Cancer Res Treat.  2023 Apr;55(2):429-441. 10.4143/crt.2022.891.

Validation and Clinical Application of ONCOaccuPanel for Targeted Next-Generation Sequencing of Solid Tumors

• Kim M
• Lee C
• Hong J
• Kim J
• Jeong JY
• Park NJY
• Kim JE
• Park JY

• KMID: 2541230
• DOI: http://doi.org/10.4143/crt.2022.891

Abstract

Purpose
Targeted next-generation sequencing (NGS) is widely used for simultaneously detecting clinically informative genetic alterations in a single assay. Its application in clinical settings requires the validation of NGS gene panels. In this study, we aimed to validate a targeted hybridization capture-based DNA panel (ONCOaccuPanel) using the Illumina MiSeq sequencing platform. The panel allows the simultaneous detection of single-nucleotide variants (SNVs), insertions, deletions, and copy number changes of 323 genes and fusions of 17 genes in solid tumors.
Materials and Methods
We used 16 formalin-fixed paraffin-embedded (FFPE) tumor samples with previously known genetic mutations and one reference material (HD827) for validation. Moreover, we sequenced an additional 117 FFPE tumor samples to demonstrate the clinical utility of this panel.
Results
Validation revealed a 100% positive percentage agreement and positive predictive value for the detection of SNVs, insertions, deletions, copy number changes, fusion genes, and microsatellite instability–high types. We observed high levels of reproducibility and repeatability (R2 correlation coefficients=0.96-0.98). In the limit of detection assessment, we identified all clinically relevant genes with allele frequencies > 3%. Furthermore, the clinical application of ONCOaccuPanel using 117 FFPE samples demonstrated robust detection of oncogenic alterations. Oncogenic alterations and targetable genetic alterations were detected in 98.2% and 27.4% cases, respectively.
Conclusion
ONCOaccuPanel demonstrated high analytical sensitivity, reproducibility, and repeatability and is feasible for the detection of clinically relevant mutations in clinical settings.

Keyword

High-throughput nucleotide sequencing; DNA mutational analysis; Genetic testing; Validation; Neoplasms

Figure

• 

  Fig. 1 Bioinformatics pipeline analysis process of ONCOaccuPanel: overall process (A) and SNV and Indel calling process (B). CNV, copy number variation; Indel, insertion and deletion; MAF, Mutation Annotation Format; MMR, mismatch repair; MSI, microsatellite instability; POLE, polymerase ɛ; SNV, single-nucleotide variant; SV, structural variation; VCF, Variant Call Format.

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  Fig. 2 Repeatability test of ONCOaccuPanel. Percentages in the vertical axis and horizontal axis denote the variant of allele frequency (VAF). Indel, insertion and deletion; SNP, single nucleotide polymorphism.

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  Fig. 3 Reproducibility test of ONCOaccuPanel. Percentages in the vertical axis and horizontal axis denote the variant of allele frequency (VAF). Indel, insertion and deletion; SNP, single nucleotide polymorphism.

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  Fig. 4 Estimating the limit of detection of OncoAccupanel; first (A) and second (B) test. Percentages in the vertical axis denote the variant of allele frequency (VAF). Exp, expected VAF; Obs, observed VAF.

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  Fig. 5 Oncogenic mutations and fusions detected in 117 clinical samples.

Reference

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