低龄大肠癌患者MSH2和MLH1基因的变异分析

目的　探讨错配修复基因MSH2、MLH1功能和结构改变在低龄大肠癌患者发病中的作用。方法　配对提取 4 2例低龄 (<5 0岁 )大肠癌患者的正常细胞和肿瘤细胞DNA ,在错配修复基因功能状态分析的基础上 ,对伴有微卫星DNA不稳定 (MSI+ )患者 ,应用变性高效液相色谱技术(DHPLC)、DNA序列分析技术及定量多重PCR技术系统分析MSH2、MLH1基因的微小突变和大片段DNA缺失。结果　 2 2 / 4 2 (5 2 .4 % )的患者肿瘤细胞检出MSI+ ,其中 10 / 4 2为高度不稳定 (MSI+ H) ,12 / 4 2低度不稳定 (MSI+ L)。对MSI+ 患者的进一步突变分析揭示 ,在 9例患者存在MSH2、MLH1基因的 8个位点遗传性单个碱基的改变 ,其中 3/ 8为新发现的点突变 ,5 / 8为基因的多态位点 ;在 2例存在MSI+ H的肿瘤组织检出MSH2基因大片段DNA(exon 1 6 )缺失或MLH1基因的错义突变Met2 4 2Ile。结论　中国人低龄大肠癌患者中MSH2、MLH1基因的遗传性和体细胞性突变为频发事件。

Mutation analysis on MSH2 and MLH1 genes in patients of colorectal cancer at early age

Objective To investigate the etiological role of function and construction alteration in mismatch repair genes MSH2 and MLH1 in the patients onset of colorectal cancers(CRC)at early ages. Methods The genomic DNA was extracted from the tumor tissues and normal colon tissues duing operation and subjected to analysis of microsatellite instability (MSI) in 42 Chinese patients aged less than 50 with CRC. Mutation screenings were performed with denaturing high-performance liquid chromatography (DHPLC) followed by DNA sequencing of DNA samples with variant peaks, and genomic deletion detection with quantitative multiplex PCR (Q-M-PCR) in the patients uncovered with MSI +. Results 22 out of the 42 (52.4%) patients investigated were microsatellite instability positive (MSI +), 10/42 MSI +-H and 12/42 MSI +-L. 8 kinds of DNA germline alterations, 5 polymorphisms and 3 novel point mutations, were found in 9 patients with MSI +. A large DNA ( exon 1-6 ) deletion in MSH2 gene and a missense mutation Met242Ile in MLH1 gene were unveiled in CRC tissues of two patients with MSI +-H. Conclusion Mutations of mismatch repair genes are frequent in Chinese patients of CRC with onset at early ages.