Preparation Method and Preliminary Functional Characterization of Chimeric Antigen Receptor Vδ1T Cells Targeting CD19 Antigen
WANG Min1, ZHANG Ping1, XU Hanying1, YANG Lin1,2*
This study aims to investigate the preparation methods of CD19-targeting CAR-Vδ1T cells and evaluate their antitumor effects against B cell-derived tumor cells both in vitro and in vivo. Leveraging the principles of CAR-T cell engineering, CD19-CAR-Vδ1T cells targeting CD19 were designed. aAPCs (artificial antigen-presenting cells) were utilized for in vitro expansion and activation of CD19-CAR-Vδ1T cells. Cytotoxic activity and anti-inflammatory cytokine secretion against CD19 antigen-positive cell lines separately were quantified using Annexin V/7-AAD apoptosis assays and CBA (cytometric bead array). Flow cytometry was employed to validate the cytotoxic activity and anti-inflammatory cytokine secretion of CD19-CAR-Vδ1T cells. Additionally, in vivo efficacy and toxicity were assessed in tumor-bearing immunodeficient mice through regular bioluminescence imaging, body mass monitoring, and survival observation. Expansion results showed that CD19-CAR-Vδ1T cells were successfully generated in vitro using aAPCs, with an average expansion fold of Vδ1T cells exceeding 10 000-fold. In vitro cytotoxicity assays demonstrated that CD19-CAR-Vδ1T cells exhibited significant antigen-specific cytotoxicity and sustained anti-tumor activity against CD19-positive tumor cells compared with the control group. In vitro anti-inflammatory cytokine secretion assays showed that CD19-CAR-Vδ1T cells displayed robust antigen-specific cytokine secretion in response to CD19-positive tumor cells compared with the control group. Furthermore, in vivo pharmacodynamic and toxicological studies in tumor-bearing immunodeficient mice revealed that CD19-CARVδ1T cell treatment not only significantly prolonged the survival of tumor-bearing mice, but also demonstrated a favorable safety profile compared with the control group. Overall, CD19-CAR-Vδ1T cells demonstrate therapeutic potential for the treatment of B cell-derived malignancies.
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