title: Apoptosis induced by intracellular ceramide accumulation in MDA-MB-435 breast carcinoma cells is dependent on the generation of reactive oxygen species.
abstract: Strategies to promote intracellular ceramide accumulation in cancer cells may have therapeutic utility because ceramide is an important second messenger during apoptosis. Exposure to cell-permeable C(6) ceramide or tricyclodecan-9-yl-xanthate (an inducer of de novo ceramide synthesis and an inhibitor of sphingomyelin synthase) caused MDA-MB-435 human breast carcinoma cells to die by apoptosis. Concomitant treatment with the ceramidase inhibitor D-erythro-2-(N-myristoylamino)-1-phenyl-1-propanol (MAPP) or the glucosylceramide synthase inhibitor 1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (PPMP) potentiated the cytotoxic effect of C(6) ceramide, indicating that C(6) ceramide-mediated cytotoxicity was antagonized by the action of ceramidases and glucosylceramide synthase. Interestingly, treatment with PPMP alone, but not MAPP alone, also induced apoptosis in MDA-MB-435 cells, suggesting that conversion to glucosylceramide rather than catabolism by ceramidases prevented endogenous ceramide from reaching cytotoxic levels. C(6) ceramide-induced apoptosis in MDA-MB-435 cells was associated with the generation of reactive oxygen species, and was inhibited by the antioxidants N-acetylcysteine and glutathione. Although mitochondrial membrane integrity was disrupted in C(6) ceramide-treated MDA-MB-435 cells, apoptosis was not mediated by caspases because there was no protective effect by the pan-caspase inhibitor z-VAD-fmk. Collectively, these findings indicate that strategies to enhance intracellular ceramide accumulation in malignant cells might offer a novel approach to the treatment of breast cancer.
authors.affiliation: Department of Pathology, Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5.
authors.forename: S Y Velda
authors.initials: SY
authors.lastname: Chan
authors.forename: Ashley L
authors.initials: AL
authors.lastname: Hilchie
authors.forename: Michael G
authors.initials: MG
authors.lastname: Brown
authors.forename: Robert
authors.initials: R
authors.lastname: Anderson
authors.forename: David W
authors.initials: DW
authors.lastname: Hoskin
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology*; D045744:Cell Line, Tumor; D002518:Ceramides / Q000037:antagonists & inhibitors / Q000378:metabolism*; D053938:DNA Fragmentation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D002518:Ceramides; D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species