P1.02-006 Arsenic Promotes Persistent Alterations in the Lung PiRNA Transcriptome to Target Epigenetic Pathways

Abstract

Chronic exposure to arsenic leads to the onset of different diseases, including lung cancer. Arsenic-induced lung tumors have been associated with a high-frequency of lung squamous-cell carcinomas among never smokers (a rare epidemiological pattern), suggesting a unique underlying biology. Epigenetic alterations are known to play a role in this process; however, detailed mechanisms are not yet fully elucidated. Piwi-interacting RNAs (piRNAs), a novel class of small non-coding RNAs (sncRNAs), play a key role in epigenetic regulation and maintenance of genome integrity. Here, we examine the impact of different arsenic species in the human piRNA transcriptome, using lung cell models mirroring chronic, low dose exposure. We also investigate the interaction network of deregulated piRNAs and identified biological pathways potentially affected. One normal lung (HBEC) and two lung cancer cell lines: A459 (adenocarcinoma) and H520 (squamous-cell carcinoma) were grown in 10 ppm of sodium arsenite (AsIII) or arsenate (AsV) for six passages. Total RNA was extracted at different time points and sequenced. piRNA expression was deduced using our custom sncRNA analysis pipeline, which interrogates >23K piRNA-encoding human loci. piRNA/DNA binding prediction was performed using two different algorithms (miRanda/ThermoBLAST). Network analysis was performed using Partek Pathways. Overall, 691 piRNAs were expressed. Persistent changes in piRNA expression over time were identified, with specific patterns associated with the different arsenic species. In HBECs (non-malignant lung tissue), 14 piRNAs were persistently upregulated and 16 downregulated in response to AsIII. Similarly, 6 were up- and 11 downregulated when the same cells were exposed to AsV. Only 1 piRNA, DQ598008, was commonly upregulated in response to both arsenic species, while 4 piRNAs were commonly downregulated. Lung cancer cell lines follow the same arsenic species-specific trends, with a high subtype-specificity indicating these species maintain a role during lung tumor development. Remarkably, we found an enrichment of genes associated with methyltransferase activities predicted to be targeted by piRNAs altered by AsIII (a biologically-relevant form of arsenic), evidencing their role in arsenic-related carcinogenic mechanisms. Arsenic induces persistent alterations in the lung sncRNA transcriptome, particularly piRNAs, impacting pathways linked to epigenetic regulation. Together, these results provide insights into sncRNA-related mechanisms in arsenic-induced lung carcinogenesis. Moreover, different arsenic species induce distinct alteration patterns, highlighting the relevance of the source of exposure. piRNAs, as with other sncRNAs, are stable in biofluids, circulating tumour cells, and archival clinical materials. Therefore, piRNAs hold great promise as potential exposure and monitoring biomarkers for arsenic-related health effects.