The JAK2 Mutation V617F Is Specifically Associated with Idiopathic Splanchnic Vein Thrombosis.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 380-380 ◽  
Author(s):  
Sandra Régina ◽  
Olivier Herault ◽  
Louis D’lteroche ◽  
Yannick Bacq ◽  
Jorge Domenenech ◽  
...  
Keyword(s):  
Risk Factors ◽  
Bone Marrow ◽  
Risk Factor ◽  
Antiphospholipid Antibodies ◽  
Jak2 V617f ◽  
Biological Data ◽  
Jak2 Mutation ◽  
Splanchnic Vein Thrombosis ◽  
Vein Thrombosis ◽  
Fv Leiden

Abstract Introduction Many risk factors have been identified in patients with portal (PVT) and hepatic venous thrombosis (HVT), including myeloproliferative disorders (MPD), i.e. polycythemia vera (PV) and essential thrombocythemia (ET), with prevalence varying between 25% and 65%. Moreover, JAK2 mutation V617F has recently been detected in 65 to 97% of cases of PV and 23 to 57% of ET. The aim of the present study was therefore to evaluate the frequency of JAK2 mutation V617F in a cohort of consecutive patients with PVT or HVT compared to a control group comprising patients with deep vein thrombosis (DVT) of the lower limbs. Patients and Methods Forty-four patients with PVT (n=42) or HVT (n=2) diagnosed between 2001 and 2005, and 44 patients (matched for age and gender) with idiopathic DVT were included. The presence of hereditary or acquired risk factors for thrombosis (including FV Leiden and FII G20210A polymorphism, defects in coagulation inhibitors, antiphospholipid antibodies and hyperhomocysteinemia) was investigated in all patients. Bone marrow (BM) biopsy and cultures of blood and BM progenitors were performed in 18 patients with PVT for whom the etiology of thrombosis was undefined. Screening for JAK2 mutation V617F was performed on genomic DNA isolated from peripheral blood cells by an allele-specific PCR and RFLP analysis using BsaXI, as previously described (Baxter et al. Lancet2005, 365:1054), combined with nano-electrophoresis (Bioanalyzer 2100, Agilent) to increase sensitivity. HEL92.1.7 and HL60 cell lines were used as positive and negative controls, respectively. Results The 2 groups were similar in terms of blood count and conventional risk factors for thrombosis (Table). JAK2 mutation V617F was detected in 8 patients with PVT (18.2%) but in none with DVT (p=0.003). These 8 subjects had normal blood counts, and no other acquired or hereditary risk factor for thrombosis except for one case exhibiting moderate hyperhomocysteinemia. Bone marrow was hyperplastic in 6/18 patients (all JAK2 V617F positive). Cultures of progenitors showed endogenous erythroid formation in 6/18 cases (all JAK2 V617F positive). The clinical follow-up confirmed the diagnosis of MPD (1 PV and 1 ET) in 2 patients. One subject died in another context and blood counts have remained normal in the 5 other patients to date. The JAK2 mutation V617F was not detected in any patient with negative bone marrow exploration (biopsy and cultures). Conclusion The JAK2 mutation V617F is frequently and specifically detected in patients with idiopathic PVT and no other risk factor for thrombosis. Whether JAK2 mutation V617F should be screened in all subjects with splanchnic vein thrombosis and should replace BM biopsy and cultures warrants further studies. Biological data in patients with PVT/HVT or DVT PVT or HVT (n=44) DVT (n=44) n % 95% CI n % 95% CI p * * Fisher exact test FV Leiden 4 9.1 2.5–21.7 11 25 13.2–40.3 NS FII 20210A 10 22.7 11.5–37.8 6 13.6 5.2–27.4 NS AT, PC, PS deficiency 7 15.9 7.9–29.3 4 9.1 2.5–21.7 NS Antiphospholipid antibodies 5 11.4 4.9–24 8 18.2 9.5–32 NS Hyperhomocysteinemia 4 9.1 2.5–21.7 4 9.1 2.5–21.7 NS JAK2 V617 mutation 8 18.2 9.5–32 0 0 0–8 0.003

