Abstract 2663: Determining the role of altered redox metabolism in lung tumorigenesis

The purpose of this study is to determine the pathways by which short-term cigarette smoke exposure induces expression of the cystine-glutamate antiporter xCT (SLC7A11) and its downstream implications in carcinogenesis. BEAS-2B lung epithelial cells and their tumorigenic counterpart, B39-TL cells, were exposed to cigarette smoke condensate (CSC) for 3-15 days. Cells were also exposed to a range of inhibitors. Erastin and buthionine sulfoximine (BSO) are inhibitors of various steps in the glutathione synthesis pathway, found to be hyperactive in xCT-overexpressing cells. Cells were also exposed to the drug mito-2-hydroxybenzoic acid (mito-2-HOBA), which scavenges isolevuglandins, reactive γ-ketoaldehydes implicated in oxidative stress and carcinogenesis. Expression of xCT and related proteins was measured using Western blotting. Glutathione levels were detected using the luciferase-based GSH-Glo assay (Promega). Oxidative stress was detected using dihydroethidium (DHE) fluorescence. It was found that CSC exposure significantly increased the expression of xCT and SLC1A5, a transporter that potentially works in synergy with xCT, and that tumorigenic B39-TL cells had significantly increased xCT and SLC1A5 expression compared to BEAS-2B cells. In addition, BEAS-2B cells exposed to CSC and a glutathione synthesis inhibitor such as erastin or BSO showed faster depletion of glutathione than cells treated with only inhibitor. Finally, it was found that BEAS-2B cells treated for 3 days with CSC did not show increased levels of superoxide compared to control cells. However, cells treated with CSC and mito-2-HOBA together showed significantly increased superoxide. These data suggest that CSC induces an antioxidant response which is ablated by mito-2-HOBA. This antioxidant response seems to at least partially rely on glutathione synthesis fueled by xCT, as evidenced by inhibitor studies. The response also seems to be activated by isolevuglandins, because removing these molecules significantly raises oxidative stress. The mechanistic link between CSC exposure, isolevuglandin generation, and xCT/glutathione induction will be clarified in further experiments. Citation Format: Alex Camai. Determining the role of altered redox metabolism in lung tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2663.