Engineering caffeine-dependent protein switches to enable functional control of CAR Ts in vivo

One obstacle associated with CAR T cells is their limited controllability after administration to the patient, which becomes particularly problematic in the case of severe toxicities. To be able to control CAR T cell function in vivo, we generated caffeine-responsive molecular switches by using high-end protein engineering. We demonstrate that these switches are clean (i.e. no background activation in the absence of caffeine) and efficiently turned on upon addition of caffeine. Furthermore, integration of these switch modules into CARs yields switchable “CaffCARs”, whose function can be precisely controlled via administration of caffeine. We show that this switch platform is modular, enabling caffeine-dependent regulation of CARs directed against different antigens (including antigens for leukemia, lymphoma and solid tumors). Importantly, these CaffCAR T cells are efficiently activated in response to caffeine concentrations that are achieved in human serum after drinking one cup of coffee. Finally, when we tested our CaffCAR T cells in a lymphoma mouse model in vivo, we observed high anti-tumor potency that could be regulated efficiently via administration of caffeine to the mice. Together, we introduce caffeine-responsive switch modules that enable functional regulation of CAR T cells in vivo with caffeine concentrations that are achieved in human serum after drinking one cup of coffee. 

Learning Objectives:

1. Define why we need functional control of CAR T cells? 

2. Describe the engineering strategy of drug-responsive molecular switches

3. Discuss optimal strategies to incorporate such switches into CARs