Promoter methylation of Wnt-antagonists in polypoid and nonpolypoid colorectal adenomas

Quirinus Jm Voorham,Renske Dm Steenbergen,Manon Van Engeland,Jerry Janssen,Bjorn J Rembacken,Chris Jj Mulder,Beatriz Carvalho,Gerrit A Meijer,Martin Kliment,Heike Grabsch,Sandra Mongera,Suzanne Snellenberg,Nicole Ct Van Grieken,Marianne Tijssen

doi:10.1186/1471-2407-13-603

Abstract

BackgroundNonpolypoid adenomas are a subgroup of colorectal adenomas that have been associated with a more aggressive clinical behaviour compared to their polypoid counterparts. A substantial proportion of nonpolypoid and polypoid adenomas lack APC mutations, APC methylation or chromosomal loss of the APC locus on chromosome 5q, suggesting the involvement of other Wnt-pathway genes. The present study investigated promoter methylation of several Wnt-pathway antagonists in both nonpolypoid and polypoid adenomas.MethodsQuantitative methylation-specific PCR (qMSP) was used to evaluate methylation of four Wnt-antagonists, SFRP2, WIF-1, DKK3 and SOX17 in 18 normal colorectal mucosa samples, 9 colorectal cancer cell lines, 18 carcinomas, 44 nonpolypoid and 44 polypoid adenomas. Results were integrated with previously obtained data on APC mutation, methylation and chromosome 5q status from the same samples.ResultsIncreased methylation of all genes was found in the majority of cell lines, adenomas and carcinomas compared to normal controls. WIF-1 and DKK3 showed a significantly lower level of methylation in nonpolypoid compared to polypoid adenomas (p < 0.01). Combining both adenoma types, a positive trend between APC mutation and both WIF-1 and DKK3 methylation was observed (p < 0.05).ConclusionsMethylation of Wnt-pathway antagonists represents an additional mechanism of constitutive Wnt-pathway activation in colorectal adenomas. Current results further substantiate the existence of partially alternative Wnt-pathway disruption mechanisms in nonpolypoid compared to polypoid adenomas, in line with previous observations.

Highlights

Nonpolypoid adenomas are a subgroup of colorectal adenomas that have been associated with a more aggressive clinical behaviour compared to their polypoid counterparts
For Secreted Frizzled-Related Protein-2 (SFRP2), Wnt Inhibitory Factor-1 (WIF-1) and DKK3 increased methylation levels were observed in all nine Colorectal cancer (CRC) cell lines, whereas SOX17 showed higher methylation levels in all but two cell lines (LS513 and LS174T) (Figure 1)
Methylation lead to decreased expression, as upon treatment with demethylating agents, an increase in expression was observed for SFRP2, DKK3 and SOX17 but not for WIF-1 (Additional file 1: Figure S1)

Summary

Introduction

Nonpolypoid adenomas are a subgroup of colorectal adenomas that have been associated with a more aggressive clinical behaviour compared to their polypoid counterparts. A substantial proportion of nonpolypoid and polypoid adenomas lack APC mutations, APC methylation or chromosomal loss of the APC locus on chromosome 5q, suggesting the involvement of other Wnt-pathway genes. Colorectal cancer (CRC) results from the accumulation of multiple alterations in the (epi) genome of the epithelial cells that line the large intestine. These events first give rise to an adenoma that, in a minority of cases progresses into an invasive and potentially metastasizing adenocarcinoma. Wnt-pathway activation represents a critical early event in colorectal tumorigenesis and primarily results from inactivating mutations in its gatekeeper APC [11,12]. Methylation plays an important role in CRC development and many genes have altered methylation patterns in the tumor compared to normal colon mucosa

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