Related Experiment Video
Updated: Jun 11, 2025
Integration of Wet and Dry Bench Processes Optimizes Targeted Next-generation Sequencing of Low-quality and Low-quantity Tumor Biopsies
Published on: April 11, 2016
ASPYRE-Lung: validation of a simple, fast, robust and novel method for multi-variant genomic analysis of actionable
Ryan T Evans1, Elizabeth Gillon-Zhang1, Julia N Brown1
1Biofidelity Inc., Morrisville, NC, United States.
Introduction:
Genomic variant testing of tumors is a critical gateway for patients to access the full potential of personalized oncology therapeutics. Current methods such as next-generation sequencing are costly and challenging to interpret, while PCR assays are limited in the number of variants they can cover. We developed ASPYRE® (Allele-Specific PYrophosphorolysis REaction) technology to address the urgent need for rapid, accessible and affordable diagnostics informing actionable genomic target variants of a given cancer. The targeted ASPYRE-Lung panel for non-small cell carcinoma covers 114 variants in 11 genes (ALK, BRAF, EGFR, ERBB2, KRAS, RET, ROS1, MET & NTRK1/2/3) to robustly inform clinical management. The assay detects single nucleotide variants, insertions, deletions, and gene fusions from tissue-derived DNA and RNA simultaneously.
Methods:
We tested the limit of detection, specificity, analytical accuracy and analytical precision of ASPYRE-Lung using FFPE lung tissue samples from patients with non-small cell lung carcinoma, variant-negative FFPE tissue from healthy donors, and FFPE-based contrived samples with controllable variant allele fractions.
Results:
The sensitivity of ASPYRE-Lung was determined to be ≤ 3% variant allele fraction for single nucleotide variants and insertions or deletions, 100 copies for fusions, and 200 copies for MET exon 14 skipping. The specificity was 100% with no false positive results. The analytical accuracy test yielded no discordant calls between ASPYRE-Lung and expected results for clinical samples (via orthogonal testing) or contrived samples, and results were replicable across operators, reagent lots, runs, and real-time PCR instruments with a high degree of precision.
Conclusions:
The technology is simple and fast, requiring only four reagent transfer steps using standard laboratory equipment (PCR and qPCR instruments) with analysis via a cloud-based algorithm. The ASPYRE-Lung assay has the potential to be transformative in facilitating access to rapid, actionable molecular profiling of tissue for patients with non-small cell carcinoma.
