Holger Cario

JAK2 haplotype is a major risk factor for the development of myeloproliferative neoplasms.

Chronic myeloproliferative neoplasms (MPNs) are a group of related conditions characterized by the overproduction of cells from one or more myeloid lineages. More than 95% of cases of polycythemia vera, and roughly half of essential thrombocythemia and primary myelofibrosis acquire a unique somatic 1849G>T JAK2 mutation (encoding V617F) that is believed to be a critical driver of excess proliferation. We report here that JAK2V617F-associated disease is strongly associated with a specific constitutional JAK2 haplotype, designated 46/1, in all three disease entities compared to healthy controls (polycythemia vera, n = 192, P = 2.9 × 10−16; essential thrombocythemia, n = 78, P = 8.2 × 10−9 and myelofibrosis, n = 41, P = 8.0 × 10−5). Furthermore, JAK2V617F specifically arises on the 46/1 allele in most cases. The 46/1 JAK2 haplotype thus predisposes to the development of JAK2V617F-associated MPNs (OR = 3.7; 95% CI = 3.1–4.3) and provides a model whereby a constitutional genetic factor is associated with an increased risk of acquiring a specific somatic mutation.

Relative response of patients with myelodysplastic syndromes and other transfusion-dependent anaemias to deferasirox (ICL670): a 1-yr prospective study

Objectives/methods:  This 1‐yr prospective phase II trial evaluated the efficacy of deferasirox in regularly transfused patients aged 3–81 yrs with myelodysplastic syndromes (MDS; n = 47), Diamond–Blackfan anaemia (DBA; n = 30), other rare anaemias (n = 22) or β‐thalassaemia (n = 85). Dosage was determined by baseline liver iron concentration (LIC).

Identification of microdeletions spanning the Diamond-Blackfan anemia locus on 19q13 and evidence for genetic heterogeneity.

Summary Diamond-Blackfan anemia (DBA) is a rare pure red-cell hypoplasia of unknown etiology and pathogenesis. A major DBA locus has previously been localized to chromosome 19q13.2. Samples from additional families have been collected to identify key recombinations, microdeletions, and the possibility of heterogeneity for the disorder. In total, 29 multiplex DBA families and 50 families that comprise sporadic DBA cases have been analyzed with polymorphic 19q13 markers, including a newly identified short-tandem repeat in the critical gene region. The results from DNA analysis of 29 multiplex families revealed that 26 of these were consistent with a DBA gene on 19q localized to within a 4.1-cM interval restricted by loci D19S200 and D19S178; however, in three multiplex families, the DBA candidate region on 19q13 was excluded from the segregation of marker alleles. Our results suggest genetic heterogeneity for DBA, and we show that a gene region on chromosome 19q segregates with the disease in the majority of familial cases. Among the 50 families comprising sporadic DBA cases, we identified two novel and overlapping microdeletions on chromosome 19q13. In combination, the three known microdeletions associated with DBA restrict the critical gene region to ∼1 Mb. The results indicate that a proportion of sporadic DBA cases are caused by deletions in the 19q13 region.

The JAK2V617F mutation is acquired secondary to the predisposing alteration in familial polycythaemia vera.

Several laboratories have recently reported an acquired mutation in Janus kinase-2 (Jak2) in patients with sporadic polycythaemia vera (PV) (Baxter et al, 2005; James et al, 2005; Kralovics et al, 2005; Levine et al, 2005). Three groups also investigated patients who had a first-degree relative with a myeloproliferative disorder (MPD) ‘raising the possibility of a constitutional mutation of Jak2 in these families’ (Baxter et al, 2005; Kralovics et al, 2005; Levine et al, 2005). From their data, which showed no evidence of the mutation in unaffected tissues, the authors concluded that the Val617Phe mutation ‘is acquired in most patients’, but ‘do not exclude the possibility that it could be inherited in a few patients’ (Baxter et al, 2005). We have therefore analysed the Jak2 mutational status in a large pedigree with familial PV that has been previously reported (Cario et al, 2003). While both affected members [unique patient numbers (UPNs) 534 and 533] displayed the heterozygous G to T mutation at base pair 1849, the obligate carrier of the familial PV trait, UPN 532, was homozygous for the wild-type allele. Thus, Jak2V617F did not represent the heritable, disease predisposing allele in our pedigree (Fig 1). This analysis supports and extends the conclusions of recent reports that the Jak2V617F mutation is acquired, even in individuals who are genetically predisposed to developing PV. Kralovics et al (2003) previously excluded the chromosomal location of Jak2 (9p) as the site of the primary familial PV locus in four families. However, they suggested a model, in which one mutation is inherited and the second (now identified as Jak2) is acquired. The inherited mutation is possibly ‘genetically heterogeneous’ (Kralovics et al, 2003), requiring linkage analyses of multiple unrelated pedigrees in order to identify all genes capable of predisposing to the development of PV.

Genetic basis of Congenital Erythrocytosis mutation update and online databases

Congenital erythrocytosis (CE), or congenital polycythemia, represents a rare and heterogeneous clinical entity. It is caused by deregulated red blood cell production where erythrocyte overproduction results in elevated hemoglobin and hematocrit levels. Primary congenital familial erythrocytosis is associated with low erythropoietin (Epo) levels and results from mutations in the Epo receptor gene (EPOR). Secondary CE arises from conditions causing tissue hypoxia and results in increased Epo production. These include hemoglobin variants with increased affinity for oxygen (HBB, HBA mutations), decreased production of 2,3‐bisphosphoglycerate due to BPGM mutations, or mutations in the genes involved in the hypoxia sensing pathway (VHL, EPAS1, and EGLN1). Depending on the affected gene, CE can be inherited either in an autosomal dominant or recessive mode, with sporadic cases arising de novo. Despite recent important discoveries in the molecular pathogenesis of CE, the molecular causes remain to be identified in about 70% of the patients. With the objective of collecting all the published and unpublished cases of CE the COST action MPN&MPNr‐Euronet developed a comprehensive Internet‐based database focusing on the registration of clinical history, hematological, biochemical, and molecular data (http://www.erythrocytosis.org/). In addition, unreported mutations are also curated in the corresponding Leiden Open Variation Database.

