Preclinical evaluation of ex vivo expanded/activated gamma delta T cells for immunotherapy of glioblastoma multiforme

We have previously shown that expanded/activated gamma delta T cells from healthy donors are cytotoxic to GBM cell lines and primary GBM explants. In this report, we examined the therapeutic effect of intracranial infusion of expanded/activated gamma delta T cells on human minimal and established U251 tumor xenografts in athymic nude mice. Immunohistochemistry was used to determine the presence of NKG2D ligands on cell lines and tumors, and blocking studies were used to determine the effect of these ligands on gamma delta T cell recognition. Expanded/activated gamma delta T cells were prepared by 18-day culture in RPMI, human serum (HS), anti-CD2, IL-12, IFN-gamma, and OKT-3. Anti-GBM activity of the cell product was assessed using in vitro cytotoxicity assays against the GBM cell line U251MG in suspension and in adherent culture. Ex vivo expanded/activated gamma delta T cells were of the effector/memory phenotype, expressed Th1 cytokines, and effectively killed U251 cells in vitro. Xenografts were prepared using a U251 cell line following transfection with a firefly luciferase gene to monitor tumor progression. Mice treated with gamma delta T cells showed slower progression of both new and established GBM xenografts versus mice that received vehicle only as determined by photon emission over time. Median survival was improved in all gamma delta T cell treated groups between 32 and 50 days by Kaplan-Meier analysis. U251 cells expressed ULBP-2 and ULBP-3, although blocking of these reduced in vitro cytotoxicity of gamma delta T cells to U251MG by only 33 and 25%, respectively. These studies show that expanded/activated gamma delta T cells can mediate killing of new or established GBM xenografts, reduce tumor progression, and constitute a potentially effective novel immunotherapeutic strategy against GBM.