title: Role of Apollon in human melanoma resistance to antitumor agents that activate the intrinsic or the extrinsic apoptosis pathways.
abstract: OBJECTIVE
  To assess the role of Apollon in melanoma resistance to intrinsic and extrinsic pathways of apoptosis and to identify strategies to reduce its expression.
  
  
  METHODS
  Apollon expression was assessed in melanoma cells in vitro and in vivo. Apollon modulation and melanoma apoptosis were evaluated by Western blot and/or flow cytometry in response to cytotoxic drugs, mitogen-activated protein/extracellular signal-regulated kinase (MEK)-, BRAF(V600E)-, and mTOR-specific inhibitors, TRAIL and anti-HLA class II monoclonal antibodies (mAb). Mitochondrial depolarization, caspase activation, apoptosis assays, and gene expression profiling were used to test effects of Apollon silencing, by siRNA, on melanoma response to antitumor agents.
  
  
  RESULTS
  Apollon was constitutively expressed by melanoma cells, in vitro and in vivo, and at higher levels than in benign melanocytic lesions. Melanoma apoptosis correlated significantly with Apollon protein downmodulation in response to cytotoxic drugs, MEK, or BRAF(V600E)-specific inhibitors. Combinatorial treatment with MEK and mTOR inhibitors and HLA class II ligation, by a specific mAb, promoted Apollon downmodulation and enhanced melanoma apoptosis. Apollon downmodulation induced by antitumor agents was caspase independent, but proteasome dependent. Knockdown of Apollon, by siRNA, triggered apoptosis and/or significantly enhanced melanoma cell death in response to cytotoxic drugs, MEK- and BRAF(V600E)-specific inhibitors, and soluble or membrane-bound TRAIL. Apollon silencing promoted mitochondrial depolarization and caspase-2, caspase-8, caspase-9, and caspase-3 activation in response to different antitumor agents and altered the profile of genes modulated by MEK or BRAF(V600E)-specific inhibitors.
  
  
  CONCLUSIONS
  Targeting of Apollon may significantly improve melanoma cell death in response to antitumor agents that trigger the intrinsic or the extrinsic apoptosis pathways.
authors.affiliation: Human Tumors Immunobiology Unit, Molecular Pharmacology Unit, Department of Experimental Oncology and Molecular Medicine, Medical Oncology Unit 3, Department of Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori and University of Milan, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Tassi
authors.forename: Marina
authors.initials: M
authors.lastname: Zanon
authors.forename: Claudia
authors.initials: C
authors.lastname: Vegetti
authors.forename: Alessandra
authors.initials: A
authors.lastname: Molla
authors.forename: Ilaria
authors.initials: I
authors.lastname: Bersani
authors.forename: Valentina
authors.initials: V
authors.lastname: Perotti
authors.forename: Marzia
authors.initials: M
authors.lastname: Pennati
authors.forename: Nadia
authors.initials: N
authors.lastname: Zaffaroni
authors.forename: Michele
authors.initials: M
authors.lastname: Milella
authors.forename: Soldano
authors.initials: S
authors.lastname: Ferrone
authors.forename: Carmelo
authors.initials: C
authors.lastname: Carlo-Stella
authors.forename: Alessandro M
authors.initials: AM
authors.lastname: Gianni
authors.forename: Roberta
authors.initials: R
authors.lastname: Mortarini
authors.forename: Andrea
authors.initials: A
authors.lastname: Anichini
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects*; D053143:Caspase 2 / Q000378:metabolism; D053148:Caspase 3 / Q000096:biosynthesis; D053181:Caspase 8 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002461:Cell Line, Transformed; D016764:Cell Polarity; D015536:Down-Regulation; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors; D020869:Gene Expression Profiling; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D008545:Melanoma / Q000188:drug therapy* / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors; D034622:RNA Interference; D034741:RNA, Small Interfering; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors
chemical_list: D000911:Antibodies, Monoclonal; D000970:Antineoplastic Agents; C401541:BIRC6 protein, human; D051034:Inhibitor of Apoptosis Proteins; D034741:RNA, Small Interfering; C546842:MTOR protein, human; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D058570:TOR Serine-Threonine Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D053143:Caspase 2; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9