CAREER: Understanding the Impact of Dephosphorylation Kinetics and Adapter Specificity on Synthetic T Cell Receptor Signaling and Function

T cells are created in bone marrow and mature in the thymus. They defend against a variety of diseases, including cancer. Some cancers disguise themselves from T-cell recognition. Others suppress the killing response of T-cells. T cells can be modified with chimeric antigen receptors (CARs). These are synthetic molecules that provide T cells with a renewed ability to sense and destroy cancer cells. CAR engineered T cells are clinically effective at fighting some blood cancers. Unfortunately, our understanding of parameters that tune how a T cell responds to a cancer cell is severely lacking. This project aims to gain a fundamental understanding of the parameters of CARs that tune T cell responses, and to characterize how these parameters affect CAR engineered T cell function and fate. This research program is integrated with an education plan that aims to support students in the community college system in the Tampa Bay area, providing mentorship and guided research experiences for students as they navigate the STEM pathway. Chimeric antigen receptors (CARs) are synthetic T cell receptors that mimic T cell receptor activation and co-stimulation. These empirically designed receptors provide a potent but unoptimized signaling response. As a result, engineered T cells are susceptible to overactivation or suppression, two common but distinct challenges to cell therapy. The influence of important parameters such as dephosphorylation kinetics and downstream adapter preferences on CAR signaling remains essentially unstudied. Optimizing receptor function to support robust T cell activation remains a trial-and-error exercise. This project will combine protein engineering, cell biology, and omics experiments to establish a fundamental understanding of how the signaling components of CARs can be tuned to yield improved responses under activating and suppressive conditions. These experiments will provide fundamental advances through 1) surveying motif landscapes for phosphorylation, dephosphorylation, and adapter binding in synthetic T cell activation; 2) quantitatively understanding how dephosphorylation kinetics influence CAR signaling; 3) mapping synthetic signaling in T cells with adapter bias. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.