Leest Paul van der, Boonstra Pieter A, Elst Arja Ter, Kempen Léon C van, Tibbesma Marco, Koopmans Jill, Miedema Anneke, Tamminga Menno, Groen Harry J M, Reyners Anna K L, Schuuring Ed
Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands.
Department of Medical Oncology, University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands.
Cancers (Basel). 2020 May 13;12(5):1222. doi: 10.3390/cancers12051222.
Circulating cell-free DNA (ccfDNA) may contain DNA originating from the tumor in plasma of cancer patients (ctDNA) and enables noninvasive cancer diagnosis, treatment predictive testing, and response monitoring. A recent multicenter evaluation of workflows by the CANCER-ID consortium using artificial spiked-in plasma showed significant differences and consequently the importance of carefully selecting ccfDNA extraction methods. Here, the quantity and integrity of extracted ccfDNA from the plasma of cancer patients were assessed. Twenty-one cancer patient-derived cell-free plasma samples were selected to compare the Qiagen CNA, Maxwell RSC ccfDNA plasma, and Zymo manual quick ccfDNA kit. High-volume citrate plasma samples collected by diagnostic leukapheresis from six cancer patients were used to compare the Qiagen CNA (2 mL) and QIAamp MinElute ccfDNA kit (8 mL). This study revealed similar integrity and similar levels of amplified short-sized fragments and tumor-specific mutants comparing the CNA and RSC kits. However, the CNA kit consistently showed the highest yield of ccfDNA and short-sized fragments, while the RSC and ME kits showed higher variant allelic frequencies (VAFs). Our study pinpoints the importance of standardizing preanalytical conditions as well as consensus on defining the input of ccfDNA to accurately detect ctDNA and be able to compare results in a clinical routine practice, within and between clinical studies.
循环游离DNA（ccfDNA）可能包含癌症患者血浆中源自肿瘤的DNA（ctDNA），并可用于无创癌症诊断、治疗预测性检测和疗效监测。癌症识别（CANCER-ID）联盟最近使用人工添加的血浆对工作流程进行的多中心评估显示出显著差异，因此仔细选择ccfDNA提取方法非常重要。在此，对从癌症患者血浆中提取的ccfDNA的数量和完整性进行了评估。选择了21份癌症患者来源的游离血浆样本，以比较Qiagen CNA、Maxwell RSC ccfDNA血浆和Zymo手动快速ccfDNA试剂盒。使用通过诊断性白细胞分离术从6名癌症患者收集的大容量柠檬酸盐血浆样本，比较Qiagen CNA（2 mL）和QIAamp MinElute ccfDNA试剂盒（8 mL）。这项研究表明，比较CNA和RSC试剂盒时，ccfDNA的完整性以及扩增的短片段和肿瘤特异性突变体的水平相似。然而，CNA试剂盒始终显示出ccfDNA和短片段的最高产量，而RSC和ME试剂盒显示出更高的变异等位基因频率（VAF）。我们的研究指出了标准化分析前条件以及就定义ccfDNA输入达成共识的重要性，以便在临床常规实践中以及临床研究内部和之间准确检测ctDNA并能够比较结果。