Thrombotic Complications in Ph'-Negative Myeloproliferative Neoplasms - 10-Year Experience of A Medical Center in Asian Area

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5171-5171
Author(s):  
Yi-Sheng Chou ◽  
Ri-Dong Bai ◽  
Chung-Jen Teng ◽  
Ying-Chung Hong ◽  
Chun-Yu Liu ◽  
...  
Keyword(s):  
Risk Factors ◽  
Bone Marrow ◽  
Risk Factor ◽  
Venous Thromboembolism ◽  
Myeloproliferative Neoplasms ◽  
Vein Thrombosis ◽  
Bone Marrow Cellularity ◽  
Reticulin Fiber ◽  
Thrombotic Complications ◽  
Marrow Cellularity

Abstract Abstract 5171 Although thrombotic complications are common in myeloproliferative neoplasms (MPN), the pertinent risk factors are not well characterized. This study aims to explore the general characteristics of MPN patients, focusing on risk factors for developing thrombotic complications. Thrombotic events were defined as relevant to the MPN if they occurred after the diagnosis of MPN or around the time of diagnosis. Results Totally 361 patients were collected, including 135 (37%) cases of polycythemia vera (PV), 167 (46%) cases of essential thrombocythemia (ET) and 59 (16%) cases of primary myelofibrosis (PMF). The positive rates of JAK2V617F mutation were 90%, 65%, and 75% for PV, ET and PMF, respectively (PV vs. ET, p<0.001). PV was associated with higher incidence of transformation to myelofibrosis compared with ET (9% vs. 3%, p=0.027) and also higher level of white blood cell count (WBC) (mean 15018/ul, 95% confidence interval (CI):13655–16381/ul; PV vs. ET, p=0.005; PV vs. PMF, p=0.001 ), higher bone marrow cellularity (bone marrow cellularity more than 60%: PV,89%; ET,70%; PV vs. ET, p=0.026; PMF,76%), and more frequent hypertension (PV, 50%; ET, 43%; PMF 19%; PV vs PMF, p<0.001; ET vs. PMF, p=0.001). ET was associated lower incidence of coagulopathy with prolonged INR in prothrombin time (ET, 19%; PV, 36%; PV vs. ET, p=0.001; PMF, 44%; ET vs. PMF, p=0.001). PMF was associated with higher incidence of leukemia transformation(PMF,15%; PV,3.7%; ET, 2.4%; PV vs. PMF, p=0.012; ET vs. PMF, p=0.001), higher incidence of coagulopathy with prolonged aPTT (PMF,51%; PV,37%;ET, 30%; ET vs. PMF, p=0.012), increased reticulin fiber(moderate increased reticulin fiber: MF,92%; PV,44%; PV vs. PMF, p<0.001; ET, 48%;ET vs. PMF,p<0.001) and splenomegaly (mean 17.3cm, 95% CI:15.8–18.4cm, p<0.001). Venous thromboembolism occurred in 6.4% of these MPN patients with incidence of 7.4%, 6.6% and 3.4% in patients of PV, ET and PMF, respectively. Arterial thrombosis developed in 20% of these MPN patients with incidence of 28%, 17%, and 10% in patients of PV, ET and PMF, respectively. (Table1) Majority of venous thromboembolism presented as deep vein thrombosis (DVT) /pulmonary embolism (PE) and portal or splenic vein thrombosis with an incidence of 1.7/1.1% and 2.2%, respectively. The most common arterial thrombotic events was ischemic stroke, followed in order by ischemic heart disease, peripheral artery occlusive disease (PAOD), and ischemic bowel disease with an incidence of 14%, 9%, 3.0% and 1.1%, respectively. (Table1) The probability of thrombosis-free survival in these 361 patients was 85%, 78% and 70% at 1, 3 and 10 year, respectively (Figure 1). In terms of arterial and venous thromembolism altogether, univariate risk factor analysis revealed several risk factors including a positive JAK2V617F, diagnosis of PV, WBC more than 16000/ul, hemoglobin level higher than 16 mg/dl, albumin level less than 4.0 g/dl, LDH higher than 250U/L, congestive heart failure(CHF),hypertension(HTN) and diabetes mellitus. With multivariate analysis, however, only WBC more than 16000/ul (Odds ratio: 3.556,95% CI:1.287–9.822,p=0.014) remained as the most significant risk factors for thromoboembolism. Conclusions Arterial thromboses are quite common in patients of MPN, especially PV. In these MPN patients, the risk of venous thrombosis including thrombosis at unusual sites is also markedly increased. The most important risk factor predisposing to vascular thrombosis (vein and artery) is WBC more than 16000/ul. Our results provide informative clinical data for management of Asian patients with MPN and facilitate further study of these disorders in this area. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Impact of acquired and genetic factors on thrombophilic phenotype in FV Leiden mutation carriers

