Chimeric antigen receptor T cell capable of conducting allograft and preparation method

A chimeric antigen receptor and allogeneic transplantation technology, applied in the field of tumor treatment, can solve the problems of inability to effectively kill tumor cells, difficult separation of T cells, mixed tumor cells, etc., and achieve the effect of facilitating tumor treatment.

Inactive Publication Date: 2016-06-08
BIOTOWNTEK CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] In view of the above-mentioned deficiencies in the prior art, the object of the present invention is to provide a chimeric antigen receptor T cell capable of allogeneic transplantation and its preparation method, aiming at solving the difficulties in the separation of existing T cells, the inability to effectively kill tumor cells and the presence of mixed positive cells. tumor cell problems

Method used

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  • Chimeric antigen receptor T cell capable of conducting allograft and preparation method
  • Chimeric antigen receptor T cell capable of conducting allograft and preparation method
  • Chimeric antigen receptor T cell capable of conducting allograft and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Allograft TRAC-inactivated T cells combined with third-generation CAR to treat tumors

[0047] In this example, CRISPR / Cas9 technology was used to knock out the gene TRAC encoding the constant region of the alpha chain of TCR. For the exons of TRAC, including exon 1, exon 2, exon 3, and exon 4, exon 1 of TRAC was knocked out using CRISPR / Cas9 technology. According to the exon sequence of TRAC, especially the sequence of exon 1, the sequence designed to express sgRNA (singleguideRNA, sgRNA) includes: Seq-TRAC-EX1-1, Seq-TRAC-EX1-2, Seq-TRAC-EX1 -3, Seq-TRAC-EX1-4, Seq-TRAC-EX1-5.

[0048] Further, the above sequence, especially the sequence of Seq-TRAC-EX1-1 is reversely complementary to obtain the reverse complementary sequence of Seq-TRAC-EX1-1 (see sequence Seq-TRAC-EX1-1-R).

[0049] Furthermore, it is also necessary to add double restriction sites BamHI and EcoRI at both ends of Seq-TRAC-EX1-1 and Seq-TRAC-EX1-1-R to facilitate the construction of entry vectors af...

Embodiment 2

[0058] Allograft TCRBC1-inactivated T cells combined with third-generation CAR to treat tumors

[0059] In this example, the CRISPR / Cas9 technology was used to knock out the gene TCRBC1 encoding the constant region of the β chain of TCR. For exons of TCRBC1, including exon 1, exon 2, exon 3 and exon 4, exon 1 of TCRBC1 was knocked out by CRISPR / Cas9 technology. According to the exon sequence of TCRBC1, especially the sequence of exon 1, the sequences designed to express sgRNA (singleguideRNA, sgRNA) include: Seq-TCRBC1-EX1-1, Seq-TCRBC1-EX1-2, Seq-TCRBC1-EX1 -3, Seq-TCRBC1-EX1-4, Seq-TCRBC1-EX1-5.

[0060] Further, the above sequence, especially the sequence of Seq-TCRBC1-EX1-1 is reversely complementary to obtain the reverse complementary sequence of Seq-TCRBC1-EX1-1 (see sequence Seq-TCRBC1-EX1-1-R).

[0061] Further, it is also necessary to add double restriction sites BamHI and EcoRI at both ends of Seq-TCRBC1-EX1-1 and Seq-TCRBC1-EX1-1-R to facilitate the construction o...

Embodiment 3

[0070] Allograft TCRBC2-inactivated T cells combined with third-generation CAR to treat tumors

[0071] In this example, the CRISPR / Cas9 technology was used to knock out the gene TCRBC2 encoding the constant region of the β chain of TCR. For the exons of TCRBC2, including exon 1, exon 2, exon 3 and exon 4, exon 1 of TCRBC2 was knocked out by using CRISPR / Cas9 technology. According to the exon sequence of TCRBC2, especially the sequence of exon 1, the sequence designed to express sgRNA (singleguideRNA, sgRNA) includes: Seq-TCRBC2-EX1-1, Seq-TCRBC2-EX1-2, Seq-TCRBC2-EX1 -3, Seq-TCRBC2-EX1-4, Seq-TCRBC2-EX1-5.

[0072] Further, the above sequence, especially the sequence of Seq-TCRBC2-EX1-1 is reversely complementary to obtain the reverse complementary sequence of Seq-TCRBC2-EX1-1 (see sequence Seq-TCRBC2-EX1-1-R).

[0073] Further, it is also necessary to add double restriction sites BamHI and EcoRI at both ends of Seq-TCRBC2-EX1-1 and Seq-TCRBC2-EX1-1-R to facilitate the cons...

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Abstract

The invention discloses a chimeric antigen receptor T cell capable of conducting allograft and a preparation method. The T cell is transformed through the gene fixed-point knockout technology, especially the CRISPR / Cas9 technology, gene knockout is conducted for the constant areas of chains alpha and beta of TCR of the T cell so that TCR of the T cell can be inactivated, and the T cell can conduct allograft without causing immunological rejection. The T cell with the inactivated TCR is combined with the CAR technology, and therefore the aim of treating specific tumors in a targeted mode through the T cell with the allograft source can be achieved. The chimeric antigen receptor T cell and the method overcome the defects that at present, T cells of patients are small in number, low in activity and long in culture time. High-quality and high-activity T cells in blood of healthy people can be used for transforming in advance, CART cells for various tumors are prepared, tumor patients do not need to wait once needing CART treatment, the precious time of treatment wait is shortened to the maximum extent.

Description

technical field [0001] The invention relates to the field of tumor treatment, in particular to a chimeric antigen receptor T cell capable of allogeneic transplantation and a preparation method thereof. Background technique [0002] Tumor has always been a major disease that plagues the world and seriously endangers human health. Therefore, it is an important research topic in the world medical community to find an effective tumor treatment method and completely conquer the tumor. At present, although the three traditional main treatment methods, namely surgery, chemotherapy and radiotherapy, are the basic methods of global tumor treatment, their therapeutic effects are limited. [0003] Among them, traditional surgical resection is the most basic and important tumor treatment method in the tumor industry. Radiation therapy is a treatment method that uses radiation to irradiate cancer tissues to inhibit and kill cancer cells. It is an adjuvant therapy for most tumors. A se...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10
CPCC07K14/705
Inventor 陈光风史秀娟刘小青
Owner BIOTOWNTEK CO LTD
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