Bioengineering and serum free expansion of blood-derived γδT cells

Background aims. Cellular immunotherapy relies on several highly variable patient-specific parameters, such as (i) cell number before and after expansion, (ii) targeting of cells to tumors, (iii) cell survival and function after infusion, and (iv) on-and off-target adverse events. Cellular approaches such as the specific expansion of gamma delta T cells as opposed to alpha beta T cells are being pursued gamma delta T cells are reasonable candidates for immunotherapy because they (i) possess intrinsic anti-tumorigenicity, (ii) require no priming, (iii) direct tumor killing via recognition of stress-responsive ligands, and (iv), as we now show, can be expanded to clinical cell doses in current Good Manufacturing Practice serum-free media (SFM). Methods. gamma delta T-cell expansion was evaluated in several SFMs. Additionally, the expanded gamma delta T cells were evaluated for their transduction efficiency using lentiviral vectors (LV). Results. Of the SFM cultures, robust expansion was only observed in OpTmizer supplemented with highdose interleukin-2. gamma delta T-cell percentages and numbers were sufficient for clinical use. Using cells from several donors, transduction efficiencies ranged from 13 to 33%, which is similar to transduction levels observed using alpha beta T cells with similar multiplicity of infection. Discussion. An optimized method of gamma delta T-cell expansion and transduction was developed that can be tested in early-phase clinical trials. With appropriate elimination of the alpha beta T cell-component, the absence of MHC-restriction affords the opportunity for use in the allogeneic setting with limited risk of graft versus host disease. Finally, the use of SFM provides clinically safer, widely applicable and potentially more efficacious cellular immunotherapy.