DOI: 10.57760/sciencedb.j00217.04539
title: Preparation of universal HER2-specific CAR-T cells expressed HLA-G and evaluation their resistance to NK cells killing effect
title: 表达HLA-G通用型HER2 CAR-T细胞的制备及其抵抗NK细胞杀伤作用的研究

description: Objective This study aimed to prepare universal human growth factor receptor 2(HER2)-specific chimeric antigen receptor T cells expressing human leukocyte antigen G(HLA-G),named"GU HER2 CAR-T",and to detect their ability to resist NK cells-induced killing effect and their ability to kill HER2-positive tumor cells.Methods Lentivirus was produced in 293-T cells by using a humanized single chain antibody P1h2 CAR lentivirus vector expressing HLA-G and targeting HER2.HER2-specific CAR-T cells expressing HLA-G(HLA-G HER2 CAR-T)were obtained by transduction of healthy human T cells with the lentivirus.The GU HER2 CAR-T cells were obtained by using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)gene-editing technology to knockout the T cell receptor(TCR)and human leukocyte antigen Ⅰ(HLA-Ⅰ)in HLA-G HER2 CAR-T cells.The killing ability of GU HER2 CAR-T cells against tumor cells was evaluated by CCK-8 assay.Allogeneic peripheral blood mononuclear cells(PBMC)immune rejection response to GU HER2 CAR-T was detected by cells co-culture assay.Luciferase assay was used to evaluate the killing capacity of NK92 cells on GU HER2 CAR-T.Results The GU HER2 CAR-T cells were prepared and could kill HER2 positive tumor cells specifically.The GU HER2 CAR-T cells could also resist immune rejection by allogeneic PBMC and resist killing by NK92 cells.Conclusion The GU HER2 CAR-T cells can resist NK cells-induced cytotoxicity,laying a foundation for improving the sustained killing of HER2 positive tumor cells by universal allogeneic CAR-T.
description: Objective This study aimed to prepare universal human growth factor receptor 2(HER2)-specific chimeric antigen receptor T cells expressing human leukocyte antigen G(HLA-G),named"GU HER2 CAR-T",and to detect their ability to resist NK cells-induced killing effect and their ability to kill HER2-positive tumor cells.Methods Lentivirus was produced in 293-T cells by using a humanized single chain antibody P1h2 CAR lentivirus vector expressing HLA-G and targeting HER2.HER2-specific CAR-T cells expressing HLA-G(HLA-G HER2 CAR-T)were obtained by transduction of healthy human T cells with the lentivirus.The GU HER2 CAR-T cells were obtained by using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)gene-editing technology to knockout the T cell receptor(TCR)and human leukocyte antigen Ⅰ(HLA-Ⅰ)in HLA-G HER2 CAR-T cells.The killing ability of GU HER2 CAR-T cells against tumor cells was evaluated by CCK-8 assay.Allogeneic peripheral blood mononuclear cells(PBMC)immune rejection response to GU HER2 CAR-T was detected by cells co-culture assay.Luciferase assay was used to evaluate the killing capacity of NK92 cells on GU HER2 CAR-T.Results The GU HER2 CAR-T cells were prepared and could kill HER2 positive tumor cells specifically.The GU HER2 CAR-T cells could also resist immune rejection by allogeneic PBMC and resist killing by NK92 cells.Conclusion The GU HER2 CAR-T cells can resist NK cells-induced cytotoxicity,laying a foundation for improving the sustained killing of HER2 positive tumor cells by universal allogeneic CAR-T.