Genetic and Epigenetic Alterations of Ph-Negative Myeloproliferative Neoplasms

Objectives and background: Genetic mutations result in abnormalities of myelopoietic proteins and lie in the basis of Ph-negative myeloproliferative neoplasms (MPNs) development and its subsequent progression. Several somatic mutations in JAK2, MPL, TET2, EZH2, ASXL1, CBL, IDH1, IDH2, IKZF1 genes were detected in chronic and blastic phase MPNs. Recent studies have revealed a number of epigenetic alterations that contribute to Ph-negative MPNs pathogenesis and determine the clinical outcome. Mutations involving the EZH2 gene are thought to result in loss of methyltransferase activity suggesting a potential role of tumor suppressor gene silencing as a mechanism in the disease progression. Decrease in ubiquitin ligase activity caused by mutations CBL gene leads to myeloid proliferation. EZH2, CBL mutations are thought to be of prognostic value in MPN’s at the time of transformation to the blastic phase but data are inconsistent and require the further verification.The goal of our research was to determine the significance of mutations genes EZH2, CBL in the diagnosis and prognosis of Ph-negative MPNs.

Methods. We have examined 102 patients with Ph-negative MPNs (45 pts with PV, 30 pts with ET and 27 pts with PMF). For all patients the detection of V617F mutation of JAK2 gene was done. V617F-negative pts with PV and pts with ET or PMF underwent the analysis of mutations in 12-th exon of JAK2 and 515 codone of MPL gene respectively. For 80 pts (30 with PV, 28 with ET and 22 with PMF) cytogenetic analysis and EZH2 mutation status were performed. Identification of CBL mutations was performed in 24 patients with available RNA samples. Mutations in 8, 10, 17, 18, 19 exons of EZH2 and RING-domen of CBL were defined by sequence analysis. V617FJAK2 mutation was detected in 44/45 (97,8%) pts with PV, 16/30 (53,3%) pts with ET and 13/27 (48,1%) pts with PMF. 538-539del-insL in 12-th exon of JAK2 was found in 1/45 (2,22%) patient with PV. W515KMPL mutation was identified in 1/30 (3,33%) pt with ET and 1/27 (3,7%) pt with PMF. 2 mutations of EZH2 gene have been found in 2 individuals with PMF (2/22). Both mutations are located in the 19 exon. The Ile713Thr mutation was detected in the patient with a del(6)(q15) karyotype which is associated with an intermediate cytogenetics risk. This patient subsequently underwent transformation from PMF to myelodysplastic syndrome in 9 months after the disease onset. Another case of mutation harboring (Thr731Asp) was detected in a patient with PMF and poor prognosis karyotype (chromosome 7 monosomy). This patient had transformation PMF to acute myeloid leukemia and died after 20 months. Homozygous mutation Q420R in CBL gene was detected in 1/24 patient with complex karyotype. Disease progression was observed after 16 months from the diagnosis.

Conclusion. Mutations in EZH2 and CBL genes could be assessed as additional prognostic markers of unfavourable prognosis in patients with BCR-ABL-negative MPNs with different chromosomal aberrations. The integration of cytogenetic and molecular analyses could be a valuable option for stratification of patients and optimising the treatment strategy.

References: Tefferi A. Novel mutations and their functional and clinical relevance in myeloproliferative neoplasms: JAK2, MPL, TET2, ASXL1, CBL, IDH and IKZF1. Leukemia 2010; 24:1128–1138.