2005 ◽  
Vol 24 (2) ◽  
pp. 141-146
Author(s):  
Valentina Djordjevic ◽  
Ljiljana Rakicevic ◽  
Predrag Miljic ◽  
Danijela Mikovic ◽  
Mirjana Kovac ◽  
...  
Keyword(s):  
Risk Factors ◽  
Risk Factor ◽  
Genetic Risk ◽  
Genetic Factors ◽  
Vein Thrombosis ◽  
Mutation Carriers ◽  
Fv Leiden ◽  
Deep Vein ◽  
The Impact ◽  
Limb Deep Vein Thrombosis

FV Leiden mutation is an important genetic risk factor for venous thromboembolsm (VTE). In this study we have analyzed clinical manifestation and the impact of other genetic and acquired risk factors in 100 patients (95 heterozygous and 5 homozygous) carriers of FV Leiden mutation. Among these patients, 91 experienced VTE, with down limb deep vein thrombosis as the most frequent manifestation. An acquired risk factor was present in 68.6% of women, whereas this was the case in 28.6% of men. FIIG20210A was present in 9.5%, MTHFR 677TT in 8.4% and both mutations in 2.1% of the heterozygous FV Leiden carriers. Our results suggest that knowledge of coexisting factors predisposing to VTE is very important for FV Leiden mutation carriers and may contribute to the prevention of VTE episodes.

scholarly journals Splanchnic vein thrombosis in myeloproliferative neoplasms: pathophysiology and molecular mechanisms of disease

2016 ◽  
Vol 8 (3) ◽  
pp. 107-118 ◽  
Author(s):  
Joan How ◽  
Amy Zhou ◽  
Stephen T. Oh
Keyword(s):  
Risk Factors ◽  
Venous Thrombosis ◽  
Blood Cells ◽  
Molecular Mechanisms ◽  
Myeloproliferative Neoplasms ◽  
Jak2 V617f ◽  
Splanchnic Vein Thrombosis ◽  
Vein Thrombosis ◽  
Disease Mechanisms ◽  
Younger Age

Myeloproliferative neoplasms (MPNs) are the most common underlying prothrombotic disorder found in patients with splanchnic vein thrombosis (SVT). Clinical risk factors for MPN-associated SVTs include younger age, female sex, concomitant hypercoagulable disorders, and the JAK2 V617F mutation. These risk factors are distinct from those associated with arterial or deep venous thrombosis (DVT) in MPN patients, suggesting disparate disease mechanisms. The pathophysiology of SVT is thought to derive from local interactions between activated blood cells and the unique splanchnic endothelial environment. Other mutations commonly found in MPNs, including CALR and MPL, are rare in MPN-associated SVT. The purpose of this article is to review the clinical and molecular risk factors for MPN-associated SVT, with particular focus on the possible mechanisms of SVT formation in MPN patients.

