D'Haene Nicky, Le Mercier Marie, De Nève Nancy, Blanchard Oriane, Delaunoy Mélanie, El Housni Hakim, Dessars Barbara, Heimann Pierre, Remmelink Myriam, Demetter Pieter, Tejpar Sabine, Salmon Isabelle
Department of Pathology, Erasme Hospital, Université Libre de Bruxelles, Brussels, Belgium.
Department of Genetics, Erasme Hospital,Université Libre de Bruxelles, Brussels, Belgium.
PLoS One. 2015 Sep 14;10(9):e0138245. doi: 10.1371/journal.pone.0138245. eCollection 2015.
Recently, Next Generation Sequencing (NGS) has begun to supplant other technologies for gene mutation testing that is now required for targeted therapies. However, transfer of NGS technology to clinical daily practice requires validation.
We validated the Ion Torrent AmpliSeq Colon and Lung cancer panel interrogating 1850 hotspots in 22 genes using the Ion Torrent Personal Genome Machine. First, we used commercial reference standards that carry mutations at defined allelic frequency (AF). Then, 51 colorectal adenocarcinomas (CRC) and 39 non small cell lung carcinomas (NSCLC) were retrospectively analyzed.
Sensitivity and accuracy for detecting variants at an AF >4% was 100% for commercial reference standards. Among the 90 cases, 89 (98.9%) were successfully sequenced. Among the 86 samples for which NGS and the reference test were both informative, 83 showed concordant results between NGS and the reference test; i.e. KRAS and BRAF for CRC and EGFR for NSCLC, with the 3 discordant cases each characterized by an AF <10%.
Overall, the AmpliSeq colon/lung cancer panel was specific and sensitive for mutation analysis of gene panels and can be incorporated into clinical daily practice.
最近，新一代测序（NGS）已开始取代其他技术用于靶向治疗所需的基因突变检测。然而，将NGS技术应用于临床日常实践需要进行验证。
我们使用Ion Torrent个人基因组测序仪对Ion Torrent AmpliSeq结肠癌和肺癌检测板进行了验证，该检测板可检测22个基因中的1850个热点区域。首先，我们使用了携带特定等位基因频率（AF）突变的商业参考标准品。然后，对51例结肠腺癌（CRC）和39例非小细胞肺癌（NSCLC）进行回顾性分析。
对于商业参考标准品，检测AF>4%的变异的灵敏度和准确性为100%。在这90例病例中，89例（98.9%）成功测序。在NGS和参考检测均有信息的86个样本中，83个样本在NGS和参考检测之间显示出一致的结果；即CRC中的KRAS和BRAF以及NSCLC中的EGFR，3例不一致的病例的AF均<10%。
总体而言，AmpliSeq结肠癌/肺癌检测板对基因检测板的突变分析具有特异性和敏感性，可纳入临床日常实践。
