Anticancer drugs mediate caspase-dependent apoptosis in the absence of CD95 signaling. (A) CD95-susceptible Jurkat or CD95-resistant Jurkat-R cells were pretreated with medium (▪ and ▧) or 100 μmol/L zVAD-fmk () for 1 hour, and subsequently stimulated with anti-CD95 (20 ng/mL), daunorubicin (Daun; 5 μg/mL), doxorubicin (Doxo; 1 μg/mL), etoposide (Etop; 25 μg/mL), or mitomycin C (Mito; 25 μg/mL). After 24 hours, induction of apoptosis was measured by propidium iodide staining of hypodiploid nuclei (upper panel) and cell death was determined by the uptake of propidium iodide in dead cells (lower panel). (B and C) stimulated were 1 × 10⁶ Jurkat or Jurkat-R cells for the indicated time with (B) daunorubicin (5 μg/mL) or with (C) different concentrations of etoposide (25 μg/mL lanes 4,8; 12.5 μg/mL lanes 3,7; 6.25 μg/mL lanes 2,6 or diluent control lanes 1,5) for 6 hours. Cellular proteins were resolved on a 8% to 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and caspase activity was detected by cleavage of the caspase-specific substrate PARP by using immunoblot analysis. Filled arrowheads (◂) indicate the uncleaved p116 and open arrowheads (◃) indicate the cleaved p89 form of PARP.