Poluben L., Chen S., Yuan X., Shumeiko O., Voznesensky O., Adam M., Fraenkel E., Rasnic R., Linial M., Klymenko S., Balk S. P., Fraenkel P. G

MEDICAL GENETICS

Scentific article

In this study we aimed to identify the spectrum of affected genes in Ukrainian patients diagnosed with primary myelofibrosis (PMF). DNA samples were obtained from peripheral blood leukocytes of 30 Ukrainian PMF patients. Using Whole Exome Sequencing, we detected previously reported and unreported sequence variants considered as pathogenic or potentially pathogenic. Canonical mutations of usual MPN-driver genes were detected in 70% of PMF patients, including JAK2V617F – in 43.3%, MPLW515 – in 10% and CALRmutations (type 1-like and 2-like) – in 16.7% of patients. Also, a non-canonical MPLP222S variant was detected in one patient negative for these mutations. However, in cell culture assay, MPLP222S expression in Ba/F3 cells did not demonstrate differences in phosphorylation of JAK/STAT signaling proteins in response to TPO stimulation compared to MPLWT expression. Overall, more pathogenic and potentially pathogenic sequence variants were found among PMF patients negative for canonical mutations in three driver genes (JAK2, MPL andCALR), compared to patients, positive for one of these mutations. The mean numbers of variants were 5 (range: 4 – 7) versus 3.3 (range: 1 – 9), respectively (p = 0.03). The most frequently affected among Ukrainian PMF patients were genes ASXL1, PEG3, EZH2, ATM, U2AF1, andCDH23in addition to JAK2, MPL andCALR. Pathogenic or potentially pathogenic sequence variants of ASXL1and EZH2genes were identified in 23.3% and 16.7% of cases, respectively. Genes JARID2, RBBP8, RTEL1, SUZ12, BRCA2, LAMB4, NF1, RBM12B, RBM43, and RBP3 were recurrently affected in PMF patients who were also positive for usual mutations in three driver genes. Among PMF patients negative for usual mutations in three driver genes recurrently affected genes were DNMT3Aand TET2in addition to mentioned earlier, and some genetic variants were identified in RTEL1, SUZ12, CBL, CUX1, FLT3LGand UMODL1genes in single cases. Also, we identified several genes affected in single cases (KIT, SF3B1, GNAS) in PMF patients negative for canonical mutations in three driver genes which may be potential MPN drivers.

primary myelofibrosis, mutation, gene.

