UC San Diego

Chimeric antigen receptor (CAR)-T cell therapies have adverse side effects such as cytokine release syndrome (CRS) due to on-target off-tumor toxicity. Responses for clinically available CAR-T therapies mainly use immunosuppressants for mitigation which lowers their overall functionality. An alternate approach will be to bioengineer cellular regulation that confers high spatiotemporal control over CAR-T cells in vivo to reduce its side effects. The lab has previously developed a protein caging system as a switch for controlling the signalling of CAR. The caging domain binds to CAR and keeps it auto inhibited. The uncaging domain which is under an external control can bind to the caging domain and free the CAR to initiate intracellular signaling. We have tested two strategies for externally controlling the uncaging of CAR by creating a dihydrofolate reductase (DHFR) system and a heat shock promoter (HSP) system. We have developed an in-vitro dimerization dependent fluorescent (ddFP) biosensor that can detect dynamics of the above protein caging system by verifying the expression and binding of the caging and uncaging domains. The results can be used to further optimize the protein caging system and create focused ultrasound (FUS) controllable CAR-T cells.