Bi-target CAR-T cell as well as preparation method and application thereof

A cell and tumor cell technology, applied in the field of double CAR-T cells and its preparation, can solve the problem of high toxicity of CAR-T technology, and achieve the effect of improving specificity of killing

Inactive Publication Date: 2018-06-29
浙江史迪姆生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of the present invention is to provide a double CAR-T cell and its preparation method and application to solve the problem of excessive toxicity in CAR-T technology

Method used

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  • Bi-target CAR-T cell as well as preparation method and application thereof
  • Bi-target CAR-T cell as well as preparation method and application thereof
  • Bi-target CAR-T cell as well as preparation method and application thereof

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Embodiment 1

[0033] 1. Experimental materials and reagents

[0034] 1.1. Cell source: T cells derived from normal human PBMC, T cells derived from PBMC of leukemia patients, human peripheral blood B cells, human acute B lymphoid leukemia cells Nalm-6 (CD19 positive); human myeloid leukemia cells K562 (CD19 negative) ).

[0035] 1.2. Initial medium: PRIM1640 (GIBCO) medium, containing 5% heat-activated human AB serum, 1% glutamine and 40IU / mL interleukin(IL)-2.

[0036] 1.3. Amplification medium: PRIM1640 medium, containing 5% heat-activated human AB serum, 1% glutamine and 300IU / mL IL-2.

[0037] 1.4. Complete medium: PRIM1640, containing 10% FBS, 10% blue chain double antibody.

[0038] 1.5. PBMC freezing solution: 90% FBS + 10% DMSO.

[0039] 1.6. Experimental reagents: PRIM1640 medium (GIBCO), IL-2 (GIBCO), AB serum (GIBCO), penicillin (GIBCO), Anti-CD3 (OKT3) (BD company), Anti-CD3-APC ( BD company), Anti-CD8-PE (BD company), Anti-CD4-FITC (BD company), anti-CD56-PerCP-Cy5.5 (BD company), Anti-...

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Abstract

The invention discloses a bi-target CAR-T cell. The bi-target CAR-T cell is characterized in that two target antigens which are in co-expression in tumor cells and are not in co-expression in normal cells are utilized, an antibody variable region of one target antigen is fused with a killing signal in a T cell, and an antibody variable region of the other target antigen is fused with a co-stimulating signal in the T cell, and two different expression cassettes are formed, so that the bi-target CAR-T cell is constructed; the constructed bi-target CAR-T cell can just identify tumor cells which can express the two target antigens at the same time and cannot identify the normal cells which just express one target antigen. The invention also discloses a preparation method and application of thebi-target CAR-T cell. According to the bi-target CAR-T cell disclosed by the invention, not only killing specificity is increased, but also killing of the normal cells is reduced, and side effects brought by an inflammatory factor storm are reduced.

Description

Technical field [0001] The invention relates to the field of biotechnology, in particular to a dual CAR-T cell and its preparation method and application. Background technique [0002] Leukemia is a malignant proliferative blood disease of hematopoietic stem cells. Under normal circumstances, the white blood cells produced by the bone marrow help the human body to fight infections. In leukemia patients, the bone marrow produces abnormal white blood cells. These malignant white blood cells expand and reject the normal blood cells in the body, causing an imbalance of immune cells in the blood and affecting the blood cells of the body. Normal function. Leukemias include acute myeloid leukemia, chronic lymphocytic leukemia, Hodgkin's leukemia, non-Hodgkin's leukemia, etc. Chronic lymphocytic leukemia usually occurs in the middle-aged and elderly population, and rarely occurs in the adolescent population. The current methods of treating leukemia mainly include chemotherapy, radioth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/867C12N15/62A61K35/17A61P35/00A61P35/02
CPCA61K35/17C07K14/7051C07K14/70521C07K16/2803C07K2319/00C07K2319/33C07K2319/74C12N5/0636C12N15/86C12N2510/00C12N2740/15043C12N2800/107
Inventor 屈彩云
Owner 浙江史迪姆生物科技有限公司
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