title: SDHB/SDHA immunohistochemistry in pheochromocytomas and paragangliomas: a multicenter interobserver variation analysis using virtual microscopy: a Multinational Study of the European Network for the Study of Adrenal Tumors (ENS@T).
abstract: Despite the established role of SDHB/SDHA immunohistochemistry as a valuable tool to identify patients at risk for familial succinate dehydrogenase-related pheochromocytoma/paraganglioma syndromes, the reproducibility of the assessment methods has not as yet been determined. The aim of this study was to investigate interobserver variability among seven expert endocrine pathologists using a web-based virtual microscopy approach in a large multicenter pheochromocytoma/paraganglioma cohort (n=351): (1) 73 SDH mutated, (2) 105 non-SDH mutated, (3) 128 samples without identified SDH-x mutations, and (4) 45 with incomplete SDH molecular genetic analysis. Substantial agreement among all the reviewers was observed either with a two-tiered classification (SDHB κ=0.7338; SDHA κ=0.6707) or a three-tiered classification approach (SDHB κ=0.6543; SDHA κ=0.7516). Consensus was achieved in 315 cases (89.74%) for SDHB immunohistochemistry and in 348 cases (99.15%) for SDHA immunohistochemistry. Among the concordant cases, 62 of 69 (~90%) SDHB-/C-/D-/AF2-mutated cases displayed SDHB immunonegativity and SDHA immunopositivity, 3 of 4 (75%) with SDHA mutations showed loss of SDHA/SDHB protein expression, whereas 98 of 105 (93%) non-SDH-x-mutated counterparts demonstrated retention of SDHA/SDHB protein expression. Two SDHD-mutated extra-adrenal paragangliomas were scored as SDHB immunopositive, whereas 9 of 128 (7%) tumors without identified SDH-x mutations, 6 of 37 (~16%) VHL-mutated, as well as 1 of 21 (~5%) NF1-mutated tumors were evaluated as SDHB immunonegative. Although 14 out of those 16 SDHB-immunonegative cases were nonmetastatic, an overall significant correlation between SDHB immunonegativity and malignancy was observed (P=0.00019). We conclude that SDHB/SDHA immunohistochemistry is a reliable tool to identify patients with SDH-x mutations with an additional value in the assessment of genetic variants of unknown significance. If SDH molecular genetic analysis fails to detect a mutation in SDHB-immunonegative tumor, SDHC promoter methylation and/or VHL/NF1 testing with the use of targeted next-generation sequencing is advisable.
authors.affiliation: 1] Department of Histopathology, King's College Hospital, London, UK [2] Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Papathomas
authors.affiliation: Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: Lindsey
authors.initials: L
authors.lastname: Oudijk
authors.affiliation: Pole of Cardiovascular Research, Institut de Recherche Expérimentale et Clinique and Division of Cardiology, Cliniques Universitaires Saint-Luc, Université Catholique de Louvain, Brussels, Belgium.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Persu
authors.affiliation: 1] Department of Anatomical Pathology, Royal North Shore Hospital, St Leonards, NSW, Australia [2] Cancer Diagnosis and Pathology Research Group, Kolling Institute of Medical Research, University of Sydney, Sydney, NSW, Australia.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Gill
authors.affiliation: Laboratory for Pathology, PAL Dordrecht, Dordrecht, The Netherlands.
authors.forename: Francien
authors.initials: F
authors.lastname: van Nederveen
authors.affiliation: Department of Pathology and Laboratory Medicine, Tufts Medical Center, Tufts University School of Medicine, Boston, MA, USA.
authors.forename: Arthur S
authors.initials: AS
authors.lastname: Tischler
authors.affiliation: 1] Institut National de la Santé et de la Recherche Médicale U1016, Institut Cochin, Centre National de la Recherche Scientifique UMR8104, Université Paris Descartes, Sorbonne Paris Cité, Rare Adrenal Cancer Network COMETE, Paris, France [2] Department of Pathology, Hôpital Pitié-Salpêtrière, Université Pierre et Marie Curie, Paris, France.
authors.forename: Frédérique
authors.initials: F
authors.lastname: Tissier
authors.affiliation: Department of Oncology, University of Turin at San Luigi Hospital, Turin, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Volante
authors.affiliation: Department of Pathology and Molecular Genetics and Research Laboratory, Hospital Universitari Arnau de Vilanova, IRBLLEIDA, University of Lleida, Lleida, Spain.
authors.forename: Xavier
authors.initials: X
authors.lastname: Matias-Guiu
authors.affiliation: Department of Medical Oncology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: Marcel
authors.initials: M
authors.lastname: Smid
authors.affiliation: 1] Paris-Centre de Recherche Cardiovasculaire (PARCC), Inserm UMR970, Hôpital Européen Georges Pompidou, Paris, France [2] Université Paris Descartes, Faculté de Médecine, Paris Cité Sorbonne, France.
authors.forename: Judith
authors.initials: J
authors.lastname: Favier
authors.affiliation: Endocrinology Unit, Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Rapizzi
authors.affiliation: Institut National de la Santé et de la Recherche Médicale U1016, Institut Cochin, Centre National de la Recherche Scientifique UMR8104, Université Paris Descartes, Sorbonne Paris Cité, Rare Adrenal Cancer Network COMETE, Paris, France.
authors.forename: Rosella
authors.initials: R
authors.lastname: Libe
authors.affiliation: Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.
authors.forename: Maria
authors.initials: M
authors.lastname: Currás-Freixes
authors.affiliation: Department of Pathology, Cliniques Universitaires Saint-Luc, Université catholique de Louvain, Institut de Recherche Expérimentale et Clinique, Brussels, Belgium.