Risk Factors, Diagnosis, Management, and Outcomes For Splanchnic Vein Thrombosis: A Retrospective Analysis

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2372-2372
Author(s):  
Benjamin A. Derman ◽  
Hau C. Kwaan
Keyword(s):  
Risk Factors ◽  
Liver Disease ◽  
Retrospective Analysis ◽  
Jak2 V617f ◽  
Diagnostic Methods ◽  
Jak2 V617f Mutation ◽  
Single Institution ◽  
Splanchnic Vein Thrombosis ◽  
Vein Thrombosis ◽  
V617f Mutation

Abstract Background There is a paucity of data on the incidence of risk factors for splanchnic vein thrombosis in current published literature. The present study is an attempt to determine the risk factors, diagnostic methods employed, treatment modalities, and outcomes in patients with splanchnic vein thrombosis in a single institution over a two-year period. Methods Retrospective chart review of patients, 18-90 years old, who were diagnosed with splanchnic vein thrombosis (SVT) at a single institution from January 1, 2010 to November 10, 2012. They were grouped as those with Budd-Chiari syndrome (BCS) and those with portal vein thrombosis (PVT), including those combined with splenic vein thrombosis (SPVT) and those with mesenteric vein thrombosis (MVT). Results Among the 246 patients studied, 21 had BCS and 225 had PVT. Associated risk factors in the order of frequency were liver disease being present in 48% of BCS, 69% of PVT, 45% of PVT+SPVT, and 52% of PVT+MVT. Next was regional cancer, being present in 24%in BCS and 47% of PVT. Third commonest was pancreatitis being present in 14% of BCS, 9% of PVT, 18% of PVT+SPVT, and 6% of PVT+MVT. Hereditary thrombophilias were found in 10% of the BCS group and 4% of PVT; however, it constituted 18% of the PVT+SPVT group, and 12% of the PVT+MVT group. 10% of patients in both the BCS and PVT groups had a liver transplant during their lifetime. The most common presenting symptom was abdominal pain occurring in 57% patients with BCS and 50% patients with PVT. The majority had laboratory findings of liver dysfunction at presentation with 86% in BCS group and 78% in PVT group. JAK2 V617F mutation, when tested, was present in 14% of those with BCS, 20% of the PVT group, 29% of those with PVT+SPVT and 22% of those with PVT+MVT. Diagnosis of SVT was most commonly made by computerized tomography (CT) with contrast (57% for BCS, 56% for PVT). Approximately 60% of BCS patients and 30% of PVT patients received either short-term or long-term anticoagulation; 20% of both groups received transjugular intrahepatic portal system (TIPS) catheterization. Recurrence of symptoms requiring a second hospitalization occurred in 24% of those with BCS and 15% of patients with PVT (36% of the PVT+SPVT and 27% of the PVT+MVT). In those patients with a greater comorbidity profile, including hypertension, diabetes, and malignancy, PVT is more likely than BCS to occur. Regional presence of inflammation or cancer, specifically underlying liver disease, hepatocellular carcinoma, pancreatic cancer, pancreatitis, as well as regional surgical procedures appear to play major role in splanchnic vein thrombosis, while hereditary thrombophilias and the JAK2 V617F mutation make up an important but small component of splanchnic vein thrombosis. Contrast-enhanced CT was the most commonly successful radiologic technique for diagnosis, though magnetic resonance imaging (MRI) provides a more accurate alternative. Anticoagulation was largely limited to patients with the most severe cases of SVT, and symptomatic recurrence was also more likely in these populations. Conclusions The present findings of risk factors associated with SVT are at variance with those in the current published literature, with higher incidence of regional cancer and lower incidence of JAK2 V617F mutation. There are, however, limitations to this study, including the fact that this is a retrospective analysis with data from a single institution. Verification of these findings has to been made in a prospective multi-institutional study involving a larger number of patients and a longer period of observation. Disclosures: No relevant conflicts of interest to declare.

