TRAIL orchestrates ThINKK-induced NK cell cytotoxicity against childhood acute lymphoblastic leukemia

BackgroundTherapeutic Inducers of Natural Killer cell Killing (ThINKK) represent a novel class of immunotherapy designed to enhance the graft-versus-leukemia effect of hematopoietic stem cell transplantation in pediatric patients with high-risk or relapse leukemia. Our previous work identified high expression of TRAIL as a key signature of Natural Killer (NK) cell stimulation by ThINKK. In this study, we aim to elucidate the mechanisms underlying acute lymphoblastic leukemia (ALL) killing by ThINNK-stimulated NK cells and to identify predictive sensitivity markers of this innovative approach. MethodsWe performed NK cell cytotoxic assays using a panel of genetically diverse ALL cell lines and patients samples. Gene deletion and gene enforced expression in sensitive or resistant cell lines were performed to demonstrate the role of TRAIL-receptors expression and death receptor signaling pathway in ALL cell death induced by ThINKK-stimulated NK cells. These findings were further validated through the analysis of primary patients samples and transcriptomic profiling of a cohort of 320 ALL patients from the CHU Sainte-Justine. ResultsWe found that ALL sensitivity to ThINKK-stimulated NK cell killing was independent of their genetic background or their HLA expression. In addition, our data revealed the dual role of TRAIL: first, a strong NK cell activating receptor that induced rapid killing of ALL expressing TRAIL-R2, and second, a death-receptor ligand inducing ALL apoptosis following sustained engagement with its receptors. The transcriptomic analysis of ALL patients samples indicated that TRAIL-R2 and TRAIL-R1 are widely expressed across ALL subtypes and are not downregulated at relapse. ConclusionThese findings support the use of TRAIL receptor expression as a biomarker of sensitivity to ThINKK immunotherapy and establish a mechanistic framework to guide patient stratification and therapeutic optimization.