hedgehog信号通路的负调控基因在早期非小细胞肺癌中的甲基化状态及其意义

摘要

目的:研究hedgehog信号通路的负调控基因生长停顿特异蛋白1(growth arrest specific1,GAS1)、人类hedgehog相互作用蛋白(human hedgehog interacting protein,HHIP)和patched1 (PTCH1)在早期非小细胞肺癌(NSCLC)中的甲基化状态并初步探讨此3个基因对早期NSCLC的诊断价值.方法:收集Ⅰ期NSCLC患者85例(NSCLC组)并以同期住院的23例非肿瘤性肺部疾病患者作为对照组.采用甲基化特异性聚合酶链反应(methylation-specific polymerase chain reaction,MSP)检测GAS1、HHIP和PTCH1在5个NSCLC细胞株A549、NCI-H1299、NCI-H460、LTEP-a-2、SPC-A-1中的甲基化状态及其在NSCLC组患者的肺癌组织中、对照组的肺部病变组织中的甲基化状态；分析3个基因诊断Ⅰ期NSCLC的敏感性和特异性.结果:hedgehog信号通路的3个负调控基因除在NCFH1299中均呈去甲基化状态外,在其他4个肺癌细胞株中均有1～2个基因呈甲基化状态.GAS1、HHIP和PTCH1在NSCLC组的甲基化率高于对照组,差异有统计学意义(P＜0.05).GAS1用于临床Ⅰ期NSCLC诊断时敏感性和特异性分别为69.41％、69.57 ％;HHIP分别为49.41％、82.61 ％;PTCH1则分别为34.12％、100.0％.结论:PTCH1、HHIP和GAS1在Ⅰ期NSCLC中存在显著的异常甲基化,提示hedgehog的异常激活可能参与了NSCLC的发生和发展.对3个基因在早期NSCLC患者血浆游离DNA中甲基化状态的进一步研究可望为NSCLC的早期诊断提供新的肿瘤分子学标志物.%Objective:To study the methylation status of three negative regulators for hedgehog pathway including growth arrest specific 1 (GAS1),human hedgehog interacting protein(H HIP) and patched1 (PTCH1),in order to investigate the value of them in the early diagnosis of stage Ⅰ non-small-cell lung cancer(NSCLC).Methods:A total of 85 patients with stage Ⅰ NSCLC (NSCLC group) and 23 patients with non-cancerous lung diseases(control group) have been enrolled in this study.Methylationspecific polymerase chain reaction(MSP) was used to detect the methylation status of three genes in five lung cancer cell lines A549,NCI-H1299,NCI-H460,LTEP-a-2 and SPC-A-1,as well as in the cancer tissue samples from NSCLC group and lung lesions from control group.The sensitivity and specificity of three genes were analyzed when the methylation detection of each of them was applied for the diagnosis of stage Ⅰ NSCLC.Results:Although in NCI-H1299,all of three genes presented unmethylation; in the other four lung cancer cell lines there were at least one to two genes presenting methylation.The methylation rates of GAS1,HHIP and PTCH1 in NSCLC group were higher than those in control group(P＜0.05).The sensitivity and specificity were 69.41％ and 69.57 ％ when the methylation of GAS1 was used for the diagnosis of stage Ⅰ NSCLC,while they were 49.41％ and 82.61 ％ for HHIP,and 34.12 ％ and 100.0 ％ for PTCH1.Conclusions:PTCH1,HHIP and GAS1 have been found abnormally hypermethylated in the tumor tissues from stage Ⅰ NSCLC patients,which suggests that the abnormal activation of hedgehog pathway may be involved in the genesis and development of NSCLC.Further study of the methylation status of these genes in the cell-free serum DNA from stage Ⅰ NSCLC patients may provide novel methylation biomarkers for the diagnosis of NSCLC at early stages.