Development, testing and validation of a targeted NGS-panel for the detection of actionable mutations in lung cancer...

Area of Science:

• Oncology
• Genomics
• Molecular Diagnostics

Background:

• Lung cancer is a genetically driven disease requiring targeted therapies.
• Biomarker testing for tumor genetic alterations is crucial for selecting appropriate treatments.
• Next-generation sequencing (NGS) technologies, including anchored multiplex PCR, amplicon-based, and hybrid capture-based PCR, are used for library generation.

Purpose of the Study:

• To evaluate the anchored multiplex PCR-based ArcherDX-Variantplex nNGMv2 panel for routine molecular testing in lung cancer.
• To assess the sensitivity, specificity, and success rate of this NGS approach in a multi-center network.

Main Methods:

• Four centers applied the ArcherDX-Variantplex nNGMv2 panel to re-analyze lung cancer samples previously tested in routine diagnostics.
• Data analysis followed centrally compiled study design and pre-defined standards.
• Panel sensitivity was determined using dilution experiments, and DNA input requirements were assessed.

Main Results:

• The nNGMv2 panel achieved a high success rate of 98.9% across 90 samples.
• Most known variants were identified, with MET exon 14 skipping variants detected using adapted filters.
• Three additional clinically relevant variants (KEAP1, STK11, TP53) were identified, missed in prior testing.
• Low DNA input (as low as 6.25 ng) was sufficient for successful analysis, with total DNA amount correlating with coverage.

Conclusions:

• Anchored multiplex PCR-based sequencing using the nNGMv2 panel is a robust and specific technology for detecting tumor genetic mutations in lung cancer.
• The Archer-Analysis pipeline facilitates user-friendly and efficient analysis for precision medicine applications.
• This method supports routine molecular testing and aids in identifying actionable mutations for personalized lung cancer treatment.