Use of m-AMSA or its derivatives in the preparation of drugs for preventing and / or treating African swine fever virus infection

Abstract

The present invention relates to the use of m-AMSA or its derivatives or their pharmaceutically acceptable salts in the preparation of drugs for preventing and / or treating African swine fever virus infection, and in the preparation of drugs for inhibiting the activity of African swine fever virus topoisomerase. The present invention first discovered and confirmed through experiments that m-AMSA can completely inhibit the activity of pP1192R to relax supercoiled DNA, and can effectively reduce the viral titer of African swine fever virus in porcine alveolar macrophages, and its viral inhibition rate can be as high as 98.56% after 48 hours of administration at the highest dosage. The above results show that the small molecule compound m-AMSA or its derivatives or their pharmaceutically acceptable salts have great potential clinical application value in treating or preventing ASFV infection.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to use of m-AMSA or its derivatives or pharmaceutically acceptable salts thereof in preparing a medicament for preventing and / or treating African swine fever virus infection. Background Art

[0002] African swine fever (ASF) is an acute, virulent, highly contagious animal disease caused by the African swine fever virus (ASFV). It is mainly transmitted between domestic pigs and wild boars, with a mortality rate of up to 100%. Since it was first discovered in Kenya in 1921, ASFV has spread globally, causing serious economic losses to the pig industry in African, European and Asian countries. ASFV is the only member of the Asfarviridae family and the only known DNA arbovirus. The virus belongs to the nucleocytoplasmic large DNA virus (NCLDVs), with a genome length of 170 to 190 kb and encoding 150 to 167 open reading frames. The high complexity of ASFV has greatly hindered the development of safe and effective vaccines and antiviral drugs.

[0003] The replication cycle of ASFV mainly occurs in cytoplasmic virus factories, and the virus has evolved an independent replication and transcription mechanism involving multiple virus-specific enzymes. Among the proteases encoded by ASFV, a type II DNA topoisomerase, pP1192R, has attracted great attention from researchers in this field.

[0004] DNA topoisomerase (Topo) is a widely-existing enzyme that plays an important role in regulating the topological structure of DNA. It ensures that life activities such as DNA replication, transcription, and chromosome separation can proceed smoothly by catalyzing the breakage and connection of DNA chains. Topoisomerase can be divided into two categories, namely topoisomerase I (Topo I) and topoisomerase II (Topo II). Among them, topoisomerase I causes the breakage of single-stranded DNA, while topoisomerase II causes the breakage of double-stranded DNA. Due to its irreplaceable role in the life cycle, topoisomerase has become one of the targets successfully applied in clinical antibacterial and anticancer treatments.

[0005] pP1192R consists of 1192 amino acids and is highly conserved in all ASFV isolates. Studies have shown that pP1192R can promote the replication and transcription of ASFV by relaxing the supercoiled structure of DNA and unwinding the ring-linked DNA. Further studies have found that transfection of ASFV-infected cells with siRNA targeting p1192R can significantly reduce the production of ASFV progeny viruses. Therefore, this protein can be used as a target for antiviral drugs in the development of anti-ASFV drugs.

[0006] m-AMSA, also known as amsacrine, has a chemical name of N-[4-(9-acridinylamino)-3-methoxyphenyl]methanesulfonamide, and its structural formula is as follows:

[0007]

[0008] It has been reported that m-AMSA has significant antitumor activity in animal tumor systems. Summary of the invention

[0009] Purpose of the Invention

[0010] The object of the present invention is to provide the use of m-AMSA or its derivatives or their pharmaceutically acceptable salts in the preparation of drugs for preventing and / or treating African swine fever virus infection.

[0011] Solution

[0012] To achieve the above object, the present invention adopts the following technical solution:

[0013] In a first aspect, the present invention provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating African swine fever virus infection;

[0014]

[0015] In formula (I),

[0016] R1 is a group selected from the group consisting of: -H, -Cl, -Br, -NO2 and -CH3;

[0017] R2 is a group selected from the following: -CH3,

[0018] R3 is -CH3 or -CH2CH3.

