Abstract 166: CpG island methylation profiling in human salivary gland adenoid cystic carcinoma

Introduction A better understanding of key molecular changes during the pathogenesis of salivary gland adenoid cystic carcinoma (ACC) could impact strategies to reduce recurrence and mortality from this cancer. Two epigenetic events involved in the pathogenesis of cancer are hypermethylation of tumor-suppressor gene promoters associated with transcriptional repression and hypomethylation associated with gene reexpression and genomic instability. The aim of this study was to identify genes (1) strongly deregulated by epigenetic CpG island methylation, (2) involved in development and survival of ACC. Experimental Procedure We analyzed 4 patients (tumor ACC and matching normal tissue) for changes in DNA methylation using the methylated CpG island amplification and microarray (MCAM) method. For validation, CpG island regions of four genes, showing highest differences in methylation with the MCAM method, were analyzed by Pyrosequencing. Results The hyper- and hypomethylation tables showed variable ratio between ACC and normal tissue: 9 - 37 fold increase in hypermethylation for 13 associated genes, and 9 −11 fold changes in hypomethylation for only two associated genes. Strongest hypomethylations occur in regions near the TSS of PARVG, and MIR339. Strongest hypermethylations occur in regions around TSS of genes encoding predominantly transcription factors: EN1, PITX1, FOXE1, LBX2, IRX1, FOXl2, FOXA1, TBX4, and GBX2, except for 4 other genes: epoxide hydrolase 3 (EPHX3, removal of mutagenic epoxides), MT1G (or MT1H, cellular processes on binding, storage, and transport of Zn /metal ions), prestin (SLC26A5, transmembrane transport,) and FNDC1 (desmoplastic response). 4 CpG island regions, showing highest hypermethylation in MCAM, and associated with EN1, PITX1, FOXE1, and TBX4 genes were validated by Pyrosequencing. This method supported the MCAM results, with an average difference of 50% in CpG hypermethylation over all 4 gene regions investigated. The resulting 13 genes associated with the highest CpG island hypermethylations in neoplastic tissue imply, that these genes are actively expressed in normal tissue and silenced in salivary gland ACC. These results are supported by literature, where afore mentioned genes with their expressed proteins are known to be involved in various types of cancers, e. g. PITX1 as activator of the tumor suppressor p53. Conclusion The 13 strongest hypermethylated genes in ACC of salivary gland, with 9 of them expressing transcription factors, and 4 expressing other genes are known to be involved in developmental, apoptotic and other cancer inducing or supporting processes. These silenced genes most probably constitute main molecular markers for quality and severity in outcome of ACC in salivary gland. These genes furthermore provide clues, for which cancer-related pathways need to be intercepted for therapeutical approaches to suppress ACC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 166.