Reporting Tumor Molecular Heterogeneity in Histopathological Diagnosis |
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| Authors: | Andrea Mafficini Eliana Amato Matteo Fassan Michele Simbolo Davide Antonello Caterina Vicentini Maria Scardoni Samantha Bersani Marisa Gottardi Borislav Rusev Giorgio Malpeli Vincenzo Corbo Stefano Barbi Katarzyna O. Sikora Rita T. Lawlor Giampaolo Tortora Aldo Scarpa |
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| Affiliation: | 1. Applied Research on Cancer Network (ARC-NET) and Department of Pathology and Diagnostics, University and Hospital Trust of Verona, Verona, Italy.; 2. Department of Surgery, University and Hospital Trust of Verona, Verona, Italy.; 3. Department of Medicine, Oncology Unit, University and Hospital Trust of Verona, Verona, Italy.; National Cancer Institute, National Institutes of Health, United States of America, |
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| Abstract: | BackgroundDetection of molecular tumor heterogeneity has become of paramount importance with the advent of targeted therapies. Analysis for detection should be comprehensive, timely and based on routinely available tumor samples.AimTo evaluate the diagnostic potential of targeted multigene next-generation sequencing (TM-NGS) in characterizing gastrointestinal cancer molecular heterogeneity.Methods35 gastrointestinal tract tumors, five of each intestinal type gastric carcinomas, pancreatic ductal adenocarcinomas, pancreatic intraductal papillary mucinous neoplasms, ampulla of Vater carcinomas, hepatocellular carcinomas, cholangiocarcinomas, pancreatic solid pseudopapillary tumors were assessed for mutations in 46 cancer-associated genes, using Ion Torrent semiconductor-based TM-NGS. One ampulla of Vater carcinoma cell line and one hepatic carcinosarcoma served to assess assay sensitivity. TP53, PIK3CA, KRAS, and BRAF mutations were validated by conventional Sanger sequencing.ResultsTM-NGS yielded overlapping results on matched fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissues, with a mutation detection limit of 1% for fresh-frozen high molecular weight DNA and 2% for FFPE partially degraded DNA. At least one somatic mutation was observed in all tumors tested; multiple alterations were detected in 20/35 (57%) tumors. Seven cancers displayed significant differences in allelic frequencies for distinct mutations, indicating the presence of intratumor molecular heterogeneity; this was confirmed on selected samples by immunohistochemistry of p53 and Smad4, showing concordance with mutational analysis.ConclusionsTM-NGS is able to detect and quantitate multiple gene alterations from limited amounts of DNA, moving one step closer to a next-generation histopathologic diagnosis that integrates morphologic, immunophenotypic, and multigene mutational analysis on routinely processed tissues, essential for personalized cancer therapy. |
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