Two novel JAK2 exon 12 mutations in JAK2V617F-negative polycythaemia vera patients

Abstract

marrow does not develop myeloid hypoplasia, during a nadir in the WBC as noted in the previously reported cases with cyclic oscillation. 2 As noted, the severe thrombocytopenia and macrocytic anemia are unusual in the chronic phase of CML. Moreover, the marrow findings of intense erythroid hyperplasia and a reduction of myeloid cells in the marrow are atypical for a diagnosis of CML. However, given the presence of the Philadelphia chromosome and typical picture Bcr-abl translocation with a striking response to imatinib mesylate, this case represents what we have termed an 'erythroid variant' of CML. To our knowledge, this appears to be the first case that has been observed and has responded to imatinib mesylate. The patient remains in complete hematologic and molecular remission for more than 6 years on continued treatment with imatinib (Figures 1-5 The Philadelphia chromosome-negative myeloproliferative diseases (MPD) are a group of late-onset, chronic haematopoietic disorders characterized by the clonal proliferation of stem and progenitor cells resulting in an increased output of mature cells of one or more blood cell lineages. A specific acquired mutation in the key haematopoietic kinase, Janus kinase-2 (JAK2), JAK2V617F, occurs within the regulatory pseudokinase JAK homology-2 (JH2) domain of the JAK2 molecule and is proposed to result in release of autoinhibition of the JAK homology-1 (JH1) kinase domain. This lesion, which occurs in exon 14 of JAK2, is associated with 495% of polycythaemia vera (PV) cases and approximately 50% of patients diagnosed with essential thrombocythaemia (ET) and idiopathic myelofibrosis (IMF), (reviewed by Kaushansky 2007). 1 The JAK2V617F mutation arises in the stem cell compartment consistent with the clonal proliferation of a multipotent progenitor that maintains longterm clonal haematopoiesis. 2 Further, the identification of a mutation in a kinase that is associated with multiple cytokine receptors (CR) is consistent with altered growth factor responses observed in PV. For example, the growth of erythropoietin (Epo)-independent 'endogenous' erythroid colonies (eBFUE) from bone marrow (BM) and peripheral blood (PB) samples in vitro is a key feature of PV and ET and can be utilized as a diagnostic tool. Further somatic mutations have been reported in JAK2V617F-negative MPD patients, including a transmembrane mutation in Mpl (Thrombopoietin receptor) in a small proportion of IMF and ET patients, 3 and more recently, a cluster of four different mutations in exon 12 of JAK2 has been described in JAK2V617F-negative PV and idiopathic erythrocytosis patients. 4,5 This group of mutations, affecting amino-acid residues F537-E543, lies in a highly conserved region proximal to the JH2 domain of JAK2 and results in altered growth factor responses in vitro and a myeloproliferative phenotype in a murine bone marrow transplant model. 4

A JAK2 Interdomain Linker Relays Epo Receptor Engagement Signals to Kinase Activation

“…All three mutations resulted in erythrocytosis, consistent with previous results (14,(22)(23)(24)26). Thus, the activating mutants we identified elicit hematological effects in vivo.…”

“…As shown in Fig. 7, animals that received V617F-transduced bone marrow cells exhibited erythrocytosis, consistent with previous observations (14,22,23,26). Recipients of K539I-transduced cells also had elevated hematocrit and higher red blood cell and reticulocyte count than mice transplanted with wild-type JAK2.…”

Rapid identification of JAK2 exon 12 mutations using high resolution melting analysis

“…However, these are both relatively rare mutant alleles, occurring in only 4 of the 50 cases reported in the literature. 9,[11][12][13][14]16,23 In contrast, the most common exon 12 mutations (F537-K539delinsL, N542-E543del, E543-D544del) were all detectable at a relative abundance of 7% or less. Secondly, patient samples with purely mutant DNA may pose a problem due to an absence of heteroduplex formation.…”

“…9,11 Individual allele-specific PCR reactions have been developed to detect the first four exon 12 mutations to be described. 9 However, additional mutations have subsequently been identified, [11][12][13][14][15][16] including duplications that might not easily be identified using an allele-specific PCR strategy. 14 It is possible that additional mutant alleles exist.…”

Aberrant signal transduction pathways in myeloproliferative neoplasms

“…18 Unlike the uniqueness of the point mutation that generates JAK2 V617F, several deletions and insertions were noted in the case of exon 12 mutations. 18,82 An attractive hypothesis is that exon 12 mutants of JAK2 favor interaction with EpoR over TpoR or G-CSFR, although a mechanistic basis for such a preference has yet to be found. Modeling of JAK2 suggests that the K539L falls in a loop in the linker region between the SH2 and the JH2 domain (Figure 1), which would be placed in space quite close to the loop represented by b4-b5 where V617 is located.…”