Liver cancer is one of the fastest increasing causes of cancer-related deaths in the U.S. and worldwide. There are approximately 700,000 deaths worldwide each year due to liver cancer. Risk factors for liver cancer include nonalcoholic fatty liver disease, diabetes, and obesity, and the rising incidence in these diseases is paralleled with an increasing incidence in liver cancer. The development of hepatocellular carcinoma, the major subtype of liver cancer, is a multistep process and often starts with chronic inflammation. However, the molecular and cellular causes of inflammation remain poorly understood. Nod2 is a bacterial innate immunity protein and Nod2 deficiency is associated with inflammatory diseases, diet-induced obesity and metabolic dysfunction, and obesity-dependent liver cancer. Preliminary data, using a mouse model for hepatocellular carcinoma, predicts that the development of liver tumorigenesis in Nod2-deficient mice on high fat diet (HFD) is associated with an inhibition of the AMP- dependent kinase (AMPK) pathway. AMPK is a master regulator of metabolic reprogramming and cell proliferation. However, the role of the AMPK pathway in liver tumorigenesis in Nod2-deficient mice has not been demonstrated. In the current project, the hypothesis that there is decreased activation of the AMPK pathway in Nod2-/- tumorigenic mice, which contributes to the development of hepatic tumors in these mice will be tested. In Aim 1, the role of Nod2 in the activation of the AMPK pathway will be determined. WT and Nod2-/- mice will be treated with the carcinogen dimethylbenz[a]anthracene (DMBA) and maintained on HFD (DMBA+HFD) to induce liver tumors and regulation of proteins in the AMPK pathway will be determined in the liver. In Aim 2, the role of the AMPK pathway in the development of obesity-dependent hepatic tumors in Nod2-/- mice will be determined. Nod2-/- DMBA+HFD mice will be treated with metformin to activate AMPK and monitored for the development of hepatic tumors. The results from these experiments will provide proof-of-concept for the role of Nod2 in activation of the AMPK pathway and for the role of this pathway in protection from the development of hepatocellular carcinoma and will provide preliminary data for future in-depth studies to determine the molecular basis of obesity-dependent liver cancer.