Abstract
Urothelial carcinoma (UC) is a common disease causing significant morbidity and mortality as well as considerable costs for health systems. Extensive aberrant methylation of DNA is broadly documented in early UC, contributing to genetic instability, altered gene expression and tumor progression. However the triggers initiating aberrant methylation are unknown. Recently we discovered that several genes encoding key enzymes of methyl group and polyamine metabolism, including Ornithine Decarboxylase 1 (ODC1), are affected by DNA methylation in early stage UC. In this study, we investigated the hypothesis that these epigenetic alterations act in a feed-forward fashion to promote aberrant DNA methylation in UC. We demonstrate that siRNA-mediated knockdown of ODC1 expression elicits genome-wide LINE-1 demethylation, induction of LINE-1 transcripts and double-strand DNA breaks and decreases viability in primary cultured uroepithelial cells. Similarly, following siRNA-mediated knockdown of ODC1, UC cells undergo double-strand DNA breaks and apoptosis. Collectively, our findings provide evidence that ODC1 gene hypermethylation could be a starting point for the onset of genome-wide epigenetic aberrations in urothelial carcinogenesis. Furthermore, LINE-1 induction enabled by ODC1 interference provides a new experimental model to study mechanisms and consequences of LINE-1 activation in the etiology and progression of UC as well as presumably other cancers.
Highlights
Urothelial carcinoma (UC), the most common cancer of the urinary bladder, is a frequent disease with yearly 549,393 new cases and 199,922 deaths worldwide[1]
During a genome-wide screening of pTa and pT1 urothelial cancer tissue samples for altered DNA methylation, we observed distinct hypermethylation at the promoters of key genes of methyl group and polyamine metabolism pathways[18]. Disturbances of these key enzymes are known to lead to grave imbalances in the delicate intracellular SAM:SAH ratio resulting in genome wide DNA methylation alterations, including genome-wide LINE-1 hypomethylation[19,20,21]
We observed LINE-1 hypomethylation in 6 pTa and 6 pT1 urothelial carcinoma samples compared to 3 samples of healthy urothelium and 4 samples of tumor-adjacent uroepithelial tissue
Summary
Urothelial carcinoma (UC), the most common cancer of the urinary bladder, is a frequent disease with yearly 549,393 new cases and 199,922 deaths worldwide[1]. Tobacco smoking is thought to contribute to 50% of all UC cases as a risk factor[3] Aromatic amines such as 2-naphthylamin and polycyclic aromatic hydrocarbons in cigarette smoke cause mutations in key cancer-related genes by forming DNA adducts[4]. The methylome which consists of all methylated cytosine-guanosine (CpG) dinucleotides, forms an elaborate, plastic and cell-type specific pattern in mammalian cells. It supports genome organization, gene regulation and preserves genome integrity by repressing transposable genetic elements[7]. It has been proposed that LINE-1 hypomethylation and activation may play a causative role in urothelial carcinogenesis by inducing genetic instability which accompanies cancer progression, especially in high-stage and high-grade cancer[15]. Non-invasive and early invasive papillary transitional cell carcinomas of stages pTa and pT1, respectively, already present a plethora of genetic aberrations including losses and gains of their genetic material[16,17], with an increase of genomic instability when tumors progress further[17]
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