title: Medium Renewal Blocks Anti-Proliferative Effects of Metformin in Cultured MDA-MB-231 Breast Cancer Cells.
abstract: Epidemiological studies indicate that metformin, a widely used type 2 diabetes drug, might reduce breast cancer risk and mortality in patients with type 2 diabetes. Metformin might protect against breast cancer indirectly by ameliorating systemic glucose homeostasis. Alternatively, it might target breast cancer cells directly. However, experiments using MDA-MB-231 cells, a standard in vitro breast cancer model, produced inconsistent results regarding effectiveness of metformin as a direct anti-cancer agent. Metformin treatments in cultured MDA-MB-231 cells are usually performed for 48-96 hours, but protocols describing renewal of cell culture medium during these prolonged treatments are rarely reported. We determined whether medium renewal protocol might alter sensitivity of MDA-MB-231 cells treated with metformin. Using the MTS assay, BrdU incorporation and Hoechst staining we found that treatment with metformin for 48-72 hours failed to suppress viability and proliferation of MDA-MB-231 cells if low-glucose (1 g/L) medium was renewed every 24 hours. Conversely, metformin suppressed their viability and proliferation if medium was not renewed. Without renewal glucose concentration in the medium was reduced to 0.1 g/L in 72 hours, which likely explains increased sensitivity to metformin under these conditions. We also examined whether 2-deoxy-D-glucose (2-DG) reduces resistance to metformin. In the presence of 2-DG metformin reduced viability and proliferation of MDA-MB-231 cells with or without medium renewal, thus demonstrating that 2-DG reduces their resistance to metformin. In sum, we show that medium renewal blocks anti-proliferative effects of metformin during prolonged treatments in low-glucose medium. Differences in medium renewal protocols during prolonged treatments might therefore lead to apparently inconsistent results as regards effectiveness of metformin as a direct anti-cancer agent. Finally, our results indicate that co-therapy with 2-DG and metformin might provide an effective strategy to overcome metformin resistance of breast cancer cells.
authors.affiliation: Faculty of Electrical Engineering, University of Ljubljana, Ljubljana, Slovenia.
authors.forename: Maruša
authors.initials: M
authors.lastname: Rajh
authors.affiliation: Faculty of Electrical Engineering, University of Ljubljana, Ljubljana, Slovenia.|Institute of Pathophysiology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors.forename: Klemen
authors.initials: K
authors.lastname: Dolinar
authors.affiliation: Institute of Pathophysiology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors.forename: Katarina
authors.initials: K
authors.lastname: Miš
authors.affiliation: Faculty of Electrical Engineering, University of Ljubljana, Ljubljana, Slovenia.
authors.forename: Mojca
authors.initials: M
authors.lastname: Pavlin
authors.affiliation: Institute of Pathophysiology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia.
authors.forename: Sergej
authors.initials: S
authors.lastname: Pirkmajer
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001943:Breast Neoplasms / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003847:Deoxyglucose / Q000494:pharmacology; D005260:Female; D006801:Humans; D008687:Metformin / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D000970:Antineoplastic Agents; D008687:Metformin; D003847:Deoxyglucose