title: MPLW515L is a novel somatic activating mutation in myelofibrosis with myeloid metaplasia.
abstract: BACKGROUND
  The JAK2V617F allele has recently been identified in patients with polycythemia vera (PV), essential thrombocytosis (ET), and myelofibrosis with myeloid metaplasia (MF). Subsequent analysis has shown that constitutive activation of the JAK-STAT signal transduction pathway is an important pathogenetic event in these patients, and that enzymatic inhibition of JAK2V617F may be of therapeutic benefit in this context. However, a significant proportion of patients with ET or MF are JAK2V617F-negative. We hypothesized that activation of the JAK-STAT pathway might also occur as a consequence of activating mutations in certain hematopoietic-specific cytokine receptors, including the erythropoietin receptor (EPOR), the thrombopoietin receptor (MPL), or the granulocyte-colony stimulating factor receptor (GCSFR).
  
  
  RESULTS
  DNA sequence analysis of the exons encoding the transmembrane and juxtamembrane domains of EPOR, MPL, and GCSFR, and comparison with germline DNA derived from buccal swabs, identified a somatic activating mutation in the transmembrane domain of MPL (W515L) in 9% (4/45) of JAKV617F-negative MF. Expression of MPLW515L in 32D, UT7, or Ba/F3 cells conferred cytokine-independent growth and thrombopoietin hypersensitivity, and resulted in constitutive phosphorylation of JAK2, STAT3, STAT5, AKT, and ERK. Furthermore, a small molecule JAK kinase inhibitor inhibited MPLW515L-mediated proliferation and JAK-STAT signaling in vitro. In a murine bone marrow transplant assay, expression of MPLW515L, but not wild-type MPL, resulted in a fully penetrant myeloproliferative disorder characterized by marked thrombocytosis (Plt count 1.9-4.0 x 10(12)/L), marked splenomegaly due to extramedullary hematopoiesis, and increased reticulin fibrosis.
  
  
  CONCLUSIONS
  Activation of JAK-STAT signaling via MPLW515L is an important pathogenetic event in patients with JAK2V617F-negative MF. The bone marrow transplant model of MPLW515L-mediated myeloproliferative disorders (MPD) exhibits certain features of human MF, including extramedullary hematopoiesis, splenomegaly, and megakaryocytic proliferation. Further analysis of positive and negative regulators of the JAK-STAT pathway is warranted in JAK2V617F-negative MPD.
authors.affiliation: Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Yana
authors.initials: Y
authors.lastname: Pikman
authors.forename: Benjamin H
authors.initials: BH
authors.lastname: Lee
authors.forename: Thomas
authors.initials: T
authors.lastname: Mercher
authors.forename: Elizabeth
authors.initials: E
authors.lastname: McDowell
authors.forename: Benjamin L
authors.initials: BL
authors.lastname: Ebert
authors.forename: Maricel
authors.initials: M
authors.lastname: Gozo
authors.forename: Adam
authors.initials: A
authors.lastname: Cuker
authors.forename: Gerlinde
authors.initials: G
authors.lastname: Wernig
authors.forename: Sandra
authors.initials: S
authors.lastname: Moore
authors.forename: Ilene
authors.initials: I
authors.lastname: Galinsky
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: DeAngelo
authors.forename: Jennifer J
authors.initials: JJ
authors.lastname: Clark
authors.forename: Stephanie J
authors.initials: SJ
authors.lastname: Lee
authors.forename: Todd R
authors.initials: TR
authors.lastname: Golub
authors.forename: Martha
authors.initials: M
authors.lastname: Wadleigh
authors.forename: D Gary
authors.initials: DG
authors.lastname: Gilliland
authors.forename: Ross L
authors.initials: RL
authors.lastname: Levine
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects / Q000473:pathology; D016026:Bone Marrow Transplantation; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects / Q000473:pathology; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000494:pharmacology; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D019337:Hematologic Neoplasms / Q000188:drug therapy / Q000503:physiopathology; D006410:Hematopoiesis / Q000235:genetics / Q000502:physiology; D006801:Humans; D053612:Janus Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D008533:Megakaryocytes / Q000187:drug effects / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D020125:Mutation, Missense; D022423:Myeloid Cells / Q000187:drug effects / Q000473:pathology; D009196:Myeloproliferative Disorders / Q000209:etiology / Q000235:genetics / Q000473:pathology; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000502:physiology; D010766:Phosphorylation / Q000187:drug effects; D017354:Point Mutation; D055728:Primary Myelofibrosis / Q000235:genetics* / Q000473:pathology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology / Q000627:therapeutic use; D011499:Protein Processing, Post-Translational / Q000187:drug effects / Q000235:genetics; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000235:genetics / Q000502:physiology; D018121:Receptors, Cytokine / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000009:adverse effects; D050791:STAT Transcription Factors / Q000502:physiology; D017422:Sequence Analysis, DNA; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D013154:Spleen / Q000473:pathology; D013922:Thrombocytosis / Q000209:etiology / Q000235:genetics / Q000473:pathology; D014158:Transcription, Genetic; D039221:mRNA Cleavage and Polyadenylation Factors / Q000235:genetics / Q000502:physiology
chemical_list: D016207:Cytokines; D015514:Oncogene Proteins, Fusion; D047428:Protein Kinase Inhibitors; D018121:Receptors, Cytokine; D011993:Recombinant Fusion Proteins; D050791:STAT Transcription Factors; D039221:mRNA Cleavage and Polyadenylation Factors; C479819:FIP1L1-PDGFRA fusion protein, human; D020796:Receptor, Platelet-Derived Growth Factor alpha; D053612:Janus Kinases