Oue Naohide, Mitani Yoshitsugu, Motoshita Junichi, Matsumura Shunji, Yoshida Kazuhiro, Kuniyasu Hiroki, Nakayama Hirofumi, Yasui Wataru
Department of Molecular Pathology, Graduate School of Biomedical Sciences, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8551, Japan.
Cancer. 2006 Mar 15;106(6):1250-9. doi: 10.1002/cncr.21754.
The authors purpose in this study was to clarify the difference in terms of clinicopathologic features between gastric cancer (GC) with high numbers of DNA methylated genes and CpG island methylator phenotype (CIMP)-positive GC as originally defined.
We analyzed DNA methylation of 12 tumor-related genes (hMLH1, MGMT, p16(INK4a), CDH1, RAR-beta, HLTF, RIZ1, TM, FLNc, LOX, HRASLS, HAND1) in 75 samples of GC from 75 patients, 25 samples of corresponding nonneoplastic mucosa from 25 patients, and 10 samples of normal gastric mucosa from 10 healthy young individuals by methylation-specific polymerase chain reaction (PCR) and bisulfite PCR. We also investigated CIMP status by examining the methylation of five MINT loci and p53 mutation status by PCR single-strand conformation polymorphism. We measured levels of expression of mRNAs for these 12 genes by quantitative reverse transcription PCR in 50 GC specimens.
The average number of methylated genes per tumor was 4.83. DNA methylation of each gene was correlated with low expression of the respective mRNA. High methylation (GC with 5 or more methylated genes) was detected in 39 (52.0%) of 75 GCs. Twenty-nine (37.8%) of 75 GCs were CIMP-positive. DNA methylation of each of the 12 genes was observed more frequently in the high-methylation group than in the low-methylation group. Methylation of 6 specific genes occurred more frequently in CIMP-positive GC than in CIMP-negative GC. Methylation of the remaining 6 genes was not correlated with CIMP-status. High methylation was found more frequently in Stage III/IV GC (26 of 40 cases, 65.0%) than in Stage I/II GC (13 of 35 cases, 37.1%, P = 0.029).CONCLUSIONS.These findings indicate that GCs with higher numbers of methylated genes have more distinct DNA methylation profiles than the originally defined CIMP-positive GCs. DNA methylation of tumor-related genes accumulates in conjunction with tumor progression.
本研究中作者的目的是阐明具有大量DNA甲基化基因的胃癌（GC）与最初定义的CpG岛甲基化表型（CIMP）阳性GC在临床病理特征方面的差异。
我们通过甲基化特异性聚合酶链反应（PCR）和亚硫酸氢盐PCR分析了75例患者的75份GC样本、25例患者的25份相应非肿瘤性黏膜样本以及10名健康年轻个体的10份正常胃黏膜样本中12个肿瘤相关基因（hMLH1、MGMT、p16(INK4a)、CDH1、RAR-β、HLTF、RIZ1、TM、FLNc、LOX、HRASLS、HAND1）的DNA甲基化情况。我们还通过检测五个MINT位点的甲基化来研究CIMP状态，并通过PCR单链构象多态性研究p53突变状态。我们在50份GC标本中通过定量逆转录PCR测量了这12个基因的mRNA表达水平。
每个肿瘤中甲基化基因的平均数量为4.83个。每个基因的DNA甲基化与各自mRNA的低表达相关。在75例GC中，39例（52.0%）检测到高甲基化（具有5个或更多甲基化基因的GC）。75例GC中有29例（37.8%）为CIMP阳性。12个基因中的每个基因的DNA甲基化在高甲基化组中比在低甲基化组中更频繁地观察到。6个特定基因的甲基化在CIMP阳性GC中比在CIMP阴性GC中更频繁地出现。其余6个基因的甲基化与CIMP状态无关。III/IV期GC（40例中的26例，65.0%）中高甲基化的发现频率高于I/II期GC（3例中的13例，37.1%，P = 0.029）。结论：这些发现表明，具有更多甲基化基因的GC比最初定义的CIMP阳性GC具有更独特的DNA甲基化谱。肿瘤相关基因的DNA甲基化随着肿瘤进展而积累。
