Analytical Validation and Clinical Implementation of a 1080-Gene Comprehensive Genomic Profiling Assay with...

Background: Comprehensive genomic profiling (CGP) via next-generation sequencing (NGS) is pivotal for precision oncology, yet many laboratories face challenges with incomplete genomic coverage, complex bioinformatics workflows, and limited integration of key biomarkers. Methods: We evaluated the analytical performance and clinical utility of a CGP assay using 119 tumor samples representing 18 types of cancer, previously analyzed with an orthogonal NGS panel. Concordance was assessed across 81 genes, covering 176 single-nucleotide variants (SNVs), eight copy number variations (CNVs), four deletions, one duplication, and four gene fusions. Limit of detection (LOD) studies employed AcroMetrix Mutant Hotspot Control and SeraSeq Lung and Brain CNV Mix. Microsatellite instability (MSI) and tumor mutational burden (TMB) were quantified. Inter- and intra-run reproducibility were evaluated to assess precision. Results: The CGP assay demonstrated high analytical performance, with >99% sensitivity, 100% specificity, and complete accuracy for variant detection. LOD studies revealed robust detection of SNVs at ≤5% variant allele frequencies (VAF) and CNVs at three copies. MSI and TMB results were consistent with clinical expectations, showing minimal bias compared to the orthogonal panel. Inter- and intra-run testing confirmed 100% reproducibility, indicating strong assay precision. Post-sequencing variant reporting was streamlined using the iCare platform, enabling direct FASTQ-to-report generation without intermediate bioinformatic steps. Conclusions: These findings support the present assay's clinical utility in personalized oncology assessment.