| dc.description.abstract | Breast cancer is the most prevalent type of cancer affecting females in world today and poses a serious public health problem worldwide. Even though the overall survival has increased significantly the last decades, the high incidence of breast cancer signifies the importance of improvements in diagnostics and treatment. Like all types of cancers, breast cancer is a result of accumulation of genetic and epigenetic alterations that leads to repression of tumor suppressor genes and activation of oncogenes. Over the past decades, several studies have highlighted alterations in DNA methylation patterns as hallmark events in many cancer types including breast cancer. Among the major types of breast cancers, the estrogen receptor positive tumors which accounts for 70 % of all breast cancers tend to display more pronounced changes in their DNA methylation landscape than the ER negative tumors when compared to normal adjacent tissue. It is well known that alterations in DNA methylation may affect the expression of genes, depending on where the changes occur. For instance, changes in DNA methylation at CpGs in cis-regulatory regions such as promotors tend to repress the expression of its associated gene. Furthermore, CpGs as far as 100 kb away from the transcription start site have been demonstrated to be associated with gene expression. Therefore, CpGs in intergenic and enhancer regions may play key roles in breast cancer pathogenesis through the regulation of expression of their associated genes. Enhancer methylation has been shown experimentally to be associated with gene expression, and these genomic regions are considered to be the most differentially methylated genomic regions during carcinogenesis and cancer progression. Enhancers are known to bind cell type specific proteins called transcription factors (TFs) which are proteins involved in the regulation of gene transcription. However, the role of DNA methylation at enhancer regions regarding TF binding and breast cancer pathogenesis is still not fully understood. Genome-wide expression-methylation quantitative trait loci (emQTL) analysis have previously been shown to identify significant correlations between the level of DNA methylation at CpG sites and gene expression due to intertumoral heterogeneity within ER positive and ER negative tumors. It has also been shown to be a valuable tool in the identification of key gene regulatory networks involved in breast cancer pathogenesis. To take this further, the same approach was applied to the ER positive breast tumors only, to investigate whether any differences within the ER positive tumors in respect to DNA methylation and gene expression could be observed. The study resulted in the identification of CpG-gene pairs in which the level of DNA methylation was significantly correlated with gene expression. Hierarchical clustering of the significant associations led to the discovery of a previously undiscovered cluster of CpG-gene associations. Gene set enrichment analysis indicated an enrichment of the genes in EMT-related processes, while the CpGs were highly enriched in enhancer regions. The CpGs in this EMT-cluster was divided into CpG-cluster A and CpG-cluster B based on whether their mean methylation value was more or less than 0.5 respectively. The CpGs in both clusters were shown to be differentially methylated among the ER positive tumors. Further characterization of the CpG-clusters by ChIP-seq peaks enrichment analysis revealed that CpG-cluster A CpGs were enriched within ChIP-seq peaks of TFs associated with EMT such as TEAD1, FOSL1, TWIST1, SIX2, YAP1 and PPARG. To investigate whether the difference in DNA methylation was associated with any phenotypic feature associated with EMT in the tumors, an EMT score was utilized and correlated with the mean methylation of CpG-cluster A CpGs. The mean methylation was negatively correlated with the EMT score, meaning that lower methylation was associated with more mesenchymal-like characteristic of the tumor samples. These findings suggest that EMT-related CpG-gene pairs discovered in this study are associated with gene regulatory networks wired by EMT related TFs through a relationship between DNA methylation at their target DNA binding regions in enhancers, and gene expression of their target genes. This study highlights the CpGs identified as potential contributors to EMT-related cancer pathogenesis in the ER positive breast tumors and constitute interesting regions for further investigations. However, these in silico findings still require better validation in vitro. The identification of cancer-causing epigenetic changes may open up possibilities of targeted treatment by utilization of technologies such as CRISPR to edit epigenetic cancer-causing mutations to inhibit tumor growth in the future. | eng |