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Application of PreS1 in preparation of hepatitis B vaccine and treatment of chronic hepatitis B

A technology for chronic hepatitis B and hepatitis B virus, applied in the field of biomedicine, can solve the problem that HBp vaccine has no good therapeutic effect

Active Publication Date: 2017-02-22
PRESONE WUHAN BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Similarly, Kazuhiro K. et al. found that using HBV protein polymerase (polymerase, HBp) as a vaccine can induce an immune response in HBV transgenic mice, but due to the defects in the quantity and quality of CTL, the HBp vaccine has no good therapeutic effect (23 )

Method used

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  • Application of PreS1 in preparation of hepatitis B vaccine and treatment of chronic hepatitis B
  • Application of PreS1 in preparation of hepatitis B vaccine and treatment of chronic hepatitis B
  • Application of PreS1 in preparation of hepatitis B vaccine and treatment of chronic hepatitis B

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Expression and purification of the vaccine

[0037] Using AAV-HBV1.3 (purchased from Beijing Wujiahe Institute of Molecular Medicine Co., Ltd.) infected mouse liver cDNA (obtained by extracting tolerant mouse liver tissue total RNA and reverse transcription) as a template, design specific primers, The HBV PRES1 (ay subtype) DNA fragment was amplified by PCR reaction, and its nucleotide sequence is:

[0038] ATGGGGCAGAATCTTTCCACCAGCAATCCTCTGGGATTCTTTCCCGACCACCAGTTGGATCCAGCCTTCAGAGCAAACACAGCAAATCCAGATTGGGACTTCAATCCCAACAAGGACACCTGGCCAGACGCCAACAAGGTAGGAGCTGGAGCATTCGGGCTGGGTTTCACCCCACCGCACGGAGGCCTTTTGGGGTGGAGCCCTCAGGCTCAGGGCATACTACAAACTTTGCCAGCAAATCCGCCTCCTGCCTCCACCAATCGCCAGACAGGAAGGCAGCCTACCCCGCTGTCTCCACCTTTGAGAAACACTCATCCTCAGGC C (SEQ ID NO: 1)

[0039] Its amino acid sequence is:

[0040] MGQNLSTSNPLGFFPDHQLDPAFRANTANPDWDFNPNKDTWPDANKVGAGAFGLGFTPPHGGLLGWSPQAQGILQTLPANPPPASTNRQTGRQPTPLSPPLRNTHPQA (SEQ ID NO: 2)

[0041] In addition, the DNA sequence of HBV PRES1 (ad subt...

Embodiment 2

[0050] Example 2: The preS1 antigen was used as a vaccine to test its effect of inducing an immune response in the body.

[0051] Use 10μg of the preS1 protein obtained in Example 1 (alone or in combination with the following adjuvants: aluminum adjuvant (purchased from InvivoGen, 1:1-1:9 mix), 10μg TLR-4 ligand MPLA adjuvant (purchased From InvivoGen), 30μgTLR-9 ligand CpG adjuvant (synthesized by Invitrogen) or Freund’s adjuvant (IFA) (purchased from Sigma) subcutaneously immunized C57BL / 6 mice (6-8 weeks, male, purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.). The immunization process is as follows figure 2 Shown in A. After immunization, the anti-preS1 antibody response in the serum was detected by ELISA ( figure 2 B); Detection of preS1-specific T cell immune response in the spleen by ELISPOT ( figure 2 C). The experimental results show that preS1 protein combined with adjuvant can induce strong antibody response and specific T cell immune res...

Embodiment 3

[0052] Example 3: Through HBV in vivo infection experiment, the ability of preS1 antigen vaccine to prevent HBV infection was verified.

[0053] By the immunization method in Example 2, 10 μg of the preS1 protein purified in Example 1 was combined with Freund’s adjuvant to subcutaneously immunize C57BL / 6 mice (6-8 weeks, male, purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd. the company). The immunization process is like image 3 As shown in A, two subcutaneous immunizations were performed at an interval of 14 days. High titer of preS1 specific antibody anti-preS1 ( image 3 B). Then infect 1x10 through the tail vein 10 vg (viral genome) AAV-HBV1.3 (purchased from Beijing Wujiahe Institute of Molecular Medicine Co., Ltd.), blood is collected every week after infection to detect HBV-related antigens in serum, including HBsAg antigen, preS1 antigen and HBV-DNA changes .

[0054] The results show that compared with the group without protein immunization, ...

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Abstract

The invention belongs to the technical field of biomedicine, relates to a hepatitis B virus (HBV) vaccine and a preparation method thereof and further relates to application of the vaccine in prevention of hepatitis B virus infection and treatment of chronic hepatitis B (CHB) infection and application of the vaccine in cooperation with an HBsAg vaccine in treatment of CHB. The vaccine comprises a hepatitis B virus envelope protein preS1 region, can effectively prevent HBV from infecting a host, has a treatment effect on chronic hepatitis virus infection, and achieves a more remarkable effect than that of a traditional HBV vaccine in the prior art.

Description

Technical field [0001] The present invention belongs to the field of biomedical technology, and relates to a hepatitis B virus (hepatitis B virus, HBV) vaccine and a preparation method thereof. The present invention also relates to the vaccine to prevent hepatitis B virus infection and treat chronic hepatitis B virus (chronic hepatitis B). , CHB) infection, and the vaccine combined with HBsAg vaccine to treat chronic hepatitis B. Background technique [0002] Hepatitis B virus (HBV) is a double-stranded DNA virus, which is a hepatotropic virus that can spread through contact with blood and body fluids. After infecting the liver, HBV can cause many diseases, including acute and chronic hepatitis, liver fibrosis, cirrhosis and liver cancer. There are approximately 240 million chronic hepatitis B patients worldwide, and approximately 1 million people die each year from cirrhosis and liver cancer caused by HBV infection (1). However, there is still no effective treatment strategy f...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/29A61K39/39A61K39/295A61P31/20A61P1/16
CPCA61K39/12A61K39/39A61K2039/55505A61K2039/55561A61K2039/55566A61K2039/55588A61K2039/70C12N2730/10134
Inventor 傅阳心边英杰彭华孙志辰刘龙超
Owner PRESONE WUHAN BIOTECHNOLOGY CO LTD
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