Detection of aberrant CpG island methylation in fecal DNA using an electrochemical DNA array chip as a molecular screening test for alimentary tract cancer

使用电化学 DNA 阵列芯片检测粪便 DNA 中的异常 CpG 岛甲基化作为消化道癌的分子筛查试验

• 批准号: 15590275
• 负责人: UEKI Takashi
• 金额: $ 2.3万
• 依托单位: KYUSHU UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590275/
• 关键词: molecular diagnosis; alimentary tract cancer; DNA methylation; fecal DNA; MSP

As a preparation of samples, DNA has been extracted from the fresh frozen sections of the colorectal, the gastric, and the esophageal cancers that have been dissected in 1997. The fresh frozen sections of both of the colorectal cancer and normal mucosa and the stool samples before and after the cancer resection from the patients who had colorectal cancers and accepted oral and written informed consent were also collected and processed.Primers for methylation-specific PCR included the CpG islands of 24 genes (p16,TSP1,hMLH1,H-cad,RARbeta,XAF1,SLIT2,GATA5,TIMP3,RASSF1A,CACNA1G,DLC1,SFRP2,WIF1,HLTF,MGMT,SLC13A5,TSLC1,KIRREL2,HRK,LOX,CDH4,ID4,and RASSF4). We already studied colorectal cancers and normal colorectal mucosa for aberrant DNA methylation of 12 genes (p16,TSP1,hMLH1,HCAD,RARbeta,XAF1,SLIT2,GATA5,TIMP3,RASSF1A,CACNA1G,DLC1,and SFRP2), and we detected aberrant methylation of at least one gene in 88.1% of cancers. We have been detecting an aberrant methylation of the remaining 12 genes in colorectal cancers and of 24 genes in the gastric and esophageal cancers. We have been examining the combination of these genes thought to be useful for the diagnosis now. We already designed and established the gene chip for CpG island of p16,one of the candidate genes for detection of colorectal cancers, and the control experiment is under going. The nest examination schedule includes DNA extraction from the fecal sample, detection of an aberrant methylation in fecal DNA using an electrochemical chip, and verifying the sensitivity and the specificity of this molecular diagnostics.

作为样品的制备，从1997年解剖的结直肠癌、胃癌和食道癌的新鲜冷冻切片中提取了DNA。同时收集并处理接受口头和书面知情同意的结直肠癌患者的结直肠癌和正常粘膜的新鲜冰冻切片以及手术前后的粪便标本，甲基化特异性PCR引物包括24个基因的CpG岛（p16、TSP 1、hMLH1、H-cad、RAR β、XAF 1、SLIT 2、GATA 5、TIMP 3、RASSF1A、CACNA1G、DLC1、SFRP 2、WIF 1、HLTF、MGMT、SLC13 A5、TSLC 1、KIRP2、HRK、LOX、CDH4、ID4和RASSF4）。我们已经研究了结直肠癌和正常结直肠粘膜中12个基因（p16、TSP 1、hMLH 1、HCAD、RAR β、XAF 1、SLIT 2、GATA 5、TIMP 3、RASSF 1A、CACNA 1G、DLC 1和SFRP 2）的异常DNA甲基化，我们在88.1%的癌症中检测到至少一个基因的异常甲基化。我们一直在检测结直肠癌中其余12个基因和胃癌和食管癌中24个基因的异常甲基化。我们一直在研究这些基因的组合，这些基因被认为对诊断有用。我们已经设计并建立了检测大肠癌候选基因之一p16的CpG岛基因芯片，并正在进行对照实验。巢式检查计划包括从粪便样本中提取DNA，使用电化学芯片检测粪便DNA中的异常甲基化，并验证该分子诊断的灵敏度和特异性。

项目成果

大腸癌切除例における複数癌関連遺伝子の異常メチル化

结直肠癌切除病例中多个癌症相关基因的异常甲基化

• 发表时间: 2005
• 期刊: 日本外科学会雑誌 第106巻
• 作者: Higuchi;Y.;Mizukami;Y.;Yoshimoto;T.;東 秀明;外園 幸司
• 通讯作者: 外園 幸司