Splanchnic vein thrombosis: Clinical manifestations, risk factors, management, and outcomes

2021 ◽  
Vol 202 ◽  
pp. 90-95
Author(s):  
Eri Kawata ◽  
Dou-Anne Siew ◽  
James Gordon Payne ◽  
Martha Louzada ◽  
Michael J. Kovacs ◽  
...  
Keyword(s):  
Risk Factors ◽  
Clinical Manifestations ◽  
Splanchnic Vein Thrombosis ◽  
Vein Thrombosis

scholarly journals Upper Extremity Deep Vein Thrombosis in Acute Leukemia and Non-Hodgkin's Lymphoma: Analysis of the California Cancer Registry

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 932-932
Author(s):  
Christina Poh ◽  
Ann M Brunson ◽  
Theresa H.M. Keegan ◽  
Ted Wun ◽  
Anjlee Mahajan
Keyword(s):  
Risk Factors ◽  
Risk Factor ◽  
Deep Vein Thrombosis ◽  
Major Bleeding ◽  
Cumulative Incidence ◽  
Steering Committee ◽  
Vein Thrombosis ◽  
California Cancer Registry ◽  
Increased Risk ◽  
Deep Vein

Background Venous thromboembolism (VTE) is a known complication in patients with acute myeloid leukemia (AML), acute lymphoid leukemia (ALL) and non-Hodgkin's lymphoma (NHL). However, the cumulative incidence, risk factors, rate of subsequent VTE and impact on mortality of upper extremity deep vein thrombosis (UE DVT) in these diseases is not well-described. Methods Using the California Cancer Registry, we identified patients with a first primary diagnosis of AML, ALL and NHL from 2005-2014 and linked these patients with the statewide hospitalization and emergency department databases to identify an incident UE DVT event using specific ICD-9-CM codes. Patients with VTE prior to or at the time of malignancy diagnosis or who were not treated with chemotherapy were excluded. We determined the cumulative incidence of first UE DVT, adjusted for the competing risk of death. We also examined the cumulative incidence of subsequent VTE (UE DVT, lower extremity deep vein thrombosis (LE DVT) and pulmonary embolism (PE)) and major bleeding after incident UE DVT. Using Cox proportional hazards regression models, stratified by tumor type and adjusted for other prognostic covariates including sex, race/ethnicity, age at diagnosis, neighborhood, sociodemographic status and central venous catheter (CVC) placement, we identified risk factors for development of incident UE DVT, the effect of incident UE DVT on PE and/or LE DVT development, and impact of incident UE DVT on cancer specific survival. The association of CVC placement with incident UE DVT was not assessed in acute leukemia patients, as all who undergo treatment were assumed to have a CVC. Results are presented as adjusted hazard ratios (HR) and 95% confidence intervals (CI). Results Among 37,282 patients included in this analysis, 6,213 had AML, 3,730 had ALL and 27,339 had NHL. The 3- and 12-month cumulative incidence of UE DVT was 2.6% and 3.6% for AML, 2.1% and 3% for ALL and 1.0% and 1.6% for NHL respectively (Figure 1A). Most (56-64%) incident UE DVT events occurred within the first 3 months of malignancy diagnosis. African Americans (HR 1.66; CI 1.22-2.28) and Hispanics (HR 1.35; CI 1.10-1.66) with NHL had an increased risk of incident UE DVT compared to non-Hispanics Whites. NHL patients with a CVC had over a 2-fold increased risk of incident UE DVT (HR 2.05; CI 1.68-2.51) compared to those without a CVC. UE DVT was a risk factor for development of PE or LE DVT in ALL (HR 2.53; CI 1.29-4.95) and NHL (HR 1.63; CI 1.11-2.39) but not in AML. The 12-month cumulative incidence of subsequent VTE after an incident UE DVT diagnosis was 6.4% for AML, 12.0% for ALL and 7.6% for NHL. 46-58% of subsequent VTEs occurred within the first 3 months of incident UE DVT diagnosis. The majority of subsequent VTEs were UE DVT which had a 12-month cumulative incidence of 4.6% for AML, 6.6% for ALL and 4.0% for NHL (Figure 1B). The 12-month cumulative incidence of subsequent LE DVT was 1.3% for AML, 1.6% for ALL and 1.9% for NHL (Figure 1C). The 12-month cumulative incidence of subsequent PE was 0.4% for AML, 4.1% for ALL and 1.8% for NHL (Figure 1D). The 12-month cumulative incidence of major bleeding after an UE DVT diagnosis was 29% for AML, 29% for ALL and 20% for NHL. Common major bleeding events included gastrointestinal (GI) bleeds, epistaxis and intracranial hemorrhage. GI bleeding was the most common major bleeding event among all three malignancies (14.2% in AML, 9.6% in ALL and 12.4% in NHL). The rate of intracranial hemorrhage was 6% in AML, 3.5% in ALL and 1.7% in NHL. A diagnosis of incident UE DVT was associated with an increased risk of cancer-specific mortality in all three malignancies (HR 1.38; CI 1.16-1.65 in AML, HR 2.16; CI 1.66-2.82 in ALL, HR 2.38; CI 2.06-2.75 in NHL). Conclusions UE DVT is an important complication among patients with AML, ALL and NHL, with the majority of UE DVT events occurring within the first 3 months of diagnosis. The most common VTE event after an index UE DVT was another UE DVT, although patients also had subsequent PE and LE DVT. UE DVT was a risk factor for development of PE or LE DVT in ALL and NHL, but not in AML. Major bleeding after an UE DVT was high in all three malignancies (&gt;20%), with GI bleeds being the most common. UE DVT in patients with AML, ALL and NHL is associated with increased risk of mortality. Disclosures Wun: Janssen: Other: Steering committee; Pfizer: Other: Steering committee.

