Epigenetic risk assessment and biomarker development for breast cancer (EpiMark)

Despite the availability of new chemotherapeutic agents and progress in screening, breast cancer is still the most common cancer affecting women in developed countries. Epigenetic changes are ubiquitous in primary breast cancer, but less is known about the dietary/lifestyle and endogenous determinants of these changes. This project`s underlying hypothesis is that environmental factors and endogenous cues trigger changes in the methylome, that these changes contribute to the onset of breast cancer, and that they can serve as biomarkers in primary and secondary prevention. The project`s three main objectives are: (1) to identify DNA methylome changes in breast cancer and surrogate tissue using a large sample of several thousand subjects from a prospective cohort, (2) to establish dietary/lifestyle and endogenous determinants of epigenetic changes, and (3) to assess the predictive value of methylation markers in clinical settings. This project is based on the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort, a study conducted in 23 centres in 10 European countries, and will use powerful epigenomic technology for biomarker discovery and validation. This study will provide unique insights into the role of risk factors in driving the epigenetic changes that are associated with breast cancer, and will reveal potential targets for biomarker discovery and future preventative strategies. Within the EpiMark project, the contribution of the group led by Christoph Bock at the CeMM Research Center for Molecular Medicine (http://epigenomics.cemm.oeaw.ac.at) and the work of the Biomedical Sequencing Facility (http://biomedical- sequencing.at) focuses on large-scale DNA methylation mapping in fresh-frozen and formalin-fixed paraffin- embedded patient samples as well as on integrative bioinformatic analysis of the generated datasets.

This project was part of a European initiative aiming to identify a robust association between changes in DNA methylation and the development of breast cancer, and to evaluate such an association as a potential biomarker. Co-ordinated by the International Agency for Research on Cancer (IARC) of the World Health Organization (WHO), the project consortium focused on the development and application of methods to analyze DNA methylation in patient samples. The CeMM contribution to the TRANSCAN EpiMark project has given rise to four main results, three of which have already led to a scientific publication in a high-profile journal: 1. We have developed a high-throughput pipeline for DNA methylation profiling in large sample sets, based on an optimized RRBS protocol (https://doi.org/10.1016/j.celrep.2015.11.024). Us- ing this protocol, we have profiled the >1,000 samples for the TRANSCAN EpiMark project as well as a large number of additional samples in collaborations with researchers worldwide. 2. Complementing the RRBS protocols focus on high sample throughput, we have developed a dedicated protocol for low-input and single-cell DNA methylation profiling termed WGBS / scWGBS (https://doi.org/10.1016/j.celrep.2015.02.001), which opens up new opportunities for dissecting tumor heterogeneity and analyzing plasma-derived cell-free DNA samples. 3. To help analyze and interpret the DNA methylation differences observed between different sample sets, we have developed the LOLA software tool (an R/Bioconductor package) for region set enrichment analysis (https://doi.org/10.1093/bioinformatics/btv612). With >3,500 user downloads, this software is already widely used for analyzing genomic region datasets. 4. Most importantly, we have used our RRBS protocol for DNA methylation profiling of >1,000 samples from a prospective breast cancer risk cohort and reference samples provided by the consortium. The resulting dataset constitutes the basis for ongoing analyses by TRANSCAN EpiMark project partners, and a consortium-level publication is planned and in preparation.