title: Melatonin-induced increase in sensitivity of human hepatocellular carcinoma cells to sorafenib is associated with reactive oxygen species production and mitophagy.
abstract: Effects of sorafenib in hepatocellular carcinoma (HCC) are frequently transient due to tumor-acquired resistance, a phenotype that could be targeted by other molecules to reduce this adaptive response. Because melatonin is known to exert antitumor effects in HCC cells, this study investigated whether and how melatonin reduces resistance to sorafenib. Susceptibility to sorafenib (10 nmol/L to 50 μmol/L) in the presence of melatonin (1 and 2 mmol/L) was assessed in HCC cell lines HepG2, HuH7, and Hep3B. Cell viability was reduced by sorafenib from 1 μmol/L in HepG2 or HuH7 cells, and 2.5 μmol/L in Hep3B cells. Co-administration of melatonin and sorafenib exhibited a synergistic cytotoxic effect on HepG2 and HuH7 cells, while Hep3B cells displayed susceptibility to doses of sorafenib that had no effect when administrated alone. Co-administration of 2.5 μmol/L sorafenib and 1 mmol/L melatonin induced apoptosis in Hep3B cells, increasing PARP hydrolysis and BAX expression. We also observed an early colocalization of mitochondria with lysosomes, correlating with the expression of mitophagy markers PINK1 and Parkin and a reduction of mitofusin-2 and mtDNA compared with sorafenib administration alone. Moreover, increased reactive oxygen species production and mitochondrial membrane depolarization were elicited by drug combination, suggesting their contribution to mitophagy induction. Interestingly, Parkin silencing by siRNA to impair mitophagy significantly reduced cell killing, PARP cleavage, and BAX expression. These results demonstrate that the pro-oxidant capacity of melatonin and its impact on mitochondria stability and turnover via mitophagy increase sensitivity to the cytotoxic effect of sorafenib.
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.
authors.forename: Néstor
authors.initials: N
authors.lastname: Prieto-Domínguez
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.
authors.forename: Raquel
authors.initials: R
authors.lastname: Ordóñez
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Fernández
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.
authors.forename: Carolina
authors.initials: C
authors.lastname: Méndez-Blanco
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.|Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB), Consejo Superior Investigaciones Científicas (CSIC) and Liver Unit-Hospital Clinic, Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Baulies
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.|Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB), Consejo Superior Investigaciones Científicas (CSIC) and Liver Unit-Hospital Clinic, Barcelona, Spain.
authors.forename: Carmen
authors.initials: C
authors.lastname: Garcia-Ruiz
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.|Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB), Consejo Superior Investigaciones Científicas (CSIC) and Liver Unit-Hospital Clinic, Barcelona, Spain.|University of Southern California Research Center for Alcohol Liver and Pancreatic Diseases and Cirrhosis, Keck School of Medicine, USC, Los Angeles, CA, USA.
authors.forename: José C
authors.initials: JC
authors.lastname: Fernández-Checa
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain.
authors.forename: José L
authors.initials: JL
authors.lastname: Mauriz
authors.affiliation: Institute of Biomedicine (IBIOMED), University of León, León, Spain. jgonga@unileon.es.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Spain. jgonga@unileon.es.
authors.forename: Javier
authors.initials: J
authors.lastname: González-Gallego
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D008550:Melatonin / Q000494:pharmacology*; D063306:Mitophagy / Q000187:drug effects*; D009363:Neoplasm Proteins / Q000378:metabolism; D009536:Niacinamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D010671:Phenylurea Compounds / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D000077157:Sorafenib
chemical_list: D009363:Neoplasm Proteins; D010671:Phenylurea Compounds; D017382:Reactive Oxygen Species; D009536:Niacinamide; D000077157:Sorafenib; D008550:Melatonin
keywords: apoptosis; hepatocarcinoma; melatonin; mitophagy; oxidative stress; sorafenib