错配修复基因1启动子甲基化状态与骨肉瘤临床病理及预后的关系

Relationship between methylation status of human mut-lhomologue 1 gene promoter and clinicopathology and prognosis of osteosarcoma

目的探讨错配修复基因1(hMLH1)启动子甲基化状态与骨肉瘤患者临床病理和预后之间的关系。方法选取于2003年至2008年在中南大学湘雅二医院行手术治疗骨肉瘤患者46例,其中男33例,女13例,年龄(20.00±9.93)岁,根据Enneking分期分为Ⅰ期27例,Ⅱ期19例。提取骨肉瘤组织中的DNA,使用巢式甲基化特异性PCR(MSP)对hMLH1基因启动子甲基化状态进行检测。采用χ^(2)检验分析hMLH1基因启动子甲基化状态与患者临床病理资料之间的关系;运用单因素分析Kaplan-Meier法对hMLH1基因启动子甲基化状态与患者总体生存、无瘤生存的关系进行分析,并对得出的生存曲线进行对数秩和(Log-rank)检验。结果在24例5年内发生复发的患者中,有16例存在hMLH1基因启动子的部分甲基化(66.67%);在22例5年内未发生复发的患者中,有4例存在hMLH1基因启动子的部分甲基化(18.18%)。经χ^(2)检验,hMLH1基因启动子的甲基化状态与骨肉瘤患者的5年无瘤生存明显相关(χ^(2)=25.956,P<0.01)。hMLH1基因启动子甲基化状态与其他临床特征的相关性分析结果表明hMLH1基因启动子甲基化状态与年龄(χ^(2)=0.054,P>0.05)、性别(χ^(2)=0.259,P>0.05)、Enneking分期(χ^(2)=2.609,P>0.05)、肿瘤的病理类型(χ^(2)=1.947,P>0.05)、病理分级(χ^(2)=0.002,P>0.05)、5年总体生存(χ^(2)=2.832,P>0.05)差异均无统计学意义。此外,通过对hMLH1基因启动子的甲基化状态与总体生存进行单因素分析,结果表明相较于hMLH1基因启动子甲基化组,非甲基化组患者的生存时间较长,但两组间差异无统计学意义(χ^(2)=1.427,P>0.05)。对hMLH1基因启动子的甲基化状态与无复发生存进行单因素分析的结果表明,相较甲基化组,hMLH1基因启动子非甲基化组患者的复发可能性更小,且两组间差异有统计学意义(χ^(2)=21.287,P<0.01)。结论hMLH1基因启动子的甲基化状态与骨肉瘤患者的无瘤生存明显相关,与患者的总体生存无相关。

Objective To discus the relationship between human mut-lhomologue 1(hMLH1)gene promoter methylation in osteosarcoma tissues and clinical pathology and prognosis of patients with osteosarcoma.Methods Totally,46 patients with osteosarcoma who underwent surgery in Xiangya Second Hospital from 2003 to 2008 were selected,including 33 males and 13 females,aged(20.00±9.93)years.They were divided into 27 cases in stageⅠand 19 cases in stageⅡaccording to Enneking staging method.The osteosarcoma tissues were collected and the DNA in the tissues was extracted.Nested methylation specific PCR was used to test hMLH1 promoter methylation status,and the relationship between hMLH1 promoter methylation and patients’clinicopathological features was analyzed byχ2 test,Univariate Kaplan-Meier and Log-rank tests were used for hMLH1 promoter methylation and overall survival analysis and disease-free survival analysis.Results Among the 24 patients who relapsed within 5 years,16 patients had partial methylation of hMLH1 gene promoter(66.67%),and among the 22 patients who did not relapse within 5 years,4 patients had partial methylation of hMLH1 gene promoter(18.18%).Chi-square test showed that the methylation status of hMLH1 gene promoter was significantly correlated with the 5-year tumor-free survival of osteosarcoma patients(χ2=25.956,P<0.01).The results showed that the methylation status of hMLH1 gene promoter had no significant difference with age(χ2=0.054,P>0.05),gender(χ2=0.259,P>0.05),Enneking stage(χ2=2.609,P>0.05),pathological type of tumor(χ2=1.947,P>0.05),pathological grade(χ2=0.002,P>0.05)and 5-year overall survival(χ2=2.832,P>0.05).In addition,the results showed that the overall survival curve of patients in hMLH1 gene promoter methylation group and non-methylation group was separated.The survival time in non-methylation group was longer than in methylation group,but there was no significant difference between the two groups(χ2=1.427,P>0.05).Univariate analysis of methylation status of hMLH1 gene promoter and relapse free survival in patients with osteosarcoma showed that the patients in non-methylation group were less likely to relapse than methylation group,and the difference was statistically significant(χ2=21.287,P<0.01).Conclusion hMLH1 gene promoter methylation is associated with 5 disease-free survival of osteosarcoma,but is not correlated with overall survival of osteosarcoma.