Transient mRNA-based CD117 CAR T cells effectively target acute myeloid leukemia in vitro for potential use as a preconditioning strategy

Background: Acute myeloid leukemia (AML) is a high-risk malignancy, particularly in patients with primary induction failure or relapsed/refractory disease. CD117 (c-Kit), expressed on both leukemic blasts and normal hematopoietic stem and progenitor cells (HSPCs), represents a potential therapeutic target but poses challenges due to the risk of severe myelotoxicity. Materials and methods: -transcribed (IVT) messenger RNA (mRNA) electroporation. The mRNA-based CD117 CAR T cells were evaluated for viability, immunophenotype, cytotoxic activity, and toxicity toward primary HSPCs using clonogenic assays, and compared with retroviral-based counterparts. Results: CD117 was expressed in AML patient samples and cell lines at varying levels. CD117 CAR T cells demonstrated potent and specific cytotoxicity against AML cells. The mRNA-based CAR T cells exhibited high transfection efficiency, good viability, and an immunophenotype similar to non-transduced T cells, and were functionally competent as early as 2 h post-electroporation. In long-term co-culture with a high tumor burden, repeated dosing of mRNA CAR T cells effectively eliminated CD117+ cells, comparable to viral vector-based CAR T cells. Notably, residual mRNA CAR T cells following AML clearance showed no detectable CAR expression and preserved HSPC colony-forming capacity. Conclusions: studies suggest the potential use of mRNA CD117 CAR T cells as a non-genotoxic preconditioning strategy for patients with high-risk or refractory AML.