[0019] In a preferred embodiment, in formula (I),

[0020] R1 is a group selected from the following: -H, -Cl, -Br;

[0021] R2 is -CH3;

[0022] R3 is -CH3.

[0023] In a further preferred embodiment, the compound is m-AMSA, and its structural formula is shown in the following formula (II):

[0024]

[0025] In a second aspect, the present invention provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof in the preparation of a medicament for inhibiting the activity of African swine fever virus topoisomerase; the compound of formula (I) is as defined in the first aspect above.

[0026] Preferably, the African swine fever virus topoisomerase is African swine fever virus pP1192R; further preferably, the African swine fever virus pP1192R is pP1192R of genotype II and / or genotype IV African swine fever virus strains, and further preferably, it is pP1192R of ASFV HLJ / 2018 strain or ASFV-R35 strain, or a homolog of pP1192R having an amino acid sequence identity of more than 80% thereto;

[0027] Preferably, the inhibition of the activity of African swine fever virus topoisomerase is the inhibition of the activity of relaxing supercoiled DNA of pP1192R.

[0028] In feasible embodiments of the above two aspects, the pharmaceutically acceptable salt of the compound is the hydrochloride of the compound; for example, the pharmaceutically acceptable salt of m-AMSA is the hydrochloride of m-AMSA.

[0029] In feasible embodiments of the above two aspects, the African swine fever virus is a genotype II and / or genotype IV African swine fever virus strain, preferably an ASFV HLJ / 2018 strain and / or an ASFV R35 strain, or an ASFV strain containing a homolog of pP1192R having an amino acid sequence identity of more than 80% with pP1192R of the ASFV HLJ / 2018 strain or the ASFV R35 strain.

[0030] Regarding the use of the first aspect above, it is feasible that the prevention and / or treatment of African swine fever virus infection includes one or more of the following:

[0031] (1) Prevent African swine fever virus infection;

[0032] (2) Prevent the spread of African swine fever virus;

[0033] (3) Inhibit the replication of African swine fever virus;

[0034] (4) Reduce, improve, alleviate or reverse the symptoms caused by African swine fever virus infection.

[0035] Regarding the use in the first aspect above, the drug is preferably a drug with the compound or a pharmaceutically acceptable salt thereof as the sole active ingredient, or a pharmaceutical composition with the compound or a pharmaceutically acceptable salt thereof as one of the active ingredients.

[0036] In a feasible embodiment, the medicament comprises a preventively and / or therapeutically effective amount of the compound or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier and / or excipient.

[0037] In a feasible embodiment, the drug is administered by one or more of the following methods: oral administration, injection, implantation, spraying and / or inhalation.

[0038] Preferably, the dosage form of the drug is one or more selected from the following: injection, oral solution, powder, tablet, granule, capsule, syrup, decoction, sustained-release preparation, enteric solvent, aerosol or suspension.

[0039] More preferably, in the dosage form of the drug, the content of m-AMSA or a pharmaceutically acceptable salt thereof per unit dosage form is 2 to 5 μg / μL, preferably 3 to 4 μg / μL.

[0040] In a third aspect, the present invention provides a method for preventing and / or treating African swine fever virus infection, the method comprising: administering a preventively and / or therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof to a subject in need; the compound of formula (I) is as defined in the first aspect above.

[0041] In a possible embodiment, the pharmaceutically acceptable salt of the compound is the hydrochloride of the compound; for example, the pharmaceutically acceptable salt of m-AMSA is the hydrochloride of m-AMSA.

[0042] In a feasible embodiment, the African swine fever virus is ASFV HLJ / 2018 strain and / or ASFV Ba71V strain.

