Int J Hematol (2008) 88:243–245 DOI 10.1007/s12185-008-0131-2
LETTER TO THE EDITOR
Simultaneous detection of JAK2 V617F mutation and Bcr-Abl translocation in a patient with chronic myelogenous leukemia Monika Ribeiro de Mello Conchon Æ Juliana Lima Costa Æ Mafalda Megumi Yoshinaga Novaes Æ Pedro Enrique Dorlhiac-Llacer Æ Dalton de Alencar Fischer Chamone Æ Israel Bendit
Received: 11 January 2008 / Revised: 7 May 2008 / Accepted: 10 June 2008 / Published online: 16 July 2008 ! The Japanese Society of Hematology 2008
Myelofibrosis and chronic myeloid leukemia (CML) are myeloproliferative disorders (MPD) of hematopoietic stem cell and characterized by clonal neoplastic proliferation of one or more of the myeloid lineage in the bone marrow. The reciprocal translocation t (9; 22) of the bcr-abl fusion gene is a key genetic marker for CML. Until recently, a point mutation at codon 617 (V617F) of the Janus kinase 2 gene (JAK2) was considered absent in CML patients with Philadelphia positive chromosome (Ph+) [1]. However, current findings have identified the coexistence of both genetic defects in patients with MPD [2, 3]. Here we present a myelofibrosis patient with CML in whom a coexistence of JAK2 V617F mutation was detectable at the initial diagnosis and despite the excessive accumulation of CML clone. In June 2007, a 52-year-old white female presented with pronounced splenomegaly. At admission, she complained of 15 kg sudden weight loss. Initial laboratory tests showed hemoglobin 100 g/L, platelets 689 9 109/L and white cell count 193 9 109/L with a differential revealing 3% myeloblasts, 9% promyelocytes, 8% myelocytes, 3% metamyelocytes, 10% band cells, 63% neutrophils, 7% eosinophils, 8% basophils, 2% lymphocytes and 7%
M. R. M. Conchon ! J. L. Costa ! M. M. Y. Novaes ! P. E. Dorlhiac-Llacer ! D. de Alencar Fischer Chamone ! I. Bendit Laboratorio de Biologia Tumoral da Disciplina de Hematologia da Faculdade de Medicina da Universidade de Sa˜o Paulo, Sao Paulo, Brazil I. Bendit (&) Department of the Tumor Biology Laboratory, Av. Dr. Eneias de Carvalho Aguiar, 155-18 andar, Cerqueira Ce´sar, Sao Paulo 05403-000, Brazil e-mail:
[email protected]
monocytes. A bone marrow biopsy disclosed megakaryocytic hyperplasia and marked marrow reticulin fibrosis (grade 3). Based on this result, a search for JAK2 V617F mutation by restriction and DNA sequence analysis in a blood sample taken at that time confirmed the presence of the homozygote mutation (Fig. 1b). Cytogenetic analysis revealed 100% Ph+ in each of 20 metaphase cells examined (Fig. 2). This was further confirmed by quantitative RT-PCR technique, which showed 130% of BCR-ABL/ BCR mRNA b2a2 transcript. The patient was initially given hydroxyurea (25 mg/kg/day) which permitted a rapid reduction of leucocytes and platelets to 37 9 109 and 398 9 109/L, respectively. As a result, hydroxyurea was substituted with imatinib at a dose of 400 mg/day and was followed. Fifteen days after initiation of imatinib, the patient achieved complete hematological response. By the end of 30 days, the leucocytes count had remained normal, but platelets count had a marked increase and rose steadily for 4–6 weeks during treatment until reached a value of 936 9 109/L. Consequently, we aimed to evaluate the molecular events underlying imatinib effect by quantifying BCR-ABL/BCR transcript. The results showed a 2-log decrease (3.7%) in mRNA transcript and suggested a gradual reduction of residual leukemic burden. Given this, the patient was placed on combinational therapy with hydroxyurea. His platelet count returned to normal levels within 10 days after treatment. The DNA sequence from blood sample collected at that time confirmed the existence of JAK2 V617F mutation and indicated approximately 50% reduction of the T peak at position 1849 (Fig. 1c). At the latest follow-up in December 2007, she remained well and evaluated as being in cytological remission. It has been suggested that the perturbation of JAK2 signaling by gain of function somatic V617F mutation of the JAK2 gene leads to dysregulation of kinase activity
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Fig. 1 JAK2 V617F mutation by DNA sequence analysis. a Wild type allele. b Sequence from sample obtained at the time of diagnosis. c Sequence of sample obtained 10 days after initiation of combinational therapy
which results in spontaneous expansion of erythroid progenitor cells [4]. This mutation has been found to account for 50% of patients with myelofibrosis, but its role on marrow fibrosis remains obscure. To the best of our knowledge, three myelofibrosis cases of Ph + CML with JAK2 V617F mutation have been published. [2, 3, 5]. In these studies, the mutation was initially detected after suppression of the Ph + CML clone by imatinib. The retrospective analysis of the pretherapeutic sample from the patient reported by Huseein et al. [3] confirmed the
