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In this Perspective, we discuss criteria for defining a new disease entity or variant of a recognized disease or disorder. We do so in the context of the current topography of the BCR::ABL-negative myeloproliferative neoplasms (MPNs) where two new variants are reported: clonal megakaryocyte dysplasia with normal blood values (CMD-NBV) and clonal megakaryocyte dysplasia with isolated thrombocytosis (CMD-IT). The cardinal feature of these variants is bone marrow megakaryocyte hyperplasia and atypia corresponding the WHO histological criteria for primary myelofibrosis (myelofibrosis-type megakaryocyte dysplasia-MTMD). Persons with these new variants have a different disease course and features from others in the MPN domain. In a broader context we suggest myelofibrosis-type megakaryocyte dysplasia defines a spectrum of related MPN variants including CMD-NBV, CMD-IT, pre-fibrotic myelofibrosis and overt myelofibrosis, which differ from polycythemia vera and essential thrombocythemia. Our proposal needs external validation and we stress the need for a consensus definition of the megakaryocyte dysplasia which is the hallmark of these disorders.

The introduction of new therapeutic agents for multiple myeloma (MM), including proteasome inhibitors, immunomodulatory drugs, and monoclonal antibodies, has improved the outcomes of patients but, in parallel, has changed the frequency and epidemiology of thrombotic events. Thrombosis is now a significant cause of morbidity and mortality in MM patients, and optimal thromboprophylaxis is far from being reached. Moving from the recognition that the above issue represents an unmet clinical need, an expert panel assessed the scientific literature and composed a framework of recommendations for improving thrombosis control in patients who are candidates for active treatment for MM. The panel generated key clinical questions using the criterion of clinical relevance through a Delphi process. It explored four domains, i.e., thrombotic risk factors and risk stratification, primary thromboprophylaxis, management of acute thrombotic events, and secondary thromboprophylaxis. The recommendations issued may assist hematologists in minimizing the risk of thrombosis and guarantee adherence to treatment in patients with MM who are candidates for active treatment.

Introduction: In 1991, we reported 18 persons with a clinical-pathologic entity and termed atypical myeloproliferative disorder because they did not meet the contemporary diagnostic criteria for a myeloproliferative neoplasm. We sought to gain further knowledge on this disease entity. Methods: This retrospective cohort study included consecutive subjects registered in the database of the Center for the Study of Myelofibrosis in Pavia, Italy, from 1998 to 2020 (June), and diagnosed with atypical myeloproliferative disorder according to our adjudicated criteria. We studied clinical, histological, cytogenetic, and molecular covariates and risks of thrombosis, disease progression, and death. Data were compared with those of concurrent subjects with prefibrotic myelofibrosis. Results: Fifteen new subjects with atypical myeloproliferative disorder were identified. Seven were male. Median age was 50 years (IQR, 41–54 years). Thirteen were diagnosed with a synchronous symptomatic or incidentally detected thrombotic event. The bone marrow showed megakaryocyte hyperplasia with dysplasia. JAK2V617F was present in 10 subjects and CALR mutation in one. No other somatic mutations were identified in next generation sequencing. After a median follow-up of 101 months (IQR, 40–160 months), no subject had disease progression or blast transformation. Incidence of post-diagnosis or recurrent thrombosis was 3.9 events (95% confidence interval, 3.5–4.0) and 5.0 events (4.6–5.6) per 100 person-years. Features of subjects with atypical myeloproliferative disorder differed markedly from those of 546 subjects with prefibrotic myelofibrosis. Conclusion: Our data indicate that these 15 persons have a distinct myeloproliferative neoplasm. We propose naming this new disorder clonal megakaryocyte dysplasia with normal blood values.

In primary myelofibrosis, extra-domain A fibronectin (EDA-FN), the result of alternative splicing of FN gene, sustains megakaryocyte proliferation and confers a pro-inflammatory phenotype to bone marrow cell niches. In this work we assessed the levels of circulating EDA-FN in plasma samples of 122 patients with primary myelofibrosis. Patients with a homozygous JAK2V617F genotype displayed the higher level of plasma EDA-FN. Increased EDA-FN levels were associated with anemia, elevated high-sensitivity C-reactive protein, bone marrow fibrosis and splanchnic vein thrombosis at diagnosis. While no correlation was observed with CD34+ hematopoietic stem cell mobilization, elevated blood level of EDA-FN at diagnosis was a predictor of large splenomegaly (over 10 cm from the left costal margin) outcome. Thus, EDA-FN expression in primary myelofibrosis may represent the first marker of disease progression, and a novel target to treat splenomegaly.

