Metformin Inhibits Migration and Invasion by Suppressing ROS Production and COX2 Expression in MDA-MB-231 Breast Cancer Cells.

BACKGROUND: Several mechanisms of action have been proposed to explain the apparent antineoplastic functions of metformin, many of which are observed at high concentrations that may not be reflective of achievable tissue concentrations. We propose that metformin at low concentrations functions to inhibit ROS production and inflammatory signaling in breast cancer, thereby reducing metastasis. METHODS: Using the highly invasive MDA-MB-231 breast carcinoma model, we ascertained the impact of metformin on cell viability by DNA content analysis and fluorescent dye exclusion. Migration and invasion assays were performed using a modified Boyden chamber assay and metastasis was ascertained using the chorioallantoic membrane (CAM) assay. PGE2 production was measured by Enzyme-Linked Immunosorbent Assay (ELISA). COX2 and ICAM1 levels were determined by flow cytometry immunoassay. RESULTS: Metformin acutely decreased cell viability and caused G2 cell cycle arrest only at high concentrations (10 mM). At 100 µM, however, metformin reduced ICAM1 and COX2 expression, as well as reduced PGE2 production and endogenous mitochondrial ROS production while failing to significantly impact cell viability. Consequently, metformin inhibited migration, invasion in vitro and PGE2-dependent metastasis in CAM assays. CONCLUSION: At pharmacologically achievable concentrations, metformin does not drastically impact cell viability, but inhibits inflammatory signaling and metastatic progression in breast cancer cells.

In-vivo evidence of nephrotoxicity and altered hepatic function in rats following administration of diglycolic acid, a metabolite of diethylene glycol.

CONTEXT: Diglycolic acid (DGA) is one of the two primary metabolites of diethylene glycol (DEG). DEG is an industrial solvent that has been implicated in mass poisonings resulting from product misuse in the United States and worldwide, with the hallmark toxicity being acute kidney injury, hepatotoxicity, encephalopathy and peripheral neuropathy. Our laboratory has generated in-vitro evidence suggesting that DGA is the metabolite responsible for the proximal tubule necrosis and decreased kidney function observed following DEG ingestion. Furthermore, we have shown that DGA specifically accumulates in kidney tissues (100× higher than peak blood concentrations) following DEG administration. OBJECTIVE: To examine renal and hepatic accumulation and dysfunction following direct administration of DGA in-vivo. We hypothesize that administration of DGA will result in renal and hepatic DGA accumulation, as well as proximal tubular necrosis and liver injury. MATERIALS AND METHODS: Adult male Wistar rats were divided into three groups dosed with 0, 100 or 300 mg/kg DGA via single oral gavage. Urine was collected every 6-12 h and blood, kidneys and liver were removed upon sacrifice at 48 h post-dosing for analysis. RESULTS: DGA accumulated significantly in both kidney and liver tissue only at 300 mg DGA/kg. DGA concentrations in the kidneys and liver correlated with renal and hepatic injury, respectively. Histopathological and clinical chemistry analysis revealed that DGA-treated animals exhibited moderate liver fatty accumulation and marked renal injury, again only at 300 mg/kg. DISCUSSION: DGA-induced kidney injury demonstrated a steep dose response threshold, where severe damage occurred only in animals given 300 mg/kg DGA, while no toxicity was observed at 100 mg/kg. CONCLUSION: These results provide evidence for in-vivo toxicity following direct administration of DGA, a metabolite of DEG. The steep dose-response threshold for toxicity suggests mechanistically that there is likely a saturable step that results in DGA accumulation in target organs.