Relationship of SNCG, S100A4, S100A9 and LCN2 gene expression and DNA methylation in bladder cancer

Microarray analysis of paired cultures of normal and cancerous urothelial cells revealed differences in cytokeratin and adhesion gene expression. Normal cells expressed autocrine growth factor genes more strongly whereas carcinoma cells were distinguished by concomitant expression of urothelial and epidermal differentiation markers. Expression of SNCG, S100A9 and LCN2 was also enhanced. In other cancers, overexpression of SNCG, LCN2 and S100A4 has been ascribed to DNA hypomethylation. We therefore investigated expression and methylation of SNCG, S100A4, S100A9 and LCN2 in urothelial cancer cell lines and tissues. SNCG and S100A4 were overexpressed in some cancer tissues and cell lines, but downregulated in others, whereas LCN2 and S100A9 were upregulated in few cancer cell lines, but regularly in tissues. Normal and cancerous urothelial cells expressing SNCG lacked promoter methylation. SNCG downregulation was associated with hypermethylation and could be reversed by the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine. S100A4 methylation at regulatory intronic sites and in the promoter region was lowest in leukocytes and fibroblasts, and denser in urothelial cells. Gene expression responded to 5-aza-2'-deoxycytidine. LCN2 promoter methylation was variable and even less consistently related to expression. The S100A9 promoter was partially methylated in nonexpressing cells, but 5-aza-2'-deoxycytidine had no effect. Our data indicate that SNCG methylation is cell type-specific and the gene is hypermethylated in some urothelial cancers. S100A4, S100A9 and LCN2 are genes with moderate CpG-density that show a less stringent relationship between DNA methylation and gene expression. Therefore, changes in methylation of these genes in cancer should be interpreted cautiously. (C) 2008 Wiley-Liss, Inc.