PROJECT SUMMARY Hepatocellular carcinoma (HCC) is the third leading cause of cancer related deaths worldwide and its incidence is increasing due to endemic obesity. Sexual dimorphism exists in HCC incidence as women have a significantly lower risk for developing HCC than men. However, the molecular mechanisms of HCC sexual dimorphism remain unclear, hindering development of better therapies for this disease. This proposal will utilize a new model that we developed of diet-induced progression of NASH to HCC that recapitulates key physiological, metabolic, histologic and transcriptomic changes observed in the human disease to examine previously unrecognized roles of sphingosine kinase 2 (SphK2), an enzyme that regulates the balance of bioactive sphingolipid metabolites, sphingosine-1-phosphate (S1P) and ceramide, in sexual dimorphism of HCC. Similar to humans, we found that on this diet, wild-type male mice, but not females, develop HCC. Strikingly, SphK2 knockout male mice have reduced tumor incidence, whereas in females, liver cancer developed only in SphK2 knockout mice. Thus, we propose that SphK2 plays a critical role in sexual dimorphism of HCC. We will test the central hypothesis that SphK2 protects females from HCC, while promoting it in males through several mutually non-exclusive mechanisms in distinct hepatic subcellular organelles. Aim 1 will examine the role of liver SphK2 in key signaling pathways that promote HCC in males and Aim 2 will define the role of SphK2 in nuclear and mitochondrial mechanisms mediating resistance of females to HCC. Our proposal utilizes a unique animal model and state-of-the-art techniques to identify novel SphK2-regulated molecular mechanisms involved in sexual disparity in diet-induced progression of NASH to HCC, and also has translational implications for the use of SphK2 inhibitors now in clinical trials. This study will lead to better understanding of sex differences in HCC important for personalized treatment strategies for this devastating disease.