FISH Studies Provide Evidence That JAK 2, Via a Myriad of Mechanisms Not Limited to Point Mutation, Has Gain of Function in Polycythemia Vera (PV), and Myelodysplasia (MDS).
Authors: Najfeld Vesna , Berkofsky-Fessler Windy , Cozza Amanda 
Published: October 2019

Abstract
The constitutively active JAK2V617F somatic point mutation is a hallmark discovery in understanding the molecular pathogenesis of Ph- chronic myeloproliferative disorders (cMPDs). It is reported as a recurrent genomic lesion occurring in up to 97% of patients (pts) with PV. It is also found in 50–70% of pts with essential thrombocythemia (ET) and 30–50% of pts with idiopathic myelofibrosis (IMF). Thus, a significant proportion of ET and IMF pts are JAK2V617F negative. However, in addition to JAK2-negative PV pts they can have recurrent chromosomal abnormalities (Vizmanos et al Leukemia 20: 534, 2006). Moreover, JAK2 has been shown to form fusion hybrid proteins with three different genes: TEL/ETV6, BCR and PCM1 in cMPDs and lymphoid neoplasms. We therefore hypothesized that pts showing 9p24 chromosomal abnormalities have additional JAK2 structural rearrangements. Among 10,000 pts with hematological malignancies we detected 15 pts (MDS=5, AML=3, NHL=5, IMF=1, MM=1) with 9p24 chromosomal rearrangements. Additionally, 23 pts: 18 with PV [10 with a normal karyotype, 4 with +9, 2 with +i(9)(p10), 1 with der(9)t(1;9), and one with del(9)(p21)] and 5 pts with IMF showing del(13q) and del(20)(q12), were also included in the study. Two JAK2 BAC FISH probes: RP11-3H3 and RP11-28A9 (provided by N. Cross, Salisbury, England) did not show structural rearrangements in 2,000 nuclei from 4 controls and mapped to the 9p24 region in 40 metaphase cells, confirming 100% specificity for the probe. A NF-E2 BAC probe (provided by H. Pahl, Freiburg, Germany) did not show rearrangements in 1,000 nuclei and mapped to the 12q13.1–13.2 region in 20 metaphase cells. Among 38 evaluated pts, FISH studies showed gain of JAK2 in 13 pts (34%): 3 to 7 copies of JAK2 were observed in 10 pts (26%) and amplification of JAK2 was demonstrated in 3 pts (8%): 2 with PV and 1 with B-cell lymphoma. Two pts with PV had two JAK2 clonal populations: gain of 4 copies of JAK2 as well as JAK2 amplification (~20 copies). The original karyotype of B-cell lymphoma pt was t(2;9)(q14;p13) but metaphase FISH revealed the complete material attached to 9p was highlighted by the JAK2 probe. Five pts demonstrated JAK2 structural rearrangements. Three MDS pts showed JAK2 translocation from 9p24 to chromosome 12: 1 pt had JAK2-NF-E2 fusion, 1pt had a JAK2-TEL/ETV6 fusion and in the 3rd pt JAK2 moved from 9p24 to 12q13, where NF-E2 was located. A pediatric MDS pt and der(9)inv(9)(p23q34)t(9;14)(q22;q13) karyotype demonstrated JAK2 translocation to chromosome 14 within close proximity to IgH locus. In the 5th pt (AML) JAK2 moved to chromosome 4, band region q27. Our observations demonstrated that numerical gain of JAK2 is present in 6 of 18 pts with PV (33%). Moreover, rare amplification of JAK2 is a recurrent phenomenon in PV (11%) and also rarely occur in other hematological malignancies. The novel observation of JAK2-NF-E2 fusion as well as TEL/ETV6-JAK2 fusion and JAK2 translocation to chromosomes 4, 12 and 14 demonstrate that JAK2 not only has multiple fusion partners but also indicate its important role in the molecular pathogenesis of cMPDs, MDS and rare B-lymphoid malignancies. These findings also support the notion that JAK2V617F mutation may not be the only molecular JAK2 genomic defect in cMPDs.