title: Apoptosis induction in human breast cancer (MCF-7) cells by a novel venom L-amino acid oxidase (Rusvinoxidase) is independent of its enzymatic activity and is accompanied by caspase-7 activation and reactive oxygen species production.
abstract: We report the elucidation of a mechanism of apoptosis induction in breast cancer (MCF-7) cells by an L-amino acid oxidase (LAAO), Rusvinoxidase, purified from the venom of Daboia russelii russelii. Peptide mass fingerprinting analysis of Rusvinoxidase, an acidic monomeric glycoprotein with a mass of ~57 kDa, confirmed its identity as snake venom LAAO. The enzymatic activity of Rusvinoxidase was completely abolished after two cycles of freezing and thawing; however, its cytotoxicity toward MCF-7 cells remained unaffected. Dose- and time-dependent induction of apoptosis by Rusvinoxidase on MCF-7 cells was evident from changes in cell morphology, cell membrane integrity, shrinkage of cells and apoptotic body formation accompanied by DNA fragmentation. Rusvinoxidase induced apoptosis in MCF-7 cells by both the extrinsic (death-receptor) and intrinsic (mitochondrial) signaling pathways. The former pathway of apoptosis operated through activation of caspase-8 that subsequently activated caspase-7 but not caspase-3. Rusvinoxidase-induced intrinsic pathway of apoptosis was accompanied by a time-dependent depolarization of the mitochondrial membrane through the generation of reactive oxygen species, followed by a decrease in cellular glutathione content and catalase activity, and down-regulation of expression of anti-apoptotic proteins Bcl-XL and heat-shock proteins (HSP-90 and HSP-70). Rusvinoxidase treatment resulted in increase of the pro-apoptotic protein Bax, subsequently leading to the release of cytochrome c from mitochondria to the cytosol and activating caspase-9, which in turn stimulated effector caspase-7. Rusvinoxidase at a dose of 4 mg/kg was non-toxic in mice, indicating that it may be useful as a model for the development of peptide-based anticancer drugs.
authors: School of Biological Sciences, University of Northern Colorado, Greeley, CO, 80639-0017, USA, akm@tezu.ernet.in.
authors: Ashis K
authors: AK
authors: Mukherjee
authors: Anthony J
authors: AJ
authors: Saviola
authors: Patrick D
authors: PD
authors: Burns
authors: Stephen P
authors: SP
authors: Mackessy
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001943:Breast Neoplasms / Q000188:drug therapy*; D053179:Caspase 7 / Q000378:metabolism; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D050830:L-Amino Acid Oxidase / Q000378:metabolism / Q000494:pharmacology*; D061986:MCF-7 Cells; D051379:Mice; D010154:Pakistan; D017382:Reactive Oxygen Species / Q000378:metabolism*; D017840:Daboia; D014757:Viper Venoms / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D017382:Reactive Oxygen Species; D014757:Viper Venoms; D050830:L-Amino Acid Oxidase; C000608416:rusvinoxidase protein, Daboia russelii russelii; D053179:Caspase 7