Abstract 4894: Human endogenous retroviral over expression in endometrial carcinoma is due to hypomethylation of the 5′LTR
Authors: Matthias Ruebner, Reiner Strick, Franziska Dietz, Stephan Ellmann, Eli Stiegler, Falk Thiel, Matthias W. Beckmann, Pamela L. Strissel
Journal: Cancer Research (2010)

Abstract The purpose of this study was to determine the expression of envelope (env) genes from five Human Endogenous Retrovirus (HERV) family members in Endometrial Carcinomas (EnCa) and correlate with CpG-methylation of the 5′LTR HERV-W in EnCa and prestages compared to matched control tissues. Over 85% of all EnCa are classified as endometrioid expressing steroid hormone receptors and involve prior stages, like hyperplasia. HERV families embody over 8% of the human genome and are derived from ancestral germ line retroviral infections. Sixteen different functional HERV-env genes are essential for cytotrophoblast cell-cell fusion during placentogenesis, but are also implicated in tumorigenesis. Aberrant methylation and hypomethylation are hallmarks of tumorigenesis. Hypomethylated 5′ LTRs of HERVs correlating with an increased expression have been shown for HERV-K and HERV-W (ovary) in different human carcinomas. Previously, we demonstrated a significant step-wise increase of the Syncytin-1 HERV-W env gene at both mRNA and protein levels for EnCa and prestages compared to controls. A pivotal role for Syncytin-1 in EnCa cell-cell fusions and cellular proliferation was shown using RNAi technology after treatment of cell lines with cAMP analogs or steroid hormones and tamoxifen, respectively. Tamoxifen also activated phosphorylation of AKT (Thr308) and ERα (Ser118) leading to Syncytin-1 induction via non-genomic ER signalling. We have established methods for absolute quantification real time PCR (qPCR) for five different HERV-env genes and methylation-specific-PCR (MSP) using bisulfite treated genomic DNA. Methylated CpGs were verified by cloning and sequencing. Gene expression results of benign and malignant tissues (EnCa n=44, polyps n=21, hyperplasia n=12) compared to control postmenopausal endometrium (n=33) showed that the HERV-env genes Syncytin-1, HERV-K, ERV-3, HERV-P(b) and Syncytin-2 were significantly up-regulated. We observed that in control endometrium the highest expressed env was ERV3; whereas in hyperplasia and EnCa Syncytin-1 was the most prominent. Methylation studies of the Syncytin-1 5′ LTR identified 20 potential CpGs where many correlated with protein binding sites, e.g. for ERα, AP-1, GATA1-3 and MeCP2. MSP results showed that the 5′LTR region was hypomethylated in tumors compared to matched controls. Cloning and sequencing of PCR fragments showed that 3-4 CpGs corresponding with protein binding sites were hypomethylated in tumors (n=10) compared to matched controls. Studies using 5′Aza-2′Cytidine treatment of EnCa cell lines resulted in a 2.5-fold increase of Syncytin-1. In conclusion, up-regulation of Syncytin-1 in EnCa is due to specific hypomethylation of the 5′ LTR. Understanding epigenetic regulation of HERV-env genes will contribute to the molecular etiology in EnCa and stepwise induction from benign prestages but also other carcinomas. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. 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 4894.