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HIV composite multi-epitope DNA vaccine and application thereof

A DNA vaccine and multi-epitope technology, applied in the field of AIDS vaccine, can solve problems such as ignoring humoral immunity

Inactive Publication Date: 2009-11-18
MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The second stage (1994-1999): characterized by emphasizing the cellular immune response to vaccines, but ignoring the role of humoral immunity

Method used

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  • HIV composite multi-epitope DNA vaccine and application thereof
  • HIV composite multi-epitope DNA vaccine and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Establishment of HIV Multi-epitope DNA Vaccine

[0036] On the basis of the HIV-1 MEGp24 epitope design completed earlier (Chinese patent application number: 03127002.6), the reverse vaccinology theory was further applied, and the authoritative American Los Alamos (Los Alamos) HIV molecular immunology database ( HIV Molecular Immunology Database), and referred to the HIV Epitope Database (HIV Epitope Database, http: / / lava.genetics.utah.edu / epitopeDBv3 / index.cfm) of Utah State University (The University of Utah), search HIV- 1 and HIV-2 CTL and CD8 T cell epitopes (http: / / hiv-web.lanl.gov / content / immunology / ctl_search). Based on the design concept of balancing the immune response and focusing on strengthening the induction of specific cellular immunity (CTL), the main structures and regulation of HIV-1 Env, Gag, Nef, Pol, Rt, Vpr, Tat and HIV-2 p24, p17 Protein-based 17 highly conserved CTL immunodominant epitopes. In terms of selection, the classic dominant ...

Embodiment 2

[0060] Example 2 Experimental Study on Mice Immunization of HIV Compound Multi-epitope DNA Vaccine

[0061] 1. Experimental grouping, mouse immunization and sampling

[0062] Twenty BALB / c female mice were randomly divided into 2 groups, 10 mice in each group, respectively pVAX1 empty plasmid control group and pVAX-MEGNp24 plasmid immunization group. Immunization was carried out three times with an interval of 14 days. Each time, 100 μg rDNA (dissolved in 100 μL sterile saline) was injected into the bilateral tibialis anterior muscle of the mice. Blood was collected 2 weeks after the second immunization, placed overnight at 4°C, centrifuged at 5000rpm for 10min, serum was collected, and stored at -20°C for testing. On the 10th day after the third immunization, the eyeballs were picked to take blood, and the cervical spine was dislocated to kill. The serum was coagulated and separated for ELISA to detect cytokines; at the same time, the spleen was aseptically collected to pre...

Embodiment 3

[0075] Example 3 Study on the immune effect of living cells of HIV compound multi-epitope DNA vaccine in vitro

[0076] 1. Sample collection:

[0077] The EDTA anticoagulated blood was sent to the laboratory for processing within 6 hours after collection, and the genetic background of the screened blood donors was HLA-0201.

[0078] 2. Human PBMC isolation:

[0079] (1) Transfer the anticoagulated blood to a 50 mL centrifuge tube and dilute it with 1640 medium (1:3).

[0080] (2) Add 15mL Ficoll Lymphocyte Layering Solution into the 50mL centrifuge tube.

[0081] (3) Slowly add the diluted blood to the surface of the layered liquid, taking care not to damage the liquid surface.

[0082] (4) Centrifuge at 700g for 30 minutes at 20°C.

[0083] (5) Aspirate the lymphocyte layer above the layered solution into a 50mL centrifuge tube, taking care not to absorb the erythrocyte layer.

[0084] (6) Add 40 mL of 1640 medium, and mix evenly by inverting. Centrifuge at 600g for 10 ...

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Abstract

The invention discloses an HIV composite multi-epitope DNA vaccine which takes HIV-1 capsid protein p24 as a vector backbone molecule and ER signal peptide as a homing sequence, and contains twenty-six highly conservative immunodominant epitopes in an HIV genome; wherein, the HIV composite multi-epitope DNA vaccine comprises twenty-nine epitopes which are three neutralizing antibody epitopes, twenty-three CTL epitopes, one HIV-1 isolate common antibody epitope, one MHC nonrestrictive T helper lymphocyte epitope and one spasmotoxin B cell epitope. The vaccine can be used for vaccination of healthy people and immunization therapy of people who are infected by AIDS virus, thus having double effects of prevention and treatment.

Description

technical field [0001] The present invention relates to an AIDS vaccine, in particular to a novel genetic engineering dual-purpose DNA vaccine for treatment and prevention based on HIV epitope, as well as its construction mode and application. Background technique [0002] The rapid spread of AIDS (Acquired Immunodeficiency Syndrome, AIDS) in the world has become one of the most serious viral diseases threatening human beings. According to the 2008 Global AIDS Epidemic Report issued by UNAIDS, 2.5 million people were newly infected with HIV in 2007. By 2008, there were 33.2 million HIV-infected people in the world, of which 22.5 million were infected. Distributed in numerous African countries south of the Sahara Desert; nearly 5 million infected people in Asia; about 1.5 million in Eastern Europe and Central Asia; about 1.7 million in Latin America; and about 2 million in North America, Western Europe, and Central and Eastern Europe. The vast majority of those living with H...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K39/00A61P31/18
Inventor 金宁一李昌李霄鲁会军田明尧金扩世申镇维金洪涛李臻付延军
Owner MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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