Lentiviral vector for expressing human CD19 and CD20 CAR (chimeric antigen receptor) genes

A technology of chimeric antigen receptors and lentiviral vectors, which can be used in virus/bacteriophage, genetic engineering, plant genetic improvement, etc., and can solve the problem of low titer of lentivirus

Inactive Publication Date: 2017-05-31
刘晓明
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  • Abstract
  • Description
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Problems solved by technology

However, the titer of lentivirus obtained by the fourth-generation vector system is not high due to the application of four kinds of plasmids.

Method used

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  • Lentiviral vector for expressing human CD19 and CD20 CAR (chimeric antigen receptor) genes
  • Lentiviral vector for expressing human CD19 and CD20 CAR (chimeric antigen receptor) genes

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Embodiment Construction

[0044] The technical solution of the present invention will be further described below, but it is not limited thereto. Any modification or equivalent replacement of the technical solution of the present invention without departing from the spirit and scope of the technical solution of the present invention should be covered by the protection scope of the present invention middle.

[0045] Such as figure 1 As shown, the construction method of the lentiviral vector expressing human CD19 and CD20 chimeric antigen receptor genes comprises the following steps:

[0046] S1: CD19 antibody light chain variable region (V H ) and heavy chain variable region (V L ) Amplification or synthesis of cDNA

[0047] 1) Cloning CD19 antibody light chain variable region (V H ) and heavy chain variable region (V L ) cDNA, namely:

[0048] a. BALB / c mice were immunized with human CD19 protein;

[0049] b. Prepare mouse anti-human CD19 hybridoma cells;

[0050] c. Screening high-affinity mous...

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Abstract

The invention discloses a lentiviral vector for expressing human CD19 and CD20 CAR (chimeric antigen receptor) genes. The lentiviral vector for expressing the human CD19 and CD20 CAR genes is constructed by reverse transcriptase, structural protein and outer membrane protein through integration, wherein the integration required by a lentivirus is provided by shuttle expression vector pLVX and an auxiliary packaging plasmid, and the shuttle expression vector pLVX contains the human CD19 and CD20 CAR genes. The invention further provides a construction method of the lentiviral vector. The constructed lentiviral vector for expressing the human CD19 and CD20 CAR genes can express the CD19 and CD20 CAR genes simultaneously and efficiently, human T cells are transformed into effector cells with specific recognition and killing effects on B cell tumor, so that an important foundation is laid for research in antibody therapy, cancer therapy and stem cell therapy, and a new direction is provided for hematological malignancy cell therapy in China.

Description

[0001] Technical field: [0002] The invention relates to the technical field of antibody genetic engineering, in particular to a lentiviral vector expressing human CD19 and CD20 chimeric antigen receptor genes and a construction method thereof. [0003] Background technique: [0004] Lymphatic T cells are the main components of cellular immunity, and using genetic engineering to make T cells express chimeric antigen receptors can transform T cells into effector cells that can specifically kill tumor cells, which is a promising strategy for tumor immunotherapy . The key to the application of chimeric antigen receptors is to determine a tumor-associated antigen, which is overexpressed or highly expressed on the surface of tumor cells, but has no or low expression in normal tissues. CD19 is expressed in B lymphocytes and filter cells. The surface protein of alveolar dendritic cells is a member of the immunoglobulin (Ig) superfamily, located on the short arm of chromosome 16 (16p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/867C12N7/01
Inventor 朱分禄刘晓明
Owner 刘晓明
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