title: Establishment of a DNA methylation marker to evaluate cancer cell fraction in gastric cancer.
abstract: BACKGROUND
  Tumor samples are unavoidably contaminated with coexisting normal cells. Here, we aimed to establish a DNA methylation marker to estimate the fraction of gastric cancer (GC) cells in any DNA sample by isolating genomic regions specifically methylated in GC cells.
  
  
  METHODS
  Genome-wide and gene-specific methylation analyses were conducted with an Infinium HumanMethylation450 BeadChip array and by quantitative methylation-specific PCR, respectively. Purified cancer and noncancer cells were prepared by laser-capture microdissection. TP53 mutation data were obtained from our previous study using next-generation target sequencing.
  
  
  RESULTS
  Genome-wide DNA methylation analysis of 12 GC cell lines, 30 GCs, six normal gastric mucosae, one sample of peripheral leukocytes, and four noncancerous gastric mucosae identified OSR2, PPFIA3, and VAV3 as barely methylated in normal cells and highly methylated in cancer cells. Quantitative methylation-specific PCR using 26 independent GCs validated that one or more of them was highly methylated in all of the GCs. Using four pairs of purified cells, we confirmed the three genes were highly methylated (85 % or more) in cancer cells and barely methylated (5 % or less) in noncancer cells. The cancer cell fraction assessed by the panel of the three genes showed good correlation with that assessed by the TP53 mutant allele frequency in 13 GCs (r = 0.77). After correction of the GC cell fraction, unsupervised clustering analysis of the genome-wide DNA methylation profiles yielded clearer clustering.
  
  
  CONCLUSIONS
  A DNA methylation marker-namely, the panel of the three genes-is useful to estimate the cancer cell fraction in GCs.
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.|Department of Gastrointestinal Surgery, Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
authors: Liang
authors: L
authors: Zong
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Naoko
authors: N
authors: Hattori
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Yukie
authors: Y
authors: Yoda
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Satoshi
authors: S
authors: Yamashita
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Hideyuki
authors: H
authors: Takeshima
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Takamasa
authors: T
authors: Takahashi
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Masahiro
authors: M
authors: Maeda
authors: Gastric Surgery Division, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan.
authors: Hitoshi
authors: H
authors: Katai
authors: Third Department of Internal Medicine, University of Toyama, 2630 Sugitani, Toyama, 930-0194, Japan.
authors: Sohachi
authors: S
authors: Nanjo
authors: Third Department of Internal Medicine, University of Toyama, 2630 Sugitani, Toyama, 930-0194, Japan.
authors: Takayuki
authors: T
authors: Ando
authors: Department of Gastrointestinal Surgery, Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
authors: Yasuyuki
authors: Y
authors: Seto
authors: Division of Epigenomics, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan. tushijim@ncc.go.jp.
authors: Toshikazu
authors: T
authors: Ushijima
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D014408:Biomarkers, Tumor / Q000235:genetics*; D045744:Cell Line, Tumor; D019175:DNA Methylation; D015972:Gene Expression Regulation, Neoplastic; D005787:Gene Frequency; D016481:Helicobacter Infections / Q000235:genetics; D006801:Humans; D009154:Mutation; D051567:Proto-Oncogene Proteins c-vav / Q000235:genetics; D013274:Stomach Neoplasms / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D014157:Transcription Factors / Q000235:genetics; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D014408:Biomarkers, Tumor; C486680:OSR2 protein, human; C000612103:PPFIA3 protein, human; D051567:Proto-Oncogene Proteins c-vav; C495901:TP53 protein, human; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; C496621:VAV3 protein, human
keywords: Cancer cell fraction; DNA methylation; Epigenetics; Gastric cancer