Xu Xinnan, Yang Yang, Li Hui, Chen Zhao, Jiang Gening, Fei Ke
a Department of Thoracic Surgery , Shanghai Pulmonary Hospital, Tongji University , Shanghai , P. R. China ;
b Beijing ACCB Biotech Ltd , Haidian District , Beijing , P. R. China ;
Scand J Clin Lab Invest. 2016 Sep;76(5):386-92. doi: 10.1080/00365513.2016.1183813. Epub 2016 May 23.
Next-generation sequencing (NGS) has been widely applied in clinical research, while its application in routine clinical molecular testing requires careful validation. The aim of our study was to assess the clinical usefulness of the NextDaySeq Lung panel on Ion Torrent™ PGM in mutation detection of actionable genes in lung cancer.
The NextDaySeq assay was evaluated by blinded comparisons to Quantitative Real-Time PCR (qPCR) assays with 188 consecutive samples from Chinese patients with non-small cell lung cancer (NSCLC) to detect mutations in EGFR, KRAS, PIK3CA and BRAF. Discordant variants were further validated by Sanger sequencing and independent qPCR and NGS assays.
Our results showed 93.3% concordance of reportable variants mutually covered in both NGS and qPCR assays, with a clinical sensitivity of 89.9%, specificity of 97.5%. Through the comparison, the NGS assays demonstrated its advantages in offering more clinical relevant information, such as detecting non-hotspot mutations and providing mutation allele frequencies (MAF) and accurate mutation sequences. The analytical sensitivity of NGS to detect mutations with low MAF needs further improvement.
The NextDaySeq Lung panel exhibited good clinical performance, strongly supporting the implementation of the NGS assay in routine clinical use to facilitate therapeutic decision-making for lung cancer patients.
下一代测序（NGS）已广泛应用于临床研究，但其在常规临床分子检测中的应用需要仔细验证。我们研究的目的是评估Ion Torrent™ PGM平台上的NextDaySeq肺癌检测板在肺癌可操作基因的突变检测中的临床实用性。
通过对188例中国非小细胞肺癌（NSCLC）患者的连续样本进行盲法比较，将NextDaySeq检测与定量实时PCR（qPCR）检测进行评估，以检测表皮生长因子受体（EGFR）、 Kirsten大鼠肉瘤病毒癌基因（KRAS）、磷脂酰肌醇-4,5-二磷酸3-激酶催化亚基α（PIK3CA）和B-Raf原癌基因（BRAF）的突变。不一致的变异通过桑格测序以及独立的qPCR和NGS检测进一步验证。
我们的结果显示，在NGS和qPCR检测中相互覆盖的可报告变异的一致性为93.3%，临床敏感性为89.9%，特异性为97.5%。通过比较，NGS检测在提供更多临床相关信息方面显示出优势，例如检测非热点突变、提供突变等位基因频率（MAF）和准确的突变序列。NGS检测低MAF突变的分析敏感性需要进一步提高。
NextDaySeq肺癌检测板表现出良好的临床性能，有力地支持了NGS检测在常规临床应用中的实施，以促进肺癌患者的治疗决策。
