Induction of acute leukemias following transplantation of human CD34⁺ cells containing knock-in MLL oncogenes. (A) Experimental scheme depicts nucleofection of CD34⁺ cells and their subsequent transplantation directly into sublethally irradiated NSG recipient mice or culture for 3 weeks in vitro prior to transplantation. (B) Kaplan-Meier plot is shown for cohorts of mice transplanted with CD34⁺ cells transfected with templates plus TALENs (n [AF9] = 25: direct inject = 19, cultured cells = 6; n [ENL2] = 7: direct inject = 5, cultured cells = 2; n [ENL7] = 8: direct inject = 5, cultured cells = 3) or template alone (control, n = 7). P < .05 was considered statistically significant. Mice were sacrificed upon signs of illness. (C) Flow cytometry profiles show representative phenotypes of various leukemias that developed in mice transplanted with CD34⁺ cells (n = 17) compared with blasts from patients with MLL translocations (AML/ALL), which display comparable phenotypes. Also shown are representative profiles of BM cells from control mice (n = 2) that received CD34⁺ cells nucleofected with knock-in construct alone (week 16 posttransplantation) and representative analysis of knock-in cells 3 weeks after cell culture prior to transplantation. l, lymphoid; m, myeloid; (Pat.), patient.