title: Metformin in combination with 5-fluorouracil suppresses tumor growth by inhibiting the Warburg effect in human oral squamous cell carcinoma.
abstract: Cancer cells show enhanced glucose consumption and lactate production even in the presence of abundant oxygen, a phenomenon known as the Warburg effect, which is related to tumor proliferation, progression and drug-resistance in cancers. Hypoxia-inducible factor-1 (HIF-1) and several members of Phosphatidylinositol-4, 5-bisphosphate 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway positively contribute to the Warburg effect, whereas AMP activated protein Kinase (AMPK) acts as a negative regulator. Targeting the regulator molecules of Warburg effect might be a useful strategy to effectively kill cancer cells. Metformin was reported to be effective against various cancers as it inhibits cell proliferation by activating AMPK, and inhibiting mTOR and HIF-1α. Several studies suggested the efficacy of metformin with 5-fluorouracil (5-FU) against esophageal and colon cancer. In this study, we evaluated the efficacy of metformin and 5-FU combined therapy against human oral squamous cell carcinoma (OSCC) in vitro and in vivo. MTT assay and TUNEL assay revealed that metformin (4 mg/ml) and 5-FU (2.5 µg/ml) combination treatment effectively inhibited cell growth and induced apoptosis in OSCC cell lines (HSC2, HSC3 and HSC4) compared to either agent alone. Lactate colorimetric assay detected decreased level of lactate in the supernatants of metformin and 5-FU treated cells compared to cells treated with metformin or 5-FU. Western blot analysis showed marked downregulation of HIF-1α and mTOR expression, and upregulation of AMPKα in cells treated with metformin and 5-FU combination treatment. Combination therapy with metformin (200 mg/kg, i.p.) and 5-FU (10 mg/kg, i.p.) for 4 weeks (5 days/week) effectively reduced HSC2 tumor growth (77.6%) compared to metformin (59.9%) or 5-FU (52%) alone in nude mice. These findings suggest that metformin and 5-FU combined therapy could exert strong antitumor effect against OSCC through the inhibition of Warburg phenomenon in tumor cells.
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, Ube 755-8505, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Harada
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, Ube 755-8505, Japan.
authors.forename: Tarannum
authors.initials: T
authors.lastname: Ferdous
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, Ube 755-8505, Japan.
authors.forename: Toyoko
authors.initials: T
authors.lastname: Harada
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, Ube 755-8505, Japan.
authors.forename: Yoshiya
authors.initials: Y
authors.lastname: Ueyama
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000096:biosynthesis; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D049109:Cell Proliferation / Q000187:drug effects; D004357:Drug Synergism; D005472:Fluorouracil / Q000008:administration & dosage*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis; D008687:Metformin / Q000008:administration & dosage*; D051379:Mice; D009062:Mouth Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000096:biosynthesis; D023041:Xenograft Model Antitumor Assays
chemical_list: C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008687:Metformin; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; D005472:Fluorouracil