CXCL8 and its cognate receptors CXCR1/CXCR2 in primary myelofibrosis.

BCR::ABL1 negative myeloproliferative neoplasms (MPN) form a distinct group of hematologic malignancies characterized by sustained proliferation of cells from multiple myeloid lineages. With a median survival of 16-35 months in patients with high-risk disease, primary myelofibrosis (PMF) is considered the most aggressive entity amongst all BCR::ABL1 MPN. Additionally, for a significant subset of patients, MPN evolve into secondary acute myeloid leukemia (AML), which has an even poorer prognosis compared to de novo AML. As the exact mechanisms of disease development and progression remain to be elucidated, current therapeutic approaches fail to prevent disease progression or transformation into secondary AML. As each MPN entity is characterized by sustained activation of various immune cells and raised cytokine concentrations within bone marrow (BM) and peripheral blood (PB), MPN may be considered to be typical inflammation-related malignancies. However, the exact role and consequences of increased cytokine concentrations within BM and PB plasma has still not been completely established. Up-regulated cytokines can stimulate cellular proliferation, or contribute to the development of an inflammation-related BM niche resulting in genotoxicity and thereby supporting mutagenesis. The neutrophil chemoattractant CXCL8 is of specific interest as its concentration is increased within PB and BM plasma of patients with PMF. Increased concentration of CXCL8 negatively correlates with overall survival. Furthermore, blockage of the CXCR1/2 axis appears to be able to reduce BM fibrosis and megakaryocyte dysmorphia in murine models. In this review, we summarize available evidence on the role of the CXCL8-CXCR1/2 axis within the pathogenesis of PMF, and discuss potential therapeutic modalities targeting either CXCL8 or its cognate receptors CXCR1/2.

Predictors for improvement in patient-reported outcomes: post hoc analysis of a phase 3 randomized, open-label study of eculizumab and ravulizumab in complement inhibitor-naive patients with paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is characterized by uncontrolled terminal complement activation leading to intravascular hemolysis (IVH), thrombosis, and impairments in quality of life (QoL). The aim of this study was to identify the clinical drivers of improvement in patient-reported outcomes (PROs) in patients with PNH receiving the complement component 5 (C5) inhibitors eculizumab and ravulizumab.This post hoc analysis assessed clinical outcomes and PROs from 246 complement inhibitor-naive patients with PNH enrolled in a phase 3 randomized non-inferiority study that compared the C5 inhibitors ravulizumab and eculizumab (study 301; NCT02946463). The variables of interest were lactate dehydrogenase (LDH) levels, a surrogate measure of IVH, and hemoglobin (Hb) levels. PROs were collected using Functional Assessment of Chronic Illness Therapy-Fatigue (FACIT-F) and European Organisation for Research and Treatment of Cancer, Quality of Life Questionnaire-Core 30 (EORTC QLQ-C30) to assess fatigue and QoL, respectively.Improvements in absolute mean LDH levels were significantly associated with improvements in mean FACIT-F score (p = 0.0024) and EORTC QLQ-C30 global health (GH) score (p < 0.0001) from baseline to day 183. Improvements in scores were achieved despite a non-significant increase in Hb levels. To understand the interaction between LDH and Hb, a regression analysis was performed: LDH response with Hb improvements was a significant predictor of improvement in fatigue. The independent effect of improved Hb did not significantly affect FACIT-F or EORTC QLQ-C30 GH scores.These findings suggest that LDH levels are an important determinant of fatigue and QoL outcomes in patients with PNH. CTR: NCT02946463, October 27, 2016.

Ruxolitinib in patients with polycythemia vera resistant and/or intolerant to hydroxyurea: European observational study.

BACKGROUND: Hydroxyurea (HU) is a commonly used first-line treatment in patients with polycythemia vera (PV). However, approximately 15%-24% of PV patients report intolerance and resistance to HU. METHODS: This phase IV, European, real-world, observational study assessed the efficacy and safety of ruxolitinib in PV patients who were resistant and/or intolerant to HU, with a 24-month follow-up. The primary objective was to describe the profile and disease burden of PV patients. RESULTS: In the 350 enrolled patients, 70% were >60 years old. Most patients (59.4%) had received ≥1 phlebotomy in the 12 months prior to the first dose of ruxolitinib. Overall, 68.2% of patients achieved hematocrit control with 92.3% patients having hematocrit <45% and 35.4% achieved hematologic remission at month 24. 85.1% of patients had no phlebotomies during the study. Treatment-related adverse events were reported in 54.3% of patients and the most common event was anemia (22.6%). Of the 10 reported deaths, two were suspected to be study drug-related. CONCLUSION: This study demonstrates that ruxolitinib treatment in PV maintains durable hematocrit control with a decrease in the number of phlebotomies in the majority of patients and was generally well tolerated.

