Vδ2 T-cell engagers bivalent for Vδ2-TCR binding provide anti-tumor immunity and support robust Vγ9Vδ2 T-cell expansion

Background: V gamma 9V delta 2 T-cells are antitumor immune effector cells that can detect metabolic dysregulation in cancer cells through phosphoantigen-induced conformational changes in the butyrophilin (BTN) 2A1/3A1 complex. In order to clinically exploit the anticancer properties of V gamma 9V delta 2 T-cells, various approaches have been studied including phosphoantigen stimulation, agonistic BTN3A-specific antibodies, adoptive transfer of expanded V gamma 9V delta 2 T-cells, and more recently bispecific antibodies. While V gamma 9V delta 2 T-cells constitute a sizeable population, typically making up similar to 1-10% of the total T cell population, lower numbers have been observed with increasing age and in the context of disease. Methods: We evaluated whether bivalent single domain antibodies (VHHs) that link V delta 2-TCR specific VHHs with different affinities could support V gamma 9V delta 2 T-cell expansion and could be incorporated in a bispecific engager format when additionally linked to a tumor antigen specific VHH. Results: Bivalent VHHs that link a high and low affinity V delta 2-TCR specific VHH can support V gamma 9V delta 2 T-cell expansion. The majority of V gamma 9V delta 2 T-cells that expanded following exposure to these bivalent VHHs had an effector or central memory phenotype and expressed relatively low levels of PD-1. Bispecific engagers that incorporated the bivalent V delta 2-TCR specific VHH as well as a tumor antigen specific VHH triggered antitumor effector functions and supported expansion of V gamma 9V delta 2 T-cells in vitro and in an in vivo model in NOG-hIL-15 mice. Conclusion: By enhancing the number of V gamma 9V delta 2 T-cells available to exert antitumor effector functions, these novel V delta 2-bivalent bispecific T cell engagers may promote the overall efficacy of bispecific V gamma 9V delta 2 T-cell engagement, particularly in patients with relatively low levels of V gamma 9V delta 2 T-cells.