Basic Study
Copyright ©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Dec 21, 2016; 22(47): 10325-10340
Published online Dec 21, 2016. doi: 10.3748/wjg.v22.i47.10325
Aging related methylation influences the gene expression of key control genes in colorectal cancer and adenoma
Orsolya Galamb, Alexandra Kalmár, Barbara Kinga Barták, Árpád V Patai, Katalin Leiszter, Bálint Péterfia, Barnabás Wichmann, Gábor Valcz, Gábor Veres, Zsolt Tulassay, Béla Molnár
Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary
Orsolya Galamb, Alexandra Kalmár, Barbara Kinga Barták, Árpád V Patai, Katalin Leiszter, Bálint Péterfia, 2nd Department of Internal Medicine, Semmelweis University, H-1088 Budapest, Hungary
Gábor Veres, 1st Department of Paediatrics, Semmelweis University, H-1083 Budapest, Hungary
Author contributions: Galamb O, Kalmár A, Péterfia B and Molnár B designed the study; Patai ÁV, Veres G and Molnár B collected the samples; Galamb O, Patai ÁV, Leiszter K, Valcz G and Veres G contributed to the collection of clinical data and histological analysis of the samples; Galamb O, Kalmár A, Barták BK and Patai ÁV performed the experiments; Galamb O, Kalmár A, Wichmann B and Valcz G analyzed the experimental data; Tulassay Z and Molnár B contributed to the design and critical review of the manuscript, obtained fundings; all authors were involved in writing the paper, made a critical revision of the manuscript for important intellectual content and had final approval of the submitted and published versions.
Supported by the National Research, Development and Innovation Office, No. KMR-12-1-2012-0216; and the Hungarian Scientific Research Fund, No. OTKA-K111743.
Institutional review board statement: All routine colonic biopsy samples from the patients were taken after informed consent and ethical permission was obtained for participation in the study.
Conflict-of-interest statement: The authors declare that no conflict of interest exists.
Data sharing statement: Additional data are available in a supplementary file.
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:
Correspondence to: Orsolya Galamb, PhD, 2nd Department of Internal Medicine, Semmelweis University, Szentkirályi str 46, H-1088 Budapest, Hungary.
Telephone: +36-1-2660926 Fax: +36-1-2660816
Received: June 22, 2016
Peer-review started: June 24, 2016
First decision: August 22, 2016
Revised: September 20, 2016
Accepted: November 13, 2016
Article in press: November 13, 2016
Published online: December 21, 2016
Processing time: 180 Days and 3 Hours

To analyze colorectal carcinogenesis and age-related DNA methylation alterations of gene sequences associated with epigenetic clock CpG sites.


In silico DNA methylation analysis of 353 epigenetic clock CpG sites published by Steve Horvath was performed using methylation array data for a set of 123 colonic tissue samples [64 colorectal cancer (CRC), 42 adenoma, 17 normal; GEO accession number: GSE48684]. Among the differentially methylated age-related genes, secreted frizzled related protein 1 (SFRP1) promoter methylation was further investigated in colonic tissue from 8 healthy adults, 19 normal children, 20 adenoma and 8 CRC patients using bisulfite-specific PCR followed by methylation-specific high resolution melting (MS-HRM) analysis. mRNA expression of age-related “epigenetic clock” genes was studied using Affymetrix HGU133 Plus2.0 whole transcriptome data of 153 colonic biopsy samples (49 healthy adult, 49 adenoma, 49 CRC, 6 healthy children) (GEO accession numbers: GSE37364, GSE10714, GSE4183, GSE37267). Whole promoter methylation analysis of genes showing inverse DNA methylation-gene expression data was performed on 30 colonic samples using methyl capture sequencing.


Fifty-seven age-related CpG sites including hypermethylated PPP1R16B, SFRP1, SYNE1 and hypomethylated MGP, PIPOX were differentially methylated between CRC and normal tissues (P < 0.05, Δβ≥ 10%). In the adenoma vs normal comparison, 70 CpG sites differed significantly, including hypermethylated DKK3, SDC2, SFRP1, SYNE1 and hypomethylated CEMIP, SPATA18 (P < 0.05, Δβ≥ 10%). In MS-HRM analysis, the SFRP1 promoter region was significantly hypermethylated in CRC (55.0% ± 8.4 %) and adenoma tissue samples (49.9% ± 18.1%) compared to normal adult (5.2% ± 2.7%) and young (2.2% ± 0.7%) colonic tissue (P < 0.0001). DNA methylation of SFRP1 promoter was slightly, but significantly increased in healthy adults compared to normal young samples (P < 0.02). This correlated with significantly increased SFRP1 mRNA levels in children compared to normal adult samples (P < 0.05). In CRC tissue the mRNA expression of 117 age-related genes were changed, while in adenoma samples 102 genes showed differential expression compared with normal colonic tissue (P < 0.05, logFC > 0.5). The change of expression for several genes including SYNE1, CLEC3B, LTBP3 and SFRP1, followed the same pattern in aging and carcinogenesis, though not for all genes (e.g., MGP).


Several age-related DNA methylation alterations can be observed during CRC development and progression affecting the mRNA expression of certain CRC- and adenoma-related key control genes.

Keywords: DNA methylation, Aging, Colorectal cancer, Adenoma, Epigenetic drift, Epigenetic clock, Secreted frizzled related protein 1

Core tip: Several age-related DNA methylation alterations could be observed during colorectal cancer (CRC) formation and progression affecting the mRNA expression of certain CRC- and adenoma-related key control genes such as hypermethylated secreted frizzled related protein 1 (SFRP1), spectrin repeat containing nuclear envelope protein 1 and hypomethylated cell migration-inducing protein. For the first time significantly lower SFRP1 methylation levels were demonstrated in colonic tissue from children (under 18 years) compared to healthy adults. The main CRC-associated signal transduction pathways, such as WNT signaling and PI3K/Akt pathways are also influenced during aging.