1. Constantinescu SN, Leroy E, Gryshkova V, Pecquet C, Dusa A. Activating Janus kinase pseudokinase domain mutations in myeloproliferative and other blood cancers. Biochem Soc Trans. 2013;41(4):1048-54. 
2. Mead AJ, Mullally A. Myeloproliferative neoplasm stem cells. Blood. 2017;129(12):1607-16. 
3. Murati A, Brecqueville M, Devillier R, Mozziconacci MJ, Gelsi-Boyer V, Birnbaum D. Myeloid malignancies: mutations, models and management. BMC Cancer. 2012;12:304.
4. Guglielmelli P, Pacilli A, Rotunno G, Rumi E, Rosti V, Delaini F, et al. Presentation and outcome of patients with 2016 WHO diagnosis of prefibrotic and overt primary myelofibrosis. Blood. 2017;129(24):3227-36. 
5. Bartels S, Lehmann U, Büsche G, Schlue J, Mozer M, Stadler J, et al. SRSF2 and U2AF1 mutations in primary myelofibrosis are associated with JAK2 and MPL but not calreticulin mutation and may independently reoccur after allogeneic stem cell transplantation. Leukemia. 2015;29(1):253-5. 
6. Milosevic Feenstra JD, Nivarthi H, Gisslinger H, Leroy E, Rumi E, Chachoua I, et al. Whole-exome sequencing identifies novel MPL and JAK2 mutations in triple-negative myeloproliferative neoplasms. Blood. 2016;127:325-32.
7. Lim K-H, Lin H-C, Chen CG-S, Wang W-T, Chang Y-C, Chiang Y-H, et al. Rapid and sensitive detection of CALR exon 9 mutations using highresolution melting analysis. Clin Chim Acta. 2015;440:133-9. 
8. Rumi E, Cazzola M. Diagnosis, risk stratification, and response evaluation in classical myeloproliferative neoplasms. Blood. 2017;129(6):680-92. 
9. Lundberg P, Karow A, Nienhold R, Looser R, Hao-Shen H, Nissen I, et al. Clonal evolution and clinical correlates of somatic mutations in myeloproliferative neoplasms. Blood. 2014;123(14):2220-8.
10. McCabe MT, Ott HM, Ganji G, Korenchuk S, Thompson C, Van Aller GS, et al. EZH2 inhibition as a therapeutic strategy for lymphoma with EZH2-activating mutations. Nature. 2012;492(7427):108-12. 
11. Tefferi A, Thiele J, Vardiman JW. The 2008 World Health Organization classification system for myeloproliferative neoplasms: order out of chaos. Cancer. 2009;115(17):3842-7.
12. Yogarajah M, Tefferi A. Leukemic Transformation in Myeloproliferative Neoplasms: a Literature Review on Risk, Characteristics, and Outcome. Mayo Clinic Proceedings. 2017;92(7):1118-28.
13. Poluben L, Puligandla M, Neuberg D, Bryke CR, Hsu Y, Shumeiko O, et al. Characteristics of myeloproliferative neoplasms in patients exposed to ionizing radiation following the Chernobyl nuclear accident. Am J Hematol. 2018. DOI: 10.1002/ajh.25307
14. Brecqueville M, Rey J, Bertucci F, Coppin E, Finetti P, Carbuccia N, et al. Mutation analysis of ASXL1, CBL, DNMT3A, IDH1, IDH2, JAK2, MPL, NF1, SF3B1, SUZ12, and TET2 in myeloproliferative neoplasms. Genes Chromosom Cancer. 2012;51(8):743-55. 
15. Nagase R, Inoue D, Pastore A, Fujino T, Hou H-A, Yamasaki N, et al. Expression of mutant Asxl1 perturbs hematopoiesis and promotes susceptibility to leukemic transformation. J Exp Med. 2018;215(6):1729-47.
16. Score J, Hidalgo-Curtis C, Jones AV, Winkelmann N, Skinner A, Ward D, et al. Inactivation of polycomb repressive complex 2 components in myeloproliferative and myelodysplastic/myeloproliferative neoplasms. Blood. 2012;119(5):1208-13. 
17. Martínez-Avilés L, Besses C, Álvarez-Larrán A, Torres E, Serrano S, Bellosillo B. TET2, ASXL1, IDH1, IDH2, and c-CBL genes in JAK2- and MPLnegative myeloproliferative neoplasms. Ann Hematol. 2012;91(4):533-41. 
18. Jara-Acevedo M, Teodosio C, Sanchez-Muñoz L, Álvarez-Twose I, Mayado A, Caldas C, et al. Detection of the KIT D816V mutation in peripheral blood of systemic mastocytosis: diagnostic implications. Mod Pathol. 2015;28(8):1138-49. 
19. Biankin AV, Waddell N, Kassahn KS, Gingras MC, Muthuswamy LB, Johns AL, et al. Pancreatic cancer genomes reveal aberrations in axon guidance pathway genes. Nature. 2012;491(7424):399-405.
20. Sashida G, Wang C, Tomioka T, Oshima M, Aoyama K, Kanai A, et al. The loss of Ezh2 drives the pathogenesis of myelofibrosis and sensitizes tumor-initiating cells to bromodomain inhibition. J Exp Med. 2016;213(8):1459-77.
21. Wan Y, Wu CJ. SF3B1 mutations in chronic lymphocytic leukemia. Blood. 2013;121(23):4627-34. 
22. Xie M, Lu C, Wang J, McLellan MD, Johnson KJ, Wendl MC, et al. Age-related mutations associated with clonal hematopoietic expansion and malignancies. Nat Med. 2014;20(12):1472-8.

«Bulletin of problems biology and medicine» Issue 4 Part 2 (147), 2018 year, 204-208 pages, index UDK 591.156:575.113:616-005.6:616.155.191

10.29254/2077-4214-2018-4-2-147-204-208