CAJ | 학술논문

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
고빈도미위성불은정성대장암"수식형"미위성변화여MLH1급KRAS기인돌변밀절상관
The"modification"Type Microsatellite Change in High Frequency Microsatellite Instability Colorectal Cancer Closely Relating to MLH1 and KRAS Mutation

万方数据

现代生物医学进展現代生物醫學進展 현대생물의학진전
PROGRESS IN MODERN BIOMEDICINE
2008年 5期 875-880 ,共6页

赵岩%张涛%张剑军%郑志超%赵宜良%前原喜彦%徐惠棉

조암%장도%장검군%정지초%조의량%전원희언%서혜면

微卫星不稳定性%DNA错配修复%突变表型%碱基置换微衛星不穩定性%DNA錯配脩複%突變錶型%堿基置換
미위성불은정성%DNA착배수복%돌변표형%감기치환
Microsatellite instability(MSI)%Rreplication errors%DNA mismatch repair%Mutator phenotype%Base substitutions

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MLHl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交变未检出.通过对突变谱的分析我们还发现,修饰型微卫星不稳定与KTAS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景解释.通过本研究,我们认为以波形为基础的微卫星不稳定新分型可能是解决目前微卫星研究领域矛盾的一个选项.一直公认为高频度微卫星不稳定性是"真正"的DNA错配修复缺陷表型,我们的研究提示实际上高频度微卫星的可能是多元的.修饰型微卫星不稳定与DNA错配修复缺陷直接关联,而跳跃型微卫星不稳定的原因尚未阐明.在高频度为微型不稳定中,携带修饰型变化的病例可以通过DNA错配修复系统缺陷来解释其病因.
본연구통과방법학적개량화관찰방식적창신시도천명저충현상적원인.미위성비전통적검측방법부능실현미위성정성검측,아소재적연구조개발료자동편단분석쌍형광표식기술,제고료미위성검측적감도화중복성,병실현료미위성편단변화장도적정량.소우6감기적미위성변화피정의위수식형미위성불은정,대우8감기적변화피정의위도약형미위성불은정,타문적전영보절연불동.전자표현위재비종류래원미위성위점기출상적증가혹감소,후자표현위거리비종류미위성편단원격부위적신파형적출현.통과연구아문발현,재DNA착배수복결함세포계급기인고제대서자발종류양본,부유수식형미위성불은정성검출;재인류DNA착배수복결함세포계련속80차전대야몰유검출도약형변화.도약형변화불능통과간단중복서렬불은정기출상적증가혹감소적루가이획득.재76례산발대장암,아문검측료미위성불은정성,KRAS기인돌변,병대고빈도미위성불은정성병례적량개주요DNA착배수복기인MSH2화MLHl진행료전장측서.아문발현,재대장암,안빈도적전통분류여안파형변화적분류유고도적일치성,고빈도미위성불은정성병례균검측도도약형표현,저빈도미위성불은정성도표현위수식형변화.재12례고빈도미위성불은정병례,유삼례검출료도약형화수식형동시존재미위성불은정적특수표형,저3례균검출KRAS적돌변,경유취적시해3례병례야동시검출료DNA착배수복기인MLH1적변이.이재기타9례고빈도미위성불은정병례,KRAS돌변급MLH1、MSH2교변미검출.통과대돌변보적분석아문환발현,수식형미위성불은정여KTAS기인12호밀마자적전환형돌변고도상관,이미위성은정적병례검출적KRAS기인12호밀마자돌변다위전환형돌변.수식형미위성불은정표형검출적고빈도전환돌변가유DNA착배수복결함적분자배경해석.통과본연구,아문인위이파형위기출적미위성불은정신분형가능시해결목전미위성연구영역모순적일개선항.일직공인위고빈도미위성불은정성시"진정"적DNA착배수복결함표형,아문적연구제시실제상고빈도미위성적가능시다원적.수식형미위성불은정여DNA착배수복결함직접관련,이도약형미위성불은정적원인상미천명.재고빈도위미형불은정중,휴대수식형변화적병례가이통과DNA착배수복계통결함래해석기병인.
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 MLH1gene, 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 MLH1 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.