Multi-targeted, NOT gated CAR-T cells as a strategy to protect normal lineages for blood cancer therapy.

Introduction: Despite advances in treatment of blood cancers, several-including acute myeloid leukemia (AML)-continue to be recalcitrant. Cell therapies based on chimeric antigen receptors (CARs) have emerged as promising approaches for blood cancers. However, current CAR-T treatments suffer from on-target, off-tumor toxicity, because most familiar blood cancer targets are also expressed in normal lineages. In addition, they face the common problem of relapse due to target-antigen loss. Cell therapeutics engineered to integrate more than one signal, often called logic-gated cells, can in principle achieve greater selectivity for tumors. Methods: We applied such a technology, a NOT gated system called Tmod™ that is being developed to treat solid-tumor patients, to the problem of therapeutic selectivity for blood cancer cells. Results: Here we show that Tmod cells can be designed to target 2-4 antigens to provide different practical and conceptual options for a blood cancer therapy: (i) mono- and bispecific activating receptors that target CD33, a well-known AML antigen expressed on the majority of AML tumors (as well as healthy myeloid cells) and CD43 (SPN), an antigen expressed on many hematopoietic cancers (and normal blood lineages); and (ii) mono- and bispecific inhibitory receptors that target CD16b (FCGR3B) and CLEC9A, antigens expressed on key normal blood cells but not on most blood cancers. Discussion: These results further demonstrate the robust modularity of the Tmod system and generalize the Tmod approach beyond solid tumors.