Tunable Universal OR-gated CAR T cells for AML

Abstract/SummaryAcute myeloid leukemia (AML) is a hematopoietic malignancy characterized by antigen heterogeneity and poor prognosis. A potential therapeutic approach to address this heterogeneity is targeting multiple surface antigens to prevent antigen escape and relapse. Chimeric antigen receptor (CAR) T cells are an adoptive cell therapy that have demonstrated remarkable clinical success in the treatment of B cell malignancies, and many efforts are underway to adapt them to myeloid malignancies. To tackle the heterogeneity of AML, logically targeting multiple antigens through an "A OR B" gated CAR circuit would be desirable. Here we combined FLT3 antigen targeting with the well characterized CD33 myeloid marker as a combinatorial OR gate approach using our split, universal, programmable (SUPRA) CAR platform. The split platform affords tunability over activation levels and multiplexed targeting that cannot be achieved through a tandem bispecific approach. We systematically characterized the specificity and sensitivity of different SUPRA CAR adapters against each target individually and in combination against a panel of target cell lines. Our results demonstrate that this CAR system can effectively target two antigens with equivalent efficacy to conventional CARs while reducing the engineering burden associated with designing CAR T cells against multiple antigens. Furthermore, we can characterize an effective dose range where off-target cytotoxicity against hematopoietic stem and progenitor cells is minimized. With the recent clinical advances in universal CAR designs, our SUPRA OR gate has the potential to provide an effective and safer solution to treating AML.