scholarly journals Is There a Role for Bone Marrow Biopsy In The Diagnosis of Masked Polycythemia Vera with Splanchnic Vein Thrombosis?

2020 ◽  
pp. 60-63
Author(s):  
Maria Cristina Scamuffa ◽  
Roberto Latagliata ◽  
Luisa Bizzoni ◽  
Maria Lucia De Luca ◽  
Federica Falco ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Polycythemia Vera ◽  
Bone Marrow Biopsy ◽  
Splanchnic Vein Thrombosis ◽  
Vein Thrombosis

Thrombophilic Genotypes in Subjects with Idiopathic Antiphospholipid Antibodies – Prevalence and Significance

1998 ◽  
Vol 79 (01) ◽  
pp. 46-49 ◽  
Author(s):  
Catello Tommasino ◽  
Giovanna D’Andrea ◽  
Luigi Iannaccone ◽  
Vincenzo Brancaccio ◽  
Maurizio Margaglione ◽  
...  
Keyword(s):  
Antiphospholipid Antibodies ◽  
Healthy Subjects ◽  
Methylenetetrahydrofolate Reductase ◽  
Thrombotic Event ◽  
Significant Proportion ◽  
Factor V ◽  
Vein Thrombosis ◽  
History Of ◽  
Fv Leiden

SummaryTo evaluate the significance of common thrombophilic genotypes in subjects with idiopathic antiphospholipid antibodies (aPL) we determined the methylenetetrahydrofolate reductase C677→ (MTHFR) and factor V A506→ G (FV Leiden) polymorphisms in 49 subjects with idiopathic aPL (57% of whom suffered spontaneous vein thrombosis), in 70 subjects with a history of spontaneous vein thrombosis and in 193 healthy subjects. The prevalence of MTHFR C677→+/+ (homozygotes) was 25%, 18% and 17% respectively amongst aPL thrombotics, non aPL thrombotics and controls and that of MTHFR C677→+/– (heterozygotes) was 53%, 59% and 53% respectively in the same groups. The prevalence of FV Leiden was higher in aPL thrombotics (14%) and in non aPL thrombotics (18%) than in controls (4%) (p ≤ = 0.05). APL thrombotics with MTHFR C677→+/+ had a lower mean age at first thrombotic event (22 ± 6 years) than aPL thrombotics with MTHFR C677→+/– and non mutated considered together (38 ± 14 years, p = 0.0004) and than non aPL thrombotics with MTHFR C677→+/+ (38 ± 14 years, p = 0.003). FV Leiden may con tribute to the hypercoagulability of a small, albeit significant proportion of thrombotic aPL subjects, whereas the association between MTHFR C677→+/+ and aPL may have an impact on age at first occlusive event and suggests a possible pathogenetic interaction.

