In vitro and in vivo cytotoxic activity of humanized CD19 CAR-T cells against Raji cell line
10.3760/cma.j.issn.0254-5101.2019.09.004
- VernacularTitle: 高亲和力人源化CD19 CAR-T细胞对Raji细胞株体外、体内杀伤活性研究
- Author:
Juanxia MENG
1
;
Nan MOU
2
;
Zhenxing YANG
2
;
Jia WANG
1
;
Xin LI
1
;
Yanyu JIANG
1
;
Ting YUAN
1
;
Qi DENG
1
Author Information
1. Department of Hematology, Tianjin First Central Hospital, Tianjin 300192, China
2. Shanghai Genbase Biotechnology Co., Ltd., Shanghai 201203, China
- Publication Type:Journal Article
- Keywords:
Murinized;
Humanized;
CD19;
CAR-T cells;
Raji cell line;
Cytotoxicity
- From:
Chinese Journal of Microbiology and Immunology
2019;39(9):662-667
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the different functions of humanized and murinized CD19 chimeric antigen receptor (CAR)-T cells against Raji cell line in vitro and in vivo.
Methods:Peripheral blood samples were collected from eight patients with lymphoma who were going to receive CD19 CAR-T cell therapy and used for the preparation of peripheral blood mononuclear cells (PBMC) as well as humanized and murinized CAR-T cells. Cell proliferation and cytotoxicity were detected with CCK-8 and LDH assays, respectively. A tumor-bearing mouse model was established by injecting BALB/c female nude mice with fluorescent Raji cells. Changes in tumor volume in these mice were observed by in vivo imaging technology. The transfection efficiency and amount of CAR-T cells in the mice were detected with flow cytometry.
Results:No statistical difference in transfection efficiency was found between humanized and murinized CAR-T cells, nor in cell proliferation at 24 h of culture in vitro(P=0.104). The proliferation of humanized CAR-T cells showed a significant increase compared with that of murinized CAR-T cells at 48 h of culture (P=0.009). Similarly, the cytotoxicity of the two types of CAR-T cells against Raji cells showed no significant difference at 24 h at any effector/target (E/T) ratio (1∶1 or 4∶1), and that of humanized CAR-T cells was higher than that of murinized CAR-T cells at both E/T ratios at 48 h (E/T ratio=1∶1, P=0.005; E/T ratio=4∶1, P=0.008). Moreover, the cytotoxicity of CAR-T cells was higher than that of PBMC in any case. Tumor volumes in mice were reduced 14 d after humanized or murinized CAR-T cell therapy, while the mice in the PBMC control group suffered tumor progression. Tumor volume began to increase in mice 21 d after murinized CAR-T cell therapy, while no significant change was observed in the mice treated with humanized CAR-T cells. All of the mice died 25 d after murinized CAR-T cell therapy, while the deaths among those under humanized CAR-T cell therapy occurred on 31 d. The proportion of CAR-T cells in mice reached the peak 7 d after receiving humanized or murinized CAR-T cell therapy, while that in the humanized group was significantly higher than that in the murinized group at any time point (P4 d=0.001, P7 d=0.000, P14 d=0.003). Murinized CAR-T cells became undetectable on 21 d, while humanized CAR-T cells on 35 d. The maximum survival time for mice in the PBMC and murinized and humanized CAR-T cell groups was 20 d, 25 d and 53 d, respectively.
Conclusions:Compared with murinized CD19 CAR-T cells, humanized CD19 CAR-T cells showed stronger proliferation potential and cytotoxicity and remained in vivo detectable for a longer period of time. This study indicated that humanized CD19 CAR-T cells were superior to murinized CD19 CAR-T cells for the treatment of B cell lymphoma.
