CAR-EBD: Modular Polypeptide Devices for the Small Molecule Regulation of Chimeric Antigen Receptor T cell Activity

Abstract

The advent of chimeric antigen receptor (CAR) T cell therapies holds immense promise for the future of cancer treatment. However, the vast majority of current CAR T cell therapies are self-guided, relying on endogenous programming to interpret novel receptor signaling, which may lead to poorly regulated and undesirable functional outputs ranging from cytokine release syndrome and neurotoxicity to incomplete tumor control and immunological exhaustion. These failures at both ends of the functional spectrum (hyperfunction and hypofunction, respectively) are the result of an incomplete mastering of the biological underpinnings of T cell activity and differentiation, combined with the inherent complexity of treating tumors with a patient-derived product – each with their own unique genetic background, immunological history, and target-tumor state. I aim to address these limitations by modifying CAR T cells so their activity may be directly regulated through the use of small molecule drugs. I have pursued several design iterations to accomplish this goal. Regulation is most successfully achieved by genetic fusion of a CAR to an estrogen-analog binding protein domain (EBD) to create CAR-EBD. These EBD regulators are small in size, highly drug-sensitive, exhibit rapid switching kinetics, are transferable between different CARs and other proteins of interest, function in vivo, and may be engineered to be selectively responsive to different estrogen analogs to provide orthogonal regulation of multiple transgenes. Having successfully demonstrated the advantageous traits of this system, there are many future applications for this technology.