Comprehensive Genomic Profiling in Cancer

CGP was performed using the FDA-approved FoundationOne CDx assay (Foundation Medicine, Cambridge, MA) in a Clinical Laboratory Improvement Amendments (CLIA)-certified and College of American Pathologists (CAP)-accredited laboratory using previously described methods.^{22 (link)} Prior to nucleic acid extraction, hematoxylin and eosin-stained slides were reviewed to confirm the presence of tumor. DNA extracted from formalin-fixed paraffin-embedded tissues underwent hybrid-capture based next generation sequencing using the FoundationOne platform which interrogates all coding exons of 324 cancer-related genes and introns from 31 genes commonly rearranged in cancer. Data were analyzed for all types of genomic alterations, including base substitutions, insertions/deletions, copy number alterations, and gene rearrangements. In addition, variant-level loss of heterozygosity (LOH), tumor mutational burden (TMB), and microsatellite instability (MSI) were determined. TMB was evaluated on up to 1.1 Mb of sequenced DNA, and MSI was assessed from DNA sequencing across 95 loci as previously described.^{23 (link),24 (link)} TMB ≥ 20 mutations/Mb was considered High (TMB-High), >10 mutations/Mb considered intermediate (TMB-Int), and 0-9 mutations/Mb to be TMB low (TMB-low).