Orally administered PD166326 rapidly reaches concentrations sufficient to inhibit Bcr/Abl tyrosine phosphorylation in vivo. (A) PD166326 was administered to Balb/c mice at the indicated doses (▪, 25 mg/kg; ▵, 35 mg/kg; •, 50 mg/kg; □, 60 mg/kg) twice a day by oral gavage, and plasma PD166326 concentrations were determined at 2, 8, and 15 hours after the last dose at steady state. The error bars indicate the standard error from 3 mice at each dose level. (B) Peripheral blood leukemia cells from mice with the CML-like disease were analyzed by anti-phosphotyrosine (ptyr) or anti-Abl Western immunoblot after a single dose of PD166326 (50 mg/kg), imatinib mesylate (100 mg/kg), or placebo. The ratio of tyrosine-phosphorylated P210 (pB-a) to P210 (B-a) signal was determined by densitometry and is shown above each lane. Ba/F3 cells expressing P210 (+) or not (-) are shown as controls at far left. Anti-actin immunoblot serves as a loading control. Note that the level of P210 expression is not necessarily identical in all animals.³³