyl, octenyl, etc.

[0043] As used herein, the term "alkynyl" refers to a hydrocarbon group formed by removing one or more hydrogen atoms from an alkane molecule, wherein the alkane has at least one carbon-carbon triple bond. "C2-C3 alkynyl" or "C2-C6 alkynyl" refers to an alkenyl group containing 2 to 3 carbon atoms, or 2 to 6 carbon atoms, respectively. Representative examples of alkynyl groups include, but are not limited to, ethynyl, 1-propynyl, 1-butynyl, heptynyl, octyynyl, etc.

[0044] The following embodiments are provided to aid in understanding the present invention. However, it should be understood that these embodiments are for illustrative purposes only and do not constitute any limitation. The actual scope of protection of the present invention is set forth in the claims. It should be understood that any modifications and changes can be made without departing from the spirit of the present invention.

[0045] Example

[0046] Example 1. Effect of naphthol AS-E on the original SARS-CoV-2 strain

[0047] 1) Experimental Objective

[0048] To evaluate the effect of naphthol AS-E on inhibiting the original SARS-CoV-2 strain at the cellular level.

[0049] 2) Main experimental materials

[0050] Naphthol AS-E (Catalog No.: 92-78-4, Purity: ≥98.0%) was supplied by MCE (MedChemExpress).

[0051] Naphthol phosphate AS-E (Catalog No.: 18228-17-6, Purity: 98.08%), MCE Corporation.

[0052] Remdesivir (lot number: 1809249-37-3, purity: 99.78%), MCE Corporation.

[0053] SARS-CoV-2 nucleoprotein antibody (Catalog No.: 40143-V08B, mouse-derived), SinoBiological.

[0054] Goat anti-mouse IgG-HRP secondary antibody (Catalog No.: 115-035-146), Jackson Biotechnology.

[0055] RIPA protein lysis buffer (catalog number: P0013B), Beyotime.

[0056] Vero E6 cells (catalog number: CRL-1586), ATCC.

[0057] The original SARS-CoV-2 strain (2019-nCoV-WIV04, IVCAS 6.7512) was preserved by the Wuhan Institute of Virology, Chinese Academy of Sciences.

[0058] 3) Experimental methods

[0059] (1) The half-maximal effective concentration (EC50) of the drug was determined using a nucleic acid quantification method. 50 The previous day, Vero E6 cells were loaded at 5 × 10⁻⁶. 448-well plates were seeded. On the second day, the drugs (naphthol AS-E, naphthol phosphate AS-E, and remdesivir) were serially diluted 3-fold at the highest concentration of 10 μM, with 3 replicates per well, for a total of 7 dilutions. The cell culture supernatant was discarded, and different concentrations of the drug (150 μL / well) were added. The plates were pretreated at 37°C for 1 hour, and then 150 μL of virus suspension containing 0.01 multiple of infection (MOI) was added to each well. A positive drug control (remdesivir), a virus control, and a normal cell control group were established. The plates were incubated at 37°C in a 5% CO2 incubator. Cytopathic effects were observed daily. Two days after infection, 140 μL of cell supernatant was collected from each well for nucleic acid extraction, and viral load was detected by quantitative RT-PCR.

[0060] (2) The anti-SARS-CoV-2 efficacy of naphthol AS-E was evaluated by Western blotting. Vero E6 cells were injected with 2×10⁻⁶ cells the day before. 5 Cell culture was seeded in 12-well plates. The next day, naphthol AS-E was serially diluted 3-fold at the highest concentration of 10 μM, resulting in three dilutions. The cell culture supernatant was discarded, and different concentrations of the drug (850 μL / well) were added. The plates were pretreated at 37°C for 1 hour, and then 150 μL of virus suspension containing 0.05 MOI was added to each well. A positive control group (Remdesivir) and a virus control group were established. The plates were incubated at 37°C in a 5% CO2 incubator. Cytopathic effects were observed daily. Two days after infection, the supernatant was discarded, and the cells were washed once with ice-cold PBS. Then, 150 μL of cell lysis buffer was added to each well to lyse the cells for 30 minutes. Viral nucleoprotein was detected by Western blotting and SARS-CoV-2 nucleoprotein antibody (Nucleocapsid).