Risk factors for thromboembolic events in patients with paroxysmal nocturnal hemoglobinuria (PNH): a nested case-control study in the International PNH Registry.

The objective of this analysis was to identify risk factors for thromboembolic events (TE) in patients with paroxysmal nocturnal hemoglobinuria (PNH) who were not treated with C5 inhibitors. Patients with PNH and a history of ≥ 1 TE at enrollment in the International PNH Registry (NCT01374360; registration date, January 2011) were each matched with up to 5 patients without TE. Multivariable analysis was performed with the following variables: percentage glycosylphosphatidylinositol (GPI)-negative cells, high disease activity (HDA), non-TE major adverse vascular event history, and recent anticoagulation. Of 2541 eligible patients, 57 with TE and 189 matched controls were analyzed. Multivariable analysis (odds ratio [95% CI]) identified the following factors as being associated with increased thrombotic risk: patients with no history of TE (with recent anticoagulation, 9.30 [1.20-72.27]), patients with history of TE (with recent anticoagulation, 8.91 [0.86-92.62]; without recent anticoagulation, 5.33 [0.26-109.57]), patients with ≥ 30% GPI-negative granulocytes (≥ 30% to < 50%, 4.94 [0.54-45.32]; ≥ 50%, 1.97 [0.45-8.55]), or patients with lactate dehydrogenase (LDH) ratio ≥ 1.5 × upper limit of normal (ULN) plus ≥ 2 HDA criteria (2-3 criteria, 3.18 [0.44-23.20]; ≥ 4 criteria, 3.60 [0.38-33.95]). History of TE, ≥ 30% GPI-negative granulocytes, and LDH ratio ≥ 1.5 × ULN with ≥ 2 HDA criteria are TE risk factors for patients with PNH. These findings will aid physicians by providing important clinical and laboratory risk factors that can be used to identify and manage patients with PNH who are at risk of developing TE.

Early high antibody titre convalescent plasma for hospitalised COVID-19 patients: DAWn-plasma.

BACKGROUND: Several randomised clinical trials have studied convalescent plasma for coronavirus disease 2019 (COVID-19) using different protocols, with different severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralising antibody titres, at different time-points and severities of illness. METHODS: In the prospective multicentre DAWn-plasma trial, adult patients hospitalised with COVID-19 were randomised to 4 units of open-label convalescent plasma combined with standard of care (intervention group) or standard of care alone (control group). Plasma from donors with neutralising antibody titres (50% neutralisation titre (NT50)) ≥1/320 was the product of choice for the study. RESULTS: Between 2 May 2020 and 26 January 2021, 320 patients were randomised to convalescent plasma and 163 patients to the control group according to a 2:1 allocation scheme. A median (interquartile range) volume of 884 (806-906) mL) convalescent plasma was administered and 80.68% of the units came from donors with neutralising antibody titres (NT50) ≥1/320. Median time from onset of symptoms to randomisation was 7 days. The proportion of patients alive and free of mechanical ventilation on day 15 was not different between both groups (convalescent plasma 83.74% (n=267) versus control 84.05% (n=137)) (OR 0.99, 95% CI 0.59-1.66; p=0.9772). The intervention did not change the natural course of antibody titres. The number of serious or severe adverse events was similar in both study arms and transfusion-related side-effects were reported in 19 out of 320 patients in the intervention group (5.94%). CONCLUSIONS: Transfusion of 4 units of convalescent plasma with high neutralising antibody titres early in hospitalised COVID-19 patients did not result in a significant improvement of clinical status or reduced mortality.

Improvement of transfusion practice and reduction in red blood cell utilization in Belgian hospitals: Results of a national survey and benchmarking.

