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Archive > Volume 32 Issue 11 > 2025,32(11):1181-1187. DOI:10.3872/j.issn.1007-385X.2025.11.011 Prev Next
Technigue and Method
Preparation method and preliminary functional characterization of chimeric costimulatory receptor-modified γδ T cells targeting the CD7 antigen
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Abstract:
[Abstract] Objective: To develop a chimeric costimulatory receptor (CCR) targeting the CD7 antigen and prepare CCR-modified γδ T cells from healthy donors for the evaluation of its in vitro and in vivo cytotoxic effects against T-cell acute lymphoblastic leukemia (T-ALL) cells. Methods: Lentiviral vectors carrying CD7-DAP10-CCR were constructed and γδ T cells in the peripheral blood of healthy individuals were transduced to prepare CCR γδ T cells targeting the CD7 antigen (CD7-DAP10-CCR-γδ T). The obtained cells were expanded in vitro using artificial antigen-presenting cells (aAPC) expressing CD64, CD86, and CD137L. The in vitro cytotoxic effects of CD7-DAP10-CCR-γδ T cells against T-ALL cells (Jurkat), CD7-deficient Jurkat cells (CD7? Jurkat), and healthy donor primary αβ T cells were detected using the Annexin Ⅴ/7-AAD assay. The experiment was performed at three effector-to-target (E∶T) ratios (1∶1, 1∶3, and 1∶10), with Jurkat cells as CD7 positive target cells, CD7? Jurkat cells as CD7 negative target cells to verify the killing specificity, and healthy donor primary αβ T cells as CD7 positive normal control cells to evaluate the off-target effects of CD7 DAP10-CCR-γδ T cells. The incubation time was 18-24 h. Furthermore, the in vivo efficacy was evaluated in an immunodeficient mouse model bearing T-ALL xenografts. In vivo imaging of tumor-bearing immunodeficient mice was regularly conducted, their body weight and length of survival monitored to evaluate in vivo efficacy of CD7-DAP10-CCR-γδ T cells in tumor-bearing immunodeficient mice. Results: CD7-DAP10-CCR-γδ T cells were successfully prepared in vitro using aAPC, achieving an average expansion fold exceeding 10 000. In vitro cytotoxicity assays demonstrated that these cells exhibited significantly high killing activity against T-ALL cells and significantly high toxicity against Jurkat cells (P < 0.01), while showing limited cytotoxicity against CD7? Jurkat cells and negligible effects on normal primary CD7-high αβ T cells. In vivo efficacy experiment on tumor-bearing immunodeficient mice indicated that treatment with CD7-DAP10-CCR-γδ T cells resulted in a significant prolongation of survival compared with the PBS control group. Conclusion: CD7-DAP10-CCR-γδ T cells can be successfully generated in vitro using aAPC. CD7-DAP10-CCR-γδ T cells demonstrate strong cytotoxicity against T-ALL cells both in vitro and in vivo, which suggests therapeutic potential of CD7-DAP10-CCR-γδ T cells against T-ALL cells.
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