Calreticulin mutation survey by high resolution melting method associated with unique presentations in essential thrombocythemic patients CALR mutation survey by HRM in ET patients
Main Article Content
Keywords
calreticulin, essential thrombocythemia, high resolution melting, JAK2, mutation
Abstract
Somatic mutations of exon 9 of calreticulin gene (CALR) were diagnosis and prognosis importance found in patients with JAK2V617F-negative essential thrombocythemia (ET). We survey CALR and JAK2 mutations in our ET patients and study the relationship between mutations and clinical presentations.
A total of 60 ET patients were enrolled in the study, and CALR mutations were studied by high resolution melting (HRM) methods and sequencing in JAK2V617F-negative group retrospectively. Clinical manifestations were reviewed retrospectively from chart records.
Twenty-one CALR mutations showed eight types of specific melting curves detected by the HRM method and sequencing validation among 26 JAK2 V617F-negative patients. Compared with JAK2 mutations, patients with CALR mutations were younger and had a higher platelet count, lower white cell counts, and lower hemoglobin levels significantly (p<0.05).
From our study, HRM methods revealed unique curve types in screening for CALR mutations screening even for complicated mutations. The mutations can be identification rapidly, and cost-effectively by HRM method than other tools. The clinical presentations of CALR mutations from JAK2 mutations showed significant differences and should be checked in ET patients.
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References
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22. Gadomska G, Bartoszewska-Kubiak A, Boinska J, Matiakowska K, Ziolkowska K, Haus O, Rosc D. Selected Parameters of Angiogenesis and the JAK2, CALR, and MPL Mutations in Patients With Essential Thrombocythemia. Clin Appl Thromb Hemost. 2018;24:1056-1060.
23. Zini R, Guglielmelli P, Pietra D, Rumi E, Rossi C, Rontauroli S, Genovese E, Fanelli T, Calabresi L, Bianchi E, Salati S, Cazzola M, Tagliafico E, Vannucchi AM, Manfredini R, investigators A. CALR mutational status identifies different disease subtypes of essential thrombocythemia showing distinct expression profiles. Blood Cancer J. 2017;7:638.
24. Giannopoulos A, Rougkala N, Loupis T, Mantzourani M, Viniou NA, Variami E, Vassilakopoulos TP, Dryllis G, Kotsianidis I, Gougopoulou T, Politou M, Konstantopoulos K, Vassilopoulos G. Detection of CALR Mutations Using High Resolution Melting Curve Analysis (HRM-A); Application on a Large Cohort of Greek ET and MF Patients. Mediterr J Hematol Infect Dis. 2019;11:e2019009.
2. Mambet C, Babosova O, Defour JP, Leroy E, Necula L, Stanca O, Tatic A, Berbec N, Coriu D, Belickova M, Kralova B, Lanikova L, Vesela J, Pecquet C, Saussoy P, Havelange V, Diaconu CC, Divoky V, Constantinescu SN. Cooccurring JAK2 V617F and R1063H mutations increase JAK2 signaling and neutrophilia in myeloproliferative neoplasms. Blood. 2018;132:2695-2699.
3. Cazzola M, Kralovics R. From Janus kinase 2 to calreticulin: the clinically relevant genomic landscape of myeloproliferative neoplasms. Blood. 2014;123:3714-3719.
4. Spanoudakis E, Papoutselis M, Bazdiara I, Lamprianidi E, Kordella X, Tilkeridis C, Tsatalas C, Kotsianidis I. The JAK2V617F Point Mutation Increases the Osteoclast Forming Ability of Monocytes in Patients with Chronic Myeloproliferative Neoplasms and Makes their Osteoclasts more Susceptible to JAK2 Inhibition. Mediterr J Hematol Infect Dis. 2018;10:e2018058.
5. Xia D, Hasserjian RP. Molecular testing for JAK2, MPL, and CALR in myeloproliferative neoplasms. Am J Hematol. 2016;91:1277-1280.
6. Lee E, Lee KJ, Park H, Chung JY, Lee MN, Chang MH, Yoo J, Lee H, Kong SY, Eom HS. Clinical Implications of Quantitative JAK2 V617F Analysis using Droplet Digital PCR in Myeloproliferative Neoplasms. Ann Lab Med. 2018;38:147-154.
7. Ojeda MJ, Bragos IM, Calvo KL, Williams GM, Carbonell MM, Pratti AF. CALR, JAK2 and MPL mutation status in Argentinean patients with BCR-ABL1- negative myeloproliferative neoplasms. Hematology. 2018;23:208-211.
8. Nangalia J, Massie CE, Baxter EJ, Nice FL, Gundem G, Wedge DC, Avezov E, Li J, Kollmann K, Kent DG, Aziz A, Godfrey AL, Hinton J, Martincorena I, Van Loo P, Jones AV, Guglielmelli P, Tarpey P, Harding HP, Fitzpatrick JD, Goudie CT, Ortmann CA, Loughran SJ, Raine K, Jones DR, Butler AP, Teague JW, O'Meara S, McLaren S, Bianchi M, Silber Y, Dimitropoulou D, Bloxham D, Mudie L, Maddison M, Robinson B, Keohane C, Maclean C, Hill K, Orchard K, Tauro S, Du MQ, Greaves M, Bowen D, Huntly BJP, Harrison CN, Cross NCP, Ron D, Vannucchi AM, Papaemmanuil E, Campbell PJ, Green AR. Somatic CALR mutations in myeloproliferative neoplasms with nonmutated JAK2. N Engl J Med. 2013;369:2391-2405.
