B24 The Role of Cigarette Smoke and miR520a in Pulmonary Frizzled 9 Expression

Abstract

Lung cancer is the deadliest cancer, and for this reason treatment is highly researched. Alternatively, chemoprevention can be used to combat lung cancer in individuals who are at high risk of diagnosis, such as cigarette smokers. Frizzled 9 (Fzd9) is required in vitro for chemopreventive effects of iloprost, a prostacyclin analogue, in the lung. In non-small cell lung cancer (NSCLC) cell lines, Fzd9 activates PPARγ, leading to inhibition of transformed growth. Cigarette smoke exposure decreases Fzd9 expression. The goal of this study is to elucidate the relationship between cigarette smoke and miRNA regulation of Fzd9 expression. NSCLC cells exposed to cigarette smoke condensate (CSC) showed decreased Fzd9 3’ UTR activity, suggesting CSC regulates Fzd9 expression through miRNA. miRNA database analysis suggested miR-95, miR-106b, and miR-520a as potential regulators of Fzd9. Immortalized human bronchial epithelial cells (HBEC) and an Fzd9-positive NSCLC cell line (A549) transfected with miR-520a oligonucleotide mimic showed decreased Fzd9 3’UTR luciferase activity. miR-520a expression increased in HBEC and NSCLC cells after CSC exposure. We have tested a miR-520a inhibitor to use for future rescue experiments in an Fzd9-negative cell line (H322). Transient overexpression of miR-520a in HBEC and A549 did not affect cell viability or proliferation, so we made a stable miR-520a expressing HBEC line that we will use for longer-duration cell assays. miR-520a may play an important role in the regulation of Fzd9 by cigarette smoke, and future experiments will characterize this relationship and potentially impact the application of iloprost chemoprevention.