[0043] Regarding the above method, the prevention and / or treatment of African swine fever virus infection includes one or more of the following:

[0044] (1) Prevent African swine fever virus infection;

[0045] (2) Prevent the spread of African swine fever virus;

[0046] (3) Inhibit the replication of African swine fever virus;

[0047] (4) Reduce, improve, alleviate or reverse the symptoms caused by African swine fever virus infection.

[0048] In a feasible embodiment, the compound (eg, m-AMSA) or a pharmaceutically acceptable salt thereof can be administered to a subject as the sole active ingredient, or can be administered to a subject as one of the active ingredients or in combination with other active ingredients.

[0049] The term "effective amount" refers to the amount or dosage of an active ingredient that provides the desired effect to the patient being diagnosed or treated by single or multiple administrations to the patient. The effective amount can be determined by the participating diagnostician as a person skilled in the art through known techniques and observations obtained under similar circumstances. In determining the effective amount or dosage of the active ingredient to be administered, the participating diagnostician should consider a variety of factors, including but not limited to: the species of the mammal; size, age and general health; the specific disease involved; the degree of involvement or severity of the disease; the response of the individual patient; the specific compound administered; the mode of administration; the bioavailability properties of the administered formulation; the selected dosage regimen; the use of concomitant drug therapy; and other relevant circumstances.

[0050] Beneficial Effects

[0051] The present invention discovers and provides new uses of m-AMSA or its derivatives or their pharmaceutically acceptable salts in preventing and / or treating African swine fever virus infection, as well as uses of m-AMSA or its derivatives or their pharmaceutically acceptable salts in preparing drugs for inhibiting the activity of African swine fever virus topoisomerase (particularly inhibiting the activity of relaxed supercoiled DNA of pP1192R).

[0052] The inventors have experimentally confirmed that m-AMSA or its derivatives or their pharmaceutically acceptable salts can target pP1192R and completely inhibit its activity of relaxing supercoiled DNA, and thus can be used as a topoisomerase inhibitor of African swine fever virus; further, the inventors have experimentally confirmed that m-AMSA can effectively inhibit the replication of African swine fever virus in porcine alveolar macrophages (PAM cells), and its IC value of inhibiting virus titer at 24 hours and 48 hours is 2.34%. 50 The results showed that the m-AMSA inhibited the replication of African swine fever virus by inhibiting the activity of pP1192R to relax supercoiled DNA.

[0053] The above results indicate that the small molecule compound m-AMSA or its derivatives or their pharmaceutically acceptable salts have great potential clinical application value in treating or preventing ASFV infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] One or more embodiments are exemplarily described by the pictures in the accompanying drawings, and these exemplary descriptions do not constitute limitations on the embodiments. The special word "exemplary" here means "used as an example, embodiment or illustrative". Any embodiment described as "exemplary" here is not necessarily interpreted as being superior or better than other embodiments.

[0055] Figure 1 The figure shows the molecular sieve and SDS-PAGE results of the purification of the ASFV-pP1192R protein expressed in Example 1.

[0056] Figure 2 The inhibitory effect (Figure A) and inhibitory mechanism (Figure B) of m-AMSA on the activity of ASFV-pP1192R to relax supercoiled DNA are shown, as described in Example 2.

[0057] Figure 3 The results of the toxicity experiment of m-AMSA on PAM cells are shown, as described in Example 3; wherein, Figure A is a bar graph of cell survival rates at various concentrations when treated with m-AMSA for 24 hours, Figure B is a bar graph of cell survival rates at various concentrations when treated with m-AMSA for 48 hours, and Figure C is a curve of drug concentration log value vs cell survival rate after m-AMSA treatment for 24 hours and 48 hours.