BACKGROUND AND OBJECTIVES: Belgian health authorities launched a national platform in 2011 to improve the quality of transfusion practices and blood use in Belgian hospitals. No data were available about the quality of hospital transfusion practice at the national level. MATERIALS AND METHODS: Three consecutive national surveys (2012, 2014 and 2016) were performed in all 111 Belgian hospitals to assess the degree of implementation of standards in four process domains related to red blood cell (RBC) transfusion: general quality aspects, ordering of RBC, electronic traceability and reporting of adverse events. The surveys were part of a methodology based on informing, feedback and benchmarking. Responses to questions were analysed semi-quantitatively, and hospitals could score 10 points on each of the domains. RESULTS: The proportion of hospitals scoring below 5 per domain decreased from 16%, 70%, 14% and 11% (2012) to 2%, 17%, 1% and 1% (2016), respectively. Similarly, scores above 7.5 increased from 25%, 1%, 23% and 36% (2012) to 64%, 30%, 68% and 81% (2016), respectively. In 2016, overall quality of transfusion practices, including the four pre-specified domains, improved continuously with an average total score (max = 40) increasing from 24.2 to 30.5 (p = 0.0005). In addition, there was a decrease in the number of distributed and transfused RBC per 1000 population between 2011 and 2019 from 47.0 to 36.5 and 43.5 to 36.1, respectively. CONCLUSION: These data show that the applied methodology was a powerful tool to improve quality of transfusion practices and to optimize utilization of RBC at the national level.

Phase III MANIFEST-2: pelabresib + ruxolitinib vs placebo + ruxolitinib in JAK inhibitor treatment-naive myelofibrosis.

Myelofibrosis (MF) is a clonal myeloproliferative neoplasm, typically associated with disease-related symptoms, splenomegaly, cytopenias and bone marrow fibrosis. Patients experience a significant symptom burden and a reduced life expectancy. Patients with MF receive ruxolitinib as the current standard of care, but the depth and durability of responses and the percentage of patients achieving clinical outcome measures are limited; thus, a significant unmet medical need exists. Pelabresib is an investigational small-molecule bromodomain and extraterminal domain inhibitor currently in clinical development for MF. The aim of this article is to describe the design of the ongoing, global, phase III, double-blind, placebo-controlled MANIFEST-2 study evaluating the efficacy and safety of pelabresib and ruxolitinib versus placebo and ruxolitinib in patients with JAKi treatment-naive MF. Clinical Trial Registration: NCT04603495 (ClinicalTrials.gov).

Myelofibrosis (MF) is a rare type of blood cancer that interferes with the process of blood cell production by the bone marrow. In patients with MF, the bone marrow becomes overactive, leading to scarring and subsequently a lack of healthy blood cells being produced. The main symptoms of MF include anemia, fatigue, weakness and pain or discomfort in the abdomen. MF is associated with a shortened life expectancy. The current go-to treatment for MF is ruxolitinib. However, ruxolitinib has shown limited efficacy in improving clinical symptoms long term; so, new safe and effective treatments are needed. Pelabresib is a novel drug currently in clinical development for treating MF. The aim of this article is to describe the design of the ongoing, global phase III MANIFEST-2 study. MANIFEST-2 is evaluating the efficacy and safety of pelabresib and ruxolitinib versus placebo and ruxolitinib in patients with MF.

Clinical outcomes in patients with Philadelphia chromosome-positive leukemia treated with ponatinib in routine clinical practice-data from a Belgian registry.

Data on clinical use of ponatinib are limited. This prospective registry aimed to evaluate outcomes of ponatinib treatment in routine practice over 3 years (2016-2019) in Belgium (NCT03678454). Patients with chronic myeloid leukemia (CML) or Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL) were treated with ponatinib per current label. Fifty patients (33 CML and 17 Ph+ ALL) were enrolled. Fifty-five percent of CML and 29% of Ph+ ALL patients had received ≥3 prior tyrosine kinase inhibitors (TKIs). Reasons for starting ponatinib were intolerance (40%), relapse or refractoriness (28%) to previous TKIs, progression (16%), or T315I mutation (16%). Median follow-up was 15 months for CML and 4.5 months for Ph+ ALL patients. Best response was a major molecular response in 58% of CML and 41% of Ph+ ALL patients. Of 20 patients who started ponatinib due to intolerance to previous TKIs, 9 (64%) CML and 4 (67%) Ph+ ALL achieved a major molecular response. Three-year estimates of overall survival were 85.3% and 85.6%, respectively, in CML and Ph+ ALL patients; estimated progression-free survival was 81.6% and 48.9%. Adverse reactions were reported in 34 patients (68%); rash (26%) and dry skin (10%) were most common. Reported cardiovascular adverse reactions included vascular stenosis (3), arterial hypertension (2), chest pain (1), palpitations (1), and vascular occlusion (1). This Belgian registry confirms results from the PACE clinical trial and supports routine ponatinib use in CML and Ph+ ALL patients who are resistant or intolerant to previous TKIs or with the T315I mutation.

