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Methylation of multiple genes as molecular markers for diagnosis of a small, well‐differentiated hepatocellular carcinoma
Author(s) -
Moribe Toyoki,
Iizuka Norio,
Miura Toshiaki,
Kimura Naoki,
Tamatsukuri Shigeru,
Ishitsuka Hideo,
Hamamoto Yoshihiko,
Sakamoto Kazuhiko,
Tamesa Takao,
Oka Masaaki
Publication year - 2009
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.24394
Subject(s) - hccs , hepatocellular carcinoma , in silico , methylation , cpg site , cancer research , biology , gene , pyrosequencing , dna methylation , bisulfite sequencing , microbiology and biotechnology , genetics , gene expression
The current study was conducted to identify robust methylation markers and their combinations that may prove useful for the diagnosis of early hepatocellular carcinoma (HCC). To achieve this, we performed in silico CpG mapping, direct sequencing and pyrosequencing after bisulfite treatment, and quantitative methylation‐specific PCR (MSP) in HCC and non‐HCC liver tissues. In the filtering group (25 HCCs), our direct sequencing analysis showed that, among the 12 methylation genes listed by in silico CpG mapping, 7 genes ( RASSF1A , CCND2 , SPINT2 , RUNX3 , GSTP1 , APC and CFTR ) were aberrantly methylated in stages I and II HCCs. In the validation group (20 pairs of HCCs and the corresponding non‐tumor liver tissues), pyrosequencing analysis confirmed that the 7 genes were aberrantly and strongly methylated in early HCCs, but not in any of the corresponding non‐ tumor liver tissues ( p < 0.00001). The results obtained using our novel quantitative MSP assay correlated well with those observed using the pyrosequencing analysis. Notably, in MSP assay, RASSF1A showed the most robust performance for the discrimination of HCC and non‐HCC liver tissues. Furthermore, a combination of RASSF1A , CCND2 and SPINT2 showed 89–95% sensitivity, 91–100% specificity and 89–97% accuracy in discriminating between HCC and non‐HCC tissues, and correctly diagnosed all early HCCs. These results indicate that the combination of these 3 genes may aid in the accurate diagnosis of early HCC. © 2009 UICC