Bergeron Daniel, Chandok Harshpreet, Nie Qian, Prego Matthew, Soucy Melissa, Kelly Kevin, Ananda Guruprasad, Hesse Andrew, Reddi Honey V
Clinical Genomics Laboratory, The Jackson Laboratory, Farmington, CT, 06032, USA.
J Mol Med (Berl). 2022 Feb;100(2):323-335. doi: 10.1007/s00109-021-02149-0. Epub 2022 Jan 10.
Whole transcriptome sequencing (RNA-Seq) has gained prominence for the detection of fusions in solid tumors. Here, we describe the development and validation of an in-house RNA-Seq-based test system (FusionSeq™ 2.0) for the detection of clinically actionable gene fusions, in formalin-fixed paraffin-embedded (FFPE) specimens, using seventy tumor samples with varying fusion status. Conditions were optimized for RNA input of 50 ng, shown to be adequate to call known fusions at as low as 20% neoplastic content. Evaluation of assay performance between FFPE and fresh-frozen (FF) tissues exhibited little to no difference in fusion calling capability. Performance analysis of the assay validation data determined 100% accuracy, sensitivity, specificity, and reproducibility. This clinically developed and validated RNA-Seq-based approach for fusion detection in FPPE samples was shown to be on par if not superior to off-the-shelf commercially offered assays. With gene fusions implicated in a variety of cancer types, offering high-quality, low-cost molecular testing services for FFPE specimens will serve to best benefit the patient and the advancement of precision medicine in molecular oncology. KEY MESSAGES: A custom RNA-Seq-based test system (FusionSeq™ 2.0) for the detection of clinically actionable gene fusions, Evaluation of assay performance between FFPE and fresh-frozen (FF) tissues exhibited little to no difference in fusion calling capability. The assay can be performed with low RNA input and neoplastic content. Performance characteristics of the assay validation data determined 100% accuracy, sensitivity, specificity, and reproducibility.
全转录组测序（RNA测序）在实体瘤融合检测中已备受瞩目。在此，我们描述了一种基于RNA测序的内部检测系统（FusionSeq™ 2.0）的开发与验证，该系统用于检测福尔马林固定石蜡包埋（FFPE）标本中具有临床可操作性的基因融合，使用了70个具有不同融合状态的肿瘤样本。针对50 ng的RNA输入量优化了条件，结果表明该输入量足以在肿瘤细胞含量低至20%时检测出已知融合。对FFPE组织和新鲜冷冻（FF）组织之间的检测性能评估显示，在融合检测能力方面几乎没有差异。对检测验证数据的性能分析确定其准确性、敏感性、特异性和可重复性均为100%。这种临床开发并验证的基于RNA测序的FFPE样本融合检测方法，即便不比市售现成检测方法更优越，至少也与之相当。鉴于基因融合与多种癌症类型相关，为FFPE标本提供高质量、低成本的分子检测服务将最大程度地造福患者，并推动分子肿瘤学精准医学的发展。关键信息：一种用于检测临床可操作性基因融合的基于定制RNA测序的检测系统（FusionSeq™ 2.0）；FFPE组织和新鲜冷冻（FF）组织之间的检测性能评估显示，在融合检测能力方面几乎没有差异；该检测可在低RNA输入量和肿瘤细胞含量下进行；检测验证数据的性能特征确定其准确性、敏感性、特异性和可重复性均为100%。
