Response of primary CML progenitor cells to treatment duration with dasatinib. Progenitor cells isolated from 3 chronic-phase CML patients (CML 1-3) and a blast-crisis CML patient with 100% Bcr-Abl^T315I (CML R) were cultured in 100 nM dasatinib with (+) or without (−) washout at 4 hours, as indicated, or in the absence of dasatinib as an untreated control. K562 cells were treated identically to primary cells for comparison (gray bars, mean ± SEM from 2 independent experiments). (A) Percentage annexin V⁺ cells at 72 hours. Drug-specific apoptosis was calculated by subtracting the percentage annexin⁺ cells in the untreated arm. (B) Proliferation over 72 hours calculated as the fold expansion of viable cells during the 72-hour culture period, expressed as a percentage of untreated control. For K562, mean ± SEM of 2 independent experiments is reported. (C) Bcr-Abl activity at 24 hours, as measured by total phosphotyrosine level per cell and expressed as a percentage of the level in untreated cells. MFI indicates mean fluorescence intensity. (D) Proliferation of granulocyte/macrophage colony-forming cells. The number of cells at 72 hours that could lead to colony formation were enumerated and expressed as a percentage of untreated control. Colony-forming cell number was assayed in triplicate and mean ± SEM reported. (E) Bcr-Abl activity at 24 hours in CML 3 as assayed by CrkL phosphorylation. Western blot of total CrkL (bottom panel) showing phosphorylated (top band) and unphosphorylated (bottom band) forms. Top and bottom bands were quantified and the percentage phosphorylated plotted. (B-E) Dotted line represents the value for the untreated control.