Hughes Edward G, Bergman Drew T, Green Donald C, Sukhadia Shrey S, Lefferts Joel A, Karrs Jeremiah X, Vyas Rutu S, Cieslak Zofia, Barbuto Jennifer N, Ognjenovic Nevena B, Palisoul Brianna E, Tsongalis Gregory J, Tafe Laura J, Shah Parth S
Laboratory for Clinical Genomics and Advanced Technologies, Department of Pathology and Laboratory Medicine, Dartmouth Hitchcock Medical Center, Lebanon, NH.
Department of Radiation Oncology and Applied Sciences, Dartmouth Hitchcock Medical Center, Lebanon, NH.
JCO Precis Oncol. 2025 May;9:e2400830. doi: 10.1200/PO-24-00830. Epub 2025 May 29.
Chromosomal rearrangements drive tumorigenesis through fusion transcripts. Clinical whole-transcriptome sequencing (WTS) enables comprehensive fusion detection, overcoming limitations of targeted methods, but its clinical application has been hindered by technical and cost barriers.
We validated a clinical bait-capture WTS assay using 78 solid tumor samples across diverse tissue types, including 59 with known fusions or oncogenic splice variants. Sensitivity was assessed against clinically reported fusions from targeted next-generation sequencing or fluorescence in situ hybridization. Analytical performance was evaluated for sensitivity, specificity, limit of detection, and reproducibility across varying RNA inputs and sequencing conditions. Clinical implementation data were also analyzed.
WTS demonstrated a sensitivity of 97.1% and a specificity of 100% for detecting clinically relevant fusions. Detection was robust across tumor types, with RNA input as low as 1 ng. Optimized sequencing thresholds of 60 million reads and 130-bp read length improved performance. The assay was highly reproducible and resistant to common contaminants. Over 6 months of clinical use (410 specimens), WTS identified 29 fusions (7% detection rate), including six clinically relevant fusions undetectable by our institution's previous targeted panel, increasing the fusion yield by 26%.
WTS provides high sensitivity, specificity, and expanded fusion detection, supporting its routine clinical adoption. Future work will explore its broader role in integrated genomic profiling and therapeutic guidance.
染色体重排通过融合转录本驱动肿瘤发生。临床全转录组测序（WTS）能够进行全面的融合检测，克服了靶向方法的局限性，但其临床应用一直受到技术和成本障碍的阻碍。
我们使用78例来自不同组织类型的实体瘤样本验证了一种临床诱饵捕获WTS检测方法，其中59例具有已知的融合或致癌剪接变体。针对靶向新一代测序或荧光原位杂交临床报告的融合，评估其敏感性。在不同RNA输入量和测序条件下，对检测方法的敏感性、特异性、检测限和重现性进行分析性能评估。还分析了临床实施数据。
WTS检测临床相关融合的敏感性为97.1%，特异性为100%。在各种肿瘤类型中检测效果良好，RNA输入量低至1 ng。优化后的测序阈值为6000万条读数和130 bp的读长可提高性能。该检测方法具有高度重现性，且对常见污染物具有抗性。在超过6个月的临床应用（410个样本）中，WTS鉴定出29种融合（检测率为7%），包括6种本机构之前的靶向检测 panel 无法检测到的临床相关融合，融合检出率提高了26%。
WTS具有高敏感性、特异性和扩展的融合检测能力，支持其在临床中的常规应用。未来的工作将探索其在综合基因组分析和治疗指导中更广泛的作用。
