17-DMAG–mediated cytotoxicity is caspase dependent. (A) Representative JC-1 flow diagram of CLL patient cells treated with vehicle or 1μM 17-DMAG for 24 hours. The percentage of cells containing intact mitochondria and depolarized mitochondria are indicated. The data are shown gated on live cells by forward and side scatter properties (top row) or ungated (bottom row). (B) Analysis of mitochondrial membrane potential in vehicle control or 1μM 17-DMAG–treated CLL at 8 and 24 hours by JC-1 staining (n = 6). Values shown in this experiment were calculated as a decrease in the JC-1 aggregate relative to the vehicle control. (C) Western blot analysis for PARP in whole cell lysate prepared from CLL patient cells treated with vehicle control or 1μM 17-DMAG for 24 hours in the presence or absence of 100μM caspase inhibitor z-VAD-fmk. Blots are probed with actin as a loading control. Results shown are representative of at least 6 patient samples. (D) Viability of CLL patient cells (n = 12) treated with vehicle control or 1μM 17-DMAG for 24 hours in the presence or absence of 100μM caspase inhibitor z-VAD-fmk determined by Ann/PI staining.