De Paolis Elisa, De Bonis Maria, Concolino Paola, Piermattei Alessia, Fagotti Anna, Urbani Andrea, Scambia Giovanni, Minucci Angelo, Capoluongo Ettore
Molecular Diagnostic and Genomic Laboratory, Fondazione Policlinico Universitario A. Gemelli IRCCS, 00168 Rome, Italy.
Division of Oncological Gynecology, Department of Women's and Children's Health, Fondazione Policlinico Universitario A. Gemelli IRCCS, 00168 Rome, Italy.
Clin Chim Acta. 2021 Feb;513:17-24. doi: 10.1016/j.cca.2020.12.001. Epub 2020 Dec 7.
With the introduction of Olaparib as target therapy for High Grade Serous Ovarian Cancer (HGSOC) patients with germline and somatic BRCA1/2 mutations, the genetic test performed on tumor tissue has become important like the germline test. In somatic testing the evaluation of Large Genomic Rearrangements (LGRs) represents the main challenge. We describe a droplet digital PCR (ddPCR) assay for the evaluation of target BRCA1 LGRs on blood and formalin-fixed paraffin-embedded (FFPE)/Fresh Frozen Tissue (FFT) samples.
We analyzed blood, FFPE and FFT samples in a validation setting of n = 78 HGSOC patients. We applied the ddPCR to BRCA1 exons 2, 20 and 21 as some of the most common BRCA1 exons involved in LGRs in our cohort of patients.
The ddPCR custom assays allowed the identification of LGRs in all sample types, including FFPE specimens. Moreover, we were able to clearly detect LGRs accounted as somatic event.
The introduction of ddPCR in a comprehensive workflow, encompassing both germline and somatic tests, represents an improvement in BRCA1/2 testing. ddPCR can overcome challenges related to BRCA testing, especially on FFPE analysis. Finally, ddPCR represents a promising alternative strategy to the established standard methods currently used in clinical setting.
随着奥拉帕尼被引入作为患有种系和体细胞BRCA1/2突变的高级别浆液性卵巢癌（HGSOC）患者的靶向治疗，对肿瘤组织进行的基因检测已变得与种系检测一样重要。在体细胞检测中，大基因组重排（LGRs）的评估是主要挑战。我们描述了一种液滴数字PCR（ddPCR）检测方法，用于评估血液以及福尔马林固定石蜡包埋（FFPE）/新鲜冷冻组织（FFT）样本中的目标BRCA1 LGRs。
我们在n = 78例HGSOC患者的验证环境中分析了血液、FFPE和FFT样本。我们将ddPCR应用于BRCA1外显子2、20和21，因为这些是我们患者队列中LGRs所涉及的一些最常见的BRCA1外显子。
ddPCR定制检测方法能够在所有样本类型中鉴定出LGRs，包括FFPE标本。此外，我们能够清晰地检测到被视为体细胞事件的LGRs。
在涵盖种系和体细胞检测的综合工作流程中引入ddPCR，代表了BRCA1/2检测的一项改进。ddPCR可以克服与BRCA检测相关的挑战，尤其是在FFPE分析方面。最后，ddPCR是临床环境中目前使用的既定标准方法的一种有前景的替代策略。
