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Preparation Method and Preliminary Functional Characterization of Chimeric Antigen Receptor γδT Cells Targeting CD7 Antigen
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Abstract:
Objective: To develop a chimeric costimulatory receptor targeting CD7 and prepare 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: γδ T cells expressing the CD7-targeting chimeric costimulatory receptor (CD7-DAP10-CCR-γδ T) were constructed. Using lentiviral transduction, the receptor was introduced into human peripheral blood-derived γδ T cells from healthy donors, followed by ex vivo expansion with artificial antigen-presenting cell (aAPC) expressing CD64, CD86, and CD137L. The cytotoxic activity of CD7-DAP10-CCR-γδ T cells against T-ALL cells (Jurkat), CD7-deficient Jurkat cells (CD7? Jurkat), and normal primary αβ T cells was assessed using the Annexin V/7-AAD assay. Furthermore, the in vivo efficacy was evaluated in an immunodeficient mouse model bearing T-ALL xenografts. Tumor burden was monitored regularly via in vivo imaging, and body weight changes and survival rates were recorded. Results: CD7-DAP10-CCR-γδ T cells were successfully expanded using aAPC, achieving an average expansion fold exceeding 10000. In vitro cytotoxicity assays demonstrated that these cells exhibited significantly high killing activity against T-ALL cells and 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 the T-ALL xenograft model, treatment with CD7-DAP10-CCR-γδ T cells resulted in a significant prolongation of survival compared to the PBS control group. Conclusion: This study establishes a robust aAPC-driven protocol for generating CD7-DAP10-CCR-γδ T cells with scalable expansion ( > 10 000-fold) and CD7-specific anti-tumor efficacy. The engineered cells exhibited selective cytotoxicity against T-ALL while sparing normal T cells, highlighting their therapeutic promise for relapsed/refractory T-ALL with mitigated on-target/off-tumor risks.
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