Recanalization of portal axis after cavoportal hemitransposition in a liver transplant recipient with complete splanchnic thrombosis.

BACKGROUND: Diffuse splanchnic thrombosis may render standard LTx difficult or even technically impossible. A 19-year-old woman with acute-on-chronic Budd-Chiari syndrome and complete splanchnic thrombosis underwent conventional LTx. Only limited anatomical portal inflow could be restored, and urgent re-transplantation for recurrent splanchnic vein thrombosis became necessary. METHODS: At re-transplant, and in addition to the reestablishment of some portal inflow through the preserved original porto (native)-portal (graft) connection, a cavoportal shunt was created (first partial via 30% tapering of the vena cava, but eventually complete by total occlusion of the vena cava). RESULTS: The postoperative course was then uneventful, and interestingly, the native portomesenteric axis gradually reopened. Two years post-transplant, the liver graft is perfused via both physiological and non-physiological sources. Liver function is normal. There is no IVC syndrome and no residual PHT. She is leading a normal life. CONCLUSION: Creation of CPHT, in addition to the preservation of portal inflow from the native splanchnic system, should be considered in patients with diffuse splanchnic thrombosis, when sufficient physiological portal inflow cannot be restored at the time of LTx, but in whom the splanchnic circulation may reopen up later.

Polycythemia vera and hydroxyurea resistance/intolerance: a monocentric retrospective analysis.

Hydroxyurea (HU) resistance or intolerance occurs in 15 to 24% of patients with polycythemia vera (PV). Resistance to HU is associated with a shortened life expectancy, intolerance has no prognostic value. We assessed the occurrence of HU resistance or intolerance comparing the original (ELNo) versus the modified European Leukemia Net (ELNm) criteria as applied in recent large clinical trials including PV patients. We retrospectively analyzed 106 patients with PV treated with HU at the University Hospitals of Leuven between 1990 and 2016 for occurrence of HU resistance/intolerance when using both ELNo as ELNm. After a mean duration of treatment of 5.1 years, when applying the ELNo 20.7% of patients had shown resistance or intolerance to HU in comparison to 39.6% when using the ELNm. When using the ELNo 4.7% of patients were resistant to HU versus 23.6% when applying the ELNm. In total, 16.0% of patients were HU intolerant. This rate was identical when using both ELNo and ELNm. 20.7% of PV patients were considered as HU-resistant or intolerant when using the original ELN criteria. However, when applying the modified ELN criteria 39.6% of PV patients were resistant or intolerant to HU. In our hands, no patient received a minimum dose of 2 g HU a day, as such the ELNm seem better adapted for daily clinical use. However, the prognostic value of HU-resistance in PV, when defined by the ELNm, still needs to be confirmed.

Interruption or Discontinuation of Tyrosine Kinase Inhibitor Treatment in Chronic Myeloid Leukaemia: A Retrospective Cohort Study (SPARKLE) in Belgium.

OBJECTIVES: To assess interruptions/discontinuations of tyrosine kinase inhibitor (TKI) treatment in Belgian patients with chronic myeloid leukaemia (CML). METHODS: This retrospective study included patients with TKI interruptions/discontinuations of ≥4 continuous weeks (no clinical trial context) between May 2013 and May 2016. Data collection took place between October 2016 and February 2017. RESULTS: All 60 participants (69 interruptions/discontinuations) had chronic-phase CML and 75% had at least a major molecular response (≥MMR) at interruption/discontinuation. Most interruptions/discontinuations occurred while on imatinib (36/69; 49%) and dasatinib (20/69; 29%). Most interruptions/discontinuations occurred due to side effects/intolerance (46/69; 67%); other reasons included a wish to conceive (6/69; 9%) and attempts to achieve treatment-free remission (TFR) (6/69; 9%). Interruptions due to side effects occurred later for imatinib- or dasatinib-treated patients than for those on nilotinib or ponatinib. Treatment was re-initiated in 62% (43/69) of cases. Most interruptions caused by side effects/intolerance were followed by treatment changes. All 4 patients with ≥MR 4.5 at interruption/discontinuation and ≥11-month follow-up who had not restarted treatment maintained the response. CONCLUSION: Although TKIs are used for long-term CML treatment, physicians sometimes recommend interruptions/discontinuations. In this study, interruptions/discontinuations were mainly caused by side effects or intolerance, rather than TFR attempts.

