Ferretti G, Curigliano G, Pastorino U, Cittadini A, Flamini G, Calabrò M G, De Pas T, Orlando L, Mandalà M, Colleoni M, Spaggiari L, Granone P L, Pagliari G, de Braud F, Fazio N, Goldhirsch A
Division of Medical Oncology, European Institute of Oncology, Milano, Italy.
Clin Cancer Res. 2000 Jun;6(6):2393-400.
A PCR-denaturant gradient gel electrophoresis (DGGE) method was developed for the detection of p53 and K-ras mutations in primary operable tumors and paired BAL samples of non-small cell lung cancer. Among 36 patients, 9 showed p53 exon V mutations in biopsies and in three paired bronchoalveolar lavage (BAL) specimens with a 33% concordance. Five patients presented p53 exon VI mutations in biopsies and in two paired BALs with a 40% concordance. No mutations were found in p53 exon VII either in biopsies or in paired BAL samples with 100% concordance. Exon VIII mutations were found in six primary tumors and in two BALs with a 33% concordance. Of 36 patients, we detected 7 (19.4%) with K-ras exon I mutations on tumor samples. DGGE analysis of DNA from BAL samples revealed three mutations distributed on K-ras exon I with a 42% overall concordance with respect to tumor tissue. Molecular screening by DGGE of p53-amplified DNA from BAL had cumulative 46.6% sensitivity, 100% specificity, and 77.7% accuracy. DGGE K-ras detection showed 43% sensitivity, 100% specificity, and 88.8% test accuracy. The method proposed demonstrated to be specific, accurate, and at relatively low cost but limited by low sensitivity in detecting the presence of neoplastic cells in patients with resectable non-small cell lung cancer.
我们开发了一种聚合酶链反应-变性梯度凝胶电泳（DGGE）方法，用于检测非小细胞肺癌可手术原发性肿瘤及配对支气管肺泡灌洗（BAL）样本中的p53和K-ras突变。在36例患者中，9例活检组织及3例配对的支气管肺泡灌洗样本显示p53外显子V突变，一致性为33%。5例患者活检组织及2例配对BAL样本显示p53外显子VI突变，一致性为40%。活检组织及配对BAL样本中均未发现p53外显子VII突变，一致性为100%。6例原发性肿瘤及2例BAL样本发现外显子VIII突变，一致性为33%。在36例患者中，我们在肿瘤样本中检测到7例（19.4%）K-ras外显子I突变。对BAL样本DNA进行DGGE分析，发现3个分布于K-ras外显子I的突变，与肿瘤组织的总体一致性为42%。对BAL样本中p53扩增DNA进行DGGE分子筛查，累积敏感性为46.6%，特异性为100%，准确性为77.7%。DGGE检测K-ras的敏感性为43%，特异性为100%，检测准确性为88.8%。所提出的方法具有特异性、准确性且成本相对较低，但在检测可切除非小细胞肺癌患者肿瘤细胞存在方面受低敏感性限制。
