PCDH18 is frequently inactivated by promoter methylation in colorectal cancer

Dan Zhou,Mingquan Cai,Weiwei Tang,Han-Xiang An,Yun Zhang,Guoqiang Su

doi:10.1038/s41598-017-03133-w

Abstract

Protocadherin18 (PCDH18) was found to be preferentially methylated and inactivated in colorectal cancer (CRC) using bioinformatics tools. However, its biologic role in tumorgenesis remains unclear. Herein, we aimed to elucidate its epigenetic regulation and biological functions in CRC. The methylation status of PCDH18 was significant higher in CRC tissues than in adjacent non-tumor tissues (median, 15.17% vs. median, 0.4438%). Expression level of PCDH18 was significantly lower in primary CRCs than in nonmalignant tissues. Importantly, methylation status of PCDH18 in cell-free DNA of CRC patients was also significantly higher than in healthy subjects. PCDH18 was readily expressed in NCM460 cells, but downregulated in 100% (4/4) of CRC cell lines by promoter methylation, despite its expression could be restored through demethylation treatment. Overexpression of PCDH18 suppressed CRC cell viability, colony formation and migration. Meanwhile, the depletion of PCDH18 by siRNA in NCM460 cells enhanced the colonogenicity and migration ability and promoted β-catenin nuclear accumulation, whereas it inhibited cell cycle arrest. These effects were associated with upregulation of phospho-GSK-3β and cyclin D1, and downregulation of caspase3 and p21. Our results suggested that PCDH18 was a putative tumor suppressor with epigenetic silencing in CRC and a potential biomarker for CRC diagnosis.

Highlights

Colorectal cancer (CRC) is ranked as the third most common cancer in males and the second in females worldwide with roughly 1.40 million new cases diagnosed in 2012 (9.7% of all cancers)[1]
In an attempt to identify Protocadherin 18 (PCDH18) involved in colorectal carcinogenesis, the Cancer Genome Altas (TCGA) expression array dataset of 145 colorectal cancer (CRC) tissue samples and 22 paired normal samples as well as Hong colorectal dataset of 70 CRC tissue samples and 12 normal samples were obtained from Oncomine database for comparative genome-wide analyses of PCDH18 expression
Analysis of these data showed that PCDH18 expression was significantly down-regulated in CRC tissues compared with normal controls (Fig. 1A)

Summary

Introduction

Colorectal cancer (CRC) is ranked as the third most common cancer in males and the second in females worldwide with roughly 1.40 million new cases diagnosed in 2012 (9.7% of all cancers)[1]. Some PCDHs including PCDH8, PCDH17 and PCDH20 were reported to be frequently silenced mainly in breast, prostatic, lung and digestive carcinomas via aberrant promoter methylation, indicating that PCDHs may function as TSGs13–15 Another studies suggested that decreased expression of PCDHs (e.g. PCDH9, PCDH10 and PCDH20) facilitated epithelial-mesenchymal transition and migration through the Wnt/β-catenin signaling pathway, revealing that PCDHs may protect against malignant transformation[16, 17]. We identified that PCDH18 was preferentially hypermethylated in colorectal cancer using bioinformatics analysis This was followed by a clinical validation study with multiple patient tissue and plasma samples that led to confirmation of PCDH18 as a potential biomarker for CRC diagnosis. We investigated the epigenetic regulation, biological function and molecular pathway of PCDH18 in CRC

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