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Open Access Research

BRAF mutation-specific promoter methylation of FOX genes in colorectal cancer

Eddy H van Roon12, Arnoud Boot12, Ashwin A Dihal12, Robert F Ernst3, Tom van Wezel2, Hans Morreau2 and Judith M Boer134*

Author Affiliations

1 Center for Human and Clinical Genetics, Leiden University Medical Center, Leiden, The Netherlands

2 Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands

3 Department of Pediatric Oncology, Erasmus MC - Sophia Children’s Hospital, Rotterdam, The Netherlands

4 Netherlands Bioinformatics Center, Nijmegen, the Netherlands

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Clinical Epigenetics 2013, 5:2  doi:10.1186/1868-7083-5-2

Published: 16 January 2013

Abstract

Background

Cancer-specific hypermethylation of (promoter) CpG islands is common during the tumorigenesis of colon cancer. Although associations between certain genetic aberrations, such as BRAF mutation and microsatellite instability, and the CpG island methylator phenotype (CIMP), have been found, the mechanisms by which these associations are established are still unclear. We studied genome-wide DNA methylation differences between colorectal tumors carrying a BRAF mutation and BRAF wildtype tumors.

Results

Using differential methylation hybridization on oligonucleotide microarrays representing 32,171 CpG-rich regions, we identified 1,770 regions with differential methylation between colorectal tumor and paired normal colon. Next, we compared the tumor/normal methylation ratios between different groups of patients. Related to CIMP, we identified 749 differentially methylated regions, of which 86% had a higher tumor/normal methylation ratio in the CIMP-positive group. We identified 758 regions with a BRAF mutation-specific methylation change, of which 96% had a higher tumor/normal methylation ratio in the BRAF mutant group. Among the genes affected by BRAF mutation-specific methylation changes, we found enrichment of several cancer-related pathways, including the PI3 kinase and Wnt signaling pathways. To focus on genes that are silenced in a tumor-specific rather than a lineage-specific manner, we used information on the epigenetic silencing mark H3K27me3 in embryonic stem (ES) cells. Among the genes showing BRAF mutation-specific promoter methylation but no H3K27me3 mark in ES cells were forkhead box (FOX) transcription factors associated with the PI3 kinase pathway, as well as MLH1 and SMO. Repression of FOXD3 gene expression in tumors could be related to its promoter hypermethylation.

Conclusions

We identified new BRAF mutation-specific methylation changes in colorectal cancer. Epigenetic downregulation of these targets may contribute to mutationally active BRAF-driven tumorigenesis, explaining its association with aberrant DNA methylation.

Keywords:
BRAF mutation; Colorectal cancer; DNA methylation; Histone pre-marking