Date of Award
Doctor of Philosophy (PhD)
Cellular, Molecular and Biomedical Sciences
Albert van der Vliet
The NADPH oxidase homolog, dual oxidase 1 (DUOX1), is an H2O2 producing transmembrane enzyme highly expressed in the airway epithelium. DUOX1-dependent redox signaling has been characterized to regulate many homeostatic processes in the lung epithelium, such as host defense, wound healing, and type II immune responses. Intriguingly, DUOX1 has been found to be suppressed in many epithelial cancers, including lung cancer, by hypermethylation of its promoter. Epigenetic silencing of DUOX1 in cancer is paradoxical to the understanding that tumors harbor elevated levels of reactive oxygen species (ROS), suggesting that DUOX1 may be a tumor suppressor.
Since DUOX1 loss occurs in many forms of lung cancer, we aimed to characterize the functional importance of DUOX1 suppression. RNAi-mediated knockdown of DUOX1 in lung epithelial cells induced features of the epithelial-to-mesenchymal transition (EMT), a characteristic of aggressive or invasive tumor cells. Indeed, DUOX1 suppression promoted the acquisition of molecular signatures associated with EMT, such as the loss of E-cadherin, and induced expression of vimentin and smooth muscle actin. Additionally, we find that DUOX1 suppression promotes the acquisition of other EMT-related features, such as enhanced levels of cancer stem cell molecular markers, cellular invasiveness, and critically, resistance to epidermal growth factor receptor (EGFR) inhibition. Importantly, overexpression of DUOX1 in DUOX1-lacking lung cancer cells promoted the recovery of epithelial characteristics, pinning DUOX1 as a critical mediator of the epithelial phenotype.
Based on prior studies demonstrating DUOX1 as an important regulator of EGFR signaling in the lung epithelium, we hypothesized that DUOX1 loss in lung cancer may impact EGFR regulation. EGFR belongs to a larger family of ErbB receptor tyrosine kinases, which are often overexpressed or mutated in many forms of lung cancer. Surprisingly, we find that lung cancer cells lacking DUOX1 have significantly altered EGFR redox regulation, specifically, kinetically enhanced cysteine oxidation-reduction dynamics. Additionally, our results demonstrate DUOX1-lacking cancer cells have altered intracellular EGFR trafficking with enhanced nuclear targeting. Indeed, we observe many oncogenic features of nuclear EGFR e.g. enhanced migratory capacity, resistance to EGFR blocking antibodies. Finally, we have uncovered that EGFR cysteine redox dynamics may regulate intracellular trafficking and/or nuclear transport, offering potentially novel avenues in the design of therapeutics.
Proper DUOX1 localization and enzymatic function in the plasma membrane requires partnership with its maturation factor, dual oxidase maturation factor 1 (DUOXA1). Preliminary findings from a newly designed DUOX1-DUOXA1 co-expression system suggests that following enzymatic activation of DUOX1, DUOXA1 dissociates from DUOX1 and potentially translocates to the nucleus, a feature not previously described in lung epithelial or cancer cells. While these preliminary results require additional experimentation, this could be a unique regulatory feature of DUOX1 and a novel role for DUOXA1.
Collectively, the research demonstrated in this dissertation characterizes the functional and mechanistic importance of DUOX1 suppression in cancer. Indeed, loss of DUOX1 expression may be an indicator of tumor aggressiveness and responsiveness to EGFR-targeted therapies, warranting its potential for use as a clinical biomarker in lung cancer.
Number of Pages
Little, Andrew Charles, "Functional and Mechanistic Consequences of Dual Oxidase 1 Suppression in Lung Cancer" (2017). Graduate College Dissertations and Theses. 810.
Available for download on Friday, September 13, 2019