Figure 3. Inhibitory effects of gilteritinib and quizartinib on FLT3^wt and FLT3-ITD.

(A and B) HEK293 cells were transfected with an FLT3^wt- or FLT3-ITD-expressing vector. After culturing overnight, cells were treated with gilteritinib or quizartinib for 1 hour. FL (25 ng/mL) was added to FLT3^wt-expressing cells simultaneously with the compounds. Cells were lysed and subjected to western blotting analysis using antibodies against total or phospho-FLT3 (Y591). The ratio of phospho/total FLT3 was calculated and is shown in the graph. (C) FLT3^wt- or FLT3-ITD-expressing Ba/F3 cells and parental cells were subjected to western blotting analysis using antibodies against FLT3, phospho-FLT3 and β-actin. (D) FLT3^wt- or FLT3-ITD-expressing Ba/F3 cells were treated with gilteritinib, quizartinib or midostaurin for 3 days. FL (25 ng/mL) was added to FLT3^wt-expressing cells simultaneously with the compounds. Cell viability was measured using the CellTiter-Glo 2.0 Assay. Three experiments were performed in triplicate, and representative data are shown as the mean ± SD. Geometric mean IC₅₀ values and 95% CIs were determined from 3 independent experiments. Abbreviations: CI, confidence interval; FL, FLT3 ligand; ITD, internal tandem duplication; wt, wild-type.