[0061] (3) The anti-SARS-CoV-2 efficacy of naphthol AS-E was evaluated using immunofluorescence. Vero E6 cells were cultured at 5 × 10⁻⁶ cells the day before. 5 Cells were seeded in 30 mm glass dishes per well. The next day, naphthol AS-E was serially diluted 3-fold at the highest concentration of 10 μM, resulting in three dilutions. The cell culture supernatant was discarded, and different concentrations of the drug (1700 μL / well) were added. The cells were pretreated at 37°C for 1 hour, and then 300 μL of virus suspension containing 0.05 MOI was added to each well. A positive control group (Remdesivir) and a virus control group were established. The cells were incubated at 37°C in a 5% CO2 incubator. Cytopathic effects were observed daily. Two days after infection, the supernatant was discarded, and the cells were washed once with ice-cold PBS, then fixed overnight with 3.7% (wt / vol) paraformaldehyde and subjected to immunofluorescence assays.

[0062] 4) Experimental Results

[0063] The efficacy of naphthol AS-E in inhibiting the original SARS-CoV-2 strain at the cellular level was determined using a nucleic acid quantification method, and the EC50 of the drug was calculated.50 The results showed that naphthol AS-E and naphthol phosphate AS-E had inhibitory effects on the original SARS-CoV-2 strain at the cellular level, and their EC50 values ​​were significantly lower than those of the original strain. 50 The concentrations were 1.04 μM and 2.25 μM, respectively. Figure 1 a). And from Figure 1 As shown in Figure a, the positive drug control (Remdesivir) can effectively inhibit the original SARS-CoV-2 strain at the cellular level, and its EC50... 50 It is 0.57 μM. Additionally, Figure 1 Immunoblotting and immunofluorescence experiments in samples b and 1c both showed that naphthol AS-E significantly inhibited SARS-CoV-2 infection at the cellular level. Clearly, naphthol AS-E effectively inhibited the original SARS-CoV-2 strain at the cellular level and exhibited an inhibitory effect comparable to that of remdesivir.

[0064] Example 2. Comparison of the original SARS-CoV-2 anti-naphthol AS-E strain and the VOC strain Delta

[0065] 1) Experimental Objective

[0066] To evaluate the difference in the inhibitory effects of naphthol AS-E on the original SARS-CoV-2 strain and the VOC strain Delta at the cellular level.

[0067] 2) Main experimental materials

[0068] Naphthol AS-E (batch number: 92-78-4, purity: ≥98.0%), MCE Corporation.

[0069] Calu-3 human lung adenocarcinoma cells (catalog number: HTB-55) from ATCC.

[0070] Cell viability assay kit (catalog number: G7572) Promega Corporation.

[0071] The original SARS-CoV-2 strain (2019-nCoV-WIV04, IVCAS 6.7512) was preserved by the Wuhan Institute of Virology, Chinese Academy of Sciences.

[0072] The SARS-CoV-2 VOC strain Delta (CSTR:16533.06.IVCAS 6.7593) was preserved by the Virus and Bacterial Culture Collection Center of Wuhan Institute of Virology, Chinese Academy of Sciences.

[0073] 3) Experimental methods

[0074] (1) The concentration at which the drug caused 50% cytotoxicity in the cells was determined using a luminescent cell viability assay kit (CC).50 Specifically, Calu-3 cells were administered at a dose of 1×10⁻⁶. 4 Cells were seeded into 96-well cell culture plates and incubated overnight at 37°C with 5% CO2. The next day, naphthol AS-E was serially diluted 3-fold at the highest concentration of 100 μM, with 8 replicates per well for a total of 7 dilutions. The culture medium was discarded, and 100 μL / well of 2% FBSDMEM maintenance medium containing different drug concentrations was added for continued incubation. Cell status was observed daily under a microscope. Cell viability was measured using a luminescent cell viability assay kit two days after drug addition. 50 μL of the luminescent solution was added to each well, and the cells were incubated at room temperature for 10 minutes. Fluorescence values ​​were then measured.

