Anti-human immunodeficiency virus (HIV) gene engineering recombinant virus and preparation method thereof, and anti-HIV gene engineering medicament

A recombinant virus and genetic engineering technology, applied in antiviral agents, biochemical equipment and methods, viruses/bacteriophages, etc., can solve the problems of limited specific binding and ineffective prevention and treatment of HIV infection, and achieve prevention and treatment Effect of HIV virus infection, simple preparation method and process, and enhanced specific binding effect

Inactive Publication Date: 2012-09-26
SHENZHEN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The technical problem to be solved by the present invention is to provide an anti-HIV antibody that can specifically bind to the HIV virus in view of the defects in the prior art that the specific binding effect between the anti-HIV antibody and the HIV virus is limited, and the effect on the prevention and treatment of HIV virus infection is not obvious. Anti-HIV genetically engineered recombinant virus for effective prevention and treatment of HIV infection, preparation method thereof, and anti-HIV genetically engineered drug

Method used

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  • Anti-human immunodeficiency virus (HIV) gene engineering recombinant virus and preparation method thereof, and anti-HIV gene engineering medicament
  • Anti-human immunodeficiency virus (HIV) gene engineering recombinant virus and preparation method thereof, and anti-HIV gene engineering medicament
  • Anti-human immunodeficiency virus (HIV) gene engineering recombinant virus and preparation method thereof, and anti-HIV gene engineering medicament

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preparation example Construction

[0023] The preparation method of the anti-HIV genetically engineered recombinant virus of the present invention comprises the following steps: A: obtain the heavy chain constant region gene of human antibody, the light chain constant region gene of human antibody, CD4 gene and CCR5 gene respectively; B: adopt the recombinant virus vector Construct the shuttle plasmid of the recombinant virus vector including the heavy chain constant region gene of human antibody, the light chain constant region gene of human antibody, CD4 gene and CCR5 gene; Co-transformation produces a recombinant virus plasmid; D: The recombinant virus plasmid formed in step C is transfected into a cell line for cultivation to obtain a recombinant virus; E: The supernatant is collected by centrifugation or the cells are purified to obtain a recombinant virus. The recombinant virus having both CD4 and CCR5 can be obtained through cell line expression, and the method is simple and convenient to operate, and is ...

Embodiment 1

[0059] PCR amplification and splicing: Extract total RNA from human fresh blood samples, reverse transcribe human cDNA, and use human cDNA as a template to amplify the heavy chain constant region of IgG1 antibody with the above primers 1 and 2, 3 and 4, 5 and 6, and 7 and 8, respectively. gene, human antibody light chain constant region gene L, CD4 gene and CCR5 gene. Then splice into CD4-IgG1, CCR5-IgG1 and CCR5-L genes by PCR method, CD4-IgG1 and CCR5-IgG1 contain flexible linker sequence 11, and at the same time, use IRES gene between CD4-IgG1 and CCR5-L by PCR method Ligated into CD4-IgG1-IRES-CCR5-L. shuttle Plasmid construction: The PCR products of CD4-IgG1, CCR5-IgG1, CCR5-L and CD4-IgG1-IRES-CCR5-L genes were digested with Sal I and Bgl II and loaded into the same digested viral vector pShuttle to construct a shuttle Plasmids pShuttle-CD4-IgG1, pShuttle-CCR5-IgG1, pShuttle-CCR5-L and pShuttle-CD4-IgG1-IRES-CCR5-L. The constructed shuttle plasmids were linearized ...

Embodiment 2

[0061] PCR amplification and splicing: Extract total RNA from human fresh blood samples, reverse transcribe human cDNA, and use human cDNA as a template to amplify the heavy chain constant region of IgG1 antibody with the above primers 1 and 2, 3 and 4, 5 and 6, and 7 and 8, respectively. gene, human antibody light chain constant region gene L, CD4 gene and CCR5 gene. Then splice into CD4-IgG1, CCR5-IgG1 and CCR5-L genes by PCR method, CD4-IgG1 and CCR5-IgG1 contain flexible linker sequence 11, and at the same time, use IRES gene between CD4-IgG1 and CCR5-L by PCR method Ligated into CD4-IgG1-IRES-CCR5-L. shuttle Plasmid construction: The PCR products of CD4-IgG1, CCR5-IgG1, CCR5-L and CD4-IgG1-IRES-CCR5-L genes were digested with Sal I and Bgl II and loaded into the same digested viral vector pdc316e to construct a shuttle Plasmids pdc316e-CD4-IgG1, pdc316e-CCR5-IgG1, pdc316e-CCR5-L and pdc316e-CD4-IgG1-IRES-CCR5-L. The constructed shuttle plasmids were linearized with ...

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Abstract

The invention relates to an anti-HIV gene engineering recombinant virus and a preparation method thereof, and an anti-HIV gene engineering medicament. The anti-HIV gene engineering recombinant virus comprises a constant region of a human antibody, and a cluster of differentiation 4 (CD4) and a chemokine receptor 5 (CCR5) which are connected with the constant region. The preparation method for the anti-HIV gene engineering recombinant virus comprises the steps as follows: obtaining a heavy chain constant region gene of the human antibody, a light chain constant region gene of the human antibody, a CD4 gene and a CCR5 gene; constructing a virus vector shuttle plasmid comprising the four genes; co-transforming the shuttle plasmid and a virus auxiliary plasmid to generate a recombinant virus plasmid; and transferring the recombinant virus plasmid to a cell strain for culturing and purifying the virus. The recombinant virus has both the CD4 and CCR5 capable of specifically binding with the CD4 site and CCR5 site of an HIV virus, an obviously enforced specific binding effect, and the function of doubly preventing the HIV virus from infecting a host cell. The preparation method is simple and practical. The anti-HIV gene engineering medicament with the recombinant virus can effectively act on the HIV virus, and can further effectively prevent and treat the infection of the HIV virus.

Description

technical field [0001] The invention belongs to the technical field of biopharmaceuticals, and relates to an anti-HIV genetically engineered virus, more specifically, to an anti-HIV genetically engineered recombinant virus, a preparation method thereof, and an anti-HIV genetically engineered drug. Background technique [0002] In the process of HIV virus infection, the HIV virus envelope first fuses with the cell membrane of the target cell. The fusion process is mainly mediated by the envelope glycoprotein gp120 and the transmembrane subunit gp41. gp120 binds to CD4 molecules and co-receptors (chemokine receptors CCR5 or CXCR4, etc.) on target cells successively, resulting in a change in the configuration of gp41, forming a core structure of 6-strand α2 helical bundles, and connecting the virus envelope with the target cells The cell membranes of the cells are drawn closer and fused, completing the infection process of the HIV virus entering the host cell. [0003] It can...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/85A61K35/76A61P31/18
Inventor 周兆平郑永唐易俊波雷明军苏庆宁
Owner SHENZHEN UNIV
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