Ultra-sensitive molecular detection of gene fusions from RNA using ASPYRE

Area of Science:

• Molecular Biology
• Oncology Diagnostics
• Genomics

Background:

• RNA analysis is crucial for identifying actionable mutations in oncology, guiding treatment decisions.
• Existing gene fusion detection methods have limitations including single-gene targeting, low sensitivity, and long turnaround times.
• Next-generation sequencing DNA-based assays for fusion detection have variable sensitivity due to large intron sizes and may not confirm RNA expression.

Purpose of the Study:

• To develop a novel RNA-based method for gene fusion detection.
• To enable concurrent assessment of DNA and RNA in a single assay run.
• To improve upon the limitations of current gene fusion detection techniques.

Main Methods:

• A pyrophosphorolysis-based method was developed for detecting gene fusions directly from RNA.
• The assay utilizes identical steps and conditions for both DNA and RNA analysis.
• The workflow is designed to be similar to standard PCR assays.

Main Results:

• The method allows for the simultaneous detection of gene fusions from RNA alongside somatic mutations in DNA.
• Achieved a limit of detection below 6 molecules per 6 µL of target volume.
• The entire assay process, from nucleic acid extraction to sample analysis, can be completed within a single day.

Conclusions:

• The developed pyrophosphorolysis-based assay provides a sensitive and rapid method for RNA-based gene fusion detection.
• This approach permits concurrent DNA and RNA analysis, streamlining oncologic diagnostics.
• The assay's efficiency and speed offer a significant advancement over existing methods, with potential for faster clinical decision-making.