Wang Yaping, Friedl Waltraut, Lamberti Christof, Jungck Matthias, Mathiak Micaela, Pagenstecher Constanze, Propping Peter, Mangold Elisabeth
Institute of Human Genetics, University Clinics Bonn, Germany.
Int J Cancer. 2003 Feb 20;103(5):636-41. doi: 10.1002/ijc.10869.
Hereditary nonpolyposis colorectal cancer (HNPCC) is often caused by a deficiency in DNA mismatch repair. By using conventional methods of mutation analysis, point mutations in the DNA mismatch repair genes MSH2 and MLH1 have been detected in up to 64% of patients suspected of HNPCC. However, large genomic deletions cannot be detected by these methods. In our study, we applied a semiquantitative multiplex PCR to detect the proportion of large deletions in patients meeting the Bethesda criteria whose tumours exhibited microsatellite instability (MSI). Of 368 unrelated patients, 180 exhibited MSI. In these patients, 68 disease-causing point mutations (38%) had previously been detected in the MSH2 and MLH1 genes by SSCP, heteroduplex analysis or DHPLC followed by direct sequencing. The remaining 112 patients (including 24 patients with rare missense or other unclarified variants) were examined for large deletions. We identified deletions in 19 patients (10.6%); 11/19 (58%) deletions were located in MSH2 and 8/19 (42%) in MLH1, respectively. The size of deletions ranged from 1 exon to a deletion of a whole gene. Five breakpoints of deletions were sequenced; Alu-repetitive elements were involved in all of them. In patients meeting the Amsterdam criteria the proportion of large deletions was 12.6%. A similar proportion of deletions was found in the group of patients with a positive family history for colorectal cancer and MSI tumours, not meeting the Amsterdam criteria. The results of our study suggest that large genomic deletions in both MSH2 and MLH1 genes play a considerable role in the pathogenesis of HNPCC and should be part of the routine HNPCC mutation detection protocols.
遗传性非息肉病性结直肠癌（HNPCC）通常由DNA错配修复缺陷引起。通过使用传统的突变分析方法，在高达64%疑似HNPCC的患者中检测到DNA错配修复基因MSH2和MLH1中的点突变。然而，这些方法无法检测到大片段基因组缺失。在我们的研究中，我们应用半定量多重PCR来检测符合贝塞斯达标准且肿瘤表现为微卫星不稳定（MSI）的患者中大片段缺失的比例。在368名无亲缘关系的患者中，180名表现出MSI。在这些患者中，先前通过单链构象多态性（SSCP）、异源双链分析或变性高效液相色谱（DHPLC）随后直接测序在MSH2和MLH1基因中检测到68个致病点突变（38%）。其余112名患者（包括24名具有罕见错义或其他未明确变异的患者）接受了大片段缺失检测。我们在19名患者（10.6%）中鉴定出缺失；19例中有11例（58%）缺失位于MSH2，8例（42%）位于MLH1。缺失大小从1个外显子到整个基因的缺失不等。对5个缺失断点进行了测序；所有断点均涉及Alu重复元件。在符合阿姆斯特丹标准的患者中，大片段缺失的比例为12.6%。在有结直肠癌家族史且肿瘤为MSI但不符合阿姆斯特丹标准的患者组中也发现了类似比例的缺失。我们的研究结果表明，MSH2和MLH1基因中的大片段基因组缺失在HNPCC的发病机制中起重要作用，应成为HNPCC突变检测常规方案的一部分。
