高频度微卫星不稳定性大肠癌“修饰型”微卫星变化与MLH1及KRAS基因突变密切相关(英文)  

The "modification" Type Microsatellite Change in High Frequency Microsatellite Instability Colorectal Cancer Closely Relating to MLH1 and KRAS Mutation

作　　者：赵岩[1] 张涛[1] 张剑军[1] 郑志超[1] 赵宜良[1] 前原喜彦[2] 徐惠棉[3] 

机构地区：[1]辽宁省肿瘤医院外三科,110042 [2]九州大学大学院医学研究院消化器综合外科 [3]中国医科大学附属第一医院肿瘤科,辽宁沈阳110001

出　　处：《现代生物医学进展》2008年第5期875-880,共6页Progress in Modern Biomedicine

基　　金：中国卫生部口川医学奖学金资助

摘　　要：本研究通过方法学的改良和观察方式的创新试图阐明这种现象的原因。微卫星非传统的检测方法仅能实现微卫星定性检测,我所在的研究组开发了自动片段分析双荧光标识技术,提高了微卫星检测的感度和重复性,并实现了微卫星片段变化长度的定量。小于6碱基的微卫星变化被定义为修饰型微卫星不稳定,大于8碱基的变化被定义为跳跃型微卫星不稳定,它们的电泳谱截然不同。前者表现为在非肿瘤来源微卫星位点基础上的增加或减少,后者表现为距离非肿瘤微卫星片段远隔部位的新波形的出现。通过研究我们发现,在DNA错配修复缺陷细胞系及基因敲除大鼠自发肿瘤样本,仅有修饰型微卫星不稳定性检出;在人类DNA错配修复缺陷细胞系连续80次传代也没有检出跳跃型变化。跳跃型变化不能通过简单重复序列不稳定基础上的增加或减少的累加而获得。在76例散发大肠癌,我们检测了微卫星不稳定性,KRAS基因突变,并对高频度微卫星不稳定性病例的两个主要DNA错配修复基因MSH2和MLH1进行了全长测序。我们发现,在大肠癌,按频度的传统分类与按波形变化的分类有高度的一致性,高频度微卫星不稳定性病例均检测到跳跃型表现,低频度微卫星不稳定性都表现为修饰型变化。在12例高频度微卫星不稳定病例,有三例检出了跳跃型和修饰型同时存在微卫星不稳定的特殊表型,这3例均检出KRAS的突变,更有趣的是该3例病例也同时检出了DNA错配修复基因MLH1的变异。而在其他9例高频度微卫星不稳定病例,KRAS突变及MLH1、MSH2突变未检出。通过对突变谱的分析我们还发现,修饰型微卫星不稳定与KRAS基因12号密码子的转换型突变高度相关,而微卫星稳定的病例检出的KRAS基因12号密码子突变多为颠换型突变。修饰型微卫星不稳定表型检出的高频度转换突变可由DNA错配修复缺陷的分子背Microsatellite instability （MSI） was defined according to the frequency of positive findings in a panel of MSI markers. High frequency MSI （ MSI-H ） was the phenotype in which repeat sequences were extraordinarily unstable, and was considered to be the bona fide phenotype of DNA mismatch repair defection. However base substitutions in some well studied oncogenes or tumor suppressors were reported to be uncommon in MSI-H tumors. To explore this obvious contradiction, the relationship between MSI and KRAS gene mutations were studied in a panel of 76 human colorectal carcinomas, the whole exon of MLH1 and MSH2 were sequenced for MSI-H tumors. KRAS gene mutation was confirmed by similar frequencies in tumors of different MSI status. Intriguingly, all of the KRAS mutant MSI-H tumors harbored sequence alterations in MLHlgene, which was a key player in DNA mismatch repair system. This implied that in MSI-H tumors carrying MMR mutations, KRAS mutation were frequently and almost exclusively occurred. Furthermore, these MMR mutants were uniformly carrying a unique ＂modification＂ ＋ ＂jumping＂ type MSI, which was different to MSI-H tumors without MLHI or MSH2 gene mutations. This study shaded lights on the heterogeneity of MSI-H tumors, and implied the connection between ＂modification＂ type MSI and DNA mismatch defection.

关 键 词：微卫星不稳定性 DNA错配修复 突变表型 碱基置换 

分 类 号：R735.34[医药卫生—肿瘤]