Coexpression of GPR56 and C3AR marks NPM1-mutated AML cells with leukemia-initiating capacity. (A) Protein coexpression of C3AR and GPR56 in 3 NPM1-mutated AML samples. Marker stains in red and isotype in gray. (B) Schematic illustration showing the experimental setup for the C3AR/GPR56 transplant experiments; the myeloid-enriched (CD3⁻CD19⁻) cells expressing C3AR from a NPM1-mutated AML case were sorted based on GPR56 expression and transplanted into immunodeficient NSGS mice, which were sacrificed at 19 weeks. (C) Leukemic engraftment in the BM of mice that had received transplantation with C3AR⁺ GPR56⁺ and C3AR⁺ GPR56⁻ cells, as the percentage of BM cells positive for human CD45 and CD33. Line at median; ∗P ≤ .05 using a nonparametric Mann-Whitney U test.