[0058] Figure 4 The inhibitory effect of m-AMSA on ASFV replication in PAM cells is shown, as described in Example 4; wherein Figure A shows the viral DNA copy number (Log 10 ) bar graph, and Figure B is the viral DNA copy number (Log 10 ), and FIG. C is a curve of the log value of drug concentration vs. relative viral DNA copy number (relative to the DMSO group) after m-AMSA treatment for 24 hours and 48 hours. DETAILED DESCRIPTION

[0059] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be clearly and completely described in conjunction with the embodiments below. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0060] In addition, in order to better illustrate the present invention, numerous specific details are provided in the specific embodiments below. It should be understood by those skilled in the art that the present invention can also be implemented without certain specific details. In some embodiments, raw materials, elements, methods, means, etc. well known to those skilled in the art are not described in detail, so as to highlight the main purpose of the present invention.

[0061] Unless explicitly stated otherwise, throughout the specification and claims, the term “comprise” or variations such as “include” or “comprising”, etc., will be understood to include the stated elements or components but not to exclude other elements or components.

[0062] The present invention is further described in detail below by way of examples.

[0063] Example 1: Expression and purification of ASFV-pP1192R protein

[0064] The full-length DNA sequence of the ASFV R35 strain pP1192R protein (Genbank: AXB50031.1) was connected to the pCAGGS vector through the restriction enzyme sites NdeI and XhoI. Among them, the Strep II tag was added to the 5' end of the ASFV-pP1192R protein coding region, and the coding sequence of the 6-histidine tag (His6-tag) and the translation stop codon were added to the 3' end.

[0065] The ASFV-pP1192R expression vector constructed as described above was transfected into Expi293F cells, and the transfected cells were placed in a CO2 constant temperature oscillator and cultured at 37°C and 140rpm / min. The cells were collected on the third day after transfection, resuspended in a protein buffer (10mM HEPES (pH7.4), 500mM NaCl, 5% (vol / vol) glycerol, 1mM PMSF), and ultrasonically disrupted to obtain soluble pP1192R protein. The obtained pP1192R protein was purified by nickel ion affinity chromatography (HisTrapTM HP, GE) and gel filtration chromatography (Superose 6Increase 10 / 300Gl, GE), replaced in a protein buffer of 10mM HEPES (pH 7.4), 150mM NaCl and 5% (vol / vol) glycerol, and then the protein purity was identified by SDS-PAGE.

[0066] The identification results are as follows Figure 1 As shown; Figure 1 The results showed that the above steps obtained a highly pure pP1192R protein with a size of about 220 kDa.

[0067] Example 2: Study on the inhibitory effect and inhibitory mechanism of m-AMSA on ASFV-pP1192R

[0068] Topo inhibitors are divided into Topo poisons and Topo catalytic inhibitors according to their mechanism of action. Topo catalytic inhibitors act on different stages of the Topo catalytic cycle by blocking a specific function of Topo or a step in the catalytic reaction. Topo poisons promote the formation of enzyme-DNA breakage complexes, causing the equilibrium reaction to tend toward the enzyme-DNA breakage complex, hindering Topo-mediated DNA reconnection reactions, leading to DNA chain breaks, affecting DNA replication, and ultimately playing a cell-killing role.

[0069] Topoisomerase has the function of relaxing DNA supercoils, so the supercoiled DNA (supercoiled plasmid pUC19) relaxation experiment was used to evaluate the inhibitory effect of m-AMSA on the catalytic activity of ASFV-pP1192R.

[0070] Specifically, the pUC19 DNA relaxation experiment (specific experimental procedures refer to Black MT, Stachyra T, Platel D, et al. Mechanism of action of the antibiotic NXL101, a novel nonfluoroquinolone inhibitor of bacterial type II topoisomerases [J]. Antimicrobial agents and chemotherapy, 2008, 52 (9): 3339-3349.) was used to evaluate the inhibitory effect of m-AMSA on the catalytic activity of ASFV-pP1192R. The principle of the experiment is as follows:

[0071] Under the catalysis of pP1192R, the topological structure of plasmid DNA will change, while its molecular weight remains unchanged. During electrophoresis, DNA molecules with different topological structures migrate at different speeds, and the order of speed is: supercoiled DNA > relaxed DNA. Based on this principle, the inhibitory activity of m-AMSA on pP1192R was studied. If m-AMSA has inhibitory activity on Topo, it can hinder the transformation of supercoiled DNA to relaxed DNA, and this change will be reflected during agarose gel electrophoresis.

