Defects in the intrinsic apoptosis pathway in CLL samples.

CLL cells from patients sensitive to fludarabine (n = 3) (panel A) or resistant to fludarabine (n = 8) (panel B) were analyzed with regard to caspase activation. Caspase-8 (12 nM) or cytochrome c (60 μM) were transduced into CLL cells with PE-BSA (4 μg/mL), in the presence or absence of the specific caspase-8 inhibitor CrmA (10 μM) or the specific caspase-9 inhibitor XIAP-BIR3 (20 nM) by electroporation (1200 V/cm and 900 μF). The control represents BSA-transduced cells. Effector caspase activity was measured by means of the fluorogenic cell-permeable substrate, CaspaTag, analysis of cells by flow cytometry, and gating on the viable cell population (7-AAD⁻) that had taken up protein (PE-BSA⁺). Data represent mean CaspaTag fluorescence (mean fluorescence intensity [MFI] ± SE).