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Updated: Aug 7, 2026
Next Generation Sequencing for the Detection of Actionable Mutations in Solid and Liquid Tumors
Published on: September 20, 2016
High-throughput detection of clinically targetable alterations using next-generation sequencing
Julie A Vendrell1, David Grand2, Isabelle Rouquette2
1CHU Montpellier, Arnaud de Villeneuve Hospital, Department of Pathology, Montpellier, France.
Abstract:
Next-generation sequencing (NGS) has revolutionized the therapeutic care of patients by allowing high-throughput and parallel sequencing of large numbers of genes in a single run. However, most of available commercialized cancer panels target a large number of mutations that do not have direct therapeutic implications and that are not fully adapted to low quality formalin-fixed, paraffin-embedded (FFPE) samples. Here, we designed an amplicon-based NGS panel assay of 16 currently actionable genes according to the most recent recommendations of the French National Cancer Institute (NCI). We developed a panel of short amplicons (<150 bp) using dual-strand library preparation. The clinical validation of this panel was performed on well-characterized controls and 140 routine diagnostic samples, including highly degraded and cross-linked genomic DNA extracted from FFPE tumor samples. All mutations were detected with elevated inter-laboratory and inter-run reproducibility. Importantly, we could detect clinically actionable alterations in FFPE samples with variant allele frequencies as low as 1%. In addition, the overall molecular diagnosis rate was increased from 40.7% with conventional techniques to 59.2% with our NGS panel, including 41 novel actionable alterations normally not explored by conventional techniques. Taken together, we believe that this new actionable target panel represents a relevant, highly scalable and robust tool that is easy to implement and is fully adapted to daily clinical practice in hospital and academic laboratories.
Insights
This study introduces a new next-generation sequencing (NGS) panel for detecting actionable cancer mutations, even in challenging formalin-fixed, paraffin-embedded (FFPE) samples. The assay improves molecular diagnosis rates and identifies novel therapeutic targets for cancer patients.
Area of Science:
- Oncology
- Genomics
- Molecular Diagnostics
Background:
- Next-generation sequencing (NGS) offers high-throughput gene analysis for cancer care.
- Current commercial cancer panels often include non-actionable mutations and struggle with degraded formalin-fixed, paraffin-embedded (FFPE) samples.
Purpose of the Study:
- To design and validate a novel amplicon-based NGS panel targeting 16 actionable genes for routine clinical use.
- To assess the panel's performance on FFPE samples and its impact on molecular diagnosis rates.
Main Methods:
- Development of a targeted NGS panel with short amplicons (<150 bp) and dual-strand library preparation.
- Clinical validation using well-characterized controls and 140 FFPE tumor samples with degraded DNA.
- Assessment of reproducibility and detection limits for actionable alterations.
Main Results:
- High inter-laboratory and inter-run reproducibility achieved.
- Detection of clinically actionable alterations in FFPE samples with variant allele frequencies as low as 1%.
- Increased molecular diagnosis rate from 40.7% to 59.2%, identifying 41 novel actionable alterations.
Conclusions:
- The developed NGS panel is a robust, scalable, and clinically relevant tool for actionable mutation detection in FFPE samples.
- The panel enhances molecular diagnosis and facilitates the identification of new therapeutic targets in daily hospital and academic laboratory practice.
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