Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay
Detection of EGFR gene mutation in lung cancer by mutant-enriched polymerase chain reaction assay
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DOI：
10.1158/1078-0432.ccr-05-0934
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发表时间：
2006-01-01
影响因子：
11.5
通讯作者：
Shimizu, N
中科院分区：
文献类型：
--
作者：
Asano, H;Toyooka, S;Shimizu, N
目的：据报道，表皮生长因子受体（EGFR）基因突变存在于非小细胞肺癌（NSCLC）中，并与肿瘤对EGFR酪氨酸激酶抑制剂的反应性相关，表明其可作为生物标志物。由于临床样本中含有肿瘤和正常细胞或基因，因此用于检测突变的高灵敏度检测方法对于临床应用至关重要。实验设计：富含突变的PCR是一种快速和灵敏的检测方法，具有选择性限制性内切酶消化。我们开发了靶向EGFR外显子19和21的mu-富集PCR检测，并应用所开发的检测方法检测108例NSCLC手术切除标本、18例计算机断层扫描（CT）引导的针吸肺活检标本和20例胸腔积液标本中的突变。此外，结果进行了比较，从直接测序和nonenriched PCR assay.Results：富集突变的2 × 103正常基因的一个突变的mu-富集PCR检测突变37例108切除肿瘤，7个CT引导下肺活检标本，和7个样本的胸水。突变的情况下，四个切除的肿瘤，两个CT引导下的肺活检，和两个胸腔积液被确定为额外的突变的情况下，被认为是正常的基础上nonenrichedassays.Conclusions：我们的研究结果表明，EGFR突变是很容易检测到的muplants富集PCR在各种临床样本。因此，富集突变体的PCR可能提供一种有价值的方法，可能检测到一小部分的突变基因在异质性标本，表明其可能用于非小细胞肺癌的临床应用。
Purpose: Mutations in the epidermal growth factor receptor (EGFR) gene have been reported to be present in non-small cell lung cancer (NSCLC) and related to the responsiveness of tumors to EGFR tyrosine kinase inhibitors, suggesting its usefulness as a biomarker. Because clinical samples contain tumor and normal cells or genes, a highly sensitive assay for detecting mutation is critical for clinical applications.Experimental Design: The mutant-enriched PCR is a rapid and sensitive assay with selective restriction enzyme digestion. We developed the mutant-enriched PCR assay targeting exons 19 and 21 of EGFR and applied the developed assay to detect mutations in 108 cases of surgically resected specimens of NSCLCs,18 samples of computed tomography (CT)-guided needle lung biopsies, and 20 samples of pleural fluid. In addition, results were then compared with those from direct sequencing and a nonenriched PCR assay.Results: The mutant-enriched PCR that was proved to enrich one mutant of 2 x 103 normal genes detected mutations in 37 cases of 108 resected tumors, seven samples of CT-guided lung biopsies, and seven samples of pleural fluid. Among mutant cases, four resected tumors, two CT-guided lung biopsies, and two pleural fluid were identified as additional mutant cases by the mutant-enriched PCR, which were considered normal based on nonenriched assays.Conclusions: Our results indicate that EGFR mutations are readily detectable by mutant-enriched PCR in various clinical samples. Thus, mutant-enriched PCR may provide a valuable method of potentially detecting a small fraction of mutant genes in heterogeneous specimens, indicating its possible use in clinical application for NSCLC.