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Updated: Jan 3, 2026
Next Generation Sequencing for the Detection of Actionable Mutations in Solid and Liquid Tumors
Published on: September 20, 2016
Development and validation of a targeted gene sequencing panel for application to disparate cancers
Mark J McCabe1,2,3, Marie-Emilie A Gauthier1,4,5, Chia-Ling Chan1
1Kinghorn Centre for Clinical Genomics, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia.
Abstract:
Next generation sequencing has revolutionised genomic studies of cancer, having facilitated the development of precision oncology treatments based on a tumour's molecular profile. We aimed to develop a targeted gene sequencing panel for application to disparate cancer types with particular focus on tumours of the head and neck, plus test for utility in liquid biopsy. The final panel designed through Roche/Nimblegen combined 451 cancer-associated genes (2.01 Mb target region). 136 patient DNA samples were collected for performance and application testing. Panel sensitivity and precision were measured using well-characterised DNA controls (n = 47), and specificity by Sanger sequencing of the Aryl Hydrocarbon Receptor Interacting Protein (AIP) gene in 89 patients. Assessment of liquid biopsy application employed a pool of synthetic circulating tumour DNA (ctDNA). Library preparation and sequencing were conducted on Illumina-based platforms prior to analysis with our accredited (ISO15189) bioinformatics pipeline. We achieved a mean coverage of 395x, with sensitivity and specificity of >99% and precision of >97%. Liquid biopsy revealed detection to 1.25% variant allele frequency. Application to head and neck tumours/cancers resulted in detection of mutations aligned to published databases. In conclusion, we have developed an analytically-validated panel for application to cancers of disparate types with utility in liquid biopsy.
Insights
A new targeted gene sequencing panel was developed for diverse cancers, including head and neck tumors, and validated for liquid biopsy applications. This molecular profiling tool demonstrates high accuracy and precision for precision oncology.
Area of Science:
- Genomic Medicine
- Cancer Genomics
- Molecular Diagnostics
Background:
- Next-generation sequencing (NGS) has transformed cancer genomics.
- Precision oncology relies on detailed molecular profiling of tumors.
- Targeted gene panels are crucial for efficient genomic analysis.
Purpose of the Study:
- To develop and validate a targeted gene sequencing panel for various cancer types.
- To assess the panel's utility in head and neck cancers.
- To evaluate the panel's application in liquid biopsy for cancer detection.
Main Methods:
- Designed a 451-gene targeted panel (2.01 Mb) using Roche/Nimblegen.
- Tested performance using 136 patient DNA samples and DNA controls.
- Assessed specificity using Sanger sequencing and liquid biopsy with synthetic ctDNA.
Main Results:
- Achieved high mean coverage (395x) with >99% sensitivity and specificity.
- Demonstrated >97% precision and detected variants down to 1.25% allele frequency in liquid biopsy.
- Identified clinically relevant mutations in head and neck tumors, aligning with databases.
Conclusions:
- Developed an analytically validated gene sequencing panel for diverse cancers.
- The panel shows significant utility for liquid biopsy applications.
- This tool supports precision oncology by enabling comprehensive tumor molecular profiling.
