title: Identification and validation of highly frequent CpG island hypermethylation in colorectal adenomas and carcinomas.
abstract: In our study, whole-genome methylation arrays were applied to identify novel genes with tumor specific DNA methylation of promoter CpG islands in pre-malignant and malignant colorectal lesions. Using a combination of Illumina HumanMethylation27 beadchips, Methylation-Sensitive High Resolution Melting (MS-HRM) analysis, and Exon arrays (Affymetrix) the DNA methylation pattern of ∼14,000 genes and their transcript levels were investigated in six normal mucosas, six adenomas and 30 MSI and MSS carcinomas. Sixty eight genes with tumor-specific hypermethylation were identified (p < 0.005). Identified hypermethylated sites were validated in an independent sample set of eight normal mucosas, 12 adenomas, 40 MSS and nine MSI cancer samples. The methylation patterns of 15 selected genes, hypermethylated in adenomas and carcinomas (FLI1, ST6GALNAC5, TWIST1, ADHFE1, JAM2, IRF4, CNRIP1, NRG1 and EYA4), in carcinomas only (ABHD9, AOX1 and RERG), or in MSI but not MSS carcinomas (RAMP2, DSC3 and MLH1) were validated using MS-HRM. Four of these genes (MLH1, AOX1, EYA4 and TWIST1) had previously been reported to be hypermethylated in CRC. Eleven genes, not previously known to be affected by CRC specific hypermethylation, were identified and validated. Inverse correlation to gene expression was observed for six of the 15 genes with Spearman correlation coefficients ranging from -0.39 to -0.60. For six of these genes the altered methylation patterns had a profound transcriptional association, indicating that methylation of these genes may play a direct regulatory role. The hypermethylation changes often occurred already in adenomas, indicating that they may be used as biomarkers for early detection of CRC.
authors.affiliation: Department of Molecular Medicine, Aarhus University Hospital, Skejby, Denmark.
authors.forename: Bodil
authors.initials: B
authors.lastname: Oster
authors.forename: Kasper
authors.initials: K
authors.lastname: Thorsen
authors.forename: Philippe
authors.initials: P
authors.lastname: Lamy
authors.forename: Tomasz K
authors.initials: TK
authors.lastname: Wojdacz
authors.forename: Lise Lotte
authors.initials: LL
authors.lastname: Hansen
authors.forename: Karin
authors.initials: K
authors.lastname: Birkenkamp-Demtröder
authors.forename: Karina D
authors.initials: KD
authors.lastname: Sørensen
authors.forename: Søren
authors.initials: S
authors.lastname: Laurberg
authors.forename: Torben F
authors.initials: TF
authors.lastname: Orntoft
authors.forename: Claus L
authors.initials: CL
authors.lastname: Andersen
mesh_terms: D000236:Adenoma / Q000235:genetics*; D000368:Aged; D002277:Carcinoma / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics*; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D006801:Humans; D008297:Male; D008875:Middle Aged; D020890:Neuregulin-1 / Q000378:metabolism
chemical_list: C094131:NRG1 protein, human; D020890:Neuregulin-1