title: Building a Robust Tumor Profiling Program: Synergy between Next-Generation Sequencing and Targeted Single-Gene Testing.
abstract: Next-generation sequencing (NGS) is a powerful platform for identifying cancer mutations. Routine clinical adoption of NGS requires optimized quality control metrics to ensure accurate results. To assess the robustness of our clinical NGS pipeline, we analyzed the results of 304 solid tumor and hematologic malignancy specimens tested simultaneously by NGS and one or more targeted single-gene tests (EGFR, KRAS, BRAF, NPM1, FLT3, and JAK2). For samples that passed our validated tumor percentage and DNA quality and quantity thresholds, there was perfect concordance between NGS and targeted single-gene tests with the exception of two FLT3 internal tandem duplications that fell below the stringent pre-established reporting threshold but were readily detected by manual inspection. In addition, NGS identified clinically significant mutations not covered by single-gene tests. These findings confirm NGS as a reliable platform for routine clinical use when appropriate quality control metrics, such as tumor percentage and DNA quality cutoffs, are in place. Based on our findings, we suggest a simple workflow that should facilitate adoption of clinical oncologic NGS services at other institutions.
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Hiemenz
authors.affiliation: Medical Scientist Training Program, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Stephan
authors.initials: S
authors.lastname: Kadauke
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: David B
authors.initials: DB
authors.lastname: Lieberman
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: David B
authors.initials: DB
authors.lastname: Roth
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Zhao
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Christopher D
authors.initials: CD
authors.lastname: Watt
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Robert D
authors.initials: RD
authors.lastname: Daber
authors.affiliation: Center for Personalized Diagnostics, Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Jennifer J D
authors.initials: JJ
authors.lastname: Morrissette
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics; D019295:Computational Biology / Q000379:methods; D005820:Genetic Testing / Q000379:methods; D023281:Genomics / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D009154:Mutation; D009369:Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D000090243:Nucleophosmin
chemical_list: D014408:Biomarkers, Tumor; C000716967:NPM1 protein, human; D000090243:Nucleophosmin