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JAK2 V617F due to a novel TG → CT mutation at nucleotides 1848–1849: diagnostic implication

Leukemia volume 21, pages 1344–1346 (2007)Cite this article

The patient was a 75-year-old Chinese lady diagnosed to have ET since 1995. She was all along put on hydroxyurea (HU) for control of platelet count. Blood counts in 2000 showed hemoglobin (Hb) 14.3 g/dl, white blood cells (WBC) 7.2 × 109/l and platelets 572 × 109/l. Over the recent few months, her platelet count decreased to within normal range so that HU was stopped. However, a gradual increase in WBC followed. Physical examination showed no hepatosplenomegaly. Recent blood counts revealed Hb 12.9 g/dl, WBC 28.9 × 109/l and platelets 430 × 109/l. Peripheral blood film showed a leucoerythroblastic picture associated with tear-drop poikilocytes, basophilia and presence of giant platelets. The lactate dehydrogenase level was raised to 1250 IU/l (reference range: 91–232 IU/l). Bone marrow aspiration yielded a dry tap. Trephine biopsy (Figure 1b) showed a hyperplastic marrow with distortion of normal architecture by underlying stromal fibrosis. Trilineage hemopoiesis was active and there was no increase in blast percentage. Megakaryocytes were markedly increased showing pleomorphism, clustering and presence of hyperlobulated or pyknotic forms. Reticulin fibers were coarsened in a diffuse manner. The diagnosis was post-thrombocythemic myelofibrosis.

High quality-genomic DNA was extracted from unfractionated bone marrow blood and JAK2 V617F mutation was detected by allele-specific PCR (AS-PCR) and PCR restriction fragment length polymorphism (PCR-RFLP) study in accordance with the protocol of Baxter et al.2 For AS-PCR the primers were reverse 5′-CTG AAT AGT CCT ACA GTG TTT TCA GTT TCA-3′, forward (mutant specific) 5′-AGC ATT TGG TTT TAA ATT ATG GAG TAT ATT-3′, and forward (internal control) 5′-ATC TAT AGT CAT GCT GAA AGT AGG AGA AAG-3′. The mutant-specific forward primer contained an intentional mismatch (G → A) at the third nt from the 3′ end (corresponding to nt 1847 of the JAK2 gene) to enhance specificity and should amplify a 203-bp product from the mutated allele, whereas the internal control forward primer should give a 364-bp product from both the mutant and wild-type alleles. For PCR-RFLP, forward primer 5′-GGG TTT CCT CAG AAC GTT GA-3′ and reverse primer 5′-TCA TTG CTT TCC TTT TTC ACA A-3′ were employed to yield a 460-bp product from JAK2 exon 14 sequence, which was then digested with BsaXI. The mutant allele should remain undigested, whereas the wild-type allele should be digested into 241-, 189- and 30-bp fragments.

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Authors and Affiliations

  1. Division of Molecular Pathology, Department of Pathology, Hong Kong Sanatorium & Hospital, Hong Kong, Hong Kong

    C L P Wong, E S K Ma, C L N Wang & H Y Lam

  2. Specialist in Hematology and Hematological Oncology, St Paul's Hospital, Hong Kong, Hong Kong

    S Y Ma

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Correspondence to E S K Ma.

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Wong, C., Ma, E., Wang, C. et al. JAK2 V617F due to a novel TG → CT mutation at nucleotides 1848–1849: diagnostic implication. Leukemia 21, 1344–1346 (2007). https://doi.org/10.1038/sj.leu.2404654

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