PROJECT SUMMARY/ABSTRACT Anti-CTLA-4 was first immune checkpoint blockade therapy approved by the FDA to treat patients with cancer. Notwithstanding its impressive clinical response in ~20% of patients with advanced melanoma, there is a clear need to improve its efficacy. We recently showed that loss of IFN- signaling genes could partially explain why the remaining ~80% patients are resistance to anti-CTLA-4 therapy. Therefore, identifying novel therapeutic targets/approaches capable of overcoming resistance and boosting efficacy of anti-CTLA-4 is of utmost importance We and others previously demonstrated that the HIF1-glycolysis axis orchestrates an essential “metabolic checkpoint” in driving effector T cell differentiation and in suppressing regulatory T cell induction, two functional outcomes closely associated with anti-CTLA-4 efficacy, suggesting activating HIF1 and glycolysis in T cells could represent an effective approach to improve therapeutic efficacy of anti-CTLA-4. To this end, we have generated interesting preliminary data showing that anti-CTLA-4 selectively upregulates HIF1 and its downstream targets, glycolytic enzymes in tumor-infiltrating T cells (TILs). And, specific deletion of HIF1 in T cells and simultaneous inhibition of glycolysis largely abolishes the therapeutic effects of anti-CTLA-4, pointing to a critical role of HIF1 and glycolysis in this therapy. Therefore, we propose that HIF1, via modulating glycolytic activity of TILs, dictates therapeutic efficacy of anti-CTLA-4, which can serve as novel therapeutic targets to promote efficacy of anti-CTLA-4. In Aim 1, we will characterize how T cell-intrinsic HIF1 governs glycolytic activity and effector functions of TILs in response to anti-CTLA-4. In Aim 2, we will target the HIF1- glycolysis pathway to promote therapeutic efficacy of anti-CTLA-4. While it is well-known that anti-CTLA-4 markedly rejuvenates effector functions of TILs, the specific underlying mechanisms are largely unknown. Insights gained from these studies will significantly advance our understanding of how anti-CTLA-4 metabolically augments anti-tumor responses and manifest novel therapeutic opportunities (e.g., targeting metabolic pathways) to improve efficacy of anti-CTLA-4.