title: Aging related methylation influences the gene expression of key control genes in colorectal cancer and adenoma.
abstract: OBJECTIVE
  To analyze colorectal carcinogenesis and age-related DNA methylation alterations of gene sequences associated with epigenetic clock CpG sites.
  
  
  METHODS
  In silicoDNA 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.
  
  
  RESULTS
  Fifty-seven age-related CpG sites including hypermethylatedPPP1R16B,SFRP1,SYNE1and hypomethylated MGP, PIPOX were differentially methylated between CRC and normal tissues (P< 0.05, Δβ ≥ 10%). In the adenomavsnormal comparison, 70 CpG sites differed significantly, including hypermethylatedDKK3,SDC2,SFRP1,SYNE1and hypomethylatedCEMIP,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 ofSFRP1promoter was slightly, but significantly increased in healthy adults compared to normal young samples (P< 0.02). This correlated with significantly increasedSFRP1mRNA 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 includingSYNE1,CLEC3B,LTBP3andSFRP1, followed the same pattern in aging and carcinogenesis, though not for all genes (e.g., MGP).
  
  
  CONCLUSIONS
  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.
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Orsolya
authors.initials: O
authors.lastname: Galamb
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Kalmár
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Barbara Kinga
authors.initials: BK
authors.lastname: Barták
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Árpád V
authors.initials: ÁV
authors.lastname: Patai
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Katalin
authors.initials: K
authors.lastname: Leiszter
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Bálint
authors.initials: B
authors.lastname: Péterfia
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Barnabás
authors.initials: B
authors.lastname: Wichmann
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Gábor
authors.initials: G
authors.lastname: Valcz
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Gábor
authors.initials: G
authors.lastname: Veres
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Tulassay
authors.affiliation: Orsolya Galamb, Barnabás Wichmann, Gábor Valcz, Zsolt Tulassay, Béla Molnár, Molecular Medicine Research Group, Hungarian Academy of Sciences, H-1088 Budapest, Hungary.
authors.forename: Béla
authors.initials: B
authors.lastname: Molnár
mesh_terms: D000236:Adenoma / Q000235:genetics* / Q000473:pathology; D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000235:genetics*; D014408:Biomarkers, Tumor / Q000235:genetics*; D001706:Biopsy; D002277:Carcinoma / Q000235:genetics* / Q000473:pathology; D016022:Case-Control Studies; D002648:Child; D002675:Child, Preschool; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000473:pathology; D018899:CpG Islands; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007223:Infant; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics; D008297:Male; D008565:Membrane Proteins / Q000235:genetics; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D000094742:Secreted Frizzled-Related Proteins
chemical_list: D014408:Biomarkers, Tumor; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C117636:SFRP1 protein, human; D000094742:Secreted Frizzled-Related Proteins
keywords: Adenoma; Aging; Colorectal cancer; DNA methylation; Epigenetic clock; Epigenetic drift; Secreted frizzled related protein 1