Abstract 1668: Association of stem cell-like phenotype with isoform specific functions of AKTs in hepatocytes

Abstract Introduction: Liver cancer is the sixth most common cancer and the third leading cause of cancer-related deaths worldwide. It is associated with abnormal activation of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, which is implicated in regulation of cancer stem cells (CSCs). The present study investigated the effects of AKT loss on stem cell-like phenotype in immortalized mouse hepatocytes. Methods: Immortalized hepatocytes were established from mice with Akt1 (Akt1−/−; Akt1-null) or Akt2 (Akt2−/−; Akt2-null) whole-body deletion. Cell morphology was assessed via microscopy. Cell proliferation was evaluated with hemacytometer count and MTT assay. Cell migration towards a chemoattractant was assessed via transwell assay. Cell viability was evaluated in response to chemotherapy treatment. The ability of cells to Results: When compared to WT cells, which exhibited cuboidal epithelial-like features, Akt1-null and Akt2-null cells demonstrated elongated mesenchymal-like morphology. Loss of either isoform was associated with a significantly decreased cell growth (p<0.05). Akt1-null cells migrated faster towards a chemoattractant in the transwell assay, were significantly more resistant to sorafenib, and formed a significantly higher number of spheres in the sphere formation assay (p<0.01). Proteomic analysis revealed that both Akt1-null and Akt2-null cells exhibited increased expression of CSC marker CD44, along with other proteins associated with liver cancer, including PDGFRA, CDH2, MMP14, and COX2 (p<0.05). Additionally, Akt1-null cells exhibited elevated levels of ANXA1 and ANXA3 (p<0.01). Conclusions: Loss of both AKT isoforms resulted in decreased cell proliferation and elevated expression of proteins with a previously reported role in liver cancer invasion, migration, and epithelial-mesenchymal transition. AKT1 loss correlated with an enhanced stem cell-like phenotype, including increased migration towards a chemoattractant, higher resistance to chemotherapy, and enhanced ability to form spheres. These results contribute to the understanding of the role of AKT isoforms in regulating stemness in liver cancer and may serve as a basis for investigation into isoform specific inhibitors. Further investigation into cellular mechanisms governing these phenotypes is required. Citation Format: Ielyzaveta Slarve, Yushan Wang, Mario Alba, Lina He, Bangyan Stiles. Association of stem cell-like phenotype with isoform specific functions of AKTs in hepatocytes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1668.