A Computational Framework for Comprehensive Genomic Profiling in Solid Cancers: The Analytical Performance of a High-Throughput Assay for Small and Copy Number Variants

SIMPLE SUMMARY: Comprehensive genomic profiling (CGP) is key to characterizing solid tumors at the molecular level and enabling personalized therapy. To this end, Fondazione Policlinico Universitario Agostino Gemelli IRCCS launched a CGP program enrolling cancer patients who were screened for nine different solid tumors (breast, colon, GIST, lung, melanoma, ovary, pancreas, prostate and thyroid). In this context, we evaluated the performance of the Illumina(®) TSO500 high-throughput assay. ABSTRACT: In January 2022, our institution launched a comprehensive cancer genome profiling program on 10 cancer types using a non-IVD solution named the TruSight Oncology 500 Assay provided by Illumina(®). The assay analyzes both DNA and RNA, identifying Single-Nucleotide Variants (SNV)s and Insertion–Deletion (InDel) in 523 genes, as well as known and unknown fusions and splicing variants in 55 genes and Copy Number Alterations (CNVs), Mutational Tumor Burden (MTB) and Microsatellite Instability (MSI). According to the current European IVD Directive 98/79/EC, an internal validation was performed before running the test. A dedicated open-source bioinformatics pipeline was developed for data postprocessing, panel assessment and embedding in high-performance computing framework using the container technology to ensure scalability and reproducibility. Our protocols, applied to 71 DNA and 64 RNA samples, showed full agreement between the TruSight Oncology 500 assay and standard approaches, with only minor limitations, allowing to routinely perform our protocol in patient screening.