title: Metformin inhibits heme oxygenase-1 expression in cancer cells through inactivation of Raf-ERK-Nrf2 signaling and AMPK-independent pathways.
abstract: Resistance to therapy is the major obstacle to more effective cancer treatment. Heme oxygenase-1 (HO-1) is often highly up-regulated in tumor tissues, and its expression is further increased in response to therapies. It has been suggested that inhibition of HO-1 expression is a potential therapeutic approach to sensitize tumors to chemotherapy and radiotherapy. In this study, we tested the hypothesis that the anti-tumor effects of metformin are mediated by suppression of HO-1 expression in cancer cells. Our results indicate that metformin strongly suppresses HO-1 mRNA and protein expression in human hepatic carcinoma HepG2, cervical cancer HeLa, and non-small-cell lung cancer A549 cells. Metformin also markedly reduced Nrf2 mRNA and protein levels in whole cell lysates and suppressed tert-butylhydroquinone (tBHQ)-induced Nrf2 protein stability and antioxidant response element (ARE)-luciferase activity in HepG2 cells. We also found that metformin regulation of Nrf2 expression is mediated by a Keap1-independent mechanism and that metformin significantly attenuated Raf-ERK signaling to suppress Nrf2 expression in cancer cells. Inhibition of Raf-ERK signaling by PD98059 decreased Nrf2 mRNA expression in HepG2 cells, confirming that the inhibition of Nrf2 expression is mediated by an attenuation of Raf-ERK signaling in cancer cells. The inactivation of AMPK by siRNA, DN-AMPK or the pharmacological AMPK inhibitor compound C, revealed that metformin reduced HO-1 expression in an AMPK-independent manner. These results highlight the Raf-ERK-Nrf2 axis as a new molecular target in anticancer therapy in response to metformin treatment.
authors.affiliation: Department of Toxicology, College of Pharmacy, Chungnam National University, Daejeon, South Korea.
authors.forename: Minh Truong
authors.initials: MT
authors.lastname: Do
authors.forename: Hyung Gyun
authors.initials: HG
authors.lastname: Kim
authors.forename: Tilak
authors.initials: T
authors.lastname: Khanal
authors.forename: Jae Ho
authors.initials: JH
authors.lastname: Choi
authors.forename: Dong Hee
authors.initials: DH
authors.lastname: Kim
authors.forename: Tae Cheon
authors.initials: TC
authors.lastname: Jeong
authors.forename: Hye Gwang
authors.initials: HG
authors.lastname: Jeong
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000737:chemistry; D002470:Cell Survival / Q000187:drug effects; D003600:Cytosol / Q000737:chemistry; D005696:Galactosidases / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000037:antagonists & inhibitors*; D006801:Humans; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007202:Indicators and Reagents; D008156:Luciferases / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D008687:Metformin / Q000494:pharmacology*; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects*; D051267:NF-E2-Related Factor 2 / Q000502:physiology*; D009369:Neoplasms / Q000201:enzymology*; D010957:Plasmids / Q000235:genetics; D012313:RNA / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection; D048490:raf Kinases / Q000187:drug effects*
chemical_list: D007004:Hypoglycemic Agents; D007202:Indicators and Reagents; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D034741:RNA, Small Interfering; D012313:RNA; D008687:Metformin; D008156:Luciferases; D051547:Heme Oxygenase-1; D048490:raf Kinases; D020928:Mitogen-Activated Protein Kinases; D005696:Galactosidases
keywords: AMP-activated protein kinase; AMPK; ARE; ERK; HO-1; Keap1; Kelch-like ECH-associated protein 1; Metformin; Nrf2; antioxidant response element; extracellular signal-regulated kinase; heme oxygenase-1; nuclear erythroid factor 2 (NE-F2)-related factor 2