Construction and functional study of CD19 CAR-T cells that secrete IL-2 locally
10.3872/j.issn.1007-385x.2025.04.002
- VernacularTitle:局部分泌IL-2的CD19 CAR-T细胞构建与功能
- Author:
Wenjuan WANG
1
;
Jing CHEN
;
Zhou HU
;
Xiuying LIU
;
Jianxun WANG
Author Information
1. 北京中医药大学 生命科学学院,北京 102400
- Publication Type:Journal Article
- Keywords:
SynNotch system;
CAR-T cell;
IL-2;
CD19
- From:
Chinese Journal of Cancer Biotherapy
2025;32(4):356-363
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the impact of locally IL-2-secreting CAR-T cells(Syn.CD19.IL-2 CAR-T cells)constructed using a novel single-vector SynNotch system on the function of conventional CD19 CAR-T cells.Methods:Building upon our previously established single-vector SynNotch system,the CD19-specific FMC63 antibody was integrated with an IL-2 expression module to prepare Syn.CD19.IL-2 CAR-T cells through T cell transduction using a self-inactivating retroviral vector.For validation experiments,cells were divided into the Syn.CD19.IL-2 CAR-T cell group and the untransduced T cells group.ELISA was used to detect IL-2 secretion levels after antigen stimulation.The CFSE staining assay was implemented to evaluate the effects of IL-2 secretion on the proliferation of CD19 CAR-T cells when CD19 CAR-T cells were co-cultured with Raji-Luc or SW620-CD19-Luc cells in the presence of Syn.CD19.IL-2 CAR-T cells.Flow cytometry was employed to detect CD69 expression and monitor the activation status of CD19 CAR-T cells when co-cultured with Raji-Luc cells in the condition of Syn.CD19.IL-2 CAR-T cells secreting IL-2.Results:The self-inactivating retroviral vector Syn.CD19.IL-2 CAR were successfully constructed and Syn.CD19.IL-2 CAR-T cells with a viral titer>1×10? copies/mL and transduction efficiency of 37.1%were generated.After antigen stimulation,SynNotch CAR-T cells demonstrated significantly elevated IL-2 secretion compared with the untransduced T cell group(P<0.001).When Syn.CD19.IL-2 CAR-T cells secreted IL-2,the CD19 CAR-T cells exhibited enhanced proliferative capacity and elevated activation level(all P<0.001).Conclusion:Successfully constructed Syn.CD19.IL-2 CAR-T cells can significantly enhance the proliferative capacity and activation ability of conventional CD19 CAR-T cells.
