Whole-exome sequencing identifies novel MPL and JAK2 mutations in triple-negative myeloproliferative neoplasms

Jelena D. Milosevic Feenstra, Harini Nivarthi, Heinz Gisslinger, Emilie Leroy, Elisa Rumi, Ilyas Chachoua, Klaudia Bagienski, Blanka Kubesova, Daniela Pietra, Bettina Gisslinger, Chiara Milanesi, Roland Jäger, Doris Chen, Tiina Berg, Martin Schalling, Michael Schuster, Christoph Bock, Stefan N. Constantinescu, Mario Cazzola and Robert Kralovics

Key Points

  • Activating mutations outside exon 10 of MPL were identified in 10% (7 of 69) of triple-negative cases of ET and PMF.

  • JAK2-V625F and JAK2-F556V were identified in 2 triple-negative cases of ET and were shown to activate JAK-STAT5 signaling.

Publisher's Note: There is an Inside Blood Commentary on this article in this issue.


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.

  • Submitted July 31, 2015.
  • Accepted September 13, 2015.
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