Drug-regulated CD33-targeted CAR T cells control AML using clinically optimized rapamycin dosing
Chimeric antigen receptor designs that incorporate pharmacologic control are desirable, however designs suitable for clinical translation are needed. We designed a fully human, rapamycin regulated, drug product for targeting CD33+ tumors called dimerization agent regulated immunoreceptor complex (DARIC33). T cell products demonstrated target specific and rapamycin dependent cytokine release, transcriptional responses, cytotoxicity, and in vivo antileukemic activity in the presence of as little as 1nM rapamycin. Rapamycin withdrawal paused DARIC33-stimulated T cell effector functions, which were restored following re-exposure to rapamycin, demonstrating reversible effector function control. While rapamycin-regulated DARIC33 T cells were highly sensitive to target antigen, CD34+ stem cell colony forming capacity was not impacted. We benchmarked DARIC33 potency relative to CD19 CAR T cells to estimate a T cell dose for clinical testing. In addition, we integrated in vitro and preclinical in vivo drug concentration thresholds for OFF-ON state transitions, as well as murine and human rapamycin pharmacokinetics, to estimate a clinically applicable rapamycin dosing schedule. A phase 1 DARIC33 trial has been initiated (PLAT-08, NCT05105152), with initial evidence of rapamycin-regulated T cell activation and anti-tumor impact. Our findings provide evidence that the DARIC platform exhibits sensitive regulation and potency needed for clinical application to other important immunotherapy targets. Overall design: DARIC33 cells and untransduced T cells derived from n = 4 healthy donors were incubated with 1nM rapamycin or media alone prior to culture alone or with CD33+ MV4-11 AML target cells. Following coculture, CD4+ and CD8+ cells were sorted and evaluated by RNA-Seq. This experiment compares transcriptomes of untransduced and DARIC33 transduced CD4+ and CD8+ T cells that were derived from four donors after 24hr incubation across two variables: with or without 1nM rapamycin and with or without activation by CD33+ MV4-11 AML target cells.  In addition, transcriptomes were assessed at the 168hr timepoint for CD4+ and CD8+ DARIC33 transduced T cells derived from 3 donors and incubated with 1nM rapamycin and activated by CD33+ MV4-11 AML target cells.