Antibody-mediated activation of Vδ1 γδ T-cells for cancer immunotherapy

Cancer remains a global leading cause of death as current treatments are often ineffective and present with high relapse-rates and severe adverse reactions. Vδ1 γδ T-cells are gaining interest as promising onco-immunotherapeutic targets due to their association with favourable prognoses in a myriad of malignancies. This thesis explores whether Vδ1-targeting bispecific engagers can improve immune responses against CD19+ haematological malignancies. Initial experiments investigated four Vδ1/CD19 bispecific engager formats, where only the BiTE format successfully enhanced Vδ1 function. The Vδ1/CD19 BiTE demonstrated specific binding to both Vδ1 TCR+ and CD19+ cells, resulting in TCR-downregulation, upregulation of T-cell activation markers 41BB, CD25 and CD107a, and enhanced cytotoxicity against CD19+ cell lines. Vδ1/CD19-treatment also increased PBMC cytotoxicity against CD19+ MEC-1 and primary CLL B-cells. To investigate mechanisms of healthy cell discrimination, differentially expressed ligands were identified from primary healthy B-cells, CLL B-cells and CD19+ cancer cell lines by RNA-analysis. Corresponding co-receptors to such ligands were identified using the literature. Following Vδ1/CD19 engager-treatment, naive PBMC-based Vδ1 T-cells increased expression of co-receptors NKG2D, CD96, DNAM-1, CD2, ICOS and PD-1 alongside CTLA-4 downregulation. Co-culture with MEC-1 cells enhanced granzyme B expression in PBMC-based Vδ1 T-cells and promoted effector-memory phenotypes. Blocking MEC-1 stress ligands reduced engager-treated Vδ1 cytotoxicity, though only under conditions of combined ligand-blockade, suggesting reduced susceptibility of these cells to immune evasion via downregulation of individual stress ligands. Engager treatment may prime naive Vδ1 T-cells for cytotoxicity by TCR-engagement but only engager-treated effector Vδ1 T-cells elicited a cytotoxic response when stimulated by plate-bound recombinant ligands, highlighting a potential lower threshold of activation for effector, over naive, Vδ1 T-cells. Findings also suggest effector Vδ1 T-cells modulate cytotoxicity through co-receptor signalling to discriminate between healthy and cancerous cells. Overall, the work presented in this thesis supports the continued investigation of Vδ1-targeting engagers as potential treatments for cancer.