title: Estimation of the fraction of cancer cells in a tumor DNA sample using DNA methylation.
abstract: Contamination of normal cells is almost always present in tumor samples and affects their molecular analyses. DNA methylation, a stable epigenetic modification, is cell type-dependent, and different between cancer and normal cells. Here, we aimed to demonstrate that DNA methylation can be used to estimate the fraction of cancer cells in a tumor DNA sample, using esophageal squamous cell carcinoma (ESCC) as an example. First, by an Infinium HumanMethylation450 BeadChip array, we isolated three genomic regions (TFAP2B, ARHGEF4, and RAPGEFL1) i) highly methylated in four ESCC cell lines, ii) hardly methylated in a pooled sample of non-cancerous mucosae, a pooled sample of normal esophageal mucosae, and peripheral leukocytes, and iii) frequently methylated in 28 ESCCs (TFAP2B, 24/28; ARHGEF4, 20/28; and RAPGEFL1, 19/28). Second, using eight pairs of cancer and non-cancer cell samples prepared by laser capture microdissection, we confirmed that at least one of the three regions was almost completely methylated in ESCC cells, and all the three regions were almost completely unmethylated in non-cancer cells. We also confirmed that DNA copy number alterations of the three regions in 15 ESCC samples were rare, and did not affect the estimation of the fraction of cancer cells. Then, the fraction of cancer cells in a tumor DNA sample was defined as the highest methylation level of the three regions, and we confirmed a high correlation between the fraction assessed by the DNA methylation fraction marker and the fraction assessed by a pathologist (r=0.85; p<0.001). Finally, we observed that, by correction of the cancer cell content, CpG islands in promoter regions of tumor-suppressor genes were almost completely methylated. These results demonstrate that DNA methylation can be used to estimate the fraction of cancer cells in a tumor DNA sample.
authors: Division of Epigenomics, National Cancer Center Research Institute, Tokyo, Japan ; Esophageal Surgery Division, National Cancer Center Hospital, Tokyo, Japan ; Division of Surgical Oncology, Department of Surgery, Nagoya University Graduate School of Medicine, Nagoya, Japan.
authors: Takamasa
authors: T
authors: Takahashi
authors: Yasunori
authors: Y
authors: Matsuda
authors: Satoshi
authors: S
authors: Yamashita
authors: Naoko
authors: N
authors: Hattori
authors: Ryoji
authors: R
authors: Kushima
authors: Yi-Chia
authors: YC
authors: Lee
authors: Hiroyasu
authors: H
authors: Igaki
authors: Yuji
authors: Y
authors: Tachimori
authors: Masato
authors: M
authors: Nagino
authors: Toshikazu
authors: T
authors: Ushijima
mesh_terms: D045744:Cell Line, Tumor; D019175:DNA Methylation / Q000235:genetics*; D004273:DNA, Neoplasm / Q000235:genetics*; D006801:Humans; D064067:Rho Guanine Nucleotide Exchange Factors / Q000235:genetics; D050656:Transcription Factor AP-2 / Q000235:genetics
chemical_list: C576602:ARHGEF4 protein, human; D004273:DNA, Neoplasm; D064067:Rho Guanine Nucleotide Exchange Factors; C493677:TFAP2B protein, human; D050656:Transcription Factor AP-2