Abstract 811: Induction of stem-like cells with malignant properties by chronic exposure of immortalized normal human urothelial cell line to arsenic

Abstract

Abstract Although there is extensive epidemiologic evidence of increased risk for the development of urothelial carcinoma (UC) associated with arsenic exposure, the mechanisms by which arsenic participates in tumorigenesis are not well understood. To gain mechanistic insights, we recently developed an in vitro arsenic model in an immortalized normal human bladder cell line (HUC1) and in this study we performed extended phenotypic and molecular characterization of the arsenic exposed in vitro model. To assess the impact of arsenic exposure at a time progressive manner, we performed MTT assay using the arsenic exposed and unexposed HUC1 cells. The growth of the HUC1 cells was increased in a time dependent manner after arsenic exposure. In soft agar assay, colonies were observed only in arsenic exposed cells. Similarly, invaded cells in invasion assay were observed only in arsenic exposed cells. To identify whether arsenic exposure induce self-renewal properties, we performed spheres forming assay and our data showed arsenic exposed cells formed more spheres under serum-free medium and non-adherence condition, and the number gradually increased with longer periods of exposure. The sphere forming ability was maintained through two subsequent serial passages, indicating of self-renewal ability. The spheroid cells showed differentiated morphology when cultured in serum containing medium. Finally, to identify epigenetic events associated with the later phenotypic alterations, we performed illumina methylation array and expression array and determined a panel of genes that are epigenetically altered that may be related to arsenic-induced tumorigenesis. Among 29 genes that exhibited reduction of promoter methylation and up-regulation of the expression, 6 genes (EDNRA, GJB2, KRT17, GDF15, S100P, and LCN2) were reported as the oncogenes. Furthermore 2 genes (GJB2 and KRT17) were maintained the alteration even in withdrawal of arsenic for 2.5 months. While, among 13 genes that exhibited induction of promoter methylation and down-regulation of the expression, one gene (TSPYL5) as a tumor suppressor gene was maintained the alteration in withdrawal of arsenic for 2.5 months. In conclusion, chronic arsenic exposure induces malignant phenotype with the acquisition of stem-like activity, and epigenetic alteration might be partial mechanism of tumorigenesis. Validation and detail molecular understanding of later genes are ongoing to determine the arsenic associated pathway alterations in the genesis of UC. Citation Format: Akira Oki, Shahnaz Begum, Mariana Brait, David Sidransky, Mohammad Obaidul Hoque. Induction of stem-like cells with malignant properties by chronic exposure of immortalized normal human urothelial cell line to arsenic. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 811. doi:10.1158/1538-7445.AM2015-811