ATRA reduces colony-forming capacity and in vivo leukemic engraftment of EVI-1–positive cells. Number of colonies generated from EVI-1–positive (A) and EVI-1–negative (B) AML cells after 7 days of treatment with the indicated concentrations of ATRA. Bars represent the average numbers of colonies from a duplicate, and error bars indicate the standard deviation. AML11 to 14 did not show colonies. (C) Flow cytometric analysis (upper panels) of cells treated with 1 µM ATRA (4 days), 0.5 µM doxorubicin (3 days), or the combination (pretreatment with ATRA) and the quantification (lower panels) of myeloid (CD33⁺/CD45^dim/CD3⁻) (blue) and lymphoid (CD33⁻/CD45^high/CD3⁺) (purple) cells, which were negative for 7-AAD. (D-E) Quantification of human myeloid cells (hCD45^dim/hCD33⁺) from BM and spleen of NSG mice injected with primary AML cells and treated with ATRA or placebo. (F) Quantitative reverse-transcription polymerase chain reaction analysis of EVI-1 expression upon 5 days of ATRA treatment of the AML cell lines NB4 and HL60 (left) and upon 7 days of ATRA treatment of EVI-1–positive AML cases (right).