SCH66336 sensitizes parental and STI571-resistant cells to STI571-induced apoptosis.

(A-E) Parental Baf/BCR-ABL-s (A) and STI571-resistant Baf/BCR-ABL-r (B), LAMA-84-r (C), AR230-r (D) cells, and Baf3 cells expressing the T315I mutant of BCR-ABL (E) were seeded at 5 × 10⁴cells/mL in cytokine-free RPMI + 10% fetal bovine serum in the presence of DMSO (●), 1 μM STI571 (▴), 5 μM SCH66 336 (▪), or a combination of both drugs (○). Viability of cells was assessed at daily intervals by dye exclusion. Results are representative of at least 3 experiments. (F, G) Parental Baf/BCR-ABL-s cells were pretreated with DMSO or 1 μM SCH66336 for 48 hours, after which cells were split into 2 groups—no further treatment (control and SCH66336 cells) or treatment for an additional 8 hours with 1 μM STI571 (STI571 and combination cells). Cells were analyzed for either annexin V staining (F) by fluorescence-activated cell sorter analysis using the Apo-Alert kit (Clontech), or they were lysed and immunoblotted with antibody against caspase-3 (G). Densitometric analysis was performed on active caspase-3 and was normalized to levels of the indicated background band (open arrowhead).