Investigation of in vivo Metabolism in Hepatic Carcinomas

Metabolic reprogramming is a common feature of cancers with many showing specific alterations in both micro and macronutrient metabolism. In this study we evaluate the utility of bioluminescent technology in evaluating the metabolism of liver cancers in vivo. We assess both nutrient uptake and nutrient sensing probes to establish the handling and/or concentrations of lipids, glucose, nicotinamide riboside, iron, copper, and ROS in both healthy mice and tumor bearing mice. Our studies showed promise for the lipid, copper, iron, and ROS probes and were able to measure metabolic differences in copper and lipid metabolism between our different cancer models./nWe evaluated both hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC) models that were established through a hydrodynamic injection. Our bioluminescent study revealed a fatty acid uptake phenotype in our ICC model, but not our HCC models, which is a rare but increasingly documented phenotype. Classical cancer lipid metabolism focuses on the upregulation of de novo lipogenesis concomitant with a decreased dependence on exogenous fatty acids. Increased expression and activity of fatty acid synthase (FASN), the rate-limiting enzyme involved in de novo lipogenesis, is required for the survival and proliferation of many tumor cells, including hepatocellular carcinoma (HCC). Another study previously demonstrated that ICC development is insensitive to FASN deprivation. Our observation that ICC maintains robust fatty acid uptake rates suggests a role of exogenous fatty acids for the growth of ICC. Fatty acid transport proteins (FATPs), now classified as solute transporter family 27 (Slc27a1-6), are major transmembrane proteins for LCFA uptake and FATP2 and -5 are robustly expressed in the liver, FATP5 being liver specific. Using genetic manipulation and in vivo bio-imaging techniques, we measured the growth of ICC in an FATP5 knock out mouse model over time. Loss of FATP5 significantly impaired ICC growth indicating that tumor growth is dependent on exogenous fatty acid uptake. This was replicated using a knockdown system in both our ICC and HCC models, and only in ICC did we see a growth impairment. We also performed a survival experiment to see how long the growth effect in ICC was maintain. Over the course of 32 week, only three out of 7 mice grew robust tumor and when assayed for lipid uptake and metabolizing genes, two showed upregulation of FASN./nMetabolomic were also performed to determine the fate and function of these fatty acids comparing ICC, HCC, and non-tumorous liver. Our data showed a robust increase in free fatty acids, structural, signaling lipids, and acyl-carnitines suggesting a differential handling of exogenous versus endogenous fatty acids. We attempted to follow up on the increased levels of acyl-carnitines, which are implicated in beta-oxidation and energy generation. We treated mice with CPT1 inhibitor etomoxir and saw no significant growth effect. While it is obvious ICC is dependent on fatty acid uptake, and the inhibition of uptake is able to prevent tumor growth, the differential handling of the downstream fatty acids is more complex. In sum, this study has identified FATP5 as a therapeutic target for the treatment of ICC and potentially other liver cancers dependent on protein mediated exogenous fatty acid uptake.

Advisor: Stahl, Andreas