[0072] The specific experimental process is:

[0073] 250 nM pP1192R, 250 ng DNA and different concentrations of m-AMSA were reacted at 37°C for 30 min, and then 1% SDS was added to terminate the reaction. Finally, the products were identified by agarose gel.

[0074] The experimental results are as follows Figure 2 As shown in Figure A; Figure 2 Figure A shows that under the action of 250nM pP1192R, 250ng of supercoiled DNA completely changes from supercoiled form to relaxed form; in contrast, the drug solvent DMSO has no effect on the relaxation of DNA mediated by pP1192R. When treated with the small molecule drug m-AMSA (purchased from Selleckchem), it can be found that as the concentration of m-AMSA increases, m-AMSA exhibits a significant inhibitory effect on the relaxation of DNA mediated by pP1192R, and this inhibitory effect is dose-dependent; when the drug concentration reaches 250μM and above, AMSA completely inhibits the activity of pP1192R in relaxing DNA, and pUC19DNA still completely maintains a supercoiled structure.

[0075] In addition, the pUC19 DNA cleavage experiment (reference Black MT, Stachyra T, Platel D, et al. Mechanism of action of the antibiotic NXL101, a novel nonfluoroquinolone inhibitor of bacterial type II topoisomerases [J]. Antimicrobial agents and chemotherapy, 2008, 52 (9): 3339-3349.) was used to explore the mechanism by which m-AMSA inhibits the activity of ASFV-pP1192R in relaxing supercoiled DNA. The principle of the experiment is as follows:

[0076] pP1192R is a type II topoisomerase that can cut double-stranded DNA. Toxins targeting pP1192R can promote the formation of a break complex. After adding proteinase K to hydrolyze pP1192R, the supercoiled DNA (pUC19DNA) has two cuts and becomes linear DNA and is released; after separation by agarose gel, the corresponding linear DNA bands can be displayed.

[0077] The specific experimental process is:

[0078] 250 nM pP1192R, 250 ng DNA and different concentrations of m-AMSA were reacted at 37°C for 30 min, 1% SDS was added to terminate the reaction, and then proteinase K was added to digest for 1 hour at 42°C. Finally, the digestion products were identified by agarose gel.

[0079] The results of the pP1192R-mediated DNA cleavage experiment are as follows Figure 2 As shown in Figure B, the results show that after adding m-AMSA at a concentration of 250 μM and above, an obvious linear plasmid band appeared, indicating that m-AMSA targeted pP1192R in the manner of a Topo poison.

[0080] Example 3: Toxicity experiment of m-AMSA on PAM cells

[0081] In this example, a CCK-8 kit (purchased from Japan Tongren Chemical Co., Ltd., catalog number: CK04) was used to determine whether m-AMSA was cytotoxic to porcine alveolar macrophages (PAM cells) through a cytotoxicity experiment, and its highest non-toxic dose was determined to provide a drug dosage reference for virus inhibition experiments at the cellular level.

[0082] The CCK-8 kit can be used to quickly and sensitively detect cell proliferation and cytotoxicity. Its working principle is as follows: the CCK-8 reagent contains water-soluble tetrazolium salt WST-8 [chemical name: 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfonic acid benzene)-2H-tetrazolium monosodium salt], which is reduced by dehydrogenase in cells under the action of electron carrier 1-methoxy-5-methylphenazinium dimethyl sulfate (1-Methoxy PMS) to a highly water-soluble yellow formazan product (Formazan dye). The depth of its color is proportional to cell proliferation and inversely proportional to cytotoxicity. For the same cells, the depth of the color is linearly related to the number of cells. Using an enzyme reader to measure the OD value at a wavelength of 450nm can indirectly reflect the number of living cells.

