Abstract A44: Endothelin A receptor-mediated biased signaling is a new player in modulating ovarian cancer progression
Authors: Teoh Jian-peng , Wang Yongchao , Tang Yaoping , Park Kyoung-mi , Kim Il-man 
Published: August 2014

Abstract
Ovarian cancer (OC) is the leading cause of gynecological cancer deaths among women in the United States. Although there has been major progress in our understanding of the biology and molecular genetics of OC, mortality rate has not been significantly improved in the past decades. Hence, introduction of new agents and refinement of existing targeted therapies through advancing biological insights are warranted. The endothelin-1 (ET-1)/endothelin A receptor (ETAR, a G protein-coupled receptor [GPCR]) axis confers pleiotropic effects on both tumor cells and tumor microenvironments, modulating epithelial to mesenchymal transition, chemo-resistance, and other tumor-associated processes through activating Gαq- and β-arrestin-mediated pathways. Because ETAR is ubiquitously expressed in the body, the current therapeutic approach to OC using ETAR antagonists without detailed knowledge of downstream signaling is prone to adverse side effects and reduced efficiency. In order to tackle the challenges ahead in terms of successful clinical application, this project seeks to employ the concept of “biased GPCR signaling”, in which β-arrestin-biased ligands and G protein-biased ligands demonstrate unique pharmacological properties that allow selective blocking of undesired complications while maintaining desired effects. The ability of biased ligands to differentially activate different signaling pathways via the same receptor raises the possibility of manipulating existing ETAR antagonists to achieve selective induction of favorable signaling pathways. The roles of individual G protein subunits such as Gαs in ETAR-mediated OC progression were therefore investigated in order to define tumor suppressive and oncogenic downstream effects. We hypothesized that Gαs-coupled signaling of ETAR confers tumor suppressive effects on OC.
Methods: To identify the roles of Gαs and Gαq in ETAR-mediated in vitro tumorigenesis, HEY (human ovarian cancer) cells endogenously overexpressing ETAR were pre-treated with pharmacological agents including forskolin (adenylate cyclase activator inducing PKA signaling), H-89 (PKA inhibitor), PMA (PKC activator), and Ro-318425 (PKC inhibitor) followed by ET-1 stimulation. We then performed cell migration, TUNEL, wound healing, and soft agar assays to test our hypothesis. Genetic gain- and loss-of-function studies using expression plasmids and lentiviral shRNAs for G proteins and β-arrestins were also performed.
Results: Consistent with previous studies, HEY cells treated with ET-1 showed an increase in cell proliferation and migration and ETAR-mediated β-arrestins signaling pathways induced OC cell migration. We also confirmed the oncogenic roles of Gαq-mediated ETAR signaling. Interestingly, Gαs-coupled ETAR signaling was found to inhibit OC cell migration, wound healing and colony formation on soft agars, but induced OC cell apoptosis. Genetic functional studies also demonstrated that ETAR-mediated Gαs signaling pathways inhibited OC cell migration, indicating its novel tumor suppressive effects in OC.
Conclusion: Our data suggest that biased signaling in ETAR is a plausible strategy to refine the existing ETAR antagonists for OC. The identification of Gαs-coupled ETAR signaling as a novel tumor suppressive signaling pathway will contribute to the development of improved ligand therapies for OC with more sustained efficacy. This novel discovery will enable a greater understanding of cancer biology, thereby facilitating the pursuit of biased signaling mechanisms in ETAR-mediated OC therapies.
Citation Format: Jian-peng Teoh, Yongchao Wang, Yaoping Tang, Kyoung-mi Park, Il-man Kim. Endothelin A receptor-mediated biased signaling is a new player in modulating ovarian cancer progression. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: From Concept to Clinic; Sep 18-21, 2013; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2013;19(19 Suppl):Abstract nr A44.