Abstract 5402: Synergism of Metformin and DFMO on colon cancer proliferation and migration.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Introduction: Metformin has been shown to inhibit cancer cell growth via activation of MAP kinase with resultant inhibition of mTOR signaling pathway. ODC, a key enzyme in polyamine biosynthesis, was found to be overexpressed in colon cancer. We determined the effect of metformin in combination with DMFO, an ODC inhibitor, on tumor proliferation and migration in human colon cancer cells. Methods: Human colon cancer cells, HCT 116 and HT 29, were treated with Metformin, DFMO, or combination. Cell proliferation assay was performed using Cell Titer Blue proliferation assay. Cell migration assay was performed by seeding colon cancer cells in 24-well plate. After overnight incubation, an artificial homogenous wound was created onto the monolayer with a plastic micropipette tip. After wounding, the migration of cells into the wound was observed up to 72 hours at various time points. Quantitative RT-PCR for ODC1 detection was performed using Assay-on-Demand Hs00159739-ml. GAPDH served as control. Western blotting was performed to determine the expression of AMP kinase, mTOR, p70S6K and 4E-BP1. Data were presented as means ± SD for the three separate experiments. For comparison between groups, the student's t test was used and p< 0.05 was considered to be statically significant. Results: There was dose-dependent (1-10 mM for DFMO and 1-20 mM for metformin) time-dependent (up to 96 hours) inhibition of cell proliferation and migration with either metformin or DFMO in both HCT 116 and HT 29 cells. Synergistic inhibition was observed with the combination treatment (5 mM DFMO and 5 mM Metformin) on both cell migration and proliferation at 24 hours compared with either agent alone. There was synergistic inhibition of ODC expression with the combined treatment compared with DFMO alone at 24 hours (p<0.01). The combination treatment also lead to significant enhancement of AMPK activation and increased phosphorylation of AMPKα at Thr-172 compared with metformin alone (p<0.01). The combination treatment resulted in significant inhibition of mTOR signaling with decreased phosphorylation of mTOR, p70S6K and 4E-BP1 in treated cancer cells, when compared with individual drug treatments. Inhibition of proliferation and migration was significantly greater in the mismatch-repair defective HCT 116 cells than HT 29. Conclusion: We have observed a synergistic effect of metformin and DFMO in suppressing human colon cancer proliferation and migration. The synergism reflects enhanced inhibition of both polyamine synthesis and mTOR signaling pathways by the drug combination. Citation Format: Yanping Zhang, Luz M. Rodriguez, Ronald Lubet, Eddy C. Hsueh. Synergism of Metformin and DFMO on colon cancer proliferation and migration. [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 5402. doi:10.1158/1538-7445.AM2013-5402