[0083] The specific experimental process is as follows:

[0084] First, 5 × 10 4 PAM cells were cultured at 37°C for 24 hours. Subsequently, the culture medium was discarded and 100 μL of gradient diluted m-AMSA was added to each well, with concentrations of 40 μM, 20 μM, 10 μM, 5 μM, 2.5 μM, 1.25 μM, and 0.625 μM, respectively. The diluent was 1640 culture medium containing 2% calf serum. Three replicate wells were made for each dilution, which was the experimental group. Since m-AMSA was dissolved in DMSO solution, a DMSO solution treatment group was set up as a control group. Next, after culturing at 37°C for 24 hours and 48 hours, the culture medium was discarded, the cells were washed twice with PBS solution, and then 100 μL of pre-prepared CCK-8 solution was added (i.e., 10 μL of CCK-8 reagent was added to every 100 μL of 1640 culture medium, wherein the CCK-8 reagent is provided in the CCK8 kit, which can be used directly or diluted with culture medium; here, in order to minimize the error caused by the residue of the reagent on the pipette, we diluted the CCK-8 reagent with culture medium). At the same time, the above CCK-8 solution was added to the cultured cells without any treatment as a blank group. The culture plate was incubated in the incubator for 2 hours, and then the absorbance at 450 nm was measured using an ELISA reader (PE, Multiscan Spectrum Enspire). The absorbance value can reflect the number of living cells, and then the cell survival rate was calculated by the following formula:

[0085] Cell survival rate % = (addition cell OD - blank OD) / (control cell OD - blank group OD) × 100%; where "addition cells" refers to the m-AMSA-treated cell group, "control cells" refers to the DMSO-treated cell group, and "blank group" refers to the cell group without any treatment (i.e., cells + culture medium + CCK-8 solution);

[0086] Finally, a bar graph was drawn with the m-AMSA treatment concentration as the horizontal axis and the cell survival rate as the vertical axis, and a drug concentration log value vs. cell survival rate curve was drawn. The experimental results are shown in Figure 3 ; Figure 3 Figure A is a bar graph of cell viability at various concentrations after m-AMSA treatment for 24 hours, Figure B is a bar graph of cell viability at various concentrations after m-AMSA treatment for 48 hours, and Figure C is a curve of drug concentration log value vs cell viability after m-AMSA treatment for 24 hours and 48 hours.

[0087] Figure 3 The results showed that high concentrations of m-AMSA showed a certain degree of cytotoxicity to PAM. When the concentration of m-AMSA was between 20 and 40 μM, the cell viability of PAM decreased to about 40% and 20% after 24 and 48 hours of administration, respectively. When the concentration of m-AMSA was 10 μM and below, administration for 24 and 48 hours did not affect or slightly affected the cell viability of PAM. At the same time, there was no significant difference in the cell morphology between the corresponding experimental group and the control group observed under a microscope. By calculation, the CC of m-AMSA at 24 hours and 48 hours 50 The results are as follows: 23.6 μM and 16.8 μM, respectively. Therefore, we selected 10 μM as the highest non-toxic dose to ensure that the antiviral effect of m-AMSA can be effectively evaluated while maintaining cell viability.

[0088] Example 4: Detection of the inhibitory ability of m-AMSA on ASFV infection at the cellular level