authors.forename: Selda
authors.initials: S
authors.lastname: Aydin
authors.affiliation: Program in Reproductive and Adult Endocrinology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Thanh
authors.initials: T
authors.lastname: Huynh
authors.affiliation: Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Klinikum der Universität München, Munich, Germany.
authors.forename: Urs
authors.initials: U
authors.lastname: Lichtenauer
authors.affiliation: Department of Internal Medicine, Section of Endocrinology, Radboud University Medical Centre, Nijmegen, The Netherlands.
authors.forename: Anouk
authors.initials: A
authors.lastname: van Berkel
authors.affiliation: Endocrinology Unit, Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy.
authors.forename: Letizia
authors.initials: L
authors.lastname: Canu
authors.affiliation: Unidade de Investigação em Patobiologia Molecular (UIPM), Instituto Português de Oncologia de Lisboa Francisco Gentil, Lisbon, Portugal.
authors.forename: Rita
authors.initials: R
authors.lastname: Domingues
authors.affiliation: Cancer Genetics, Kolling Institute of Medical Research, Royal North Shore Hospital, University of Sydney, Sydney, NSW, Australia.
authors.forename: Roderick J
authors.initials: RJ
authors.lastname: Clifton-Bligh
authors.affiliation: Department of Pathomorphology, Jagiellonian University Medical College, Krakow, Poland.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Bialas
authors.affiliation: Laboratory of Human Molecular Genetics, de Duve Institute, Université Catholique de Louvain, Brussels, Belgium.
authors.forename: Miikka
authors.initials: M
authors.lastname: Vikkula
authors.affiliation: Department of Pathology, Technische Universität Dresden, Dresden, Germany.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Baretton
authors.affiliation: Department of Oncology, University of Turin at San Luigi Hospital, Turin, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Papotti
authors.affiliation: Division of Pathological Anatomy, University of Florence, Florence, Italy.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Nesi
authors.affiliation: 1] Paris-Centre de Recherche Cardiovasculaire (PARCC), Inserm UMR970, Hôpital Européen Georges Pompidou, Paris, France [2] Université Paris Descartes, Faculté de Médecine, Paris Cité Sorbonne, France [3] Service d'Anatomie Pathologique, Hôpital Européen Georges-Pompidou, Assistance Publique Hôpitaux de Paris (AP-HP), Paris, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Badoual
authors.affiliation: Program in Reproductive and Adult Endocrinology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Karel
authors.initials: K
authors.lastname: Pacak
authors.affiliation: Institute of Clinical Chemistry and Laboratory Medicine and Department of Medicine III, University Hospital, Technische Universität Dresden, Dresden, Germany.
authors.forename: Graeme
authors.initials: G
authors.lastname: Eisenhofer
authors.affiliation: Department of Internal Medicine, Section of Endocrinology, Radboud University Medical Centre, Nijmegen, The Netherlands.
authors.forename: Henri J
authors.initials: HJ
authors.lastname: Timmers
authors.affiliation: Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Klinikum der Universität München, Munich, Germany.
authors.forename: Felix
authors.identifier: 0000000178263984
authors.initials: F
authors.lastname: Beuschlein
authors.affiliation: 1] Institut Cochin, Université Paris Descartes, INSERM U1016, CNRS UMR8104, Paris, France [2] Department of Endocrinology, Referral Center for Rare Adrenal Diseases, Assistance Publique Hôpitaux de Paris, Hôpital Cochin, Paris, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Bertherat
authors.affiliation: 1] Endocrinology Unit, Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy [2] Istituto Toscano Tumori (ITT), Florence, Italy.
authors.forename: Massimo
authors.initials: M
authors.lastname: Mannelli
authors.affiliation: 1] Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain [2] Centre for Biomedical Network Research on Rare Diseases (CIBERER), Madrid, Spain.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Robledo
authors.affiliation: 1] Paris-Centre de Recherche Cardiovasculaire (PARCC), Inserm UMR970, Hôpital Européen Georges Pompidou, Paris, France [2] Université Paris Descartes, Faculté de Médecine, Paris Cité Sorbonne, France.
authors.forename: Anne-Paule
authors.initials: AP
authors.lastname: Gimenez-Roqueplo
authors.affiliation: Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: Winand Nm
authors.initials: WN
authors.lastname: Dinjens
authors.affiliation: Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: Esther
authors.initials: E
authors.lastname: Korpershoek
authors.affiliation: 1] Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands [2] Department of Pathology, Reinier de Graaf Hospital, Delft, The Netherlands [3] Department of Pathology, University Medical Center Utrecht, Princess Maxima Center for Pediatric Oncology, Utrecht, The Netherlands.
authors.forename: Ronald R
authors.initials: RR
authors.lastname: de Krijger
mesh_terms: D000310:Adrenal Gland Neoplasms / Q000235:genetics*; D014408:Biomarkers, Tumor / Q000032:analysis / Q000235:genetics; D042963:Electron Transport Complex II / Q000032:analysis* / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry / Q000592:standards*; D008853:Microscopy / Q000379:methods; D009154:Mutation; D015588:Observer Variation; D010235:Paraganglioma / Q000235:genetics*; D010673:Pheochromocytoma / Q000235:genetics*; D013385:Succinate Dehydrogenase / Q000032:analysis* / Q000235:genetics; D019113:Telepathology / Q000379:methods
chemical_list: D014408:Biomarkers, Tumor; D042963:Electron Transport Complex II; C511268:SDHA protein, human; C108095:SDHB protein, human; D013385:Succinate Dehydrogenase