Quantitative DNA methylation analyses reveal stage dependent DNA methylation and association to clinico-pathological factors in breast tumors
1 Department of Clinical Molecular Biology and Laboratory Science (EpiGen), Akershus University hospital, Division of Medicine, 1476 Lørenskog, Norway
2 Institute for Clinical Medicine, Faculty of Medicine, University of Oslo, 0372 Oslo, Norway
3 Department of Genetics, Institute for Cancer Research, OUS Radiumhospitalet Montebello, 0310 Oslo, Norway
4 Laboratory for Epigenetics, Centre National de Génotypage, CEA – Institut de Génomique, 91000 Evry, France
5 Department of Surgery, Uppsala University Hospital, Uppsala, Sweden
6 Department of Surgery, Akerhus University Hospital, Oslo, Norway
7 Section of Oncology, Institute of Medicine, University of Bergen and Department of Oncology, Haukeland University Hospital, N5021 Bergen, Norway
BMC Cancer 2013, 13:456 doi:10.1186/1471-2407-13-456Published: 5 October 2013
Aberrant DNA methylation of regulatory genes has frequently been found in human breast cancers and correlated to clinical outcome. In the present study we investigate stage specific changes in the DNA methylation patterns in order to identify valuable markers to understand how these changes affect breast cancer progression.
Quantitative DNA methylation analyses of 12 candidate genes ABCB1, BRCCA1, CDKN2A, ESR1, GSTP1, IGF2, MGMT, HMLH1, PPP2R2B, PTEN, RASSF1A and FOXC1 was performed by pyrosequencing a series of 238 breast cancer tissue samples from DCIS to invasive tumors stage I to IV.
Significant differences in methylation levels between the DCIS and invasive stage II tumors were observed for six genes RASSF1A, CDKN2A, MGMT, ABCB1, GSTP1 and FOXC1. RASSF1A, ABCB1 and GSTP1 showed significantly higher methylation levels in late stage compared to the early stage breast carcinoma. Z-score analysis revealed significantly lower methylation levels in DCIS and stage I tumors compared with stage II, III and IV tumors. Methylation levels of PTEN, PPP2R2B, FOXC1, ABCB1 and BRCA1 were lower in tumors harboring TP53 mutations then in tumors with wild type TP53. Z-score analysis showed that TP53 mutated tumors had significantly lower overall methylation levels compared to tumors with wild type TP53. Methylation levels of RASSF1A, PPP2R2B, GSTP1 and FOXC1 were higher in ER positive vs. ER negative tumors and methylation levels of PTEN and CDKN2A were higher in HER2 positive vs. HER2 negative tumors. Z-score analysis also showed that HER2 positive tumors had significantly higher z-scores of methylation compared to the HER2 negative tumors. Univariate survival analysis identifies methylation status of PPP2R2B as significant predictor of overall survival and breast cancer specific survival.
In the present study we report that the level of aberrant DNA methylation is higher in late stage compared with early stage of invasive breast cancers and DCIS for genes mentioned above.