Recent updates in the classification of central nervous system (CNS) tumors have increased the need for molecular testing. Assessment of multiple alterations in parallel, complex combinations of gene sequence and chromosomal changes, as well as therapy prediction by identification of actionable mutations are the major challenges. We here report on a customized next generation sequencing (NGS)-based DNA panel assay that combines diagnostic and predictive testing and -as a comprehensive approach- allows for simultaneous single nucleotide variant (SNP) / small insertion/deletion (InDel), copy number variation (CNV) and loss of heterozygosity (LOH) detection. We analyzed formalin-fixed and paraffin-embedded (FFPE) DNA from a total of 104 patients with CNS tumors. After amplicon capture-based library preparation, sequencing was performed on the relatively cost-efficient Illiumina MiniSeq platform and evaluated with freely available bioinformatical tools. 57 genes for exonic SNP/InDel calling (19 of those in intronic regions for CNV analysis), 3 chromosomal arms and 4 entire chromosomes for CNV and LOH analysis were covered. Results were extensively validated. Our approach yielded high accuracy, sensitivity and specificity. It led to refined diagnoses in a relevant number of analyzed cases, reliably enabled complex subclassifications (e.g. for medulloblastomas) and identified actionable targets for clinical use. Thus, our single-platform approach is an efficient and powerful tool to comprehensively support molecular testing in neurooncology. Future functionality is guaranteed as novel upcoming biomarkers can be easily incorporated in a modular panel design.

中文翻译：

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全面的 DNA panel 下一代测序方法支持神经肿瘤学的诊断和治疗预测。


中枢神经系统 (CNS) 肿瘤分类的最新更新增加了对分子检测的需求。对基因序列和染色体变化的并行、复杂组合的多重改变进行评估，以及通过识别可操作的突变来预测治疗是主要的挑战。我们在此报告基于定制的下一代测序 (NGS) 的 DNA 组合检测，该检测结合了诊断和预测测试，并且作为一种综合方法，允许同时进行单核苷酸变异 (SNP)/小插入/缺失 (InDel)、拷贝数变异（CNV）和杂合性丢失（LOH）检测。我们分析了总共 104 名中枢神经系统肿瘤患者的福尔马林固定石蜡包埋 (FFPE) DNA。在基于扩增子捕获的文库制备之后，在相对经济高效的 Iliumina MiniSeq 平台上进行测序，并使用免费提供的生物信息工具进行评估。涵盖了用于外显子 SNP/InDel 调用的 57 个基因（其中 19 个基因位于用于 CNV 分析的内含子区域）、用于 CNV 和 LOH 分析的 3 个染色体臂和 4 个完整染色体。结果得到了广泛验证。我们的方法具有很高的准确性、灵敏度和特异性。它导致了对相关数量的分析病例的精确诊断，可靠地实现了复杂的子分类（例如髓母细胞瘤）并确定了临床使用的可行目标。因此，我们的单一平台方法是全面支持神经肿瘤学分子测试的高效而强大的工具。由于即将推出的新型生物标志物可以轻松地融入模块化面板设计中，因此未来的功能得到了保证。