Abstract 3257: CTLA-4 blockade drives loss of regulatory T cell functional stability in glycolysis defective tumors

Only a limited fraction of patients derives durable clinical benefit upon immune checkpoint blockade. Deepening our understanding of mechanisms of response and resistance to these therapies is thus needed to optimize their employment in rational combinations. Cellular energy metabolism reprogramming is a critical hallmark of cancer. High glucose consumption and lactate production by tumor cells restrict nutrient availability in the tumor microenvironment (TME) for effector T cells, which also rely on glycolysis to replicate and function. In addition, immune checkpoints and co-stimulatory molecules are emerging as important regulators of T cell metabolism. Exploiting the capacity of immune checkpoint blockade to perturb T cell metabolism in combination with inhibition of tumor glycolysis may thus be a rational and more effective anti-cancer approach. We investigated the link between tumor glycolysis and immune cell function using RNA sequencing data sets from patients treated with checkpoint blockade and in tumor:T cell co-culture systems. We then studied the effects of checkpoint blockade in syngeneic glycolysis-defective murine tumor models and explored mechanisms underlying anti-tumor activity. We found that expression of glycolysis-related genes is inversely correlated with infiltration of most immune cell types in melanomas from patients before CTLA-4 blockade. However, after treatment, immune-related and glycolysis-related genes were more often co-expressed, suggesting that anti-CTLA-4 may partially restore immune cell fitness in the glycolytic TME. To directly assess the effect of tumor metabolism on T cell function, we co-cultured activated T cells with the highly glycolytic murine mammary carcinoma 4T1 and observed that tumor cells, or similar non-toxic concentrations of exogenous lactate, significantly limited T cell activation and viability. We thus investigated whether inhibition of tumor glycolysis could potentiate the activity of CTLA-4 blockade. We found that neoadjuvant treatment with anti-CTLA-4 significantly prolonged survival in mice bearing glycolysis defective 4T1 tumors, where lactate dehydrogenase A (LDH-A) - the critical enzyme controlling lactate production in aerobic glycolysis - was knocked down (4T1-KD). Intriguingly, tumor protection was associated with intratumoral regulatory T cell (Treg) functional phenotypic destabilization towards IFN-gamma and TNF-alpha producing Tregs. By mimicking the LDH-A-KD and control 4T1 TME in vitr