[0075] (2) The EC of the drug was determined by nucleic acid quantification method. 50 The day before, Calu-3 cells were loaded at 5 × 10⁻⁶. 4 48-well plates were seeded. The next day, naphthol AS-E was serially diluted 3-fold at the highest concentration of 10 μM, with 3 replicates per well for a total of 6 dilutions. Cell culture supernatant was discarded, and different concentrations of the drug (150 μL / well) were added. The plates were pretreated at 37°C for 1 hour, followed by the addition of 150 μL of virus suspension containing 0.01 MOI to each well. Positive drug control (Remdesivir), virus control, and normal cell control were established. The plates were incubated at 37°C in a 5% CO2 incubator. Cytopathic effects were observed daily. Two days after infection, 140 μL of cell supernatant was collected from each well for nucleic acid extraction, and viral load was detected by quantitative RT-PCR.

[0076] (3) Selectivity index (SI) = CC 50 / EC 50 .

[0077] 4) Experimental Results

[0078] First, the cytotoxicity of different concentrations of naphthol AS-E on Calu-3 cells was determined by luminescence immunoassay. The cytotoxicity (CC) of the drug was then calculated. 50 ()( Figure 2 a and 2b). The results showed that naphthol AS-E had an effect on CC in Calu-3 cells. 50 Greater than 100 μM. Then, using nucleic acid quantification, the effect of naphthol AS-E on inhibiting the original SARS-CoV-2 strain and VOC strain Delta at the cellular level was determined, and the EC50 of the drug was calculated. 50 The results showed that naphthol AS-E had an inhibitory effect on the original SARS-CoV-2 strain and the VOC strain Delta at the Calu-3 cell level, and its EC50... 50 The concentrations were 1.12 μM and 1.71 μM, respectively. Figure 2 a and 2b). Ultimately, according to CC50 and EC 50 The selectivity index (SI) of naphthol AS-E for the original SARS-CoV-2 strain and VOC strain at the cellular level was calculated to be 89.29 and 58.48, respectively. Furthermore, from... Figure 2 As shown in a and 2b, the inhibitory efficacy of naphthol AS-E against different SARS-CoV-2 strains at the cellular level was comparable. Naphthol AS-E effectively inhibited different SARS-CoV-2 strains at the cellular level.

[0079] Example 3. SARS-CoV-2 infection enhances the transcriptional activity of the naphthol AS-E target gene CREB.

[0080] 1) Experimental Objective

[0081] To evaluate the effect of SARS-CoV-2 on the cyclic AMP-response element-binding protein (CREB) of the naphthol AS-E target gene.

[0082] 2) Main experimental materials

[0083] Naphthol AS-E (Catalog No.: 92-78-4, Purity: ≥98.0%) was supplied by MCE.

[0084] Calu-3 human lung adenocarcinoma cells (catalog number: HTB-55), ATCC.

[0085] SARS-CoV-2 nucleoprotein antibody (Catalog No.: 40143-V08B, mouse-derived), SinoBiological.

[0086] Internal control antibody β-actin (mouse-derived, 66009-1-Ig), Proteintech.

[0087] Tumor suppressor Bcl-2 antibody (rabbit-derived, #2876), CST Biotechnology.

[0088] Phosphorylated p-Bcl-2 (Ser70) antibody (rabbit source, #2827), CST Biotechnology.

[0089] CREB-binding protein CBP antibody (rabbit, #7389), CST Biotechnology.

[0090] CREB antibody (rabbit, 12208-1-AP), Proteintech.

