The effect of CD123-CAR–expressing T cells on normal and leukemic progenitor cells in vitro. (A-B) CD34⁺ CB cells (n = 3) were CD34-immunomagnetically selected and cocultured with either CD19- or CD123-specific pair-matched T cells from a healthy donor or media alone (untreated) for 4 hours at an E:T of 25:1. The cells were then plated in semisolid methylcellulose progenitor culture for 14 to 18 days and scored for the presence of BFU-E (A), CFU-GM (B) and colonies. Colony numbers (left) and normalized colony formation percentages (right) are presented. Percentages are normalized to CD19-specific T cell controls. Data represent mean values + SEM for 3 different CB samples. (C) CD34⁺ primary AML patient samples (AML 493, 519, or 545) were immunomagnetically selected and cocultured with either CD19 or CD123-specific CAR T cells from a healthy donor or media alone (untreated) for 4 hours at an E:T of 25:1. The cells were then plated in semisolid methylcellulose progenitor culture for 14 to 18 days and scored for the presence of leukemia colony-forming units (CFU-L). Colony numbers (left) and normalized colony formation percentages (right) are presented. Percentages are normalized to CD19-specific T cell controls. Data represent mean values + SEM for 3 different primary AML patient samples. *P < .05 using the unpaired Student t test comparing 26292 and 32716 to CD19R.