Cultivation of Megakaryocytes On a Collagen Matrix: A More Sensitive and Reproducible Test for the Diagnosis of Patients with Essential Thrombocythaemia.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4970-4970
Author(s):  
Adrian Emanuel Schmidt ◽  
Patricia Darlington ◽  
Lucie Kopfstein ◽  
Elisabeth Ischi ◽  
Elisabeth Oppliger Leibundgut ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Platelet Count ◽  
Negative Predictive Value ◽  
Predictive Value ◽  
Healthy Subjects ◽  
Myeloproliferative Neoplasms ◽  
Jak2 V617f ◽  
Essential Thrombocythaemia ◽  
Jak2 Mutation ◽  
Spontaneous Proliferation

Abstract Abstract 4970 Background Essential thrombocythaemia (ET) is one of the chronic myeloproliferative neoplasms (MPN), along with polycythaemia vera (PV), primary myelofibrosis (PMF) and chronic myeloid leukaemia (CML). Their common feature is excessive proliferation of a certain stem or progenitor cell in the bone marrow; in the case of ET, the megakaryocytic lineage is affected. Clinical manifestations include thrombotic events and haemorrhage. Diagnosis of ET according to new WHO-criteria requires a sustained high platelet count, bone marrow biopsy showing proliferation of the megakaryocytic lineage with large and mature morphology, demonstration of JAK2 V617F (although only present in about 50% of patients with ET) or another clonal marker and explicit exclusion of other myeloid and myeloproliferative neoplasms as well as signs of reactive thrombocytosis. Additionally, spontaneous proliferation of megakaryocytes obtained from peripheral blood can be detected in in vitro culture assays. Presently, we use agar as a matrix for megakaryocyte cultivation, although this assay has never been validated in connection with ET. The identification of megakaryocytic colonies grown on agar can sometimes be quite difficult. Our aims were therefore to technically evaluate the use of a collagen based matrix and to investigate its suitability to identify patients with ET. Patients and Methods We have examined 63 patients (26 with ET, 21 with PV, 8 with myelofibrosis [MF; including PMF and post-ET/PV-MF], 6 with secondary or idiopathic erythrocytosis and 2 with secondary thrombocytosis; mean age=59.8, male=33, female=30, mean platelet count 457 G/l) and 5 healthy subjects. Following informed consent, both clinical and laboratory data was collected. Medication intake, phlebotomies, smoking habits and regular haemogram results were noted in order to recognise possible confounding factors influencing laboratory results. Results of megakaryocyte cultivation on both agar and collagen matrixes were recorded, considering both spontaneous growth and growth stimulated by megakaryocyte derived growth factor (MDGF). Results Based on our collagen culture results we were able to define 2 or more spontaneously grown megakaryocyte colonies as the most optimal cut-off for the identification of patients with MPN (sensitivity 71%, specificity 100% with positive and negative predictive values of 100% and 45%, respectively). Compared to the agar culture results (where a specificity and a positive predictive value of 100% were demonstrated at a cut-off value of ≥ 10 CFU-Mega) we found a higher accuracy and better reproducibility. In addition, we observed an improved negative predictive value (45% with collagen versus 25% with agar cultures) reducing false negative results. Healthy subjects and patients with secondary thrombocytosis showed no significant spontaneous megakaryocyte proliferation. In patients with MF, we observed strong spontaneous and MDGF-stimulated growth of megakaryocytic colonies. At a cut-off value of ≥ 50 CFU-Mega (after stimulation with MDGF), the collagen assay showed a sensitivity of 100% and a specifity of 70% for this special form of MPN, resulting in a negative predictive value of 100%. We found no confounding clinical or laboratory parameters such as medication intake (particularly cytoreductive treatment with hydroxyurea) or phlebotomies influencing our culture results, and no significant effect of the Jak2-V617F mutation on the growth behaviour of megakaryocytic colonies. Conclusion The results of this ongoing study imply that the collagen based assay is more sensitive, specific, time efficient and user friendly regarding the detection of spontaneous proliferation of megakaryocytes than the currently used agar based culture assay. In addition, the collagen based assay also has the great advantage that it allows isolation of single megakaryocytic colonies for further analyses, for example PCR-based identification of a JAK2 mutation. Furthermore, the collagen based assay facilitates the diagnosis of patients with MPN, especially in cases where conventional diagnostic criteria are lacking, such as in ET without a JAK2 mutation. Ultimately, the new assay may well be able to detect transformation from PV/ET to MF. Disclosures No relevant conflicts of interest to declare.

