PO-399 Genome wide histone modification patterns at the promoter regions are distinct in low and high grade gliomas

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IntroductionFast and accurate detection of BRCA1 and BRCA2 germline alterations is at utmost importance in both prevention and therapeutic aims. High-throughput NGS is increasingly being applied in clinical diagnosis for genetic testing that reduces the cost and turnaround time. As several commercial multiplex amplicon-based targeted NGS kits and bioinformatics solutions are now available for BRCA testing, the aim of this study was to evaluate the best approach for the identification of single-nucleotide variants (SNVs), insertion/deletion variants (indels) and copy number variations (CNVs).Material and methodsFour assays (BRCA HC, BRCATumour, Access Array BRCA, and Ion AmpliSeq BRCA) and two bioinformatics software programs (Sophia DDM platform and SeqNext software) were tested on a training set of 28 previously genotyped samples. The most relevant solution was then blindly tested on a larger cohort of 152 samples.Results and discussionsIn the training set, the BRCATumour and Access array BRCA panels provide the highest performance in terms of coverage uniformity. For the Ion AmpliSeq BRCA panel, exons 20 and 23 of BRCA2 showed a systematic drop in coverage and it also displayed an important issue of low-level of overlapping amplicons. Moreover, false positive variants were only reported using this assay independently of the bioinformatics analysis. Except for the Access Array BRCA, the analysis of CNVs has been successfully performed in the assays. Regardless of the enrichment kit used for library preparation, Sophia DDM outperforms SeqNext for CNV analysis. Our analysis performed on the training set suggests that, in terms of variant detection accuracy in BRCA1 and BRCA2, the BRCATumour and the BRCA HC panels are the most appropriate solutions. As the BRCA HC panel features lower coverage uniformity, the BRCATumour panel was preferred for the validation step. As for the training set, a good/excellent accuracy was achieved by the selected workflow (BRCATumour and Sophia DDM) for BRCA alterations detection in the independent validation set.ConclusionTo our knowledge, a direct and systematic comparison of these Amplicon–based panel NGS methods has so far never been performed. Among the four solutions, that BRCA HC and BRCATumour panels exhibited suitable data for SNVs/indels and CNV analysis. Interestingly, the use of the BRCA Tumour Kit combined with the Sophia DDM platform provides a powerful tool for reliable detection of genetic alterations in BRCA1 or BRCA2.