Erythrocytosis associated with a novel missense mutation in the BPGM gene

The ERYTHROPOIETIN (EPO) gene is regulated by the transcription factor Hypoxia Inducible Factor-α (HIF-α). In this pathway, Prolyl Hydroxylase Domain protein 2 (PHD2) hydroxylates two prolyl residues in HIF-α, which in turn promotes HIF-α degradation by the von Hippel Lindau (VHL) protein. Evidence that HIF-2α is the important isoform for EPO regulation in humans comes from the recent observation that mutations in the HIF2A gene are associated with cases of erythrocytosis. We report here a new erythrocytosis-associated mutation, p.Asp539Glu, in the HIF2A gene. Similar to all reported cases, the affected residue is in close vicinity and C-terminal to the primary hydroxylation site in HIF-2α, Pro531. This mutation, however, is notable in producing a rather subtle amino acid substitution. Nonetheless, we find that this mutation compromises binding of HIF-2α to both PHD2 and VHL, and we propose that this mutation is the cause of erythrocytosis in this individual.

Clinical and molecular characterisation of a prospectively collected cohort of children and adolescents with polycythemia vera.

The clinical, haematological, molecular and treatment data of eight paediatric patients with polycythemia vera (PV) were collected prospectively. One patient developed PV after treatment for large‐cell anaplastic lymphoma. Budd‐Chiari syndrome was diagnosed in two patients, necessitating orthotopic liver transplantation in one and transjugular portosystemic shunting in the other. The remaining patients presented with non‐specific symptoms. Endogenous erythroid colonies were detected in all cases examined. The JAK2V617F mutation was found in six patients; two patients displayed JAK2 exon 12 mutations, including one novel mutation (JAK2H538‐K539delinsI). CD177 (PRV‐1) mRNA expression was increased in three of five patients tested.

Familial polycythemia vera with Budd–Chiari syndrome in childhood

Summary. Polycythemia vera is a myeloproliferative disorder that, in most cases, occurs sporadically with a median age at presentation of 60 years. Familial cases are very rare and usually manifest in elderly family members. The Budd–Chiari syndrome, characterized by the obstruction and occlusion of the suprahepatic veins, is a rare typical complication in polycythemia vera patients. To date, only two children or adolescents with polycythemia vera and Budd–Chiari syndrome have been described. Here, we report an 11‐year‐old girl with Budd–Chiari syndrome as the initial symptom of familial polycythemia vera, which was also found in the girls grandmother. Details of the diagnostic procedures used and the clinical course are reported. The patient underwent orthotopic liver transplantation and is being treated with hydroxyurea. The available literature on familial polycythemia vera and polycythemia vera in childhood with and without Budd–Chiari syndrome is reviewed.

Gene Panel Sequencing Improves The Diagnostic Work-Up Of Patients With Idiopathic Erythrocytosis And Identifies New Mutations

Erythrocytosis is a rare disorder characterized by increased red cell mass and elevated hemoglobin concentration and hematocrit. Several genetic variants have been identified as causes for erythrocytosis in genes belonging to different pathways including oxygen sensing, erythropoiesis and oxygen transport. However, despite clinical investigation and screening for these mutations, the cause of disease cannot be found in a considerable number of patients, who are classified as having idiopathic erythrocytosis. In this study, we developed a targeted next-generation sequencing panel encompassing the exonic regions of 21 genes from relevant pathways (~79 Kb) and sequenced 125 patients with idiopathic erythrocytosis. The panel effectively screened 97% of coding regions of these genes, with an average coverage of 450×. It identified 51 different rare variants, all leading to alterations of protein sequence, with 57 out of 125 cases (45.6%) having at least one of these variants. Ten of these were known erythrocytosis-causing variants, which had been missed following existing diagnostic algorithms. Twenty-two were novel variants in erythrocytosis-associated genes (EGLN1, EPAS1, VHL, BPGM, JAK2, SH2B3) and in novel genes included in the panel (e.g. EPO, EGLN2, HIF3A, OS9), some with a high likelihood of functionality, for which future segregation, functional and replication studies will be useful to provide further evidence for causality. The rest were classified as polymorphisms. Overall, these results demonstrate the benefits of using a gene panel rather than existing methods in which focused genetic screening is performed depending on biochemical measurements: the gene panel improves diagnostic accuracy and provides the opportunity for discovery of novel variants.

Erythrocytosis in children and adolescents-classification, characterization, and consensus recommendations for the diagnostic approach.

During recent years, the increasing knowledge of genetic and physiological changes in polycythemia vera (PV) and of different types of congenital erythrocytosis has led to fundamental changes in recommendations for the diagnostic approach to patients with erythrocytosis. Although widely accepted for adult patients this approach may not be appropriate with regard to children and adolescents affected by erythrocytosis. The “congenital erythrocytosis” working group established within the framework of the MPN&MPNr‐EuroNet (COST action BM0902) addressed this question in a consensus finding process and developed a specific algorithm for the diagnosis of erythrocytosis in childhood and adolescence which is presented here. Pediatr Blood Cancer 2013;60:1734–1738.