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Chimeric antigen receptor targeting HER2, expression vector and application thereof

A technology of chimeric antigen receptors and expression vectors, applied in the field of tumor immunology drugs, can solve problems such as adverse reactions, patient death, cytokine storm, etc.

Active Publication Date: 2020-01-14
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 CAR-T cells can effectively kill HER2-positive breast cancer, it also brings adverse reactions of cytokine storm, which can lead to death of patients in severe cases.

Method used

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  • Chimeric antigen receptor targeting HER2, expression vector and application thereof
  • Chimeric antigen receptor targeting HER2, expression vector and application thereof
  • Chimeric antigen receptor targeting HER2, expression vector and application thereof

Examples

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Embodiment 1

[0024] An embodiment of the present invention provides a chimeric antigen receptor targeting HER2, such as figure 1 As shown, the chimeric antigen receptor includes: sequentially connected signal peptide (human CD8α signal peptide, CD8α leader), single-chain antibody targeting HER2 (HER2 scFv), lengthened CD8α hinge region (human CD8α hinge region, CD8αHinge), Transmembrane region (human CD8α transmembrane region, CD8α transmembrane), co-stimulatory factors, intracellular signal peptide (human CD3ζ intracellular signal peptide, CD3ζsignal), P2A linking peptide, IL2 signal peptide (IL2signal) and anti-PD1 single-chain antibody secretion region (aPD1 scFv), 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 HER2 is shown in SEQ ID NO: 2 in the sequence listing, and CD8α is lengthened The nucleotide sequence of the hinge region is shown in SEQ ID NO: 3 in the sequence listi...

Embodiment 2

[0031] The present invention provides an expression vector, which includes the vector and the chimeric antigen receptor provided in Example 1. The vector can be a lentiviral vector.

[0032] The following briefly introduces the preparation method of the expression vector, as follows:

[0033] Amplify and obtain the CAR01 gene sequence by PCR (Polymerase Chain Reaction, polymerase chain reaction), respectively add restriction site Xba I and restriction site EcoR I to the two ends of the CAR01 gene sequence to obtain the product to be digested, Carry out the double digestion reaction of Xba I and EcoR I with the lentiviral vector plasmid pCDH-EF1-MCS-T2A-copGFP, respectively, to obtain the digestion fragment containing CAR01 and the digestion fragment containing ppCDH-EF1-MCS-T2A-copGFP fragment. The enzyme digestion reaction conditions are as follows: the enzyme digestion temperature is 37° C., and the enzyme digestion time is 30 min. Enzyme digestion system (total volume 50...

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Abstract

The invention discloses a chimeric antigen receptor targeting HER2 and an expression vector and application thereof, and belongs to the field of tumor immune drugs. The chimeric antigen receptor targeting HER2 includesa signal peptide, a single-chain antibody targeting HER2, an extended CD8 alpha hinge region, a transmembrane region, an inducible co-stimulater, an intracellular signal peptide, a P2A linker peptide, an IL2 signal peptide, and an anti-PD1 single-chain antibody secretion region, wherein the signal peptide, the single-chain antibody targeting HER2, the extended CD8 alpha hinge region, the transmembrane region, the inducible co-stimulater, the intracellular signal peptide, the P2A linker peptide, the IL2 signal peptide, and the anti-PD1 single-chain antibody secretion region are connected in sequence, the nucleotide sequence of the signal peptide is shown as SEQ ID NO:1in a sequence table, the nucleotide sequence of the single-chain antibody targeting HER2 is shown as SEQ ID NO:2in the sequence table, the nucleotide sequence of the extended CD8 alpha hinge region is shown as SEQ ID NO:3in the sequence table, the nucleotide sequence of the transmembrane region is shown as SEQ ID NO:4in the sequence table,the nucleotide sequence of the intracellular signal peptide is shown as SEQ ID NO:5in the sequence table, the nucleotide sequence of the P2A linker peptide is shownas SEQ ID NO:6 in the sequence table, and the nucleotide sequence of the IL2 signal peptide is shown as SEQ ID NO:7in the sequence table. The chimeric antigen receptor has high antitumor activity while producing fewer cytokines.

Description

technical field [0001] The present invention relates to the field of tumor immune drugs, in particular to a chimeric antigen receptor targeting HER2 and its expression vector and application. Background technique [0002] Malignant tumors are one of the diseases that seriously threaten human health. Among them, breast cancer is one of the most common malignant tumors affecting women's health, and HER2-positive breast cancer patients account for 15% to 30% of the total number of breast cancer patients. [0003] Chimeric antigen receptor (CAR) technology is a cell therapy technology that has developed very rapidly in recent years, and is used to treat HER2-positive breast cancer. At present, although CAR-T cells can effectively kill HER2-positive breast cancer, it also brings adverse reactions of cytokine storm, which can lead to death of patients in severe cases. Therefore, there is an urgent need for a safe chimeric antigen receptor for the treatment of HER2-positive breast...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/867A61K39/00A61P35/00A61P15/14
CPCA61P15/14A61P35/00A61K39/001106A61K39/001111A61K39/00114A61K2039/812C07K14/55C07K14/7051C07K14/70517C07K14/70578C07K16/2818C07K16/32C07K2317/622C07K2319/02C07K2319/03C07K2319/33C07K2319/74C12N15/86C12N2740/15043C12N2800/107
Inventor 张坤
Owner 华夏源细胞工程集团股份有限公司
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