Abstract A037: Exploring the therapeutic effectiveness of EO3001 in the treatment of clear cell ovarian cancers with ARID1A mutations
Authors: Ding Yuchen , Li Lucy Yuqin , Zhang Joyce Yuhan , Kabeer Farhia , Lam TszYin , Kalantari Forouh , Bacha Jeffrey , Brown Dennis , Kanekal Sarath , Sankar Neil , Lizardo Michael , EL-Naggar Amal , Huntsman David 
Published: March 2024

Abstract
Introduction: Clear Cell Ovarian Cancer (CCOC) is an inherently chemoresistant subtype of ovarian cancer, accounting for approximately 10% of all epithelial ovarian carcinoma (EOC) in north America and up to 30% EOC in patients in Japan. Recent evidence suggests that hematogenous spread plays a crucial role in CCOC metastasis. However, this aspect has been largely overlooked due to the absence of an appropriate model. ARID1A, a subunit of the SWI/SNF complex, is mutated in around 65% of CCOC cases and has been identified as a key oncogenic factor in CCOC transformation. Mutations in the ARID1A gene result in an accumulation of reactive oxygen species (ROS) and a decrease in the antioxidant glutathione (GSH) due to the downregulation of SLC7A11, a transporter responsible for importing the GSH precursor cystine. Consequently, cancer cells with ARID1A mutations become highly reliant on oxidative phosphorylation (OXPHOS) as an energy source. EO3001, a synthetic small molecule drug that has been studied in various cancers, can selectively transport extracellular Cu(II) to mitochondria. In mitochondria, Cu(II) can accumulate and induce mitochondrial ROS, thereby triggering cuproptosis. Recent studies have shown that ARID1A-deficient cells are more sensitive to EO3001. The Pulmonary Metastasis Assay (PuMA), an ex vivo model initially established for sarcomas, has been proven to provide a richer microenvironment compared to traditional 2D and 3D culture models. In PuMA, cancer cells labeled with fluorescent tags are inoculated into lung sections of mice inflated with agarose gel. The cells are then treated with drugs and/or different conditions, and the growth of cancer cells is measured and quantified by the fluorescence intensity of the lung sections. Methods: In this study, we generated isogenic CCOC cell lines (RMG-1, OVCA429, and JHOC-5 -/+ ARID1A loss) using CRISPR-Cas9. We then evaluated the therapeutic effects of EO3001 on the tumorigenic potential and cell growth of these cells in vitro and ex vivo: To assess the effects of EO3001, we performed multiple in vitro assays, including cell viability, cell proliferation, total and mitochondrial ROS levels, OXPHOS activities, migration, and invasion assays on CCOC cells with and without ARID1A loss, treated with or without EO3001 under various stress conditions. Additionally, we established and optimized the PuMA assay on CCOC cell lines to study the effects of EO3001 on the interactions between cancer cells and surrounding normal tissues. Conclusions: While there is a notable distinction in the response to EO3001 between CCOC -/+ ARID1A loss cell lines in both in vitro and ex vivo models, it is noteworthy that hypoxia significantly diminishes the effects of EO3001, potentially attributed to alterations in glycolysis and OXPHOS. Therefore, taking advantage of the susceptibility arising from the dependence on OXPHOS in ARID1A-deficient CCOC, the utilization of EO3001 could potentially present a highly encouraging and viable approach for the therapeutic management of these patients.
Citation Format: Yuchen Ding, Lucy Yuqin Li, Joyce Yuhan Zhang, Farhia Kabeer, TszYin Lam, Forouh Kalantari, Jeffrey Bacha, Dennis Brown, Sarath Kanekal, Neil Sankar, Michael Lizardo, Amal EL-Naggar, David Huntsman. Exploring the therapeutic effectiveness of EO3001 in the treatment of clear cell ovarian cancers with ARID1A mutations [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A037.