Ruxolitinib for the treatment of inadequately controlled polycythemia vera without splenomegaly: 80-week follow-up from the RESPONSE-2 trial.

RESPONSE-2 is a phase 3 study comparing the efficacy and safety of ruxolitinib with the best available therapy (BAT) in hydroxyurea-resistant/hydroxyurea-intolerant polycythemia vera (PV) patients without palpable splenomegaly. This analysis evaluated the durability of the efficacy and safety of ruxolitinib after patients completed the visit at week 80 or discontinued the study. Endpoints included proportion of patients achieving hematocrit control (< 45%), proportion of patients achieving complete hematologic remission (CHR) at week 28, and the durability of hematocrit control and CHR. At the time of analysis, 93% (69/74) of patients randomized to ruxolitinib were receiving ruxolitinib; while in the BAT arm, 77% (58/75) of patients crossed over to ruxolitinib after week 28. No patient remained on BAT by week 80. Among patients who achieved a hematocrit response at week 28, the probability of maintaining response up to week 80 was 78% in the ruxolitinib arm. At week 80, durable CHR was achieved in 18 patients (24%) in the ruxolitinib arm versus 2 patients (3%) in the BAT arm. The safety profile of ruxolitinib was consistent with previous reports. These data support that ruxolitinib treatment should be considered also as a standard of care for hydroxyurea-resistant/hydroxyurea-intolerant PV patients without palpable splenomegaly.

Diagnosis and management of PNH: Review and recommendations from a Belgian expert panel.

Despite its considerable morbidity and mortality, paroxysmal nocturnal haemoglobinuria (PNH) is still underdiagnosed. Patients with PNH can suffer from cardiovascular, gastrointestinal, neurological or haematological symptoms and refer to several specialists. The aim of this paper is to review the diagnosis and the management of PNH patients, with the primary focus on identifying high-risk groups. Additionally, the implementation and prognostic value of the defined high-risk groups will be commented on and the management of PNH patients is discussed from a Belgian perspective. Finally, based on the available data, recommendations are provided. Eculizumab is a potent C5 complement inhibitor and reduces intravascular haemolysis and thrombosis in PNH patients and improves their quality of life. As thrombosis is the main cause of death in PNH patients, identifying high-risk PNH patients in need of therapy is essential. Currently, novel complement inhibitors are in development and the first data seem promising. Another challenge in PNH is to identify new markers to assess the thrombotic risk to achieve a better risk-based prophylactic anti-thrombotic management. Finally, because of the low prevalence of the disease, PNH patients should be included in the prospective PNH registry, which will offer new insights on the natural course of the disease and the impact of treatment of PNH.

Disease and treatment characteristics of polycythemia vera patients in Belgium: Results from a scientific survey.

OBJECTIVE: The current survey aimed to gather predefined disease parameters and treatment strategies to characterize the polycythemia vera (PV) patient population in Belgium. METHODS: Cross-sectional data from PV patients, seen at least once between May 2014 and May 2015 at 10 sites in Belgium, were collected in aggregated form and analyzed descriptively and quantitatively. RESULTS: Data from 343 PV patients were collected. Of these, 174 (50.7%) were male and 256 (74.6%) were ≥60 years of age. Ninety-two (26.8%) had a prior history of thrombotic events. Considerable proportions of patients had increased hematological parameters (hematocrit > 45% [31.2%], leukocytes > 10 × 109 /L [33.3%], and platelet > 400 × 109 /L [38.2%]). Most patients had non-palpable spleen (284, 87.7%) and no phlebotomies during the past 6 months (197, 57.4%). Low-dose aspirin was given as thrombosis prophylaxis in 249 (72.6%) patients, while 232 (67.6%) received hydroxyurea (HU) as cytoreductive treatment. Forty-one patients (12.0%) were reported as resistant and/or intolerant to HU. Seventeen patients (5.0%) received ruxolitinib in the context of clinical trials. CONCLUSION: This survey provides better insight into the characteristics of Belgian PV patients and currently used treatment strategies. It shows that 232 (67.6%) PV patients continue to receive HU despite being potentially HU-resistant.

