Reversible ON- and OFF-switch chimeric antigen receptors controlled by lenalidomide

Two chemical genetic control systems were engineered to regulate CAR T cell function with lenalidomide. Flipping the switch on T cells Cell-based treatments such as chimeric antigen receptor (CAR) T cells have been finding increasing applications against cancer and other disorders. However, these treatments can have side effects, and it is difficult to stop the activity of an overactive cell-based therapy once it is inside a patient. To improve regulation of CAR T cell treatment, Jan et al. developed a switch-based system for activating and inactivating these cells. T cells with an OFF switch were deactivated using the drug lenalidomide, whereas those with an ON switch required both lenalidomide and a target antigen for activation, allowing control over the timing of T cell activation. Cell-based therapies are emerging as effective agents against cancer and other diseases. As autonomous “living drugs,” these therapies lack precise control. Chimeric antigen receptor (CAR) T cells effectively target hematologic malignancies but can proliferate rapidly and cause toxicity. We developed ON and OFF switches for CAR T cells using the clinically approved drug lenalidomide, which mediates the proteasomal degradation of several target proteins by inducing interactions between the CRL4CRBN E3 ubiquitin ligase and a C2H2 zinc finger degron motif. We performed a systematic screen to identify “super-degron” tags with enhanced sensitivity to lenalidomide-induced degradation and used these degradable tags to generate OFF-switch degradable CARs. To create an ON switch, we engineered a lenalidomide-inducible dimerization system and developed split CARs that required both lenalidomide and target antigen for activation. Subtherapeutic lenalidomide concentrations controlled the effector functions of ON- and OFF-switch CAR T cells. In vivo, ON-switch split CARs demonstrated lenalidomide-dependent antitumor activity, and OFF-switch degradable CARs were depleted by drug treatment to limit inflammatory cytokine production while retaining antitumor efficacy. Together, the data showed that these lenalidomide-gated switches are rapid, reversible, and clinically suitable systems to control transgene function in diverse gene- and cell-based therapies.