title: Mechanisms by which low glucose enhances the cytotoxicity of metformin to cancer cells both in vitro and in vivo.
abstract: Different cancer cells exhibit altered sensitivity to metformin treatment. Recent studies suggest these findings may be due in part to the common cell culture practice of utilizing high glucose, and when glucose is lowered, metformin becomes increasingly cytotoxic to cancer cells. In low glucose conditions ranging from 0 to 5 mM, metformin was cytotoxic to breast cancer cell lines MCF7, MDAMB231 and SKBR3, and ovarian cancer cell lines OVCAR3, and PA-1. MDAMB231 and SKBR3 were previously shown to be resistant to metformin in normal high glucose medium. When glucose was increased to 10 mM or above, all of these cell lines become less responsive to metformin treatment. Metformin treatment significantly reduced ATP levels in cells incubated in media with low glucose (2.5 mM), high fructose (25 mM) or galactose (25 mM). Reductions in ATP levels were not observed with high glucose (25 mM). This was compensated by enhanced glycolysis through activation of AMPK when oxidative phosphorylation was inhibited by metformin. However, enhanced glycolysis was either diminished or abolished by replacing 25 mM glucose with 2.5 mM glucose, 25 mM fructose or 25 mM galactose. These findings suggest that lowering glucose potentiates metformin induced cell death by reducing metformin stimulated glycolysis. Additionally, under low glucose conditions metformin significantly decreased phosphorylation of AKT and various targets of mTOR, while phospho-AMPK was not significantly altered. Thus inhibition of mTOR signaling appears to be independent of AMPK activation. Further in vivo studies using the 4T1 breast cancer mouse model confirmed that metformin inhibition of tumor growth was enhanced when serum glucose levels were reduced via low carbohydrate ketogenic diets. The data support a model in which metformin treatment of cancer cells in low glucose medium leads to cell death by decreasing ATP production and inhibition of survival signaling pathways. The enhanced cytotoxicity of metformin against cancer cells was observed both in vitro and in vivo.
authors.affiliation: Cancer Biology Research Center, Sanford Research/USD, Sioux Falls, South Dakota, United States of America.
authors.forename: Yongxian
authors.initials: Y
authors.lastname: Zhuang
authors.affiliation: Sanford School of Medicine, The University of South Dakota, Vermillion, South Dakota, United States of America.
authors.forename: Daniel K
authors.initials: DK
authors.lastname: Chan
authors.affiliation: Cancer Biology Research Center, Sanford Research/USD, Sioux Falls, South Dakota, United States of America.
authors.forename: Allison B
authors.initials: AB
authors.lastname: Haugrud
authors.affiliation: Cancer Biology Research Center, Sanford Research/USD, Sioux Falls, South Dakota, United States of America.
authors.forename: W Keith
authors.initials: WK
authors.lastname: Miskimins
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001786:Blood Glucose; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects*; D055423:Diet, Ketogenic; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology*; D006019:Glycolysis / Q000187:drug effects; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D008687:Metformin / Q000494:pharmacology*; D051379:Mice; D009369:Neoplasms / Q000378:metabolism; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D001786:Blood Glucose; D007004:Hypoglycemic Agents; D000255:Adenosine Triphosphate; D008687:Metformin; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases; D005947:Glucose