M. R. M. Conchon et al.
heterozygous JAK2 V617F clone in 5% of alleles. In our case, myelofibrosis was established before initiation of imatinib and a coexistence of JAK2 V617F mutation were detectable at the first visit in 100% of the cells. The occurrence of JAK2 V617F allele burden was reflected by elevated levels of platelets even before complete suppression of the CML clone. Recent studies have suggested two possibilities to account for expression of BCR-ABL and JAK2 V617F anomalies in a single patient. The first possibility is that there are two clones each having BCR-ABL and JAK2 V617F mutation [3, 6]. Another possibility is that a single clone concurrently possesses both BCR-ABL and JAK2 V617F. As for this mechanism, Kramer et al. [2] postulated that BCR-ABL mutation had occurred in a preexisting JAK2 V617F-positive clone. In addition, Bocchia et al. [7] recently reported a case with polycythemia vera, which transformed into the CML phenotype. In this patient, they confirmed that JAK2 V617F and BCR-ABL were concurrently detected at a single cell level. Moreover, they found that, although BCR-ABL was not detected during a major molecular response induced by imatinib, JAK2 V617F was still detected in the peripheral blood of this patient. This result again suggests that a pre-existing JAK2 V617F-positive clone acquired BCR-ABL translocation at a certain point. In our case, JAK2 V617F positive cells were detected in 100% of the cells before the treatment with imatinib. Then, although imatinib was effective in drastically reducing BCR-ABL-positive cells, a marked increase of platelet was observed during this period, to which hydroxyurea was effective. This result together with the constant detection of mutant allele even during the combinational treatment with imatinib and hydroxyurea, suggested that the remaining JAK2 V617F-positive cells caused this thrombocytosis, and that the BCR-ABL
Fig. 2 The G-banded karyotype was determined as 46XX, t(9;22) (q34;q11)[20] . Arrows indicate chromosomes involved in translocation
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JAK2 V617F mutation in Ph + CML patient
mutation may be a second leukemogenic event that occurred in the JAK2 V617F-positive clone. Together with previous reports, we suggest that screening for JAK2 V617F mutation should be considered in any Ph + CML patients with chronic MPD.
References 1. Scott LM, Campbell PJ, Baxter EJ, et al. The V617F JAK2 mutation is uncommon in cancers and in myeloid malignancies other than the classic myeloproliferative disorders. Blood. 2005;106:2920–1. 2. Kramer A, Reiter A, Kruth J, et al. JAK2–V617F mutation in a patient with Philadelphia-chromosome-positive chronic myeloid leukaemia. Lancet Oncol. 2007;8:658–60.
245 3. Hussein K, Bock O, Seegers A, Flasshove M, et al. Myelofibrosis evolving during imatinib treatment of a chronic myeloproliferative disease with coexisting BCR-ABL translocation and JAK2V617F mutation. Blood. 2007;109:4106–7. 4. Baxter EJ, Scott LM, Campbell PJ, et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet. 2005;365:1054–61. 5. Bornhauser M, Mohr B, Oelschlaegel U, et al. Concurrent JAK2 (V617F) mutation and BCR-ABL translocation within committed myeloid progenitors in myelofibrosis. Leukemia. 2007;21:1824–6. 6. Busche G, Hussein K, Bock O, Kreipe H. Insights into JAK2– V617F mutation in CML. Lancet Oncol. 2007;8:863–4. 7. Bocchia M, Vannucchi AM, Gozzetti A, et al. Insights into JAK2– V617F mutation in CML. Lancet Oncol. 2007;8:864–6.
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