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Updated: Jul 3, 2026
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Comparative Lesions Analysis Through a Targeted Sequencing Approach
Published on: November 5, 2019
Segmental Copy Number Variant Detection Using an Amplicon-Based Next-Generation Sequencing Panel for Integrated
Kajeetha Sarvananthan1, Stephanie Santos1, Brent Saylor1
1Molecular Diagnostics Division, Pathology and Laboratory Medicine, London Health Sciences Centre, London, Ontario.
The Journal of Molecular Diagnostics : JMD
|July 1, 2026
Summary
This study introduces a custom analysis for detecting copy number variants (CNVs) in glioma using next-generation sequencing (NGS). The method accurately identifies key biomarkers, improving diagnostic efficiency and reducing costs.
Area of Science:
- Molecular oncology
- Genomics
- Cancer diagnostics
Background:
- Next-generation sequencing (NGS) is crucial for molecular oncology, detecting sequence variants (SVs) and copy number variants (CNVs).
- Amplicon-based gene panels, while not primarily designed for segmental chromosomal CNVs, can infer them.
- Accurate CNV detection is vital for classifying gliomas and guiding treatment.
Purpose of the Study:
- To describe and validate a custom analysis for CNV detection using the amplicon-based Oncomine Comprehensive Assay v3 (OCAv3).
- To assess the assay's performance for critical glioma biomarkers: 1p/19q co-deletion, +7/-10, EGFR amplification, and CDKN2A/B homozygous deletion.
- To evaluate the clinical utility and workflow impact of integrated SV and CNV analysis in glioma diagnostics.
Main Methods:
- Developed a custom CNV calling algorithm using vendor-normalized gene-level CNV data with inter-sample normalization.
- Assessed assay performance across three cohorts: discovery (N=52), validation (N=39) against FISH, and clinical (N=53).
- Defined positive thresholds for 100% specificity and permissive thresholds for reflex FISH to ensure 100% sensitivity; excluded samples with <30% tumor cellularity.
Main Results:
- The custom analysis achieved 100% concordance with FISH testing in the validation cohort for all evaluated glioma biomarkers.
- The OCAv3 assay enabled integrated classification of glioma subtypes, particularly oligodendroglioma and glioblastoma, by simultaneously assessing SVs and CNVs.
- Implementation in a clinical cohort reduced FISH studies by 90%, shortened turnaround times, and decreased molecular testing costs.
Conclusions:
- The custom amplicon-based NGS analysis provides accurate and efficient detection of critical glioma CNV biomarkers.
- Integrated SV and CNV analysis using OCAv3 streamlines glioma classification and improves diagnostic workflow.
- This approach offers a cost-effective and time-saving alternative to traditional methods, enhancing molecular oncology diagnostics.
