There is a dire need for new treatment approaches to glioblastoma (GBM), a uniformly lethal brain cancer. We previously reported that targeting the lipid kinase Diacylglycerol kinase-α (DGKα) has therapeutic potential against GBM; we demonstrated direct actions on cancer cells and on angiogenesis, while others have identified a role for DGKα in T cell reactivity. In addition, we identified an abandoned medication safe in humans, ritanserin, as a novel DGKα inhibitor that can be repurposed to accelerate clinical translation of this therapeutic strategy. Our new preliminary data indicate in an immunocompetent mouse model of GBM that intermittent low dosing of ritanserin is dramatically more effective than similar cumulative doses done in a near- continuous fashion. Furthermore, other new data suggest a novel immunologic mechanism for DGKα inhibition involving bosting macrophage and microglia function, and also show synergistic activity in combining a DGKα inhibitor and anti-PD-1 checkpoint inhibition or temozolomide in mouse GBM models. This revised proposal focuses on developing these novel findings with three Specific Aims. The first Aim investigates the effects of DGKα inhibitors on macrophages and microglia, testing a putative mechanism as well. It also tests the hypothesis that the anti-mesenchymal activity of DGKα inhibition will reduce expression of immunosuppressive proteins in GBM cells. Aim 2 will dissect in syngeneic mouse models of GBM the immunologic and other effects of intermittent low dosing of DGKα inhibition, identifying mechanisms for its curative potential in mouse GBM and comparing treatment schedules. While our new data show striking efficacy of intermittent ritanserin in a mouse GBM model, we anticipate that combination therapy will be necessary to effectively treat GBM in patients; therefore Aim 3 tests combinations of intermittent ritanserin with other agents—anti-PD1 checkpoint inhibition and temozolomide—based on strong rationales and preliminary data. The most effective combination will be advanced for clinical investigation. Successful completion of the proposed studies will establish critical therapeutic and mechanistic aspects of DGKα inhibition, alone and in combination, helping to place it in the GBM armamentarium but with broader implications for oncology and other areas as well.