title: Synergistic induction of apoptosis in neuroblastoma cells using a combination of cytostatic drugs with interferon-gamma and TRAIL.
abstract: The majority of high-risk neuroblastomas lack the expression of caspase-8 due to gene silencing which suggest a mechanism for the selection of tumour cells that are refractory to multiple cytotoxic drugs including tumour necrosis factor-related apoptosis-inducing ligand (TRAIL). Inhibitors of DNA methyltransferases and IFN-gamma induce expression of caspase-8, and sensitise some neuroblastoma cells to TRAIL-mediated apoptosis. Here we demonstrate that a combination of cytostatic drugs with IFN-gamma and TRAIL synergistically induces neuroblastoma cell death, which may have implications for future therapy of children with neuroblastoma. Treatment of neuroblastoma cells with IFN-gamma induced caspase-8 expression in all cell lines investigated. In five of the neuroblastoma cell lines (SHEP-1, SK-N-AS, SK-N-FI, SH-SY-5Y and Kelly), IFN-gamma promoted TRAIL-mediated cleavage of caspase-8, initiating a caspase cascade involving caspase-7 and PARP followed by apoptosis. IFN-gamma-mediated facilitation of apoptosis was inhibited by the pan-caspase inhibitor zVAD-fmk and the caspase-8 specific inhibitor zIEDT-fmk, indicating an important role of caspase-8 in mediating sensitation by IFN-gamma in neuroblastoma cells. In three of the cell lines [SK-N-BE(2), SK-N-DZ and IMR-32] caspase-8 expression was induced by IFN-gamma, but the cells were still resistant to TRAIL-mediated apoptosis. The pattern of basal TRAIL receptor expression, decoy receptors, FLIP and FADD could not be correlated with resistance or sensitivity to TRAIL-induced apoptosis. Importantly, treatment of neuroblastoma cell lines with cytostatic drugs increased apoptosis in the TRAIL-sensitive cell lines whereas the resistant cell lines were susceptible to TRAIL-mediated apoptosis in the presence of the anticancer drugs. The mechanism of the increased susceptibility to apoptosis might results from drug-mediated up-regulation of the death receptors DR4 and DR5.
authors.affiliation: Department of Woman and Child Health, Karolinska Institutet, Stockholm, Sweden. john.inge.johnsen@kbh.ki.se
authors.forename: John I
authors.initials: JI
authors.lastname: Johnsen
authors.forename: Ingvild
authors.initials: I
authors.lastname: Pettersen
authors.forename: Frida
authors.initials: F
authors.lastname: Ponthan
authors.forename: Baldur
authors.initials: B
authors.lastname: Sveinbjørnsson
authors.forename: Trond
authors.initials: T
authors.lastname: Flaegstad
authors.forename: Per
authors.initials: P
authors.lastname: Kogner
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins; D053181:Caspase 8; D061945:Caspase Inhibitors; D020169:Caspases / Q000494:pharmacology*; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology*; D008024:Ligands; D008562:Membrane Glycoproteins / Q000494:pharmacology*; D009447:Neuroblastoma / Q000473:pathology*; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053221:TNF-Related Apoptosis-Inducing Ligand; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
chemical_list: D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D016898:Interferon-alpha; D008024:Ligands; D008562:Membrane Glycoproteins; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D018124:Receptors, Tumor Necrosis Factor; D053221:TNF-Related Apoptosis-Inducing Ligand; C105190:TNFRSF10A protein, human; C505639:TNFRSF10B protein, human; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; C505530:CASP8 protein, human; D053181:Caspase 8; D020169:Caspases