Nagasaka Takeshi, Tanaka Noriaki, Cullings Harry M, Sun Dong-Sheng, Sasamoto Hiromi, Uchida Takuyuki, Koi Minoru, Nishida Naoshi, Naomoto Yoshio, Boland C Richard, Matsubara Nagahide, Goel Ajay
Department of Gastroenterological Surgery and Surgical Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Okayama City, Okayama 700-8558, Japan.
J Natl Cancer Inst. 2009 Sep 16;101(18):1244-58. doi: 10.1093/jnci/djp265. Epub 2009 Aug 21.
The development of noninvasive screening tests is important to reduce mortality from gastrointestinal neoplasia. We sought to develop such a test by analysis of DNA methylation from exfoliated cancer cells in feces.
We first analyzed methylation of the RASSF2 and SFRP2 gene promoters from 788 primary gastric and colorectal tissue specimens to determine whether methylation patterns could act as stage-dependent biomarkers of gastrointestinal tumorigenesis. Next, we developed a novel strategy that uses single-step modification of DNA with sodium bisulfite and fluorescence polymerase chain reaction methodology to measure aberrant methylation in fecal DNA. Methylation of the RASSF2 and SFRP2 promoters was analyzed in 296 fecal samples obtained from a variety of patients, including 21 with gastric tumors, 152 with colorectal tumors, and 10 with non-neoplastic or inflammatory lesions in the gastrointestinal lumen.
Analysis of DNA from tissues showed presence of extensive methylation in both gene promoters exclusively in advanced gastric and colorectal tumors. The assay successfully identified one or more methylated markers in fecal DNA from 57.1% of patients with gastric cancer, 75.0% of patients with colorectal cancer, and 44.4% of patients with advanced colorectal adenomas, but only 10.6% of subjects without neoplastic or active diseases (difference, gastric cancer vs undiseased = 46.5%, 95% confidence interval (CI) = 24.6% to 68.4%, P < .001; difference, colorectal cancer vs undiseased = 64.4%, 95% CI = 53.5% to 75.2%, P < .001; difference, colorectal adenoma vs undiseased = 33.8%, 95% CI = 14.2% to 53.4%, P < .001).
Methylation of the RASSF2 and SFRP2 promoters in fecal DNA is associated with the presence of gastrointestinal tumors relative to non-neoplastic conditions. Our novel fecal DNA methylation assay provides a possible means to noninvasively screen not only for colorectal tumors but also for gastric tumors.
开发非侵入性筛查测试对于降低胃肠道肿瘤的死亡率至关重要。我们试图通过分析粪便中脱落癌细胞的DNA甲基化来开发这样一种测试。
我们首先分析了788份原发性胃和结肠直肠组织标本中RASSF2和SFRP2基因启动子的甲基化情况，以确定甲基化模式是否可作为胃肠道肿瘤发生的阶段依赖性生物标志物。接下来，我们开发了一种新策略，该策略使用亚硫酸氢钠对DNA进行单步修饰和荧光聚合酶链反应方法来测量粪便DNA中的异常甲基化。在从各种患者获得的296份粪便样本中分析了RASSF2和SFRP2启动子的甲基化情况，这些患者包括21例胃肿瘤患者、152例结肠直肠肿瘤患者以及10例胃肠道腔中非肿瘤性或炎性病变患者。
对组织DNA的分析显示，仅在晚期胃和结肠直肠肿瘤中，两个基因启动子均存在广泛甲基化。该检测方法成功地在57.1%的胃癌患者、75.0%的结肠直肠癌患者和44.4%的晚期结肠直肠腺瘤患者的粪便DNA中鉴定出一种或多种甲基化标志物，但在无肿瘤或活动性疾病的受试者中仅为10.6%（差异，胃癌与无疾病者相比=46.5%，95%置信区间(CI)=24.6%至68.4%，P<.001；差异，结肠直肠癌与无疾病者相比=64.4%，95%CI=53.5%至75.2%，P<.001；差异，结肠直肠腺瘤与无疾病者相比=33.8%，95%CI=14.2%至53.4%，P<.001）。
相对于非肿瘤性疾病，粪便DNA中RASSF2和SFRP2启动子的甲基化与胃肠道肿瘤的存在相关。我们新的粪便DNA甲基化检测方法提供了一种不仅可用于非侵入性筛查结肠直肠肿瘤，还可用于胃肿瘤的可能方法。
