Article
Author: Borkhardt, Arndt ; Möbs, Markus ; Hamelmann, Stefan ; Ball, Markus ; Goering, Wolfgang ; Blanc, Eric ; Mock, Andreas ; Martis-Thiele, Mihaela ; Meier-Kolthoff, Jan ; Hartmann, Nils ; Pfarr, Nicole ; Mayr, Eva-Maria ; Hölscher, Florian ; Klauschen, Frederick ; Hirsch, Daniela ; Kumbrink, Jörg ; Murtagh, Justin ; Evert, Matthias ; Werner, Martin ; Niewöhner, Jakob ; Lehmann, Ulrich ; Fink, Annette ; Horny, Kai ; Immel, Alexander ; Stenzinger, Albrecht ; Meier-Kolthoff, Jan P. ; Gerstenmaier, Uwe ; Hirsch, Steffen ; Begemann, Matthias ; Gaisa, Nadine T ; Merkelbach-Bruse, Sabine ; Greif, Philipp A ; Wulf, Anna-Lena ; Moskalev, Evgeny A ; Dikow, Nicola ; Lehmann, Kjong-Van ; Siebolts, Udo ; Ruckert, Christian ; Matysiak, Uta ; Jelting, Yvonne ; Seillier, Lancelot ; Ferrazzi, Fulvia ; Eberhardt, Timo ; Hofmann, Winfried ; Junk, Stefanie V ; Malek, Nisar ; Junk, Stefanie V. ; Jonigk, Danny ; Beule, Dieter ; Sahm, Felix ; Kloth, Michael ; Fürstberger, Axel ; Gaisa, Nadine T. ; Banan, Rouzbeh ; Reuter-Jessen, Kirsten ; Tischler, Verena ; Jurmeister, Phillip ; Wolter, Steffen ; Roth, Wilfried ; Sivalingam, Sugirthan ; Goldschmid, Hannah ; Claus, Rainer ; Schroeder, Christopher ; Ossowski, Stephan ; Doll, Julia ; Schirmacher, Peter ; Groß, Thomas ; Junk, Stefanie ; Göbel, Kirsten ; Rehker, Jan ; Dietmaier, Wolfgang ; Kirmse, Santina ; Menzel, Michael ; Maurus, Katja ; Auber, Bernd ; Schaaf, Christian P ; Benary, Manuela ; Dintner, Sebastian ; Horst, David ; Tögel, Lars ; Yasin, Layal ; Lassmann, Silke ; Schmidt, Gunnar ; Brüchle, Nadina Ortiz ; Kutz, Oliver ; Ströbel, Philipp ; Märkl, Bruno ; Schaaf, Christian P. ; Jacob, Anne ; Kazdal, Daniel ; Meier-Kolthoff, Jan P ; Romanovsky, Eva ; Weichert, Wilko ; Falkenberg, Kim ; Prause, Rebecca ; Marienfeld, Ralf ; Neumann, Olaf ; Blank, Cornelia ; Armeanu-Ebinger, Sorin ; Budczies, Jan ; Basitta, Patrick ; Greif, Phillip ; Adam, Eugen ; Jung, Andreas ; Appenzeller, Silke ; Bartels, Stephan ; Haack, Tobias ; Ott, Alexander ; Joosten, Maria ; Elbracht, Miriam ; William, Doreen ; Carta, Maria Giulia ; Aust, Daniela ; Moskalev, Evgeny A. ; Greif, Philipp A. ; Meyer, Robert ; Trautmann, Marcel ; Maurer, Angela ; Siemanowski-Hrach, Janna ; Rosenthal, Tessa ; Dönitz, Jürgen
INTRODUCTIONWhole Exome Sequencing (WES) has emerged as an efficient tool in clinical cancer diagnostics to broaden the scope from panel-based diagnostics to screening of all genes and enabling robust determination of complex biomarkers in a single analysis.METHODSTo assess concordance, six formalin-fixed paraffin-embedded (FFPE) tissue specimens and four commercial reference standards were analyzed by WES as matched tumor-normal DNA at 21 NGS centers in Germany, each employing local wet-lab and bioinformatics. Somatic and germline variants, copy-number alterations (CNAs), and complex biomarkers were investigated. Somatic variant calling was performed in 494 diagnostically relevant cancer genes. The raw data were collected and re-analyzed with a central bioinformatic pipeline to separate wet- and dry-lab variability.RESULTSThe mean positive percentage agreement (PPA) of somatic variant calling was 76 % while the positive predictive value (PPV) was 89 % in relation to a consensus list of variants found by at least five centers. Variant filtering was identified as the main cause for divergent variant calls. Adjusting filter criteria and re-analysis increased the PPA to 88 % for all and 97 % for the clinically relevant variants. CNA calls were concordant for 82 % of genomic regions. Homologous recombination deficiency (HRD), tumor mutational burden (TMB), and microsatellite instability (MSI) status were concordant for 94 %, 93 %, and 93 % of calls, respectively. Variability of CNAs and complex biomarkers did not decrease considerably after harmonization of the bioinformatic processing and was hence attributed mainly to wet-lab differences.CONCLUSIONContinuous optimization of bioinformatic workflows and participating in round robin tests are recommended.