Fig. 3: Efficacy of gilteritinib against first- or second-generation ALK-TKI–resistant single mutants.

a IC₅₀ calculated from the viability analysis of Ba/F3 cells carrying single mutations conferring resistance to first-generation or second-generation ALK-TKIs. Cells were treated with lorlatinib, alectinib, or gilteritinib for 72 h. N = 3 independent samples examined over three independent experiments and data presented as mean values ± SD. b The suppression of phospho-ALK in Ba/F3 cells carrying single mutations conferring resistance to first-generation or second-generation ALK-TKIs was evaluated via western blotting. Cells were treated with the indicated concentrations of gilteritinib for 3 h (n = 2). c The suppression of phospho-ALK and its downstream signals in MCC-003 cells harboring EML4-ALK I1171N was evaluated via western blotting. Cells were treated with the indicated concentrations of drugs for 6 h (n = 2). d Flow cytometric analysis of apoptosis using Annexin-V and propidium iodide staining after 72 h of treatment with 100 nM lorlatinib or gilteritinib in MCC-003 cells. The percentage of cells undergoing apoptosis is shown in red. e MCC-003 cells were subcutaneously transplanted into BALB/c nu/nu mice. When the average tumor volume reached ~150 mm³, the mice were randomized to treatment with vehicle control, alectinib (30 mg/kg), or gilteritinib (30 mg/kg) treatment group once daily for 5 days/week via oral gavage. Tumor volumes were measured three times a week (n = 8 per treatment group). Data are presented as mean values ± SD. The significance of differences on day 14 was calculated using the two-sided Mann–Whitney U test (P value: Vehicle vs. gilteritinib treatment group, 0.0002; Alectinib vs. gilteritinib treatment group, 0.0002). f Phospho-ALK and its downstream signals in MCC-003 tumor samples obtained from vehicle-, alectinib-, or gilteritinib-treated mice were evaluated via western blotting (n = 2).