An Exome Capture-Based RNA-Sequencing Assay for Genome-Wide Identification and Prioritization of Clinically Important...

Area of Science:

• Oncology
• Genomics
• Bioinformatics

Background:

• Gene fusions are critical drivers in hematologic, solid, and central nervous system tumors.
• Accurate detection of these fusions is essential for diagnosis and targeted therapy.

Purpose of the Study:

• To develop and validate an exome capture-based RNA-sequencing assay for comprehensive gene fusion detection.
• To establish a robust bioinformatic pipeline for prioritizing clinically relevant fusions.

Main Methods:

• Utilized Twist Comprehensive Exome capture with fresh or formalin-fixed samples.
• Employed a consensus approach with four fusion callers (Arriba, FusionCatcher, STAR-Fusion, Dragen) and custom software.
• Developed a filtering and ranking algorithm based on read support, consensus, gene involvement, and database cross-referencing.

Main Results:

• The assay demonstrated high efficacy in identifying known clinically relevant fusions, ranking them first in 94% of evaluated samples.
• A significant variation in call numbers was observed among individual callers, underscoring the need for a consensus and ranking approach.
• Successfully detected pathogenic gene fusions in three diagnostically challenging pediatric cancer cases.

Conclusions:

• The developed RNA-sequencing assay provides a powerful tool for genome-wide, nontargeted gene fusion detection across diverse tumor types.
• The integrated bioinformatic platform effectively prioritizes candidate fusions, enhancing diagnostic utility.
• This approach is particularly valuable for investigating complex pediatric cancers where fusion identification is critical.