The role of p38 MAPK activation in auranofin‐induced apoptosis of human promyelocytic leukaemia HL‐60 cells

In a previous study, we reported an antileukaemic activity of auranofin (AF), demonstrating its dual effects: on the induction of apoptotic cell death and its synergistic action with retinoic acid on cell differentiation. In this study, we investigated the downstream signalling events of AF‐induced apoptosis to determine the molecular mechanisms of AF activity. Treatment of HL‐60 cells with AF induced apoptosis in a concentration‐ and time‐dependent manner. Western blot analysis showed that AF‐induced apoptosis was accompanied by the activation of caspase‐8, caspase‐9, and caspase‐3, and the release of cytochrome c from the mitochondria. The phosphorylation and kinase activities of p38 mitogen‐activated protein kinase (p38 MAPK) increased gradually until 12 h after AF (2 μM) treatment, and p38 MAPK was also activated concentration‐dependently. Pretreatment with SB203580, a specific inhibitor of p38 MAPK, significantly blocked DNA fragmentation and the cleavage of procaspase‐8, procaspase‐3, and poly‐ADP‐ribose polymerase (PARP), whereas SB203580 alone had no effect. Reactive oxygen species (ROS) were also detected within 1 h after AF treatment, and the antioxidant N‐acetyl‐L‐cysteine (NAC) effectively protected the cells from apoptosis by inhibiting the phosphorylation of p38 MAPK and the activation of caspases. These results suggest that ROS generation and the subsequent activation of p38 MAPK are essential for the proapoptotic effects of AF in human promyelocytic leukaemia HL‐60 cells. British Journal of Pharmacology (2005) 146, 506–513. doi:10.1038/sj.bjp.0706360