Basic Study
Copyright ©The Author(s) 2018. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Nov 28, 2018; 24(44): 5013-5024
Published online Nov 28, 2018. doi: 10.3748/wjg.v24.i44.5013
Involvement of methylation-associated silencing of formin 2 in colorectal carcinogenesis
Dao-Jiang Li, Zhi-Cai Feng, Xiao-Rong Li, Gui Hu
Dao-Jiang Li, Xiao-Rong Li, Gui Hu, Department of Gastrointestinal Surgery, the Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China
Zhi-Cai Feng, Department of Burns and Plastic Surgery, the Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China
Author contributions: Li DJ, Li XR, and Hu G contributed to conceptualization and design of the study, analyzed the data, and coordinated the research; Li DJ and Feng ZC performed the majority of the experiments and analyzed the data; all authors drafted the article and made revisions related to the intellectual content of the manuscript, and approved the final version of the article to be published.
Supported by the National Nature Science Foundation of China, No. 81773130; the Fundamental Research Funds for the Central Universities of Central South University(the Key Projects of Postgraduate Independent Exploration and Innovation of Central South University, No. 2018zzts050); and the New Xiangya Talent Projects of the Third Xiangya Hospital of Central South University, No. JY201508.
Institutional review board statement: This study was reviewed and approved by the Ethics Committee of Central South University. Informed consent was obtained from all individual participants included in the study.
Conflict-of-interest statement: To the best of our knowledge, no conflict of interest exists.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Corresponding author to: Gui Hu, PhD, Surgeon, Department of Gastrointestinal Surgery, the Third Xiangya Hospital, Central South University, No. 172, Tongzi Po Road, Changsha 410013, Hunan Province, China. xy3zzz@csu.edu.cn.
Telephone: +86-13908472002 Fax: +86-731-88618832
Received: August 20, 2018
Peer-review started: August 20, 2018
First decision: October 11, 2018
Revised: October 14, 2018
Accepted: November 7, 2018
Article in press: November 7, 2018
Published online: November 28, 2018
Abstract
AIM

To investigate whether promoter methylation is responsible for the silencing of formin 2 (FMN2) in colorectal cancer (CRC) and to analyze the association between FMN2 methylation and CRC.

METHODS

We first identified the expression levels and methylation levels of FMN2 in large-scale human CRC expression datasets, including GEO and TCGA, and analyzed the relationship between the expression and methylation levels. Then, the methylation levels in four CpG regions adjacent to the FMN2 promoter were assessed by MethylTarget™ assays in CRC cells and in paired colorectal tumor samples and adjacent nontumor tissue samples. Furthermore, we inhibited DNA methylation in CRC cells with 5-Aza-2’-deoxycytidine and assessed the expression of FMN2 by qRT-PCR. Last, the association between FMN2 methylation patterns and clinical indicators was analyzed.

RESULTS

A statistically significant downregulation of FMN2 expression in large-scale human CRC expression datasets was found. Subsequent analysis showed that a high frequency of hypermethylation occurred in the FMN2 gene promoter in CRC tissues; operating characteristic curve analysis revealed that FMN2 gene methylation had a good capability for discriminating between CRC and nontumor tissue samples (AUC = 0.8432, P < 0.0001). MethylTarget™ assays showed that CRC cells and tissues displayed higher methylation of these CpG regions than nontumor tissue samples. Correlation analysis showed a strong inverse correlation between methylation and FMN2 expression, and the inhibition of DNA methylation with 5-Aza significantly increased endogenous FMN2 expression. Analysis of the association between FMN2 methylation patterns and clinical indicators showed that FMN2 methylation was significantly associated with age, N stage, lymphovascular invasion, and pathologic tumor stage. Notably, the highest methylation of FMN2 occurred in tissues from cases of early-stage CRC, including cases with no regional lymph node metastasis (N0), cases in stages I and II, and cases with no lymphovascular invasion, but the methylation level began to decrease with tumor progression. Additionally, FMN2 promoter hypermethylation was more common in patients > 60 years old and in colon cancer tissue.

CONCLUSION

FMN2 promoter hypermethylation may be an important early event in CRC, most likely playing a critical role in cancer initiation, and can serve as an ideal diagnostic biomarker in elderly patients with early-stage colon cancer.

Keywords: Formin 2, Colorectal cancer, Methylation, Methylation-associated silencing, Early-stage cancer

Core tip: Colorectal cancer (CRC) is the leading cause of cancer death in the world. We identified a statistically significant downregulation of formin 2 (FMN2) expression in large-scale human CRC expression datasets and our clinical samples. Then, we first showed that a high frequency of hypermethylation occurred in the FMN2 gene promoter, which is responsible for the downregulation of FMN2 expression. Additionally, the highest methylation of FMN2 occurred in tissues from cases of early-stage CRC and patients > 60 years old. FMN2 hypermethylation may be an important early event in CRC and can serve as an ideal diagnostic biomarker in elderly patients with early-stage CRC.