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Double-target chimeric antigen receptor targeting CD19 and CD20 as well as expression vector and application thereof

A technology of chimeric antigen receptors and expression vectors, which is applied in the field of tumor immunology drugs, can solve the problems of tumor cell cancer recurrence, loss of CD19 expression, antigen escape, etc., achieve strong anti-tumor activity, avoid immune escape, and reduce the risk of recurrence Effect

Active Publication Date: 2020-04-10
华夏源细胞工程集团股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, although CD19 CAR-T cells have achieved great success, as many as two-thirds of patients who receive CD19 CAR-T cell therapy will experience loss of CD19 expression, leading to antigen escape and tumor cell persistence. Growth that manifests as cancer recurrence

Method used

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  • Double-target chimeric antigen receptor targeting CD19 and CD20 as well as expression vector and application thereof
  • Double-target chimeric antigen receptor targeting CD19 and CD20 as well as expression vector and application thereof
  • Double-target chimeric antigen receptor targeting CD19 and CD20 as well as expression vector and application thereof

Examples

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Embodiment 1

[0024] An embodiment of the present invention provides a dual-target chimeric antigen receptor targeting CD19 and CD20, such as figure 1 As shown, the dual-target chimeric antigen receptor includes: sequentially connected signal peptide (human CD8α signal peptide, CD8α leader), single-chain antibody targeting CD19 (CD19 scFv), single-chain antibody targeting CD20 (CD20 scFv), extended CD8α hinge region (human CD8α hinge region, CD8α Hinge), transmembrane region (human CD8α transmembrane region, CD8α transmembrane), co-stimulatory factors and intracellular signal peptide (human CD3ζ intracellular signal peptide, CD3ζ signal ), the nucleotide sequence of the signal peptide is shown in SEQ ID NO: 1 in the sequence listing, the nucleotide sequence of the single-chain antibody targeting CD19 is shown in SEQ ID NO: 2 in the sequence listing, and the single-chain antibody targeting CD20 The nucleotide sequence of the chain antibody is shown in SEQ ID NO: 3 in the sequence listing, th...

Embodiment 2

[0032] The present invention provides an expression vector, which includes the vector and the dual-target chimeric antigen receptor provided in Example 1. Specifically, the vector may be a lentiviral vector, a retroviral vector, an electrotransduction vector or a Sleeping Beauty transposon vector. In this embodiment, the vector is a lentiviral vector.

[0033] The following briefly introduces the preparation method of the expression vector, as follows:

[0034]Amplify and obtain the CART02 gene sequence by PCR (Polymerase Chain Reaction, polymerase chain reaction), add restriction site Xba I and restriction site EcoR I respectively at the two ends of CART02 gene sequence, obtain the thing to be digested, Carry out the double enzyme digestion reaction of Xba I and EcoR I with the lentiviral vector plasmid pCDH-EF1-MCS-T2A-copGFP, respectively, to obtain the enzyme-digested fragment containing CART02 and the enzyme-digested fragment containing pCDH-EF1-MCS-T2A-copGFP fragment....

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Abstract

The invention discloses a CD19 and CD20 targeting double-target chimeric antigen receptor and an expression vector and an application thereof, which belong to the field of tumor immune drugs. The double-target chimeric antigen receptor comprises a signal peptide, a single-chain antibody targeting CD19, a single-chain antibody targeting CD20, a CD8 alpha hinge region, a transmembrane region, a costimulatory factor and an intracellular signal peptide which are connected in sequence. The double-target chimeric antigen receptor can be used for specifically identifying tumor cells expressed by twosingle targets, namely a CD19 antigen and a CD20 antigen, can be used for identifying tumor cells co-expressed by CD19 antigen and CD20 antigen, the double-target CAR-T has stronger anti-tumor activity, and can prevent positive tumor cells expressed by low-abundance antigen from generating immune escape, so that the recurrence risk is reduced.

Description

technical field [0001] The invention relates to the field of tumor immunity drugs, in particular to a dual-target chimeric antigen receptor targeting CD19 and CD20 and its expression vector and application. Background technique [0002] Malignant tumors are one of the diseases that seriously threaten human health. According to incomplete statistics, the annual incidence of malignant tumors in China has reached more than 4 million cases. In the past, most of them were middle-aged and elderly people, and gradually spread to young patients. This may be related to factors such as modern people's high life pressure, increased environmental pollution, and unhealthy living habits. [0003] Chimeric antigen receptor (CAR) technology is a cell therapy technology that has developed very rapidly in recent years. At present, although CD19 CAR-T cells have achieved great success, as many as two-thirds of patients who receive CD19 CAR-T cell therapy will experience loss of CD19 expressio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/867A61K39/00A61P35/00A61P35/02
CPCC07K16/2803C07K16/2887C07K14/70517C07K14/7051C07K14/70578C12N15/86A61K39/001112A61K39/001124A61P35/00A61P35/02C07K2317/622C07K2317/31C07K2319/02C07K2319/03C07K2319/33C07K2319/74C12N2740/15043C12N2800/107A61K2039/5156A61K2039/804
Inventor 张坤
Owner 华夏源细胞工程集团股份有限公司
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