Sefrioui David, Beaussire Ludivine, Perdrix Anne, Clatot Florian, Michel Pierre, Frebourg Thierry, Di Fiore Frédéric, Sarafan-Vasseur Nasrin
Normandie Univ, UNIROUEN, Inserm U1245, IRON group, Rouen University Hospital, Normandy Centre for Genomic and Personalized Medicine, Department of Hepatogastroenterology, F 76000 Rouen, France.
Normandie Univ, UNIROUEN, Inserm U1245, IRON group, Rouen University Hospital, Normandy Centre for Genomic and Personalized Medicine, F 76000 Rouen, France.
Clin Biochem. 2017 Nov;50(16-17):963-966. doi: 10.1016/j.clinbiochem.2017.06.005. Epub 2017 Jun 20.
In standard pre-analytical conditions, an isolation step is required for circulating tumor DNA (ctDNA) analysis. The need for this step remains unclear with the development of ultrasensitive detection technologies such as digital PCR (dPCR). The aim of our study was to evaluate the ctDNA detection by dPCR platform either directly from plasma (plasma group, PG) or after an isolation step (isolation group, IG).
We included 17 patients corresponding to a selection of 43 blood samples in metastatic colorectal cancer patients. For each sample, ctDNA was analyzed with or without isolation step (IG and PG, respectively) using KRAS, NRAS and BRAF mutations identified from the tumor tissue. ctDNA detection was performed after a preamplication step using dPCR platform (QuantStudio™ 3D Digital PCR System). ctDNA detection rate and mutant allelic frequencies (MAF) were compared between IG and PG.
Our results showed a detection rate at 93% in IG vs. 88% in PG. The concordance rate between the two groups was 91% (39/43) for ctDNA detection with the four discordant cases occurring in patients with low MAF (<0.5%). The mean value of MAF were 16.9±18.9 and 18.5±18.9 for IG and PG, respectively (p=0.24). The correlation coefficient r for MAF was 0.82 between the two methods (p<0.0001).
In conclusion, our results show that direct detection of ctDNA from unpurified plasma is a feasible approach, particularly from sample with high MAF (>0.5%).
在标准的分析前条件下，循环肿瘤DNA（ctDNA）分析需要一个分离步骤。随着数字PCR（dPCR）等超灵敏检测技术的发展，这一步骤的必要性仍不明确。我们研究的目的是评估通过dPCR平台直接从血浆中（血浆组，PG）或经过分离步骤后（分离组，IG）检测ctDNA的情况。
我们纳入了17例患者，对应于从转移性结直肠癌患者中选取的43份血样。对于每个样本，分别在有或无分离步骤的情况下（分别为IG和PG），使用从肿瘤组织中鉴定出的KRAS、NRAS和BRAF突变来分析ctDNA。在预扩增步骤后，使用dPCR平台（QuantStudio™ 3D数字PCR系统）进行ctDNA检测。比较IG和PG之间的ctDNA检测率和突变等位基因频率（MAF）。
我们的结果显示，IG组的检测率为93%，而PG组为88%。两组之间ctDNA检测的一致率为91%（39/43），4例不一致的病例发生在MAF较低（<0.5%）的患者中。IG组和PG组的MAF平均值分别为16.9±18.9和18.5±18.9（p = 0.24）。两种方法之间MAF的相关系数r为0.82（p<0.0001）。
总之，我们的结果表明，直接从未纯化的血浆中检测ctDNA是一种可行的方法，特别是对于MAF较高（>0.5%）的样本。
