Performance characteristics of a targeted sequencing platform for simultaneous detection of single nucleotide variants, insertions/deletions, copy number alterations, and gene fusions in cancer genome

Context. - An increasing number of molecular laboratories are implementing next-generation sequencing platforms to identify clinically actionable and relevant genomic alterations for precision oncology. Objective. - To describe the validation studies as per New York State-Department of Health (NYS-DOH) guidelines for the Oncomine Comprehensive Panel v2, which was originally tailored to the National Cancer Institute Molecular Analysis for Therapy Choice (NCIMATCH) trial. Design. - Accuracy, precision, and reproducibility were investigated by using 130 DNA and 18 RNA samples from cytology cell blocks; formalin-fixed, paraffin-embedded tissues; and frozen samples. Analytic sensitivity and specificity were tested by using ATCC and HapMap cell lines. Results. - High accuracy and precision/reproducibility were observed for single nucleotide variants and insertion/ deletions. We also share our experience in the detection of gene fusions and copy number alterations from an amplicon-based sequencing platform. After sequencing analysis, variant annotation and report generation were performed by using the institutional knowledgebase. Conclusions. - This study serves as an example for validating a comprehensive targeted next-generation sequencing assay with both DNASeq and RNASeq components for NYS-DOH.