title: Bcl-2 and Mn-SOD antisense oligodeoxynucleotides and a glutamine-enriched diet facilitate elimination of highly resistant B16 melanoma cells by tumor necrosis factor-alpha and chemotherapy.
abstract: Mitochondrial glutathione (mtGSH) depletion increases sensitivity of Bcl-2-overexpressing B16 melanoma (B16M)-F10 cells (high metastatic potential) to tumor necrosis factor-alpha (TNF-alpha)-induced oxidative stress and death in vitro. In vivo, mtGSH depletion in B16M-F10 cells was achieved by feeding mice (where the B16M-F10 grew as a solid tumor in the footpad) with an L-glutamine (L-Gln)-enriched diet, which promoted in the tumor cells an increase in glutaminase activity, accumulation of cytosolic L-glutamate, and competitive inhibition of GSH transport into mitochondria. L-Gln-adapted B16M-F10 cells, isolated using anti-Met-72 monoclonal antibodies and flow cytometry-coupled cell sorting, were injected into the portal vein to produce hepatic metastases. In l-Gln-adapted invasive (iB16M-Gln+) cells, isolated from the liver by the same methodology and treated with TNF-alpha and an antisense Bcl-2 oligodeoxynucleotide, viability decreased to approximately 12%. iB16M-Gln+ cell death associated with increased generation of O2*- and H2O2, opening of the mitochondrial permeability transition pore complex, and release of proapoptotic molecular signals. Activation of cell death mechanisms was prevented by GSH ester-induced mtGSH replenishment. The oxidative stress-resistant survivors showed an adaptive response that includes overexpression of manganese-containing superoxide dismutase (Mn-SOD) and catalase activities. By treating iB16M-Gln+ cells with a double anti- antisense therapy (Bcl-2 and SOD2 antisense oligodeoxynucleotides) and TNF-alpha, metastatic cell survival decreased to approximately 1%. Chemotherapy (taxol plus daunorubicin) easily removed this minimum percentage of survivors. This contribution identifies critical molecules that can be sequentially targeted to facilitate elimination of highly resistant metastatic cells.
authors.affiliation: Department of Physiology, University of Valencia, 17 Av. Blasco Ibañez, 46010 Valencia, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Benlloch
authors.forename: Salvador
authors.initials: S
authors.lastname: Mena
authors.forename: Paula
authors.initials: P
authors.lastname: Ferrer
authors.forename: Elena
authors.initials: E
authors.lastname: Obrador
authors.forename: Miguel
authors.initials: M
authors.lastname: Asensi
authors.forename: José A
authors.initials: JA
authors.lastname: Pellicer
authors.forename: Julian
authors.initials: J
authors.lastname: Carretero
authors.forename: Angel
authors.initials: A
authors.lastname: Ortega
authors.forename: José M
authors.initials: JM
authors.lastname: Estrela
mesh_terms: D000821:Animal Feed; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D003131:Combined Modality Therapy; D004195:Disease Models, Animal; D019008:Drug Resistance, Neoplasm; D015316:Genetic Therapy; D005973:Glutamine / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D008297:Male; D008545:Melanoma / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008928:Mitochondria / Q000378:metabolism; D009368:Neoplasm Transplantation; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D018384:Oxidative Stress; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D012983:Soft Tissue Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D013482:Superoxide Dismutase / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D005973:Glutamine; D013482:Superoxide Dismutase; D005978:Glutathione