The ultimate goal of immune-engineering is to harness the exquisite specificity of the immune system to prevent and treat disease. However, it is becoming increasingly clear that cellular metabolic programming is not merely a consequence of immune cell activation but rather an integral component of promoting differentiation and function. The generation of cells with robust effector function is intimately dependent upon the generation of nucleic acid, lipid and protein substrates, and energy. Likewise, persistence or the induction of immunologic “memory” is dependent upon metabolic reprogramming that fuels long term survival. In this context successful engineering of immune cells will entail modulation, programming and even reprogramming of energy and substrate-generating metabolic programs. Initially based on our studies dissecting the role of the mTOR pathway in regulating immune responses our lab has defined specific nodes/targets to regulate metabolic programming necessary to fuel effector function and long term persistence. TR&D 3 seeks to exploit these findings to target these critical nodes in order to engineer metabolic reprogramming of cells, thus maximizing function and persistence. To this end we will pursue the following Aims: Aim 1. Employing synthetic biology, genetically egineer cells with enhanced mTORC1 activity by knocking down/out/mutating TSC2, leading to enhanced effector function characterized by more robust glycolytic reprogramming. Deliverable: The creation of a reprogrammed “stock” effector cell that can then be modified for an array of cellular therapies. Aim 2. By regulating glutamine metabolism, formulate growth and differentiation conditions that promote the generation of cells epigenetically programmed to persist when adoptively transferred in vivo and to maximally respond upon rechallenge. Deliverable: Metabolically optimized media for producing robust and persistent effector cells. Aim 3. Based on novel findings regarding the ability of SGK1 to promote both a memory and effector T cell phenotype, develop small molecule inhibitors of SGK1 that can metabolically reprogram T cells both ex vivo and in vivo. Deliverable: Small molecule inhibitors to enhance efficacy of Adoptive Cellular Therapy.