Distinct Bioenergetic Features of Human Invariant Natural Killer T Cells Enable Retained Functions in Nutrient-Deprived States

Invariant natural killer T (iNKT) cells comprise a unique subset of lymphocytes that are primed for activation and possess innate NK-like functional features. Currently, iNKT cell-based immunotherapies remain in early clinical stages, and little is known about the ability of these cells to survive and retain effector functions within the solid tumor microenvironment (TME) long-term. In conventional T cells (T(CONV)), cellular metabolism is linked to effector functions and their ability to adapt to the nutrient-poor TME. In contrast, the bioenergetic requirements of iNKT cells – particularly those of human iNKT cells – at baseline and upon stimulation are not well understood; neither is how these requirements affect effector functions such as production of cytokines and cytolytic proteins. We find that unlike T(CONV), human iNKT cells are not dependent upon glucose or glutamine for these effector functions upon stimulation with anti-CD3 and anti-CD28. Additionally, transcriptional profiling revealed that stimulated human iNKT cells are less glycolytic than T(CONV) and display higher expression of fatty acid oxidation (FAO) and adenosine monophosphate-activated protein kinase (AMPK) pathway genes. Furthermore, stimulated iNKT cells displayed higher mitochondrial mass and membrane potential relative to T(CONV). Real-time Seahorse metabolic flux analysis revealed that stimulated human iNKT cells utilize fatty acids as substrates for oxidation more than stimulated T(CONV.) Together, our data suggest that human iNKT cells possess different bioenergetic requirements from T(CONV) and display a more oxidative metabolic program relative to effector T(CONV). Importantly, iNKT cell-based immunotherapeutic strategies could co-opt such unique features of iNKT cells to improve their efficacy and longevity of anti-tumor responses.