title: Whole-exome sequencing identifies novel MPL and JAK2 mutations in triple-negative myeloproliferative neoplasms.
abstract: Essential thrombocythemia (ET) and primary myelofibrosis (PMF) are chronic diseases characterized by clonal hematopoiesis and hyperproliferation of terminally differentiated myeloid cells. The disease is driven by somatic mutations in exon 9 of CALR or exon 10 of MPL or JAK2-V617F in >90% of the cases, whereas the remaining cases are termed "triple negative." We aimed to identify the disease-causing mutations in the triple-negative cases of ET and PMF by applying whole-exome sequencing (WES) on paired tumor and control samples from 8 patients. We found evidence of clonal hematopoiesis in 5 of 8 studied cases based on clonality analysis and presence of somatic genetic aberrations. WES identified somatic mutations in 3 of 8 cases. We did not detect any novel recurrent somatic mutations. In 3 patients with clonal hematopoiesis analyzed by WES, we identified a somatic MPL-S204P, a germline MPL-V285E mutation, and a germline JAK2-G571S variant. We performed Sanger sequencing of the entire coding region of MPL in 62, and of JAK2 in 49 additional triple-negative cases of ET or PMF. New somatic (T119I, S204F, E230G, Y591D) and 1 germline (R321W) MPL mutation were detected. All of the identified MPL mutations were gain-of-function when analyzed in functional assays. JAK2 variants were identified in 5 of 57 triple-negative cases analyzed by WES and Sanger sequencing combined. We could demonstrate that JAK2-V625F and JAK2-F556V are gain-of-function mutations. Our results suggest that triple-negative cases of ET and PMF do not represent a homogenous disease entity. Cases with polyclonal hematopoiesis might represent hereditary disorders.
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Jelena D
authors.identifier: http://orcid.org/0000-0001-7420-3298
authors.initials: JD
authors.lastname: Milosevic Feenstra
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Harini
authors.initials: H
authors.lastname: Nivarthi
authors.affiliation: Department of Internal Medicine I, Division of Hematology and Blood Coagulation, Medical University of Vienna, Vienna, Austria;
authors.forename: Heinz
authors.identifier: http://orcid.org/0000-0002-0295-7629
authors.initials: H
authors.lastname: Gisslinger
authors.affiliation: Ludwig Institute for Cancer Research, Brussels, Belgium; de Duve Institute, Université Catholique de Louvain, Brussels, Belgium;
authors.forename: Emilie
authors.initials: E
authors.lastname: Leroy
authors.affiliation: Department of Hematology Oncology, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Policlinico San Matteo, Pavia, Italy;
authors.forename: Elisa
authors.initials: E
authors.lastname: Rumi
authors.affiliation: Ludwig Institute for Cancer Research, Brussels, Belgium; de Duve Institute, Université Catholique de Louvain, Brussels, Belgium;
authors.forename: Ilyas
authors.initials: I
authors.lastname: Chachoua
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Klaudia
authors.initials: K
authors.lastname: Bagienski
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria; Center of Molecular Biology and Gene Therapy, Department of Internal Medicine-Hematology and Oncology, University Hospital Brno and Medical Faculty Masaryk University, Brno, Czech Republic; and.
authors.forename: Blanka
authors.initials: B
authors.lastname: Kubesova
authors.affiliation: Department of Hematology Oncology, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Policlinico San Matteo, Pavia, Italy;
authors.forename: Daniela
authors.initials: D
authors.lastname: Pietra
authors.affiliation: Department of Internal Medicine I, Division of Hematology and Blood Coagulation, Medical University of Vienna, Vienna, Austria;
authors.forename: Bettina
authors.initials: B
authors.lastname: Gisslinger
authors.affiliation: Department of Hematology Oncology, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Policlinico San Matteo, Pavia, Italy;
authors.forename: Chiara
authors.initials: C
authors.lastname: Milanesi
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Roland
authors.initials: R
authors.lastname: Jäger
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Doris
authors.initials: D
authors.lastname: Chen
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Tiina
authors.initials: T
authors.lastname: Berg
authors.affiliation: Department of Internal Medicine I, Division of Hematology and Blood Coagulation, Medical University of Vienna, Vienna, Austria;
authors.forename: Martin
authors.initials: M
authors.lastname: Schalling
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Michael
authors.initials: M
authors.lastname: Schuster
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria;
authors.forename: Christoph
authors.initials: C
authors.lastname: Bock
authors.affiliation: Ludwig Institute for Cancer Research, Brussels, Belgium; de Duve Institute, Université Catholique de Louvain, Brussels, Belgium;
authors.forename: Stefan N
authors.initials: SN
authors.lastname: Constantinescu
authors.affiliation: Department of Hematology Oncology, Fondazione Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Policlinico San Matteo, Pavia, Italy; Department of Molecular Medicine, University of Pavia, Pavia, Italy.
authors.forename: Mario
authors.identifier: http://orcid.org/0000-0001-6984-8817
authors.initials: M
authors.lastname: Cazzola
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria; Department of Internal Medicine I, Division of Hematology and Blood Coagulation, Medical University of Vienna, Vienna, Austria;
authors.forename: Robert
authors.initials: R
authors.lastname: Kralovics
mesh_terms: D056915:DNA Copy Number Variations; D004252:DNA Mutational Analysis; D059472:Exome; D023281:Genomics / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D053614:Janus Kinase 2 / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D009196:Myeloproliferative Disorders / Q000235:genetics* / Q000378:metabolism; D055728:Primary Myelofibrosis / Q000235:genetics / Q000378:metabolism; D053628:Receptors, Thrombopoietin / Q000235:genetics* / Q000378:metabolism; D013920:Thrombocythemia, Essential / Q000235:genetics / Q000378:metabolism; D049951:X Chromosome Inactivation / Q000235:genetics
chemical_list: D053628:Receptors, Thrombopoietin; C082218:MPL protein, human; D053614:Janus Kinase 2