Cell-cycle arrest and induction of apoptosis in Kit-mutant cell lines following AP23464 treatment. (A, left) Juxtamembrane mutant HMC-1 and activation-loop mutant D816V Ba/F3 and P815 cell lines were analyzed for cell-cycle distribution by propidium iodide (PI) staining following 24-hour AP23464 treatment. Ba/F3 cells were used as a control. (Right) The percentages of cells in the G₀/G₁ (▦), S(▪), and G₂/M (□) phases were calculated by fitting flow cytometry data with ModFit software. Cells in sub-G₁ were not included in the calculation. Three independent assays were averaged to generate the graphs. Representative histograms are shown. For histograms, x axis indicates DNA content and y axis indicates cell count; for bar graphs, y axis indicates the percentage of cells. (B) Juxtamembrane mutant HMC-1 and activation-loop mutant D816V Ba/F3 and P815 m-D814Y cell lines were assessed for annexin-V binding and PI incorporation as a measure of apoptosis following 48-hour AP23464 treatment. Ba/F3 cells are shown as a control. Three independent assays were averaged to generate graphs. Representative dot plots are shown. Error bars depict standard deviation of the mean. FITC indicates fluorescein isothiocyanate.