[0089] 5×10 5PAM cells were inoculated in a 24-well cell culture plate and cultured at 37°C for 24 hours. The ASFV HLJ / 2018 strain was diluted with 1640 medium to an MOI of 0.1, added to the PAM cell culture plate from which the medium was discarded, and infected at 37°C for 1 hour. After infection, the virus solution was discarded, and the PAM cells were washed twice with PBS to remove residual viruses. Gradient-diluted m-AMSA was added to the PAM cell culture plate, with concentrations of 10μM, 5μM, 2.5μM, 1.25μM, 0.625μM, and 0.3125μM, respectively. The diluent was 1640 medium containing 2% calf serum. Since m-AMSA was dissolved in DMSO solution, DMSO solution was also used as the control group in this experiment. The cells were cultured at 37°C and 5% CO2 for 24 and 48 hours, and the cell supernatant was collected. The viral genomic DNA in the supernatant was extracted using a kit, and then the viral copy number in the supernatant was absolutely quantified using fluorescent quantitative PCR. The target gene detected was ASFV-VP72, and the primer and probe sequences used were referenced in King, DP et al. (2003) Development of a PCR assay with internal amplification control for the detection of African swine fever virus. J. Virol. Methods, 107, 53-61, Section 2.4.1.

[0090] A bar graph was drawn with the m-AMSA treatment concentration as the horizontal axis and the viral DNA copy number as the vertical axis, and a curve of the drug concentration log value vs the relative viral DNA copy number was drawn. The experimental results are shown in Figure 4 ; Figure 4 Figure A shows the viral DNA copy number (Log 10 ) bar graph, and Figure B is the viral DNA copy number (Log 10 ), and FIG. C is a curve of the log value of drug concentration vs. relative viral DNA copy number (relative to the DMSO group) after m-AMSA treatment for 24 hours and 48 hours.

[0091] Figure 4 The results showed that with the gradual increase in the dosage, m-AMSA showed a significant inhibitory effect on the replication of ASFV in PAM cells. At 24 hours and 48 hours after drug treatment, the IC 50When the dosage was 10 μM, the highest non-toxic dose of PAM cells, the number of viral nucleic acid copies decreased by 88.41% and 98.56% at 24 hours and 48 hours after m-AMSA treatment, respectively.

[0092] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of a compound of formula (II) or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and / or treating African swine fever virus infection; 2. The use according to claim 1, characterized in that The pharmaceutically acceptable salt of the compound is the hydrochloride of the compound; And / or, the African swine fever virus is a genotype II and / or genotype IV African swine fever virus strain.

3. The use according to claim 2, characterized in that: The African swine fever virus is an ASFV HLJ / 2018 strain or an ASFV R35 strain, or an ASFV strain containing a homolog of pP1192R having an amino acid sequence identity of more than 80% with pP1192R of the ASFV HLJ / 2018 strain or the ASFV R35 strain.

4. The use according to claim 1, characterized in that The prevention and / or treatment of African swine fever virus infection may include one or more of the following: (1) Prevent African swine fever virus infection; (2) Prevent the spread of African swine fever virus; (3) Inhibit the replication of African swine fever virus; (4) Reduce, improve, alleviate or reverse the symptoms caused by African swine fever virus infection; And / or, the prevention and / or treatment of African swine fever virus infection is achieved by inhibiting the activity of African swine fever virus pP1192R in relaxing supercoiled DNA.

5. The use according to claim 1, characterized in that: The drug is a drug with the compound or a pharmaceutically acceptable salt thereof as the sole active ingredient, or a pharmaceutical composition with the compound or a pharmaceutically acceptable salt thereof as one of the active ingredients.

6. The use according to claim 1, characterized in that: The medicament comprises a preventively and / or therapeutically effective amount of the compound or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier and / or excipient.

7. The use according to claim 1, characterized in that The administration method of the drug is one or more selected from the following: oral administration, injection, implantation, spraying and / or inhalation.

8. The use according to claim 7, characterized in that The dosage form of the drug is one or more selected from the following: injection, oral solution, powder, tablet, granule, capsule, syrup, decoction, sustained-release preparation, enteric solvent, aerosol or suspension; And / or, in the dosage form of the drug, the content of the compound or a pharmaceutically acceptable salt thereof in each unit dosage form is 2 to 5 μg / μL.

9. The use according to claim 8, characterized in that In the dosage form of the drug, the content of the compound or its pharmaceutically acceptable salt per unit dosage form is 3-4 μg / μL.

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