Abstract 4639: Do cation-selective transporters help or hurt the antitumor efficacy of metformin in breast cancer

Introduction Metformin is effective against many cancers including breast cancer. Activation of intracellular adenosine monophosphate-activated protein kinase (AMPK) is implied in metformin anticancer efficacy. Due to its hydrophilicity and positive charge, metformin relies on cation-selective transporters for cellular uptake. Accordingly, metformin cellular uptake, AMPK activation and anti-proliferative effects increased when organic cation transporter 3 (OCT3) was expressed in BT20 cells (BT20-OCT3) that have very low levels of cation-selective transporters (Cai et al., AAPS Abstract W4368, 2013). Our previous studies showed that human breast cancer cell lines and tumors exhibit wide variability in expression of metformin transporters, e.g. OCT1-3, plasma monoamine transporter (PMAT), and multidrug and toxin extrusion (MATE) transporters 1-2 (Zhang et al., AACR Abstract 9800, 2012). This study aims to assess the pivotal role of transporters in metformin antitumor effects using xenograft mice with tumors from BT20 and OCT3-BT20 cells. Methods BT20 cells stably transfected with OCT3 were produced, and OCT3 activity verified by [14C]metformin (50µM) uptake in presence/absence of 50µM famotidine (OCT3 and MATE1 inhibitor) or 500 µM quinidine (pan transporter inhibitor). Optimum metformin dose for tumor study was assessed by a dose-range pharmacokinetic study in mice dosed intraperitoneally (IP) with 15, 30, 50, 100, and 150mg/kg [14C]metformin. Xenograft mice were produced using 2×106 BT20/OCT3-BT20 cells. Doxorubicin (DOX; 4mg/kg) or DOX + 100mg/kg metformin was given IP every 5 days, and tumor volumes measured over 25 days. Results Metformin plasma AUC0-24hr was linear up to 100 mg/kg (AUC0-24hr of 95, 160, 366, 770 and 510 µmol*hr/L at 15, 30, 50, 100 and 150 mg/kg, respectively); hence, this dose was used for antitumor efficacy study. Surprisingly, tumors were larger in DOX + metformin group compared to DOX (45.3 vs. 23.5 mm3) in mice with OCT3-BT20 tumors. In mice with BT20 tumors, tumors were smaller in DOX + metformin group compared to DOX (20.5 vs. 61.8 mm3). Since BT20 tumors grew slowly, tumor measurements were possible in only 2 mice/group. Hence, the more aggressive MCF-7 human breast cancer cell was used to develop an OCT3-expressing (OCT3-MCF7) clone in which metformin uptake was 4.5-fold higher than in MCF-7 cells (65.66 vs. 15.17 pmol/mg protein/min). Metformin antitumor effect in xenograft mice with MCF-7 and OCT3-MCF7 tumors is under evaluation. Conclusion In vitro data imply that OCT3 expression in human breast cancer cells enhances metformin cellular uptake and anti-proliferative effect. In vivo xenograft mice studies show that BT20 tumors over-expressing OCT3 are less responsive to metformin + DOX than native BT20 tumors; however, further studies are needed to show statistical significance. Citation Format: Hao Cai, Muhammad Wahajuddin, Ruth Everett, Dhiren R. Thakker. Do cation-selective transporters help or hurt the antitumor efficacy of metformin in breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4639. doi:10.1158/1538-7445.AM2014-4639