Ruxolitinib for the treatment of inadequately controlled polycythaemia vera without splenomegaly (RESPONSE-2): a randomised, open-label, phase 3b study.

BACKGROUND: In the pivotal RESPONSE study, ruxolitinib, a Janus kinase (JAK)1 and JAK2 inhibitor, was superior to best available therapy at controlling haematocrit and improving splenomegaly and symptoms in patients with polycythaemia vera with splenomegaly who were inadequately controlled with hydroxyurea. In this study, we assessed the efficacy and safety of ruxolitinib in controlling disease in patients with polycythaemia vera without splenomegaly who need second-line therapy. METHODS: RESPONSE-2 is a randomised, open-label, phase 3b study assessing ruxolitinib versus best available therapy in patients with polycythaemia vera done in 48 hospitals or clinics across 12 countries in Asia, Australia, Europe, and North America. Eligible patients (aged ≥18 years) with polycythaemia vera, no palpable splenomegaly, and hydroxyurea resistance or intolerance were stratified by their hydroxyurea therapy status (resistance vs intolerance) and randomly assigned (1:1) by an interactive response technology provider using a validated system to receive either oral ruxolitinib 10 mg twice daily or investigator-selected best available therapy (hydroxyurea [at the maximum tolerated dose], interferon or pegylated interferon, pipobroman, anagrelide, approved immunomodulators, or no cytoreductive treatment). Investigators and patients were not masked to treatment assignment; however, the study sponsor was masked to treatment assignment until database lock. The primary endpoint was the proportion of patients achieving haematocrit control at week 28. Analyses were done according to an intention-to-treat principle, including data from all patients randomly assigned to treatment. This study is registered with ClinicalTrials.gov (NCT02038036) and is ongoing but not recruiting patients. FINDINGS: Between March 25, 2014, and Feb 11, 2015, of 173 patients assessed for eligibility, 74 patients were randomly assigned to receive ruxolitinib and 75 to receive best available therapy. At randomisation, best available therapy included hydroxyurea (37 [49%] of 75 in the best available therapy group), interferon or pegylated interferon (ten [13%] of 75), pipobroman (five [7%] of 75), lenalidomide (one [1%] of 75), no treatment (21 [28%] of 75), and other (one [1%] of 75). Haematocrit control was achieved in 46 (62%) of 74 ruxolitinib-treated patients versus 14 (19%) of 75 patients who received best available therapy (odds ratio 7·28 [95% CI 3·43-15·45]; p<0·0001). The most frequent haematological adverse events of any grade were anaemia (ten [14%] of 74 in the ruxolitinib group vs two [3%] of 75 in the best available therapy group) and thrombocytopenia (two [3%] vs six [8%]). No cases of grade 3-4 anaemia or thrombocytopenia occurred with ruxolitinib; one patient (1%) reported grade 3-4 anaemia and three patients (4%) reported grade 3-4 thrombocytopenia in the group receiving best available therapy. Frequent grade 3-4 non-haematological adverse events were hypertension (five [7%] of 74 vs three [4%] of 75) and pruritus (0 of 74 vs two [3%] of 75). Serious adverse events occurring in more than 2% of patients in either group, irrespective of cause, included thrombocytopenia (none in the ruxolitinib group vs two [3%] of 75 in the best available therapy group) and angina pectoris (two [3%] of 74 in the ruxolitinib group vs none in the best available therapy group). Two deaths occurred, both in the best available therapy group. INTERPRETATION: RESPONSE-2 met its primary endpoint. The findings of this study indicate that ruxolitinib could be considered a standard of care for second-line therapy in this post-hydroxyurea patient population. FUNDING: Novartis.

The transcription factor GATA1 regulates NBEAL2 expression through a long-distance enhancer.

