Abstract 5432: Modulation of glycolysis potentiates the cytotoxic effects of metformin in breast cancer cells.
Authors: Zhuang Yongxian , Chan Daniel K. , Miskimins W. Keith 
Published: August 2014

Abstract
Metformin hydrochloride is an oral anti-hyperglycemic drug used in the management of Type 2 diabetes. Recently it has been widely studied as an anti-cancer agent. Our previous findings indicate that metformin can inhibit cell proliferation and induce cell death in most breast cancer cell lines, including MCF7. However, MDAMB231 and SKBR3 cells are resistant to the cytotoxic effects of metformin. Commonly used tissue culture medium (DMEM) has high glucose levels (25 mM) compared to physiological conditions. Our previous studies with metformin were performed in this high glucose medium. We now report that reducing glucose levels in tissue culture medium, or inhibiting glycolysis, significantly enhances the sensitivity of breast cancer cells to metformin cytotoxicity. In low glucose conditions ranging from 0 to 5 mM MCF7, MDAMB231 and SKBR3 cells are all sensitive to metformin treatment. However, when glucose is increased to 10 mM or above, all of these cell lines are less responsive to metformin treatment. MCF7 cells are significantly more sensitive to metformin than the other two cell lines at all glucose concentrations. Supplementing the medium with fructose or galactose does not rescue the cells from death caused by metformin. A one day treatment with metformin significantly reduces ATP levels in cells incubated in medium with low glucose (2.5 mM), fructose (25 mM) or galactose (25 mM), but not in medium with high glucose (25 mM). This suggested that lowering glucose potentiates the effects of metformin on induction of cell death by reducing glycolytic flux. Glucose enters glycolysis after phosphorylation by hexokinase. In most cancer cells hexokinase II is overexpressed. 2-deoxyglucose (2DG), which reduces glycolysis by inhibiting hexokinase II, also significantly sensitizes breast cancer cells to metformin treatment. It has been documented that hexokinase II is localized to the outer mitochondrial membrane through its interaction with VDAC. This interaction facilitates efficient glycolytic metabolism and can also prevent apoptotic cell death. After metformin treatment in low glucose, or high fructose or galactose, we find that hexokinase II is translocated from the cytoplasm to the nucleus. Additionally, under low glucose conditions metformin significantly decreases active phosphorylated AKT. AKT has been shown to phosphorylate hexokinase II and to enhance its mitochondrial localization to prevent cell death. Thus, our data supports a model in which metformin treatment of breast cancer cells in low glucose leads to relocalization of hexokinase in an AKT-dependent manner, leading to inhibition of ATP production and cell death.
Citation Format: Yongxian Zhuang, Daniel K. Chan, W. Keith Miskimins. Modulation of glycolysis potentiates the cytotoxic effects of metformin in breast cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5432. doi:10.1158/1538-7445.AM2013-5432