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Recombinant plasmid DNA vaccine composition for treating Hepatitis B

A technology of DNA vaccines and recombinant plasmids, which is applied in the field of biomedicine and can solve problems such as the unsatisfactory immune effect

Active Publication Date: 2011-11-09
BEIJING KAWIN TECH SHARE HLDG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] Although the preliminary clinical trial results of DNA vaccine are satisfactory, its immune effect has not yet reached the ideal level. Therefore, how to enhance the immune effect has become the key to DNA vaccine research, especially how to ensure that DNA vaccine can simultaneously obtain a stronger effect. The immune effect of HBsAg and HBcAg

Method used

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  • Recombinant plasmid DNA vaccine composition for treating Hepatitis B
  • Recombinant plasmid DNA vaccine composition for treating Hepatitis B
  • Recombinant plasmid DNA vaccine composition for treating Hepatitis B

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Example 1 Construction of recombinant plasmid pRec2.0-preS2S-C (pS2S-C)

[0077] (1) First construct the recombinant plasmid vector backbone pOE-EKS:

[0078] Using pDRVISV1.0 as a template, Primer1 and Primer2 as primers, among which primer 2 is a primer phosphorylated at the 5' end, amplified to obtain a replicon region (Ori) with a size of 748bp, and introducing EcoRI, KpnI and SwaI restriction sites at the same time point;

[0079] Primer1: 5'-GGAATTCGGGGTACCATTTAAATTTGAACGTTCGCAAtATGTGAGCAAAAGGCCAGC-3'

[0080] Primer2: 5'-CGGCGCGCGCCGAAAACGACGATTGCGAACGTTCAACCCGTAGAAAAAGATCAAAGG-3'

[0081] Using pDRVISV1.0 as a template, using Primer3 and Primer4 as primers, among which primer 3 is a primer phosphorylated at the 5' end, amplified to obtain a Kanna resistance marker gene (Kan) with a size of 1056bp, and introducing an EcoRI restriction site ;

[0082] Primer3: 5'-tcgtcgttttcggcgcgcgccgTTGAACGTTCGCAAtTCAAGTCAGCGTAATGCTC-3'

[0083] Primer4: 5'-GGAATTCGGCGCGCG...

Embodiment 2

[0103] Example 2 Construction of recombinant plasmid pRec2.0-C-preS2S (pC-S2S)

[0104] Using the pRec2.0-PreS2S constructed in Example 1 as a template, using Primer11 and Primer12 as primers, amplify the PreS2S gene with a size of 873bp, and pass HindIII+XbaI double digestion and HindIII+XbaI double digestion vector pcDNA3 .1 connect, construct and obtain pcDNA3.1-PreS2S;

[0105] Primer11: 5'-CCCAAGCTTGCCGCCACCATGCAGTGGAACTC-3'

[0106] Primer12: 5'-GCTCTAGAATCAGATGTAAACCCAC-3'

[0107] The PreS2S expression cassette was excised from the plasmid pcDNA3.1-PreS2S by BglII+PvuII double enzyme digestion, and cloned into pRec2.0-C which was digested by BglII+EcoRV double enzymes, and the recombinant plasmid pRec2.0-C-PreS2S (pC -S2S).

Embodiment 3

[0108] Example 3 Containing the core protein gene of hepatitis B and not containing the Core protein gene to the humoral and cellular immunity of healthy Balb / c mice

[0109] 6-8-week-old Balb / c female mice were purchased from the Animal Breeding Center of the Chinese Academy of Medical Sciences and kept in clean grade. 24 mice were divided into 4 groups, 3 immunized groups were given pRec2.0 empty vector group, recombinant plasmid DNA vaccine pRec2.0-S2S group, recombinant plasmid DNA vaccine pRec2.0-S2S-C (pS2S-C) , recombinant plasmid DNA vaccine pRec2.0-C-S2S (pC-S2S) for immunization, the dose is 50 μg / monkey, and the immunization procedure is to immunize once in the 0th, 2nd, and 4th weeks, and the rats are killed in the 6th week to get serum and separate lymphocytes.

[0110] Humoral immunity test: take mouse serum in strict accordance with the instructions of Beijing Yuande Biotechnology Co., Ltd. kit.

[0111] 1. Cellular immunity detection: mouse lymphocyte prepara...

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Abstract

The invention relates to a recombinant plasmid DNA vaccine composition for treating Hepatitis B. The composition contains a recombinant vaccine plasmid which simultaneously carries Hepatitis B middle protein S2S coding gene and Hepatitis B Core protein coding gene as well as a recombinant adjuvant plasmid, wherein the adjuvant plasmid is the adjuvant plasmid of recombinant interleukin 2 or recombinant human interferon gamma.

Description

technical field [0001] The invention relates to a recombinant plasmid DNA vaccine composition for treating hepatitis B disease, which belongs to the field of biomedicine. Background technique [0002] Hepatitis B seriously endangers human health, and there is no effective treatment at present. The main reason for the chronicity of hepatitis B is that the body lacks persistent specific cellular immunity and humoral immunity after hepatitis B virus (HBV) infection. [0003] HBV coat protein is composed of large protein LS (S1S2S antigen, encoded by pre-S1-pre-S2-S gene), medium protein MS (S2S antigen, encoded by pre-S2-S gene), small protein S (S antigen, encoded by S gene coding) composed of three molecules, among which the pre-S2 antigen has a small molecular weight and strong antigenicity, and involves the adsorption of hepatitis B virus to host cells. Therefore, the vaccine containing the pre-S2 antigen should be more effective in protecting the vaccinated. [0004] HBcA...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K39/39A61P1/16A61P31/20
Inventor 李鼎峰刘勇周德胜张春丽
Owner BEIJING KAWIN TECH SHARE HLDG
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