Elsevier

The Lancet

Volume 365, Issue 9464, 19–25 March 2005, Pages 1054-1061
The Lancet

Fast track — Mechanisms of Disease
Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders

https://doi.org/10.1016/S0140-6736(05)71142-9Get rights and content

Summary

Background

Human myeloproliferative disorders form a range of clonal haematological malignant diseases, the main members of which are polycythaemia vera, essential thrombocythaemia, and idiopathic myelofibrosis. The molecular pathogenesis of these disorders is unknown, but tyrosine kinases have been implicated in several related disorders. We investigated the role of the cytoplasmic tyrosine kinase JAK2 in patients with a myeloproliferative disorder.

Methods

We obtained DNA samples from patients with polycythaemia vera, essential thrombocythaemia, or idiopathic myelofibrosis. The coding exons of JAK2 were bidirectionally sequenced from peripheral-blood granulocytes, T cells, or both. Allele-specific PCR, molecular cytogenetic studies, microsatellite PCR, Affymetrix single nucleotide polymorphism array analyses, and colony assays were undertaken on subgroups of patients.

Findings

A single point mutation (Val617Phe) was identified in JAK2 in 71 (97%) of 73 patients with polycythaemia vera, 29 (57%) of 51 with essential thrombocythaemia, and eight (50%) of 16 with idiopathic myelofibrosis. The mutation is acquired, is present in a variable proportion of granulocytes, alters a highly conserved valine present in the negative regulatory JH2 domain, and is predicted to dysregulate kinase activity. It was heterozygous in most patients, homozygous in a subset as a result of mitotic recombination, and arose in a multipotent progenitor capable of giving rise to erythroid and myeloid cells. The mutation was present in all erythropoietin-independent erythroid colonies.

Interpretation

A single acquired mutation of JAK2 was noted in more than half of patients with a myeloproliferative disorder. Its presence in all erythropoietin-independent erythroid colonies demonstrates a link with growth factor hypersensitivity, a key biological feature of these disorders.

Relevance to practice

Identification of the Val617Phe JAK2 mutation lays the foundation for new approaches to the diagnosis, classification, and treatment of myeloproliferative disorders.

Introduction

The human myeloproliferative disorders represent a range of clonal haematological malignant diseases, with three main members: polycythaemia vera, essential thrombocythaemia, and idiopathic myelofibrosis.1 These three disorders are all thought to reflect transformation of a multipotent haemopoietic stem cell,2, 3 but their molecular pathogenesis remains obscure. The absence of definitive diagnostic tests and the scarcity of randomised controlled trials make management of these diseases especially challenging.4, 5

The myeloproliferative disorders are characterised by overactive haemopoiesis, with increased production of red cells and platelets the major feature of polycythaemia vera and essential thrombocythaemia, respectively. The main clinical complication in these two disorders is thrombosis, although haemorrhage can also happen. In the longer term, a few patients with polycythaemia vera and essential thrombocythaemia might develop myelofibrosis or acute myeloid leukaemia. Late-stage idiopathic myelofibrosis is characterised by bone-marrow fibrosis, cytopenia, and splenomegaly, and can also transform to acute myeloid leukaemia.

Erythroid progenitor cells from patients with polycythaemia vera form colonies in the absence of erythropoietin.6 Subsequently, investigators showed that both erythroid and myeloid progenitor cells from these patients are hypersensitive to several different growth factors7, 8 and that similar abnormalities are present in a proportion of patients with essential thrombocythaemia and idiopathic myelofibrosis.8, 9, 10 These observations suggest that the primary abnormality in myeloproliferative disorders might be in a signalling component downstream of multiple growth-factor receptors.11

Several lines of evidence suggest that kinases, particularly tyrosine kinases, represent attractive candidate genes. The BCR-ABL tyrosine kinase fusion protein causes the related disorder chronic myeloid leukaemia,12 and several tyrosine kinases are altered in patients with other myeloid malignant diseases,13, 14 including FLT3 in acute myeloid leukaemia, PDGFRβ in chronic myelomonocytic leukaemia, PDGFRα in chronic eosinophilic leukaemia, and KIT in systemic mastocytosis.

JAK2 is a cytoplasmic tyrosine kinase with a key role in signal transduction from multiple haemopoietic growth-factor receptors and so is an especially attractive candidate gene.15, 16 As part of our continuing assessment of protein kinase genes in patients with a myeloproliferative disorder, we sequenced the coding exons of JAK2 in peripheral-blood granulocytes from patients with polycythaemia vera, essential thrombocythaemia, or idiopathic myelofibrosis.

Section snippets

Study population

Our study was approved by Addenbrooke's NHS Trust local research ethics committee. We recruited patients from the myeloproliferative disorders clinic at Addenbrooke's Hospital and other haematology clinics. Written informed consent was obtained from every patient. We prospectively gathered data for baseline characteristics and diagnostic information, and we defined clinical events according to standard criteria17 by retrospective case-note review. Patients' diagnoses were defined according to

Results

In 66 of 140 patients with a myeloproliferative disorder, a G→T alteration was detected in exon 12 of JAK2 by sequence analysis of peripheral-blood granulocytes—53 of 73 patients with polycythaemia vera, six of 51 with essential thrombocythaemia, and seven of 16 with idiopathic myelofibrosis (figure 1; table 1). This alteration was within the pseudokinase (JH2) domain and changed a highly conserved valine at position 617 to phenylalanine (figure 2). In 30 patients with the Val617Phe mutation,

Discussion

The myeloproliferative disorders were classified as a spectrum of related diseases in 1951 by Dameshek.22 The prescience of this insight is emphasised by the results presented here, which show that a single mutation is shared by patients with polycythaemia vera, essential thrombocythaemia, and idiopathic myelofibrosis and underline the many similarities between these disorders. The observation that the Val617Phe alteration is seen in only 50% of patients with essential thrombocythaemia or

Glossary

10K single nucleotide polymorphism (SNP) array
10 000 SNP-containing DNA fragments are arranged onto a chip and hybridised with sample DNA
Metaphase FISH
Hybridisation of fluorescently labelled DNA to metaphase chromosomes
Microsatellite PCR
PCR amplification of highly polymorphic tandem repeat sequences

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    These authors contributed equally to this study

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