title: DNA methylation alterations of AXIN2 in serrated adenomas and colon carcinomas with microsatellite instability.
abstract: BACKGROUND
  Recent work led to recognize sessile serrated adenomas (SSA) as precursor to many of the sporadic colorectal cancers with microsatellite instability (MSI). However, comprehensive analyses of DNA methylation in SSA and MSI cancer have not been conducted.
  
  
  METHODS
  With an array-based methylation sensitive amplified fragment length polymorphism (MS-AFLP) method we analyzed 8 tubular (TA) and 19 serrated (SSA) adenomas, and 14 carcinomas with (MSI) and 12 without (MSS) microsatellite instability. MS-AFLP array can survey relative differences in methylation between normal and tumor tissues of 9,654 DNA fragments containing all NotI sequences in the human genome.
  
  
  RESULTS
  Unsupervised clustering analysis of the genome-wide hypermethylation alterations revealed no major differences between or within these groups of benign and malignant tumors regardless of their location in intergenic, intragenic, promoter, or 3' end regions. Hypomethylation was less frequent in SSAs compared with MSI or MSS carcinomas. Analysis of variance of DNA methylation between these four subgroups identified 56 probes differentially altered. The hierarchical tree of this subset of probes revealed two distinct clusters: Group 1, mostly composed by TAs and MSS cancers with KRAS mutations; and Group 2 with BRAF mutations, which consisted of cancers with MSI and MLH1 methylation (Group 2A), and SSAs without MLH1 methylation (Group 2B). AXIN2, which cooperates with APC and β-catenin in Wnt signaling, had more methylation alterations in Group 2, and its expression levels negatively correlated with methylation determined by bisulfite sequencing. Within group 2B, low and high AXIN2 expression levels correlated significantly with differences in size (P = 0.01) location (P = 0.05) and crypt architecture (P = 0.01).
  
  
  CONCLUSIONS
  Somatic methylation alterations of AXIN2, associated with changes in its expression, stratify SSAs according to some clinico-pathological differences. We conclude that hypermethylation of MLH1, when occurs in an adenoma cell with BRAF oncogenic mutational activation, drives the pathway for MSI cancer by providing the cells with a mutator phenotype. AXIN2 inactivation may contribute to this tumorigenic pathway either by mutator phenotype driven frameshift mutations or by epigenetic deregulation contemporary with the unfolding of the mutator phenotype.
authors: Yuta
authors: Y
authors: Muto
authors: Takafumi
authors: T
authors: Maeda
authors: Department of Surgery, Saitama Medical Center, Jichi Medical University, 1-847, Amanuma-cho, Omiya-ku, Saitama 330-8503, Japan. ksuzbnhm@yahoo.co.jp.
authors: Koichi
authors: K
authors: Suzuki
authors: Takaharu
authors: T
authors: Kato
authors: Fumiaki
authors: F
authors: Watanabe
authors: Hidenori
authors: H
authors: Kamiyama
authors: Masaaki
authors: M
authors: Saito
authors: Kei
authors: K
authors: Koizumi
authors: Yuichiro
authors: Y
authors: Miyaki
authors: Fumio
authors: F
authors: Konishi
authors: Sergio
authors: S
authors: Alonso
authors: Manuel
authors: M
authors: Perucho
authors: Toshiki
authors: T
authors: Rikiyama
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics*; D000236:Adenoma / Q000235:genetics* / Q000473:pathology; D000368:Aged; D000369:Aged, 80 and over; D060466:Axin Protein / Q000235:genetics*; D002277:Carcinoma / Q000235:genetics* / Q000473:pathology; D003110:Colonic Neoplasms / Q000235:genetics* / Q000473:pathology; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008297:Male; D053842:Microsatellite Instability; D008875:Middle Aged; D000070957:MutL Protein Homolog 1; D009687:Nuclear Proteins / Q000235:genetics*; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*
chemical_list: C491761:AXIN2 protein, human; D048868:Adaptor Proteins, Signal Transducing; D060466:Axin Protein; C501486:MLH1 protein, human; D009687:Nuclear Proteins; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D000070957:MutL Protein Homolog 1