New use of compound r-7050 for preventing or treating african swine fever

Abstract

The present application belongs to the technical field of African swine fever treatment, and particularly relates to a new use of compound R-7050 for preventing or treating African swine fever. The present application surprisingly finds that the compound R-7050 can reduce the replication level of the African swine fever virus, inhibit the expression of the structural protein of the African swine fever virus, has the effect of inhibiting the infection of the African swine fever virus, can be used for preparing a medicine or an adjuvant against the infection of the African swine fever virus, and is used for inhibiting the replication of the African swine fever virus. However, the present application is not limited to the African swine fever virus, and can also be used for inhibiting the replication of other viruses, and preparing a medicine or an adjuvant against the infection of the viruses.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biotechnology, and particularly relates to a new use of a compound R-7050 for preventing or treating African swine fever. BACKGROUND

[0002] African swine fever (ASF) is an acute and severe infectious disease caused by African swine fever virus (ASFV) infection, characterized by fever and hemorrhage of all organs in pigs, and the mortality rate of domestic pigs is as high as 100%. Since there is no effective vaccine and specific treatment drug so far, once the African swine fever epidemic occurs, it can only be controlled by culling, but this method not only causes economic losses, but also cannot meet the needs of large-scale pig farming in China. Therefore, how to effectively control the ASF epidemic is one of the great challenges facing the world pig industry at present, and is also a major strategic issue urgently needed to be solved in the prevention and control of ASF in China.

[0003] P30 and p72 are key structural proteins in ASFV, which are the main structural proteins of virus particles and important surface antigens, closely related to host cell tropism, pathogenicity and immunogenicity, and are involved in virus internalization and virus invasion of host cells.

[0004] R-7050 is a TNF-alpha receptor antagonist with cell permeability. The application unexpectedly finds that R-7050 can reduce the replication level of African swine fever virus and inhibit the expression of African swine fever virus structural proteins, has the effect of inhibiting African swine fever virus infection, and can be used as an inhibitor of African swine fever virus for preventing or treating African swine fever. SUMMARY

[0005] In view of the above technical problems, the application provides a new use of a compound R-7050 for preventing or treating African swine fever, which specifically comprises the following contents.

[0006] In a first aspect, the application provides a use of a compound R-7050 or a pharmaceutically acceptable salt thereof in the preparation of a medicine for preventing viral infection, and the structure of the compound R-7050 is shown in the following formula (I):

[0007]

[0008] Preferably, the virus is African swine fever virus.

[0009] Preferably, the compound R-7050 or the pharmaceutically acceptable salt thereof is added into a pharmaceutically acceptable carrier and / or excipient to form any one of the following dosage forms: tablets, sprays, granules, capsules, oral liquids, injections, suspensions.

[0010] Preferably, the dosage form comprises a powder injection, a capsule, a tablet, a suspension.

[0011] In a second aspect, the present application provides a use of a compound R-7050 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating viral infection, wherein the compound R-7050 has the following structural formula (I):

[0012]

[0013] Preferably, the virus is African swine fever virus.

[0014] Preferably, the compound R-7050 or a pharmaceutically acceptable salt thereof is added into a pharmaceutically acceptable carrier and / or adjuvant to form any one of a tablet, a spray, a granule, a capsule, an oral liquid, an injection, a suspension.

[0015] Preferably, the dosage form comprises a powder injection, a capsule, a tablet, a suspension.

[0016] The present application has the following beneficial effects: The present application surprisingly finds that the compound R-7050 can reduce the replication level of African swine fever virus, inhibit the expression of structural proteins of African swine fever virus, and has the effect of inhibiting African swine fever virus infection, and can be used for preparing a medicament or adjuvant for resisting African swine fever virus infection, and for inhibiting the replication of African swine fever virus. However, the present application is not limited to African swine fever virus, and can also be used for inhibiting the replication of other viruses, and for preparing a medicament or adjuvant for resisting viral infection. BRIEF DESCRIPTION OF DRAWINGS

[0017] Figure 1 R-7050 can inhibit the replication of ASFV; wherein A is the expression level of WB detection of ASFV structural proteins P72 and P30; B is the transcription level of RT-qPCR detection of ASFV B646L, CP204L genes; C is the virus titer HAD 50 Detection results

[0018] Figure 2 Effects of different concentrations of R-7050 on the activity of 3D4 / 21 cells. DETAILED DESCRIPTION

[0019] In order to make the technical means, creative features, purposes and effects achieved by the present application easy to understand, the present application is further described below in combination with specific embodiments. However, the protection scope of the present application is not limited to the following embodiments.

[0020] The experiments described in the following embodiments are obtained with biological safety permission and African swine fever laboratory activity permission:

[0021] According to the requirements of biosafety level 3 laboratory (BSL-3) and African swine fever related biosafety, the Lanzhou Veterinary Research Institute of Chinese Academy of Agricultural Sciences was reported by the Biosafety Committee of Lanzhou Veterinary Research Institute, the Experimental Animal Ethics Committee, the Biosafety Committee of Chinese Academy of Agricultural Sciences, the Experimental Animal Ethics Committee of Lanzhou Veterinary Research Institute, and the Biosafety Committee of Lanzhou Veterinary Research Institute, and obtained the permission of the Ministry of Agriculture and Rural Affairs for carrying out high pathogenic ASFV pathogen and animal research, and has been recorded in the Ministry of Agriculture and Rural Affairs, which meets the requirements of national biosafety level.

