血浆中多基因联合甲基化检测在肺癌诊断中的应用

摘要

目的：探究血浆中CDH13、RASSF13A、DLEC13、SEPT13、RUNX13等抑癌基因启动子甲基化及其联合检测在肺癌诊断中的价值。从中选出诊断效能高的组合。方法：采用巢式甲基化特异性PCR（ nest methylation specific PCR，nMSP）法，检测106例健康人血浆样本、106例肺癌组织和癌旁组织以及其对应的106例术前血浆样本、中基因启动子区的甲基化状态。对血浆基因组DNA修饰后进行多重置换扩增（ multiple displacement amplifica-tion， MDA），以解决血浆DNA模板不足的问题。结果：肺癌组织样本中的CDH13、RASSF13A、DLEC13、SEPT13、RUNX13基因启动子甲基化率分别为51.9％、44.3％、54.7％、36.8％、24.5％。对应的血浆样本中的 CDH13、RASSF1A、DLEC1、SEPT9、RUNX3基因启动子甲基化率分别为46.2％、41.5％、50.9％、31.1％、19.8％。 Kappa一致性检验结果表明肺癌组织与血浆的甲基化检出率一致。 CDH13、DLEC1、RASSF1A、SEPT9组合对肺癌的诊断效能明显高于其它组，准确度ACC为82.08％， Youden指数为0.6415（灵敏度为79.49％，特异度为81.13％）。结论：血浆多基因联合甲基化检测有望应用于肺癌早期诊断。%[ ABSTRACT] AIM: To determine the aberrant methylation status in the gene promoter regions of CDH13, RASSF1A, DLEC1, SEPT9 and RUNX3 by detecting the plasma specimens and the value of their combined detection for di-agnosis of lung cancers.METHODS:Nest methylation specific PCR ( nMSP) was used to detect the promoter methylation status of the 5 genes in the plasma from 106 normal controls, lung cancer tissues, lung benign tissues and the plasma from 106 patients with lung cancers.Multiple displacement amplification ( MDA) was used to amplify modified genomic DNA to solve the problem of insufficient of plasma DNA template.RESULTS: The positive rates of promoter methylation of CDH13, RASSF1A, DLEC1, SEPT9 and RUNX3 in the lung cancer tissues were 51.9%, 44.3%, 54.7%, 36.8%, 24.5%, respectively, and those in the plasma were 46.2%, 41.5%, 50.9%, 31.1%, 19.8%, respectively.The re-sults of the Kappa consistency check showed that the lung cancer tissues and the plasma had obviously coherence in the methylation status of the 5 gene promoter regions.Combination of DLEC1, CDH13, RASSF1A, and SEPT9 had a higher di-agnostic efficiency than the others, as their ACC value was 0.8208 and youden index was 0.6415 ( with the sensitivity of 81.13% and the specificity of 83.02%) .CONCLUSION:Combination detection of promoter methylation of lung cancer-related genes in the plasma is expected to apply to the early diagnosis of lung cancer.