Clofilium, a potassium channel blocker, induces apoptosis of human promyelocytic leukemia (HL-60) cells via Bcl-2-insensitive activation of caspase-3

We have demonstrated that clofilium, a potassium channel blocker, induces a poptosis on human promyelocytic leukemia (HL-60) cells. Cells treated with clofilium led to suppression of viability and proliferation in both time an d concentration-dependent manners. Nuclear DAPI staining and electronmicros copic examination revealed typical nuclear features of apoptosis in cells t reated with clofilium that was further verified in DNA fragmentation analys is. Flow cytometry analysis with FITC-annexin V and propidium iodide (PT) r evealed that apoptotic cell population with Annexin V+/PI- increased gradua lly from <2% at 0 h, to 20% at 4 h and 29% at 16 h after exposure to 10 mu M clofilium in HL-60 cells. Furthermore, fluorometric immunosorbent enzyme assay for activity of caspase-3 showed approximately a 10-fold increase of activity in cells treated with 10 mu M of clofilium for 2-3 h compared with the basal level of its activity in untreated control cells. Immunoblotting analysis revealed proteolytic cleavage of caspase-3 and subsequent cleavag e of PARP, However, there was no significant change of Eel 2 and Bar protei ns. These results indicate that clofilium exerts antiproliferative action a nd growth inhibition on HL-60 through induction of apoptosis which is media ted via Bcl-2-insensitive activation of caspase-3, and suggest chemotherape utic and cytostatic potentials of this compound in human leukemias. (C) 199 9 Elsevier Science Ireland Ltd. All rights reserved.