A Novel Mutation in MPL (Y252H) Results in Increased Thrombopoietin Sensitivity and Moderate Thrombocytosis in a Pediatric Patient with Essential Thrombocytosis

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2817-2817
Author(s):  
Michele Lambert ◽  
Jing Jiang ◽  
Wei Tong
Keyword(s):  
Bone Marrow ◽  
Platelet Count ◽  
Bone Marrow Cells ◽  
Novel Mutation ◽  
Missense Variant ◽  
Jak2 V617f ◽  
Jak2 Mutation ◽  
Hematopoietic Malignancies ◽  
Healthy Control ◽  
Pediatric Populations

Abstract Abstract 2817 Myeproliferative neoplams (MPNs) constitute a group of hematopoietic malignancies that feature enhanced proliferation and/or survival of one or more myeloid lineage cells, including Essential Thrombocythemia (ET). MPN development is rare in children with an estimated annual incidence of ET of 0.09/106 children. The WHO defines ET as persistent platelet count >600 k/mcL in the absence of known cause of reactive/secondary thrombocytosis. The JAK2 V617F mutation is most commonly reported in both children and adults with ET although the reported frequency varies in pediatric populations from 0 to 36% of patients. Mutations in the thrombopoietin receptor (TPO receptor or MPL) intracellular domain, specifically W515K/L, have also been reported in both adult and pediatric ET patients. Here we report a novel mutation in the MPL extracellular domain, Y252H, causing mild thrombocytosis. The patient presented at 2 years of age with a platelet count of 765 k/mcL. During the 3-year follow-up period, she possessed platelet counts between 600–700k/mcL, without any obvious indication of reactive/secondary thrombocytosis. Because of the persistently increased platelet count, her bone marrow was evaluated and it demonstrated increased numbers of megakaryocytes with focal clustering. JAK2 mutation analysis was negative and cytogenetics did not show any clonal abnormalities. Sequencing of the MPL gene showed a missense variant at c.754 T>C resulting in a tyr252his amino acid substitution. To investigate if this Y252H mutation in MPL dysregulates TPO/MPL- mediated cell growth, we introduced it into cytokine-dependent BaF3 cells. Cells stably expressing the mutant MPL allele showed increased proliferation to TPO, in particular at low concentration, in comparison to cells expressing wildtype (WT) MPL. Upon cytokine withdrawal, BaF3 cells expressing the MPL Y252H mutant survived better than that of WT MPL. Primary bone marrow cells from this patient along with the healthy control were subjected to colony forming unit -megakaryocyte (CFU-meg) assays in response to a serial dose of TPO. The Y252H MPL bone marrow showed significantly increased megakaryocyte colonies at low dose of TPO when compared to control bone marrow (17.5 ± 2.5 colonies versus 4.75 ± 1.1 colonies at 15 ng/mL TPO, p<0.001). These results are consistent with the clinically mild thrombocytosis. In summary, our results suggest a novel MPL mutation, Y252H, results in pediatric ET. Further evaluation of the mechanisms of increased TPO sensitivity imparted by this mutation should contribute to our understanding of the molecular pathogenesis of ET. Disclosures: Lambert: Cangene: Honoraria.

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