Gray platelet syndrome is named after the gray appearance of platelets due to the absence of α-granules. It is caused by recessive mutations in NBEAL2, resulting in macrothrombocytopenia and myelofibrosis. Though using the term gray platelets for GATA1 deficiency has been debated, a reduced number of α-granules has been described for macrothrombocytopenia due to GATA1 mutations. We compared platelet size and number of α-granules for two NBEAL2 and two GATA1-deficient patients and found reduced numbers of α-granules for all, with the defect being more pronounced for NBEAL2 deficiency. We further hypothesized that the granule defect for GATA1 is due to a defective control of NBEAL2 expression. Remarkably, platelets from two patients, and Gata1-deficient mice, expressed almost no NBEAL2. The differentiation of GATA1 patient-derived CD34+ stem cells to megakaryocytes showed defective proplatelet and α-granule formation with strongly reduced NBEAL2 protein and ribonucleic acid expression. Chromatin immunoprecipitation sequencing revealed 5 GATA binding sites in a regulatory region 31 kb upstream of NBEAL2 covered by a H3K4Me1 mark indicative of an enhancer locus. Luciferase reporter constructs containing this region confirmed its enhancer activity in K562 cells, and mutagenesis of the GATA1 binding sites resulted in significantly reduced enhancer activity. Moreover, DNA binding studies showed that GATA1 and GATA2 physically interact with this enhancer region. GATA1 depletion using small interfering ribonucleic acid in K562 cells also resulted in reduced NBEAL2 expression. In conclusion, we herein show a long-distance regulatory region with GATA1 binding sites as being a strong enhancer for NBEAL2 expression.

Anaplastic lymphoma kinase-positive anaplastic large cell lymphoma with the variant RNF213-, ATIC- and TPM3-ALK fusions is characterized by copy number gain of the rearranged ALK gene.

Anaplastic lymphoma kinase (ALK)-positive anaplastic large cell lymphoma is characterized by 2p23/ALK aberrations, including the classic t(2;5)(p23;q35)/NPM1-ALK rearrangement present in ~80% of cases and several variant t(2p23/ALK) occurring in the remaining cases. The ALK fusion partners play a key role in the constitutive activation of the chimeric protein and its subcellular localization. Using various molecular technologies, we have characterized ALK fusions in eight recently diagnosed anaplastic large cell lymphoma cases with cytoplasmic-only ALK expression. The identified partner genes included EEF1G (one case), RNF213/ALO17 (one case), ATIC (four cases) and TPM3 (two cases). Notably, all cases showed copy number gain of the rearranged ALK gene, which is never observed in NPM1-ALK-positive lymphomas. We hypothesized that this could be due to lower expression levels and/or lower oncogenic potential of the variant ALK fusions. Indeed, all partner genes, except EEF1G, showed lower expression in normal and malignant T cells, in comparison with NPM1 In addition, we investigated the transformation potential of endogenous Npm1-Alk and Atic-Alk fusions generated by clustered regularly interspaced short palindromic repeats/Cas9 genome editing in Ba/F3 cells. We found that Npm1-Alk has a stronger transformation potential than Atic-Alk, and we observed a subclonal gain of Atic-Alk after a longer culture period, which was not observed for Npm1-Alk Taken together, our data illustrate that lymphomas driven by the variant ATIC-ALK fusion (and likely by RNF213-ALK and TPM3-ALK), but not the classic NPM1-ALK, require an increased dosage of the ALK hybrid gene to compensate for the relatively low and insufficient expression and signaling properties of the chimeric gene.

A haemovigilance team provides both significant financial and quality benefits in a University Hospital.

BACKGROUND: Haemovigilance is the process of surveillance of blood transfusion procedures including unexpected hazards and reactions during the transfusion pathway in both donors and recipients. The haemovigilance team aims to increase blood transfusion safety and to decrease both morbidity and mortality in donors and recipients. The team collects data about transfusion reactions and incidents, instructs the involved health workers and assures the tracing of blood components. The haemovigilance team at the University Hospitals Leuven has played a pioneering role in the development of haemovigilance in Belgium Although the literature about safety and quality improvements by haemovigilance systems is abundant, there are no published data available measuring their financial impact in a hospital. Therefore, we studied the costs and returns of the haemovigilance team at the University Hospitals Leuven. MATERIAL AND METHODS: This study has a descriptive explorative design. Research of the current costs and returns of the haemovigilance team were based upon data from the Medical Administration of the hospital. Data were analyzed descriptively. RESULTS: The haemovigilance team of the University Hospitals Leuven is financially viable: the direct costs are covered by the annual financial support of the National Public Health Service. The indirect returns come from two important tasks of the haemovigilance team itself: correction of the electronic registration of administered blood component and improvement of the return of conform preserved blood components to the blood bank. DISCUSSION: Besides safety and quality improvement, which are obviously their main goals, the haemovigilance team also implies a financial benefit for the hospital.