[0091] Phosphorylated p-CREB (Ser133) antibody (rabbit, 28792-1-AP), Proteintech.

[0092] Protein kinase A (PKA) antibody (mouse-derived, 67491-1-Ig), Proteintech.

[0093] Phosphorylated p-PKA (Thr197) antibody (rabbit source, #5661), CST Biotechnology.

[0094] The original SARS-CoV-2 strain (2019-nCoV-WIV04, IVCAS 6.7512) was preserved by the Wuhan Institute of Virology, Chinese Academy of Sciences.

[0095] 3) Experimental methods

[0096] (1) The effect of SARS-CoV-2 on the activation of the naphthol target gene CREB was evaluated by Western blotting. Calu-3 cells were inoculated at 2×10⁻⁶ cells per day prior to the study. 5 / wells were seeded into 12-well plates. The cell culture supernatant was discarded the next day, and 1000 μL of virus suspension containing 0.05 MOI was added to each well. A normal cell control group was also included. The plates were incubated at 37°C in a 5% CO2 incubator. Cytopathic effects were observed daily. Two days after infection, the supernatant was discarded, and the cells were washed once with ice-cold PBS. Then, 150 μL of cell lysis buffer was added to each well to lyse the cells for 30 minutes. The expression and activation of different target proteins were detected by Western blotting and corresponding antibody assays.

[0097] (2) The effect of SARS-CoV-2 on the interaction between the naphthol target genes CREB and CBP was assessed using a co-immunoprecipitation method. Calu-3 cells were injected with 1×10⁻⁶ cells the day before. 6 Inoculate 10 cm plates per well. On the second day, aspirate the cell culture supernatant, add 10 mL of virus suspension containing 0.05 MOI to each well, and set up a normal cell control group. Incubate at 37°C in a 5% CO2 incubator. Observe cytopathic effects daily. Two days after infection, discard the supernatant, wash the cells once with ice-cold PBS, and then add 1 mL of cell lysis buffer to each plate to lyse the cells for 30 minutes. Perform immunoprecipitation experiments using CREB or IgG antibodies and detect interacting CBP proteins using the corresponding antibodies.

[0098] 4) Experimental Results

[0099] First, Western blotting was used to detect the effect of SARS-CoV-2 on the activation of the naphthol target gene CREB. The results showed that phosphorylation of CREB, PKA, and Bcl-2 was significantly enhanced in human lung adenocarcinoma Calu-3 cells after infection with SARS-CoV-2. Figure 3a). Then, the interaction between CREB and its transcriptional cofactor CBP during SARS-CoV-2 infection was determined using immunoprecipitation. The results showed that SARS-CoV-2 infection significantly enhanced the interaction between CREB and CBP. Figure 3 (b) Clearly, SARS-CoV-2 infection enhances the transcriptional activity of the naphthol AS-E target gene CREB. Furthermore, given naphthol AS-E's high clinical safety and bioavailability, it can be safely and effectively used against SARS-CoV-2, and consequently, for the treatment of COVID-19 infection.

[0100] The technical solutions of the present invention are not limited to the specific embodiments described above. Any technical modifications made in accordance with the technical solutions of the present invention fall within the protection scope of the present invention.

Claims

1. Use of the following compounds or pharmaceutically acceptable salts thereof in the preparation of medicaments for the treatment or prevention of coronavirus infection. in, The coronavirus is SARS-CoV-2 or its Delta variant.

2. The use according to claim 1, characterized in that, The pharmaceutically acceptable salt of the compound is at least one of phosphate, hydrochloride, sulfate and maleate.

3. The use according to claim 2, characterized in that, The pharmaceutically acceptable salt of the compound is a phosphate.

4. The use according to claim 1, characterized in that, The drug is selected from at least one dosage form chosen from tablets, capsules, granules, suspensions, solutions, and inhalers.

5. The use according to claim 4, characterized in that, The solution is an injectable preparation.

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