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

Abstract

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 A(2) 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.

Store-operated Ca2+ Entry-associated Regulatory factor (SARAF) Plays an Important Role in the Regulation of Arachidonate-regulated Ca2+ (ARC) Channels

“…4, treatment with ATRA alone does not significantly increase the percentage of apoptotic cells (a slight increase was detected after treatment with 15 M ATRA). As previously reported (22), inhibition of PLA2 using 10 M AACOCF3, a concentration that has been previously used to inhibit AA release in neuroblastoma cell lines (23), significantly enhance the percentage of apoptotic cells at all the ATRA concentrations tested (Fig. 4).…”

p38MAPK-Dependent Sensitivity of Ewing's Sarcoma Family of Tumors to Fenretinide-Induced Cell Death

“…Collectively, these data show that fenretinide is a potent inducer of apoptosis in ESFT cell lines and that in vitro the ESFT cells are more sensitive to fenretinide than the three neuroblastoma cell lines examined. The mechanism of fenretinide-induced apoptosis has been extensively examined in the neuroblastoma SH-SY5Y cells (23,24,30); therefore, we have used these cells to compare the mechanism of response with ESFT cells.…”

“…At 24 hours, cells were collected and viable cell number was determined by the trypan blue exclusion assay. ESFT cells were exposed to the inhibitors alone at previously published doses (23,29) to determine whether the inhibitor had any toxic effects; for PD146176, MK886, baicalein, AACOCF3, and nordihydroguairetic acid, the highest dose tolerated by ESFT was also evaluated. The effects of diaphenylene iodonium and AEBSF on fenretinide-induced cell death were not assessed, as these agents alone were toxic to ESFT cells at doses required to inhibit ROS accumulation.…”

“…Unlike cell death mediated by the classic retinoids, fenretinidemediated cell death is largely independent of retinoic acid receptors (18,19), although accumulation of reactive oxygen species (ROS) has been described in many different tumor cell types following exposure to fenretinide (12,20). Although the mechanisms of fenretinide-mediated ROS production are not well characterized, the mitochondrial electron transport chain (21,22) and lipoxygenase enzymes (23,24) have both been implicated. The ability of antioxidants to prevent fenretinide induced cell death suggests a critical role for ROS in the apoptotic signal transduction (21).…”

Regulation of the apoptosis–necrosis switch

“…In addition to regulating directly the 'apoptotic' machinery, see Figure 4, NO finely regulates the NMDA receptors, and therefore excitotoxicity in neuronal cells (Lipton et al, 1993), including neuroblastoma cells exposed to the gp120 HIV protein (Corasaniti et al, 1992(Corasaniti et al, , 1994(Corasaniti et al, , 1995Bagetta et al, 1993;Clementi et al, 1996;Maccarrone et al, 2000), or even tamoxifen (Maccarrone et al, 1998) or fenretinide (Lovat et al, 2003). Via S-nitrosylation or nitration of proteins, NO can be a bifunctional modulator of cell death capable of either triggering or inhibiting cell demise.…”