尿沉淀细胞DNA的甲基化谱式分析和膀胱癌的诊断

Methylation profiling in urine sediments and detection of bladder cancer

目的:寻找并评估通过对尿沉淀细胞DNA甲基化分析检出膀胱癌的基因组合。方法:甲基化特异性PCR(methylation specific polymerase chain reaction,MSP)。结果:对3株膀胱癌细胞系进行的20个基因启动子区域CpG岛甲基化状态分析发现,除BRCA1以外其他基因至少在细胞株的1个等位基因呈高甲基化状态。对40例膀胱癌组和3组非膀胱癌对照组的尿沉淀细胞DNA进行了6个基因甲基化状态分析发现,SALL3在膀胱癌组甲基化率为67.5%(27/40),MT1A为50.0%(20/40),HPP1为50.0%(20/40),MYOD1为37.5%(15/40),BRCA1为32.5%(13/40),TMS1为5.0%(2/40)。这6个基因在3个非膀胱癌对照组,即非肿瘤性泌尿生殖系统疾病患者23例、脑外科患者6例和正常人群7例均呈去甲基化状态,所以可以受试基因中任一个基因呈现高甲基化状态作为膀胱癌的指征,该法检出膀胱癌的敏感性为90.0%(36/40)。结论:对这5个基因在尿沉淀细胞DNA中甲基化状态的检测可望有效地检出膀胱癌。

OBJECTIVE： To identify and evaluate the informative set of genes for DNA methylation-based detection of bladder cancer in urine sediments. METHODS： Methylation specific PCR was used in the test. RESULTS： Ninety of 20 genes tested in three bladder cancer cell lines （T24,5637 and SCaBER） had at least one allele methylated. Six genes were MSP profiled in urine sediments from 40 cases of bladder cancer and the following controls., the healthy volunteers （7 cases）, non-cancerous urinary diseases （23 cases） and the neurological diseases （6 cases）. While no methylated were observed in the control groups, the methylated incidence in bladder cancer group was： SALL3 is 67.5%（27/40）, MT1A is 50.0% （20/40）, HPP1 is 50.0% （20/40）, MYOD1 is 37.5%（15/40）, BRCA1 is 32.5%（13/40）, and TMS1 is 5.0%（2/40）, respectively. By using any gene being hypermethylated of the 5 most frequently methylated genes tested as the positive indicator, over 90 % （36/40 cases） bladder cancer cases in this retrospective study were detectable. CONCLUSION： MSP profiling of these 5 genes in urine sediments is a potentially robust method for the non-invasive detection of bladder cancer.