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CAR-CD123T2 chimeric antigen receptor T cell and application thereof

A CAR-CD123T2, chimeric antigen receptor technology, applied in receptors/cell surface antigens/cell surface determinants, animal cells, blood cell cancer vaccines, etc., can solve the problem of not fully considering the mobilization of T cell adaptive immune responses and Innate immune response, etc.

Inactive Publication Date: 2020-03-24
HEFEI CASTEM CELL & REGENERATIVE MEDICINE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The CAR molecules targeting CD123 involved in the prior art do not fully consider the use of CAR molecules to mobilize T cell adaptive immune responses and innate immune responses

Method used

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  • CAR-CD123T2 chimeric antigen receptor T cell and application thereof
  • CAR-CD123T2 chimeric antigen receptor T cell and application thereof
  • CAR-CD123T2 chimeric antigen receptor T cell and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0039] Preparation of CAR-CD123T2 Chimeric Antigen Receptor T Cells

[0040] 1. Construction of new CAR molecules

[0041] Such as figure 1 As shown, the extracellular part of the new CAR molecule includes GM-CSF signal peptide, part of scFv and CD28, the transmembrane region is from CD28, the intracellular part is the signaling domain of CD28 and CD3zeta, and the last part is the added TLR2 The intracellular domain TIR (626~784a.a.);

[0042] 2. Preparation and transfection efficiency of CAR-CD123T2 chimeric antigen receptor T cells

[0043] Such as figure 2As shown, the constructed novel CAR molecule was transfected into human T cells by GFP-Luciferase (GL) to obtain CAR-CD123T2 cells, that is, CAR-CD123T2 chimeric antigen receptor T cells.

Embodiment 2

[0045] Application of CAR-CD123T2 chimeric antigen receptor T cells in the treatment of AML

[0046] 1. Selection of AML cell lines

[0047] Select CD123-positive cell lines and CD123-negative cell lines, and detect whether the AML cell lines express CD123 antigen by flow cytometry. According to the experimental results, three cell lines, K562, AML3, and Molm13, were selected. image 3 The result graph of detecting whether the AML cell line expresses CD123 antigen by flow cytometry;

[0048] 2. Treatment of cell lines

[0049] K562GL, AML3GL, and MOLM13GL target cell lines were constructed by infecting K562, AML3, and Molm13 with GFP-Luciferase (GL) virus, and the expression of CD123 and GFP in these three cell lines was detected. K562GL was used as a negative control group, and AML3GL and Molm13GL Two cell lines were used as the positive experimental group, Figure 4 It is the expression result graph of CD123 and GFP in K562GL, AML3GL and MOLM13GL target cell lines;

[00...

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Abstract

The invention discloses a CAR-CD123T2 chimeric antigen receptor T cell. The CAR-CD123T2 chimeric antigen receptor T cell is formed by constructing a novel CAR molecule which takes CD123 as a target point and contains a TLR2 signal transmission structural domain, and then transfecting human T cells through lentivirus. The CAR-CD123T2 chimeric antigen receptor T cell is provided, a GM-CSF signal peptide and the scFv region of a CD123 monoclonal antibody 32716 are selected, Vh and V1 are connected through a G4S linker, an extracellular segment, a transmembrane region and an intracellular signal segment of the CD28 part are selected, a CD3zeta chain is added, and the intracellular signal segment of TLR2 is integrated to construct the novel CAR molecule. Adaptive immune response brought by activation of CD28 and CD3Z and inherent immune response caused by a TLR2 signal domain are effectively combined, CAR-T is modified from the aspect of optimizing a CAR-CD123 structure, and a new method can be provided for treating AML.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a CAR-CD123T2 chimeric antigen receptor T cell and an application thereof. Background technique [0002] Acute myeloid leukemia (AML) is characterized by the malignant clonal proliferation of primitive and immature myeloid cells in the bone marrow and peripheral blood, and a group of myeloid hematopoietic stems that severely damage normal hematopoiesis with the rapid accumulation of leukemia cells. / group of cell malignancies. AML is the most common type of acute leukemia in adults with the highest mortality rate and poor prognosis. [0003] Once AML is diagnosed, standard chemotherapy will be used, and some patients may benefit from allogenic hematopoietic stem cell transplantation (AlloHSCT). The 5-year survival rate of AML patients is 21.4%, while the 5-year survival rate of patients over 75 years old is only 1%. The initial remission rate of AML patients under 60 ...

Claims

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Application Information

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IPC IPC(8): C12N5/10C12N15/62A61K39/00A61P35/02
CPCA61K2039/5156A61P35/02A61K39/001119A61K2039/804C07K14/535C07K14/7051C07K14/70521C07K14/70596C07K16/2866C07K2317/622C07K2319/02C07K2319/03C07K2319/33C07K2319/74C12N5/0636C12N2510/00
Inventor 李鹏魏新茹徐有娣程琳姚瑶王素娜吴绮婷龙有国陈冬梅林思妙
Owner HEFEI CASTEM CELL & REGENERATIVE MEDICINE CO LTD
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