Mechanisms of free-radical induction in relation to fenretinide-induced apoptosis of neuroblastoma

The mechanisms of fenretinide‐induced cell death of neuroblastoma cells are complex, involving signaling pathways mediated by free radicals or reactive oxygen species (ROS). The aim of this study was to identify mechanisms generating ROS and apoptosis of neuroblastoma cells in response to fenretinide. Fenretinide‐induced ROS or apoptosis of SH‐SY5Y or HTLA 230 neuroblastoma cells were not blocked by Nitro l‐argenine methyl ester (l‐NAME), an inhibitor of nitric oxide synthase. Flavoprotein‐dependent superoxide‐producing enzymes such as NADPH oxidase were also not involved in fenretinide‐induced apoptosis or ROS generation. Similarly, ketoconazole, a cytochrome P450 inhibitor, and inhibitors of cyclooxygenase (COX) were also ineffective. In contrast, inhibition of phospholipase A2 or lipoxygenases (LOX) blocked the induction of ROS and apoptosis in response to fenretinide. Using specific inhibitors of LOX, blocking 12‐LOX but not 5‐ or 15‐LOX inhibited both fenretinide‐induced ROS and apoptosis. The effects of eicosatriynoic acid, a specific 12‐LOX inhibitor, were reversed by the addition of the 12‐LOX products, 12 (S)‐hydroperoxyeicosatetraenoic acid and 12 (S)‐hydroxyeicosatetraenoic acid. The targeting of 12‐LOX in neuroblastoma cells may thus be a novel pathway for the development of drugs inducing apoptosis of neuroblastoma with improved tumor specificity. J. Cell. Biochem. 89: 698–708, 2003. © 2003 Wiley‐Liss, Inc.