Analysis of the DNA methylation level of cancer-related genes in colorectal cancer and the surrounding normal mucosa

Background: Two molecular pathways promote the development of colorectal cancer (CRC). One is termed microsatellite stable (MSS) whereas the other is characterized by microsatellite instability (MSI or MIN). In addition, the CpG island methylation phenotype is known to be an important alteration as a third molecular type. Thus, DNA methylation is thought to provide potential biomarkers for assessment of cancer risk in normal mucosa. In addition, it is also known that colonic location is an important parameter in the development of CRC. Methods: We examined the surrounding normal mucosa in three parts of the colon. Next, we quantified DNA methylation levels of SFRP1, SFRP2, SFRP5, DKK2, DKK3, mir34b/c, RASSF1A, IGFBP7, CDKN2A, and MLH1 in isolated cancerous glands and crypts of normal colorectal mucosa adjacent to CRCs using a pyrosequencer. Results: DNA methylation levels of SFRP1, SFRP2, DKK2, and mir34b/c were significantly higher in CRCs with an MSS phenotype than in those with an MSI phenotype. The average level of methylation in normal crypts did not decrease with the distance from the tumor, irrespective of microsatellite status or the tumor location. DNA methylation levels in SFRP1 and SFRP2 genes in normal crypts were significantly higher in left-side than right-side CRC with an MSS phenotype. Finally, the genes were classified into three types based on the methylation frequencies in normal crypts, including type I (SFRP1 and SFRP2I), type II (DKK2 and mir34b/c), and type III (others). Conclusions: Our results showed that DNA methylation of SFRP1 and SFRP2 might be useful to predict cancer risk of surrounding normal mucosa. In addition, a field effect may be present in CRC, affecting both adjacent and non-adjacent normal mucosa.