Abstract 261: Blocking AKT2 downregulates SOX9/Notch signaling inhibiting PTEN-null driven liver tumorigenesis

Abstract Primary liver cancer, including hepatocellular carcinoma (HCC) and Cholangiocarcinoma (CCA), arise from the neoplastic transformation of hepatocytes and cholangiocytes in the liver. Their association with liver cancer stem-like cell (LCSC) makers presents a more aggressive behavior and high lethal rate. PTEN is a tumor suppressor gene that negatively regulates the PI3K/AKT pathway. Downregulation of Pten is observed in up to 70% of the cases in CCA and 50% of the cases in HCC. In mouse models lacking PTEN, CCA develops in 100% of the mice at 12 months of age regardless of whether the whole liver cells (LiPTEN, PtenloxP/loxP; Alb-Cre+), hepatocytes-only (HpPTEN, PtenloxP/loxP; R26RYFP; AAV8-TBG-Cre), or cholangiocytes-only (ChoPten, PtenloxP/loxP; R26RYFP; SOX9-CreERT) are targeted. Treatment of these mice with DDC at 3 months to obstruct the bile duct leads to an earlier CCA oncogenesis at 6 months of age. Using YFP to follow the lineage of tumor cells in the HpPTEN and ChoPTEN mice, we found that PTEN loss allows for transdifferentiation from cholangiocytes to hepatocytes or vice versa. To explore the mechanism for how PTEN regulates cell fate, we focused on SOX9, a transcription factor that plays a critical role in embryonic development and is believed to be the downstream target of NOTCH signaling. Hyperactivation of the SOX9 and NOTCH signaling are observed at the tumor stage of all the PTEN loss mouse models. We showed that knocking down SOX9 with siRNA reduced the sphere-forming ability of liver cells established from the mice mentioned above, indicating SOX9 promotes LCSC proliferation and sustains the tumorigenic potential of these liver cells. Gain-of-function assays of the Notch signaling through exposure to Jag1 ligand-coated extracellular matrix or transfection of Notch intracellular domain (NICD) both induced the expression of SOX9 significantly. The treatment of Pten-deleted mice with DAPT to inhibit NOTCH signaling showed concurrently decreased expression of Notch target Hes1 and SOX9. We also addressed the roles of AKTs in this PTEN-regulated CCA development. Our data show that deletion of Akt2 in the LiPTEN mice (LiPTEN-A2, Akt2loxP/loxP; PtenloxP/loxP; Alb-Cre+) significantly attenuates tumor development. Compared with the LiPTEN mice, loss of AKT2 in LiPTEN-A2 mice robustly and significantly reduces the expression of SOX9/NOTCH signaling and manifests a bile duct malformation at 12 months of age. Together, this finding suggests NOTCH activation regulates SOX9 and collaborates with PTEN loss to drive liver cancer progression, and inhibiting AKT2 specifically can block HCC-CCA formation through SOX9/NOTCH downregulation. Citation Format: Qi Tang, Jingyu Chen, Guo Zhang, Ielyzaveta Slarve, Ni Zeng, Lina He, Diala Alhousari, Phillip Nguyen, Shunan Hu, Shefali Chopra, Liyun Yuan, Bangyan L. Stiles. Blocking AKT2 downregulates SOX9/Notch signaling inhibiting PTEN-null driven liver tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 261.