An immunosuppressive circuit between HSPC-like cells and cytotoxic lymphocytes in younger patients. (A) UMAP of scRNA-Seq data for normal immune cells in 18 KMT2A-r patients. Number of sequenced cells is indicated. (B) NK T cells are the major source of IFNG in patients. Shown are violin plots of IFNG expression in various immune cell populations in both KMT2A-r patients and healthy donors. HD, healthy donor; PT, patient. (C) Upregulated IFNG signaling in the HSPC-like population in younger patients. Left, barplot showing younger patients have a higher fraction of NK T cells expressing IFNG. Right, barplot showing IFGN receptor 2 gene (IFNGR2) was expressed higher in HPSC-like cells in the PB of younger infants with ALL compared with normal BM HSC/MPPs. (D) Activating and suppressive signaling pathways for NK cell cytotoxicity induced by IFNG signaling in blasts. Y-axis, potential for inducing NK cell cytotoxicity based on combined normalized expression of genes in activating and suppressive pathways in HSPC-like cells, CD19⁺ blasts, and normal HSC/MPPs. (E) Schematic overview of blast and NK cell coculture experiment. (F) Percentage of HSPC-like cells and CD19⁺ blasts killed by NK cells. Dots represent different patient samples. P values were computed using Student t-test. (G) Predicted L-R interactions between the leukemia cell containing HSPC-like population (left panel) or CD19⁺ blasts (right panel) and 2 major classes of cytotoxic lymphocytes, CD8⁺ T cells and NK cells. Red, blue, and gray arcs, suppressive, activating, and unknown interactions, respectively. (H) Cytotoxicity scores of NK and CD8⁺ T cells based on combined activating and suppressive signaling between the cytotoxic lymphocyte and the HSPC-like/blast populations via L-R pairs (see supplemental Methods for details). L-R pairs were based on those in panel E.