Pharmacokinetic and pharmacodynamic effects of KW-2449 in FLT3-activated leukemia. (A) MOLM-13 cells (107 cells/mouse) were subcutaneously inoculated into 2 SCID mice. Ten days after inoculation, KW-2449 at 20 mg/kg was orally administered twice every 12 hours. The concentrations of KW-2449 in plasma and tumor were sequentially analyzed by LC/MS/MS after the final administration of KW-2449. (B) The transition of FLT3 and STAT5 phosphorylation levels in tumor were analyzed by Western blotting. For analysis of FLT3 and its phosphorylated form (P-FLT3), the blots of immunoprecipitated FLT3 SDS-PAGE samples were analyzed by antiphospho-tyrosine (P-Tyr) antibody and anti-FLT3 antibody. For detection of STAT5 and its phosphorylated form (P-STAT5), total tumor lysate SDS-PAGE samples were analyzed by anti–P-STAT5 antibody and anti-STAT5 antibody as primary antibodies. Dephosphorylations of FLT3 and STAT5 in MOLM-13 were observed until 12 hours after the last administration. Results from 2 mice at each point are shown.