9. Clinton A, McMullin MF. The Calreticulin gene and myeloproliferative neoplasms. J Clin Pathol. 2016;69:841-845.
10. Zhang X, Pan J. Somatic mutations of calreticulin in patients with myelodysplastic/myeloproliferative neoplasms-unclassifiable. Ann Hematol. 2017;96:1949-1951.
11. Misawa K, Yasuda H, Araki M, Ochiai T, Morishita S, Shirane S, Edahiro Y, Gotoh A, Ohsaka A, Komatsu N. Mutational subtypes of JAK2 and CALR correlate with different clinical features in Japanese patients with myeloproliferative neoplasms. Int J Hematol. 2018;107:673-680.
12. Lundberg P, Karow A, Nienhold R, Looser R, Hao-Shen H, Nissen I, Girsberger S, Lehmann T, Passweg J, Stern M, Beisel C, Kralovics R, Skoda RC. Clonal evolution and clinical correlates of somatic mutations in myeloproliferative neoplasms. Blood. 2014;123:2220-2228.
13. Pronier E, Cifani P, Merlinsky TR, Berman KB, Somasundara AVH, Rampal RK, LaCava J, Wei KE, Pastore F, Maag JL, Park J, Koche R, Kentsis A, Levine RL. Targeting the CALR interactome in myeloproliferative neoplasms. JCI Insight. 2018;3.
14. Keaney T, O'Connor L, Krawczyk J, Abdelrahman MA, Hayat AH, Murray M, O'Dwyer M, Percy M, Langabeer S, Haslam K, Glynn B, Mullen C, Keady E, Lahiff S, Smith TJ. A novel molecular assay using hybridisation probes and melt curve analysis for CALR exon 9 mutation detection in myeloproliferative neoplasms. J Clin Pathol. 2017;70:662-668.
15. Didone A, Nardinelli L, Marchiani M, Ruiz ARL, de Lima Costa AL, Lima IS, Santos NM, Sanabani SS, Bendit I. Comparative study of different methodologies to detect the JAK2 V617F mutation in chronic BCR-ABL1 negative myeloproliferative neoplasms. Pract Lab Med. 2016;4:30-37.
16. Leszczynska A, Grzenkowicz-Wydra J, Chmielewska-Gorycka L, Bieniaszewska M, Hellmann A. Detection of JAK2 Exon 12 Mutations in JAK2 V617F-Negative Polycythemia Vera Patients by Cloning Technique. Acta Haematol. 2016;136:123-128.
17. Chi J, Nicolaou KA, Nicolaidou V, Koumas L, Mitsidou A, Pierides C, Manoloukos M, Barbouti K, Melanthiou F, Prokopiou C, Vassiliou GS, Costeas P. Calreticulin gene exon 9 frameshift mutations in patients with thrombocytosis. Leukemia. 2014;28:1152-1154.
18. Bilbao-Sieyro C, Santana G, Moreno M, Torres L, Santana-Lopez G, Rodriguez-Medina C, Perera M, Bellosillo B, de la Iglesia S, Molero T, Gomez-Casares MT. High resolution melting analysis: a rapid and accurate method to detect CALR mutations. PLoS One. 2014;9:e103511.
19. Pavlov I, Hadjiev E, Alaikov T, Spassova S, Stoimenov A, Naumova E, Shivarov V, Ivanova M. Calreticulin Mutations in Bulgarian MPN Patients. Pathol Oncol Res. 2018;24:171-174.
20. Hsiao HH, Yang MY, Liu YC, Lee CP, Yang WC, Liu TC, Chang CS, Lin SF. The association of JAK2V617F mutation and leukocytosis with thrombotic events in essential thrombocythemia. Exp Hematol. 2007;35:1704-1707.
21. Rumi E, Pietra D, Ferretti V, Klampfl T, Harutyunyan AS, Milosevic JD, Them NC, Berg T, Elena C, Casetti IC, Milanesi C, Sant'antonio E, Bellini M, Fugazza E, Renna MC, Boveri E, Astori C, Pascutto C, Kralovics R, Cazzola M, Associazione Italiana per la Ricerca sul Cancro Gruppo Italiano Malattie Mieloproliferative I. JAK2 or CALR mutation status defines subtypes of essential thrombocythemia with substantially different clinical course and outcomes. Blood. 2014;123:1544-1551.
22. Gadomska G, Bartoszewska-Kubiak A, Boinska J, Matiakowska K, Ziolkowska K, Haus O, Rosc D. Selected Parameters of Angiogenesis and the JAK2, CALR, and MPL Mutations in Patients With Essential Thrombocythemia. Clin Appl Thromb Hemost. 2018;24:1056-1060.
23. Zini R, Guglielmelli P, Pietra D, Rumi E, Rossi C, Rontauroli S, Genovese E, Fanelli T, Calabresi L, Bianchi E, Salati S, Cazzola M, Tagliafico E, Vannucchi AM, Manfredini R, investigators A. CALR mutational status identifies different disease subtypes of essential thrombocythemia showing distinct expression profiles. Blood Cancer J. 2017;7:638.
24. Giannopoulos A, Rougkala N, Loupis T, Mantzourani M, Viniou NA, Variami E, Vassilakopoulos TP, Dryllis G, Kotsianidis I, Gougopoulou T, Politou M, Konstantopoulos K, Vassilopoulos G. Detection of CALR Mutations Using High Resolution Melting Curve Analysis (HRM-A); Application on a Large Cohort of Greek ET and MF Patients. Mediterr J Hematol Infect Dis. 2019;11:e2019009.