中国人遗传性非息肉病性结直肠癌家系错配修复基因突变的研究

目的探讨中国人遗传性非息肉病性结直肠癌(HNPCC)家系中hMLH1、hMSH2基因遗传性突变情况。方法取14个符合中国人HNPCC标准的HNPCC家系肿瘤先证者外周血DNA样本,聚合酶链反应扩增hMLH1、hMSH2基因共35个外显子,应用变性高效液相色谱技术(DHPLc)结合DNA测序法检测突变。结果14个家系中共发生41个大肠癌和19个肠外恶性肿瘤,其中胃癌是最常见的肠外恶性肿瘤类型。14个患者中检测到分属于9个家系的12个遗传性单个碱基改变,其中8％为无义突变,25％为错义突变,其余42％为单核苷酸多态,17％为内含子区的单碱基改变,8％为同义突变。结论(1)应用DHPLc成功检测到hMLH1、hMSH2基因杂合性突变。(2)符合中国人HNPCC标准家系约有1／3可检出hMLH1、hMSH2基因遗传杂合性突变,其中错义突变较多见。

Mutation detection of mismatch repair genes in hereditary nonpolyposis colorectal cancer by denaturing high-performance liquid chromatography

OBJECTIVES: To establish DHPLC method in detecting mutations of mismatch repair genes, hMLH1 and hMSH2, and to identify germline mutations of hMLH1 and hMSH2 in HNPCC kindreds fulfilling Chinese HNPCC criteria. METHODS: Fourteen peripheral blood DNA samples from 14 unrelated HNPCC probands fulfilling Chinese HNPCC criteria were obtained respectively. PCR amplified 35 exons of two main mismatch repair genes, hMLH1 and hMSH2. DHPLC followed by DNA sequencing was used to detect and confirm mutations. RESULTS: a total of 41 colorectal cancers and 19 extracolonic tumors were developed in 14 HNPCC kindreds, and gastric cancer was the most common extracolonic tumor type. Twelve single nucleotide changes were identified by DHPLC in 14 probands. Among them, three were missense mutations, one was a nonsense mutation. Other single nucleotide changes included five single nucleotide polymorphisms, two intron single nucleotide changes, one synonymous mutation. hMLH1 EXON19 CODON749 TAC-->TAG (Tyr-->X), hMSH2 EXON12 CODON629 CAA-->CGA (Gln-->Arg) and hMSH2 EXON15 CODON839 CAT-->CGT (His-->Arg) were new discovered mutations. CONCLUSIONS: (1) DHPLC was considered to be highly effective, convenient technique with consistent results for the mutation detection of hMLH1 and hMSH2 genes. (2) Valid mutations of hMLH1 and hMSH2 genes were identified in about one-third HNPCC kindreds fulfilling Chinese HNPCC criteria and missense mutation was the most common mutational types in this cohort of families.