[0022] Example 1 Influence of R-7050 on ASFV

[0023] PAM cells were uniformly plated in cell culture plates at 1x10 6 / mL for 12 h of stable adhesion, then 1, 2, and 4 μM concentrations of R-7050 were added to the PAM cells for 12 h, with DMSO as a blank control, then ASFV was inoculated into the PAM cells at MOI = 1, and the cells and supernatant were collected at 24 hpi for WB and RT-qPCR detection, and HAD was also determined. 50 To show the situation of virus replication.

[0024] According to the primer sequences designed according to the ASFV CP204L gene sequence (GenBank: MK333184.1), ASFV B646L gene sequence (GenBank: MK333180.1), and pig GAPDH gene sequence (GenBank: NM001206359.1) in the NCBI database:

[0025]

[0026] RNA in the treated cell sample was extracted using the TRIzol method, and after measuring the RNA concentration, DNA Eraser was added to remove the possible DNA in the RNA sample. In a 10 μL system, 2 μL 5xgDNA Eraser Buffer, 1 μL gDNA Eraser, and 7 μL RNA were mixed uniformly and placed in a 42°C PCR instrument for 2 min. Then the DNA-removed RNA was added to a 20 μL reaction system, and cDNA was obtained by reverse transcription as a template for SYBR Green Real-time PCR. The reaction program was as follows: 95°C for 3 min, 95°C for 15 s, 60°C for 1 min, and 40-45 cycles of amplification under the above conditions. The obtained results were analyzed to obtain the transcription of virus genes CP204L and B646L under different treatment conditions.

[0027] The treated samples were collected, repeatedly freeze-thawed and lysed for HAD 50Assay: PAM was first cultured in 96-well plates, and the sample lysate was inoculated into 8 repeated samples according to the 10-fold serial dilution method. Before the cytopathic effect of ASFV in PAM, 1% fresh pig red blood cells were added to each well with a final concentration, and then the cells were incubated at 37°C, and hemadsorption (HAD) of red blood cells around the infected cells was observed within 7 days after infection (dpi). The method of Reed and Muench was used to calculate 50% HAD (HAD 50 ) to show the level of virus replication.

[0028] The results are shown in Figure 1 , where A is the expression level of ASFV structural proteins P72 and P30 detected by WB; B is the transcription level of ASFV B646L, CP204L genes detected by RT-qPCR; C is the virus titer HAD 50 detection results. The results show that, compared with the control group, compound R-7050 treatment can dose-dependently inhibit the transcription level of ASFV B646L, CP204L genes and the expression level of the encoded structural proteins P72 and P30, and reduce the virus titer, indicating that the compound R-7050 can significantly inhibit the replication of ASFV.

[0029] Example 2 Safety

[0030] The R-7050 compound with increasing concentration was added to the culture medium of 3D4 / 21 cells, incubated for 12 h, and then CCK-8 was added and incubated for 2-4 h, and then the cell survival rate and proliferation were calculated by reading the values at OD 450 wavelength.

[0031] The results are shown in Figure 2 , and the cell survival rate gradually decreased under the treatment of R-7050 with a concentration higher than 40 μM, and there was no significant difference in the cell survival rate under the treatment of R-7050 with a concentration less than 20 μM, so the treatment of R-7050 with a concentration less than 40 μM is a safe working concentration for 3D4 / 21 cells, and the safety is good.

[0032] In summary, the R-7050 described in the present application can dose-dependently inhibit the transcription of ASFV B646L, CP204L genes and the expression level of the encoded structural proteins P72 and P30, and significantly inhibit the replication of ASFV, and can be used as an inhibitor of African swine fever virus for the prevention or treatment of African swine fever.

[0033] The application is illustrated by the above examples to show the detailed features and methods, but the application is not limited to the above detailed features and methods, i.e. it does not mean that the application must rely on the above detailed features and methods to be implemented. It should be understood by those skilled in the art that any improvement of the application, equivalent replacement of the selected components, addition of auxiliary components, selection of specific modes, etc. fall within the protection scope and disclosure scope of the application.

Claims

1. The use of compound R-7050 or a pharmaceutically acceptable salt thereof in the preparation of a drug for the prevention of African swine fever virus infection, wherein the structural formula of compound R-7050 is shown in formula (Ⅰ): （Ⅰ）。 2. The application as described in claim 1, characterized in that, The compound R-7050 or a pharmaceutically acceptable salt thereof, when added to a pharmaceutically acceptable carrier and / or excipients, can be formulated into any dosage form, such as tablets, sprays, granules, capsules, oral liquids, injections, or suspensions.

3. The application as described in claim 2, characterized in that, The dosage forms include powder for injection, capsules, tablets, and suspensions.

4. The use of compound R-7050 or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating African swine fever virus infection, wherein the structural formula of compound R-7050 is shown in formula (Ⅰ): （Ⅰ）。 5. The application as described in claim 4, characterized in that, The compound R-7050 or a pharmaceutically acceptable salt thereof, when added to a pharmaceutically acceptable carrier and / or excipients, can be formulated into any dosage form, such as tablets, sprays, granules, capsules, oral liquids, injections, or suspensions.

6. The application as described in claim 5, characterized in that, The dosage forms include powder for injection, capsules, tablets, and suspensions.

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