Department of Thoracic and Visceral Organ Surgery, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi 371-8511, Japan.
J Mol Diagn. 2010 Mar;12(2):257-64. doi: 10.2353/jmoldx.2010.090105. Epub 2010 Jan 21.
Recent evidence indicates that the presence of epidermal growth factor receptor (EGFR) or KRAS mutations in non-small cell lung cancer (NSCLC) can predict the response of the tumor to gefinitib. However, it is difficult to detect these mutations using formalin-fixed, paraffin-embedded (FFPE) tissues because the fixation process and aging can damage the DNA. In this study, we describe our work in adapting the Smart Amplification Process version 2 (SmartAmp2) to detect EGFR or KRAS mutations in DNA extracted from FFPE tissues. We were able to detect these mutations in 37 (97%) of 38 FFPE lung cancer tissue samples within 60 minutes with the SmartAmp2 assay and to confirm the correlation between EGFR mutations in FFPE tissues and gefitinib responsiveness. All mutations had previously been confirmed in the 38 samples using DNA extracted from frozen tissues. Electrophoresis results indicated that PCR analysis was not reliable for DNA extracted from FFPE tissue when primers with a long amplicon (>300 bp) were used. This study confirms that the SmartAmp2 assay is suitable for use with DNA extracted from FFPE as well as frozen tissues.
最近的证据表明，表皮生长因子受体（EGFR）或 KRAS 突变存在于非小细胞肺癌（NSCLC）中，可以预测肿瘤对吉非替尼的反应。然而，由于固定过程和老化会损害 DNA，因此使用福尔马林固定、石蜡包埋（FFPE）组织很难检测这些突变。在这项研究中，我们描述了我们在将 Smart Amplification Process version 2（SmartAmp2）适应于检测从 FFPE 组织中提取的 DNA 中的 EGFR 或 KRAS 突变方面的工作。我们能够在 60 分钟内通过 SmartAmp2 检测到 38 个（97%）FFPE 肺癌组织样本中的这些突变，并证实了 FFPE 组织中的 EGFR 突变与吉非替尼反应性之间的相关性。所有突变在之前均已使用从冷冻组织中提取的 DNA 在 38 个样本中得到证实。电泳结果表明，当使用长扩增子（>300bp）的引物时，从 FFPE 组织中提取的 DNA 时，PCR 分析不可靠。这项研究证实，SmartAmp2 检测法适用于从 FFPE 组织和冷冻组织中提取的 DNA。
