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Construction and application of TanCAR

A technology of E13K.R66D.S69D.R109K and extracellular segment, which is applied in the field of CART therapy and can solve problems such as specificity needs to be improved

Pending Publication Date: 2022-06-03
SHAANXI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the affinity of IL13 for highly expressed IL13Rα2 in glioma cells is higher than its affinity for IL13Rα1 normally expressed in somatic cells, IL13 CAR still has off-target toxicity caused by the combination of IL13 and IL13Rα1, so its specificity remains to be determined. improve

Method used

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  • Construction and application of TanCAR
  • Construction and application of TanCAR
  • Construction and application of TanCAR

Examples

Experimental program
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Embodiment Construction

[0036] This example presents a TanCAR, which is constructed on the basis of the third-generation CAR backbone. The extracellular segment of the TanCAR that recognizes the target molecule on the surface of the target cell consists of IL13 in the base sequence shown by NO1 in the nucleotide sequence table. (E13K.R66D.S69D.R109K) mutant nucleic acid sequence and EphA2 scFv nucleic acid sequence with the optimized restriction enzyme cleavage site shown in the base sequence shown in NO2 in the nucleotide sequence table and the addition of linker, wherein, IL13 (E13K. The R66D.S69D.R109K) sequence is located at the 5' end of the extracellular segment that recognizes the target molecule on the surface of the target cell, and the EphA2scFv sequence is located at the 3' end of the extracellular segment that recognizes the target molecule on the target cell surface. Nucleic acid sequence spacing.

[0037] The construction method of the above TanCAR, the steps are as follows:

[0038] 1...

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Abstract

The invention discloses a TanCAR, which is constructed on the basis of a third-generation CAR skeleton and consists of an extracellular segment part for identifying target molecules on the surface of a target cell, a transmembrane part and an intracellular signal transduction part. An extracellular segment part for recognizing a target molecule on the surface of a target cell in the TanCAR consists of an IL13 (E13K.R66D.S69D.R109K) mutant sequence with a base sequence as shown in NO1 in a nucleotide sequence table and an EphA2scFv sequence as shown in NO2 in the nucleotide sequence table; an IL13 (E13K.R66D.S69D.R109K) mutant sequence is located at the 5'end of an extracellular segment part for recognizing a target molecule on the surface of a target cell, an EphA2scFv sequence is located at the 3 'end of the extracellular segment part for recognizing the target molecule on the surface of the target cell, and the two segments of sequences are connected through a GGGS linker. The construction method comprises the following three steps: synthesizing a gene of a third-generation CAR molecule, constructing a third-generation CAR skeleton vector, constructing a novel IL13 (E13K.R66D.S69D.R109K) CAR vector, and constructing a novel IL13 (E13K.R66D.S69D.R109K)-EphA2scFv-TanCAR vector. The construction method comprises the following steps: synthesizing a gene of the third-generation CAR molecule; researches show that the T cells prepared based on the TanCAR can be used for cellular immunotherapy in tumors.

Description

technical field [0001] The invention belongs to the technical field of CART treatment, in particular to the construction and application of a TanCAR. Background technique [0002] Malignant tumors seriously endanger human life and health. At present, the conventional treatment for malignant tumors is still surgery, radiotherapy and chemotherapy. The curative effect of this conventional treatment on most tumors is still not very satisfactory. For most chemotherapy drugs, its therapeutic index is still very low, that is, the The therapeutic dose is closer to the toxic dose. Therefore, this treatment is usually accompanied by obvious toxic effects. Therefore, the method of selectively killing tumor cells mainly relies on the specific markers of tumor cells, and the current targeted therapy of tumors has become a research hotspot. In recent years, with the rapid development of biotechnology, CAR-T (Chimeric antigen receptor T cell, CAR-T) therapy has become a very promising c...

Claims

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Application Information

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IPC IPC(8): C07K19/00C07K14/54C07K16/28C12N15/62C12N15/24C12N15/13C12N15/867C12N5/10A61K39/00A61P35/00
CPCC07K16/2866C07K14/5437C07K14/7051C12N15/86C12N5/0636A61K39/001119A61K39/001116A61P35/00C07K2317/622C07K2319/02C07K2319/03C07K2319/33C07K2319/74C12N2740/15043C12N2800/107C12N2510/00A61K2039/5156
Inventor 夏海滨郑晓晶赵俊丽
Owner SHAANXI NORMAL UNIV
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