Characterization of Donor Variability for γδ T Cell ex vivo Expansion and Development of an Allogeneic γδ T Cell Immunotherapy

Gamma delta (gamma delta) T cells recently emerged as an attractive candidate for cancer immunotherapy treatments due to their inherent cytotoxicity against both hematological and solid tumors. Moreover, gamma delta T cells provide a platform for the development of allogeneic cell therapies, as they can recognize antigens independent of MHC recognition and without the requirement for a chimeric antigen receptor. However, gamma delta T cell adoptive cell therapy depends on ex vivo expansion to manufacture sufficient cell product numbers, which remains challenging and limited by inter-donor variability. In the current study, we characterize the differences in expansion of gamma delta T cells from various donors that expand (EX) and donors that fail to expand, i.e., non-expanders (NE). Further, we demonstrate that IL-21 can be used to increase the expansion potential of NE. In order to reduce the risk of graft vs. host disease (GVHD) induced by an allogeneic T cell product, alpha beta T cell depletions must be considered due to the potential for HLA mismatch. Typically, alpha beta T cell depletions are performed at the end of expansion, prior to infusion. We show that gamma delta T cell cultures can be successfully alpha beta depleted on day 6 of expansion, providing a better environment for the gamma delta T cells to expand, and that the alpha beta T cell population remains below clinically acceptable standards for T cell-depleted allogeneic stem cell products. Finally, we assess the potential for a mixed donor gamma delta T cell therapy and characterize the effects of cryopreservation on gamma delta T cells. Collectively, these studies support the development of an improved allogeneic gamma delta T cell product and suggest the possibility of using mixed donor gamma delta T cell immunotherapies.