Inhibition of FLT3/ITD kinase activity induces apoptosis in FLT3/ITD-32D cells. (A) Inhibitory effects of AG1296 on the viability of parental 32D, WtFLT3-32D, and FLT3/ITD-32D cells. Cells were cultured with medium in the presence or absence of IL-3 (1.0 ng/mL) or FL (50 ng/mL). AG1296 was added to the culture medium at the indicated concentrations, and viable cells were counted by the trypan blue dye–exclusion assay after 48-hour culture. (B) Dephosphorylation of FLT3 and STAT5a in FLT3/ITD-32D cells treated with AG1296. Cells were treated with AG1296 at 0.3, 1.0, and 3.0 μ for 4 hours. Each cell lysate was immunoprecipitated by anti-FLT3 antibody or anti-STAT5a antibody, and the immunocomplex was analyzed by immunoblotting with the indicated antibodies. (C) Effect of IL-3 on FLT3/ITD-32D treated with AG1296. IL-3 (1.0 ng/mL) and AG1296 (1.0 μM) were used. The data shown are means and SDs of 3 independent experiments.