A kind of medicine for preventing or treating hepatitis B virus infection and its preparation method and application

A hepatitis B virus and drug technology, applied in the direction of drug combination, antiviral agent, pharmaceutical formula, etc., can solve the problem that cccDNA cannot be directly degraded, and achieve the effect of diversification of antiviral mechanism and resistance to drug resistance

Active Publication Date: 2022-06-21
佛山病原微生物研究院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] Existing nucleoside analogs can effectively inhibit HBV reverse transcription, but cannot directly degrade cccDNA

Method used

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  • A kind of medicine for preventing or treating hepatitis B virus infection and its preparation method and application
  • A kind of medicine for preventing or treating hepatitis B virus infection and its preparation method and application
  • A kind of medicine for preventing or treating hepatitis B virus infection and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] The anti-HBV activity of human SOX9 was evaluated by overexpression experiments.

[0067] The application method of a kind of SOX9 protein provided in the embodiment of the present invention is used for preventing or treating hepatitis B virus infection, and its steps are:

[0068] 1. Experimental materials:

[0069] 1.1 Cells, plasmids:

[0070] HepG2 cells (from ATCC) and pHBV1.3 plasmid. The total mRNA extracted from HepG2 cells was used as a template, and cDNA was first formed by reverse transcription, and then the cDNA was used as a template to amplify the complete SOX9 gene fragment with specific primers. The primer sequences are as follows:

[0071] SOX9CDS Forward: 5'-CGGGATCCATGAATCTCCTGGACCC-3';

[0072] SOX9CDS Reverse: 5'-GGAATTCTCAAGGTCGAGTGAGC-3'.

[0073] The SOX9 gene was cloned into the vector pcDNA3.1 (purchased from addgene) by specific restriction endonucleases (BamHI and EcoRI) and ligated to obtain an overexpression plasmid of pcDNA3.1-SOX9, wh...

Embodiment 2

[0093] The anti-HBV activity of human SOX9 was evaluated by interference experiments.

[0094] An application method of human SOX9 provided in the embodiment of the present invention in the preparation of a medicine for treating or preventing hepatitis B virus infection, the steps of which are:

[0095] 1. Experimental materials:

[0096] The interfering RNA package was purchased from Guangzhou Ribo Company.

[0097] 2. Experimental method

[0098] Same as Example 1.

[0099] 3. Experimental results:

[0100] like image 3 As shown, a schematic diagram of the interference effect of the interfering RNA provided in the embodiment of the present invention.

[0101] In the figure, A is a schematic diagram showing that si-SOX9 has a better interference effect. The interference rate is 50%.

[0102] Schematic representation of B and C demonstrating that interfering with SOX9 in cells HepG2 and Huh7 promotes secretion of e- and s-antigens of HBV. Both e-antigen and s-antigen ...

Embodiment 3

[0110] The application method of a kind of human SOX9 protein provided in the embodiment of the present invention is used for preventing or treating hepatitis B virus infection, and its steps are:

[0111] A. Evaluation of the anti-HBV activity of SOX9 protein:

[0112] B. Co-transfection of human SOX9 overexpression plasmid (constructed in our laboratory) or small interfering RNA (synthesized by Guangzhou Ribo Company) and plasmid pHBV1.3 (type D, see below for construction method) capable of expressing HBV into hepatocytes The expression levels of HBV secreted antigens HBeAg and HBsAg (Shanghai Kehua) in the cell culture supernatant and the level of nucleocapsid-related DNA, an intermediate of HBV replication in cells, were detected in HepG2 (from ATCC) or Huh7 (from ATCC).

[0113] The results of this experiment are as figure 2 and image 3 shown.

[0114] Both overexpression and interference experiments proved that SOX9 could down-regulate the expression levels of HBeA...

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Abstract

The invention belongs to the technical field of antiviral drugs, and discloses a drug for preventing or treating hepatitis B virus infection and a preparation method, wherein an overexpression plasmid or small interfering RNA of SOX9 is co-transformed with a plasmid pHBV1.3 capable of expressing HBV Transfect into the liver cell line HepG2 or Huh7; detect the expression levels of HBV secreted antigens HBeAg and HBsAg in the cell culture supernatant and the level of HBV replication intermediate nucleocapsid-related DNA in the cells. Human SOX9 provided by the present invention is a cytokine expressed constitutively in the human body, and has no toxic and side effects on the human body at physiological concentrations; human SOX9 directly acts on the virus envelope, or affects virus adsorption, or changes intracellular signaling pathways , so as to resist viruses, because of its diversified anti-virus mechanisms, it is not easy for viruses to develop drug resistance.

Description

technical field [0001] The invention belongs to the technical field of antiviral drugs, and in particular relates to a drug for preventing or treating hepatitis B virus infection, a preparation method and application thereof. Background technique [0002] At present, the existing technologies commonly used in the industry are as follows: [0003] Hepatitis B virus (HBV) infection is one of the main factors causing liver cancer, which seriously threatens human life and health. Worldwide, 364 million people are infected with chronic hepatitis B virus (HBV), which kills 800,000 people each year. my country is a high-incidence area of ​​hepatitis B virus infection, with about 120 million chronic hepatitis B patients, and about 300,000 people die of liver cancer caused by chronic hepatitis B every year. HBV belongs to the family Hepadnaviridae, and its genome is partially double-stranded circular DNA. The core particle of HBV is 28nm in diameter and is icosahedral. Infectious v...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K38/17A61P1/16A61P31/20C12N15/85C12N15/66
Inventor 吴建国杨桦邬开朗谭秋萍刘映乐刘为勇
Owner 佛山病原微生物研究院
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