Epigenetic Changes in Basal Cell Carcinoma Affect SHH and WNT Signaling Components

Tjinta Brinkhuizen,Karin Van Den Hurk,Manon Van Engeland,Maurice A M Van Steensel,Joep P De Hoon,Véronique J L Winnepenninckx,Ariënne M Van Marion,Jürgen Veeck,Javier S Castresana

doi:10.1371/journal.pone.0051710

Tjinta Brinkhuizen, Karin Van Den Hurk + Show 7 more

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https://doi.org/10.1371/journal.pone.0051710

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Abstract

BackgroundThe genetic background of Basal Cell Carcinoma (BCC) has been studied extensively, while its epigenetic makeup has received comparatively little attention. Epigenetic alterations such as promoter hypermethylation silence tumor suppressor genes (TSG) in several malignancies.ObjectiveWe sought to analyze the promoter methylation status of ten putative (tumor suppressor) genes that are associated with Sonic Hedgehog (SHH), WNT signaling and (hair follicle) tumors in a large series of 112 BCC and 124 healthy control samples by methylation-specific PCR.ResultsGene promoters of SHH (P = 0.016), adenomatous polyposis coli (APC) (P = 0.003), secreted frizzled-related protein 5 (SFRP5) (P = 0.004) and Ras association domain family 1A (RASSF1A) (P = 0.023) showed significantly more methylation in BCC versus normal skin. mRNA levels of these four genes were reduced for APC and SFRP5 in BCC (n = 6) vs normal skin (n = 6). Down regulation of SHH, APC and RASSF1A could be confirmed on protein level as well (P<0.001 for all genes) by immunohistochemical staining. Increased canonical WNT activity was visualized by β-catenin staining, showing nuclear β-catenin in only 28/101 (27.7%) of BCC. Absence of nuclear β-catenin in some samples may be due to high levels of membranous E-cadherin (in 94.1% of the samples).ConclusionsWe provide evidence that promoter hypermethylation of key players within the SHH and WNT pathways is frequent in BCC, consistent with their known constitutive activation in BCC. Epigenetic gene silencing putatively contributes to BCC tumorigenesis, indicating new venues for treatment.

Highlights

Basal cell carcinoma (BCC) accounts for 75% of all skin cancers and is the most frequent malignancy in Caucasians
A total of nine candidate tumor suppressor genes (TSG) and one oncogene was examined in a series of primary Basal Cell Carcinoma (BCC) (n = 112) of three different subtypes and normal skin (n = 124)
Our data showed that promoters of four genes were significantly more frequently methylated in BCC tissue versus normal skin: Sonic Hedgehog (SHH) (44/ 100 (40.0%) vs. 30/122 (24.6%), P = 0.016), adenomatous polyposis coli (APC) (64/110 (58.2%) vs. 47/124 (37.9%), P = 0.003), secreted frizzled-related protein 5 (SFRP5) (52/109 (47.7%) vs. 28/100 (28.0%), P = 0.004), and Ras association domain family 1A (RASSF1A) (52/112 (46.4%) vs. 39/124 (31.3%), P = 0.023

Summary

Introduction

Basal cell carcinoma (BCC) accounts for 75% of all skin cancers and is the most frequent malignancy in Caucasians. Its incidence is rising by 3–8% each year [1], resulting in an average lifetime risk for Caucasians of developing BCC of 30% [2,3]. In order to develop one, a thorough understanding of BCC pathobiology will be required and the past few years have witnessed considerable progress in this respect. The genetic background of Basal Cell Carcinoma (BCC) has been studied extensively, while its epigenetic makeup has received comparatively little attention. Epigenetic alterations such as promoter hypermethylation silence tumor suppressor genes (TSG) in several malignancies

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