Abstract 2248: Both rpS6 and 4EBP1 are required for the carcinogenesis induced by coactivation of AKT and Ras

Abstract

Abstract AKT and Ras pathways are frequently concurrently hyperactivated in cancer. We have shown that co-expression of constitutively active forms of AKT (Myr-AKT) and activated Ras (N-RasV12) can rapidly induce liver tumors in mice. mTORC1 is the main downstream effector of AKT. mTORC1 phosphorylates rpS6 and 4EBP1, eukaryotic initiation factor eIF4E binding protein. Rapamycin is a well-characterized drug that partially inhibits mTORC1 activity. It can efficiently inhibit the phosphorylation of rpS6 but not the phosphorylation of 4EBP1. An unphophorylable mutant of 4EBP1, 4EBP1A4, is able to continuously bind to eIF4E and to inhibit eIF4E activity. Whether both of the two downstream effectors of mTORC1, rpS6 and 4EBP1, are required for hepatocarcinogenesis is unknown. The purpose of this study is to investigate the role of these two downstream effectors in hepatocarcinogenesis induced by co-activation of AKT and Ras. To determine the role of rpS6 in AKT/Ras-driven hepatocarcinogenesis, we treated AKT/Ras mice with rapamycin or vehicle daily for 7 weeks. After 7 weeks of vehicle treatment, all of the AKT/RAS/Veh mice developed large liver tumors. Lesions occupied ∼70-80% of the liver parenchyma. Histologically, the lesions consisted of large hepatocellular carcinonas (HCCs) and adenomas (HCAs). Tumor cells showed high proliferative activity, as indicated by Ki-67 staining. By contrast, after 7 weeks of Rapamycin treatment, none of the AKT/RAS/Rapa mice developed tumor. Small clusters of lipid-rich preneoplastic cells occupied 5-15% of the liver tissue. Few hepatocytes and non-parenchymal cells showed proliferative activity. Western blot analysis confirmed that Rapamycin inhibited the activation of rpS6 but had no effect on phosphorylated/inactivated levels of 4EBP1. These result indicated that Rapamycin treatment efficiently inhibited the progression of AKT/RAS induced liver tumor via inhibiting the rpS6 pathway. To determine the role of 4EBP1 in AKT/Ras-driven hepatocarcinogenesis, we overexpressed 4EBP1A4 or 4EBP1WT along with AKT and Ras into the mouse liver. Seven weeks after hydrodynamic injection, AKT/Ras/4EBP1WT mice developed large liver tumors which were equivalent to those developed in AKT/RAS/Veh mice. However, only few very small nodules developed in the livers of AKT/Ras/4EBP1A4 mice. The nodules consisted only of HCA and represented less than 5% of the lesional tissue. Notably, the proliferation in tumor cells of AKT/Ras/4EBP1A4 was more robust than in AKT/RAS/Rapa mice. The AKT/Ras/4EBP1A4 mice finally developed large liver tumors around 20 weeks after hydrodynamic injection. These results indicated that 4EBP1A4 efficiently delayed the AKT/RAS induced liver tumor. Altogether, our result indicates that the two main downstream effectors of mTORC1, rpS6 and 4EBP1, are both required for hepatocarcinogenesis induced by co-activation of AKT and Ras. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2248. doi:1538-7445.AM2012-2248