Abstract 4783: ADO is essential for redox homeostasis in liver cancer

Abstract

Abstract 2-Aminoethanethiol dioxygenase (ADO) is a thiol dioxygenase that plays a direct role in both metabolism and protein stability. Oxidation of the metabolite cysteamine produces hypotaurine, while oxidation of N-terminal cysteines targets protein substrates for N-degron pathway-mediated degradation. Despite these known functions, the (patho)physiological roles of ADO remain obscure. By analyzing TCGA datasets we discovered that high ADO expression is associated with poor outcome for patients with hepatocellular carcinoma (HCC) (HR 1.2, p<0.001). HCC is linked to viral hepatitis, alcohol, metabolic syndrome, non-alcoholic fatty liver disease and chronic inflammation. It is the third leading cause of cancer-related deaths worldwide and is significantly more common in males than females. With poor outcomes, there is a need to better understand HCC biology and vulnerabilities. To assess the functional roles of ADO, we created ADO knockout mice as well as two liver cancer cell line models where ADO was depleted by siRNA, doxycycline-inducible shRNA, or CRISPR/Cas9. While male ADO knockout mice were viable, fertile and healthy, depletion of ADO in liver cancer models significantly reduced cancer cell proliferation and colony formation. ADO was also essential for supporting xenograft growth when implanted subcutaneously in immune-compromised mice. Taken together, this suggested that ADO depletion represents a cancer-specific vulnerability. Comprehensive metabolomic, transcriptomic and proteomic characterization of HCC cells isogenic for ADO demonstrated that loss of ADO resulted in dysregulation of glutathione, ascorbate, polyamine and proline metabolism. Consistent with this, ADO depleted cells had high levels of reactive oxygen species (ROS) measured by CellROX flowcytometry, and an increased ratio of oxidized to reduced glutathione, indicative of oxidative stress. Since mitochondria represent a substantial source of ROS, we assessed mitochondrial mass and function using MitotrackerTM flow cytometry and Seahorse stress tests, respectively. ADO depleted cancer cells had higher mitochondrial mass and higher maximal respiration rates compared to control cells. Finally, exogenously supplied antioxidants could rescue the survival of HCC cells with ADO depletion, demonstrating that the observed loss of viability is due to oxidative stress. This work shows that ADO is essential for redox homeostasis in liver cancer models and suggests that interfering with ADO function may represent a novel targeting strategy for HCC. Citation Format: Sandy C-E Lee, Andrea H. Pyo, Helia Mohammadi, Ji Zhang, Anna Dvorkin-Gheva, Lucie Malbeteau, Stephen Chung, Shahbaz Khan, Thomas Kislinger, Julie A. Reisz, Courtney Jones, Marianne Koritzinsky. ADO is essential for redox homeostasis in liver cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4783.