PROJECT SUMMARY Although hepatocellular carcinoma (HCC) is the most common form of liver cancer, there are few therapies that are effective in treating this malignant disease. With less than 10% of patients who have HCC surviving beyond 5 years, it is important to identify additional therapeutic vulnerabilities. Metabolic alterations are a key feature of HCC pathogenesis, and represent targets that may reveal liabilities in HCC. Glucose-6-Phosphatase (G6PC) is recurrently downregulated in HCC and the magnitude of this downregulation correlates with patient outcome. Sequencing studies of HCCs reveal that somatic mutations occur in a subset of these malignancies. Additional evidence for G6PC involvement in HCC comes from Glycogen Storage Disease Type 1a (GSD1a), where germline mutations in G6PC lead to benign and malignant forms of liver cancer without cirrhosis. The functional relevance of recurrent mutations in G6PC or loss of G6PC expression to HCC pathogenesis in unclear. I hypothesize that loss of G6PC stimulates anabolic metabolic pathways required for tumor cell growth and are sufficient to drive hepatocyte proliferation and tumor initiation. The Specific Aims of this proposal aim to (1) determine how loss of G6PC alters metabolism to support tumor growth, (2) test whether G6PC can increase proliferation, and (3) explore ways in which G6PC loss influences liver tumors initiation and progression in mouse HCC models. We will use G6PC deficient in vitro and in vivo models that faithfully recapitulate aspects of GSD1a biology in patients, and stable isotope tracing to dissect how loss of G6PC rewires metabolic pathways. Altogether, these Aims will elucidate how G6PC loss influences non-cirrhotic development of HCC, and will provide insight into Glycogen Storage Disease Type 1a. We hope to identify metabolic liabilities in HCC pathogenesis that may provide new therapeutic strategies for this deadly disease.