Laboratory of Molecular Pathology and Cancer Genomics, College of Pharmacy, Seoul National University, Seoul, Korea.
The Center for Anti-Cancer Companion Diagnostics, Bio-MAX/N-Bio, Seoul National University, Seoul, Korea.
Sci Rep. 2018 Jan 11;8(1):543. doi: 10.1038/s41598-017-18642-x.
In clinical translational research and molecular in vitro diagnostics, a major challenge in the detection of genetic mutations is overcoming artefactual results caused by the low-quality of formalin-fixed paraffin-embedded tissue (FFPET)-derived DNA (FFPET-DNA). Here, we propose the use of an 'internal quality control (iQC) index' as a criterion for judging the minimum quality of DNA for PCR-based analyses. In a pre-clinical study comparing the results from droplet digital PCR-based EGFR mutation test (ddEGFR test) and qPCR-based EGFR mutation test (cobas EGFR test), iQC index ≥ 0.5 (iQC copies ≥ 500, using 3.3 ng of FFPET-DNA [1,000 genome equivalents]) was established, indicating that more than half of the input DNA was amplifiable. Using this criterion, we conducted a retrospective comparative clinical study of the ddEGFR and cobas EGFR tests for the detection of EGFR mutations in non-small cell lung cancer (NSCLC) FFPET-DNA samples. Compared with the cobas EGFR test, the ddEGFR test exhibited superior analytical performance and equivalent or higher clinical performance. Furthermore, iQC index is a reliable indicator of the quality of FFPET-DNA and could be used to prevent incorrect diagnoses arising from low-quality samples.
在临床转化研究和分子体外诊断中，检测基因突变的一个主要挑战是克服由福尔马林固定石蜡包埋组织（FFPET）衍生 DNA（FFPET-DNA）质量低引起的人为结果。在这里，我们建议使用“内部质量控制（iQC）指数”作为判断基于 PCR 分析的 DNA 最低质量的标准。在一项临床前研究中，比较了基于液滴数字 PCR 的 EGFR 突变检测（ddEGFR 检测）和基于 qPCR 的 EGFR 突变检测（cobas EGFR 检测）的结果，建立了 iQC 指数≥0.5（iQC 拷贝数≥500，使用 3.3ng 的 FFPET-DNA [1000 个基因组当量]），表明超过一半的输入 DNA 可扩增。使用该标准，我们对 ddEGFR 和 cobas EGFR 检测用于检测非小细胞肺癌（NSCLC）FFPET-DNA 样本中的 EGFR 突变进行了回顾性比较临床研究。与 cobas EGFR 检测相比，ddEGFR 检测表现出优越的分析性能和等效或更高的临床性能。此外，iQC 指数是 FFPET-DNA 质量的可靠指标，可用于防止因低质量样本导致的误诊。
