Clinical Validation of a Next-Generation Sequencing Genomic Oncology Panel via Cross-Platform Benchmarking against Established Amplicon Sequencing Assays.
Next-generation sequencing (NGS) genomic oncology profiling assays have emerged as key drivers of personalized cancer care and translational research. However, validation of these assays to meet strict clinical standards has been historically problematic because of both significant assay complexity and a scarcity of optimal validation samples. Herein, we present the clinical validation of 76 genes from a novel 1212-gene large-scale hybrid capture cancer sequencing assay (University of Chicago Medicine OncoPlus) using full-data comparisons against multiple clinical NGS amplicon-based assays to yield dramatic increases in per-sample data comparison efficiency compared with previously published validations. Using a sample set of 104 normal, solid tumor, and hematopoietic malignancy specimens, head-to-head NGS data analyses allowed for 6.8 million individual clinical base call comparisons, including 2729 previously confirmed variants, with 100% sensitivity and specificity. University of Chicago Medicine OncoPlus showed excellent performance for detection of single-nucleotide variants, insertions/deletions up to 52 bp, and FLT3 internal tandem duplications of up to 102 bp or larger. Highly concordant copy number variant and ALK/RET/ROS1 gene fusion detection were also observed. In addition to underlining the efficiency of NGS validation via full-data benchmarking against existing clinical NGS assays, this study also highlights the degree of performance similarity between hybrid capture and amplicon assays that is attainable with the application of strict quality control parameters and optimized computational analytics.
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Sabah
S
Kadri
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Bradley C
BC
Long
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Ibro
I
Mujacic
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Chao J
CJ
Zhen
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Michelle N
MN
Wurst
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Shruti
S
Sharma
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Nadia
N
McDonald
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Nifang
N
Niu
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Sonia
S
Benhamed
Department of Pathology, the Institute for Genomics and Systems Biology, The University of Chicago, Chicago, Illinois.
Jigyasa H
JH
Tuteja
Department of Medicine, The University of Chicago, Chicago, Illinois.
Tanguy Y
TY
Seiwert
Department of Pathology, the Institute for Genomics and Systems Biology, The University of Chicago, Chicago, Illinois.
Kevin P
KP
White
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Megan E
ME
McNerney
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Carrie
C
Fitzpatrick
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Y Lynn
YL
Wang
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois.
Larissa V
LV
Furtado
Division of Genomic and Molecular Pathology, The University of Chicago, Chicago, Illinois. Electronic address: jsegal5@bsd.uchicago.edu.
Jeremy P
JP
Segal
D019985:Benchmarking; D056915:DNA Copy Number Variations; D004252:DNA Mutational Analysis / Q000592:standards*; D005787:Gene Frequency; D050939:Gene Fusion; D052138:Genes, Neoplasm; D023281:Genomics; D059014:High-Throughput Nucleotide Sequencing / Q000592:standards*; D006801:Humans; D057230:Limit of Detection; D009154:Mutation; D009369:Neoplasms / Q000235:genetics; D012015:Reference Standards; D012680:Sensitivity and Specificity