This article presents the results of group discussion among an ad hoc constituted panel of experts aimed at identifying and addressing unmet clinical needs (UCNs) in the management of infectious risk associated with eculizumab or new terminal complement inhibitors (CIs) in paroxysmal nocturnal hemoglobinuria (PNH). With the Delphi technique, the most clinically relevant UCNs in PNH patients candidate to or on terminal CI were selected. They resulted to be: optimizing the infection prevention measures; developing non pharmacological infectious risk-mitigation strategies; improving the management of disease exacerbation during infectious complications. For each of these issues consensus opinions were provided and, when appropriate, proposals for advancement in clinical practice were addressed. The hope is that this comprehensive overview will serve to improve the practice of CIs therapy and inform the design and implementation of new studies in the field.

Not available.

Different bodies of evidence support the existence of a common origin of hematopoietic and endothelial lineages; moreover, recent studies have indicated the presence of a hemogenic endothelium and a common hemato-endothelial precursor both in the embryo and in the cord blood. Conversely, to our knowledge, there is no evidence of such bipotential cells in human postnatal tissues or blood. In this study, we investigated the presence and phenotype of "transitional" cells in different tissues of patients with primary myelofibrosis (PMF). Using confocal microscopy and flow cytometry, we identified a rare cell population in the bone marrow and spleen of patients with PMF, which coexpresses the endothelial marker CD144 (vascular endothelial (VE)-cadherin), the pan-hematopoietic marker CD45, the early myeloid marker CD33, and CD34, a common endothelial and hematopoietic antigen.

Introduction: About 15% of people with a myeloproliferative neoplasm (MPN) are identified as MPN, unclassifiable using the 2016 WHO classification. Methods: We tested whether persons with platelet concentration ≥450 × 10E+9/L, bone marrow megakaryocyte morphology typical of prefibrotic/early myelofibrosis (pre-MF), and no minor criteria of pre-MF should be classified as a distinct MPN subtype, clonal megakaryocyte dysplasia with isolated thrombocytosis (CMD-IT). Results: 139 subjects meet these criteria who we compared with primary myelofibrosis (PMF) including 402 with pre-MF and 521 with overt myelofibrosis. CMD-IT subjects were more likely female and younger. They had lower frequencies of JAK2V617F compared with persons with PMF (55% vs. 70%; p < 0.001) and higher frequencies of CALR mutations (37% vs. 17%; p < 0.001). They also had lower frequency of variations associated with JAK2V617F susceptibility, JAK2 46/1 (35% vs. 47%; p = 0.021), and VEGFA rs3025039 (12% vs. 17%; p = 0.030). Subjects with CMD-IT had lower incidences of thrombotic events compared with those with pre-MF (9.7% vs. 26%; p < 0.001) and longer survival (median, not reached vs. 23 years; HR = 0.34 (0.10, 0.30); p < 0.001). Conclusion: Our data indicate CMD-IT is a distinct MPN subtype and should be included in the classification of myeloid neoplasms.

The expression of the CXCR4 chemokine receptor on CD34-positive blood cells is reduced in persons with primary myelofibrosis (PMF). We analyzed the relevance of cytofluorimetric assessment of the percentage of CD34-positive blood cells that had a positive CXCR4 surface expression (CD34/CXCR4-se) in a large cohort of subjects with myeloproliferative neoplasms. Mean CD34/CXCR4-se was lower in subjects with PMF compared with those with essential thrombocythemia (ET) or polycythemia vera (PV). A cutoff value of 39% was associated with a diagnosis of pre-fibrotic PMF vs. ET with a positive predictive value of 97%. In PMF male sex, older age, and MPL mutation were independent correlates of reduced CD34/CXCR4-se and associated with a briefer interval to development of severe anemia, large splenomegaly, thrombocytopenia, leukopenia, elevated CD34-positive blood cells, blast transformation and death. We constructed a prognostic model including age >65 years, hemoglobin 50 × 106/L, and CD34/CXCR4-se

Approximately one-fourth of patients with essential thrombocythemia or primary myelofibrosis carry a somatic mutation of the calreticulin gene (CALR), the gene encoding for calreticulin. A 52-bp deletion (type I mutation) and a 5-bp insertion (type II mutation) are the most frequent genetic lesions. The mechanism(s) by which a CALR mutation leads to a myeloproliferative phenotype has been clarified only in part. We studied the interaction between calreticulin and store-operated calcium (Ca2+) entry (SOCE) machinery in megakaryocytes (Mks) from healthy individuals and from patients with CALR-mutated myeloproliferative neoplasms (MPNs). In Mks from healthy subjects, binding of recombinant human thrombopoietin to c-Mpl induced the activation of signal transducer and activator of transcription 5, AKT, and extracellular signal-regulated kinase 1/2, determining inositol triphosphate–dependent Ca2+ release from the endoplasmic reticulum (ER). This resulted in the dissociation of the ER protein 57 (ERp57)-mediated complex between calreticulin and stromal interaction molecule 1 (STIM1), a protein of the SOCE machinery that leads to Ca2+ mobilization. In Mks from patients with CALR-mutated MPNs, defective interactions between mutant calreticulin, ERp57, and STIM1 activated SOCE and generated spontaneous cytosolic Ca2+ flows. In turn, this resulted in abnormal Mk proliferation that was reverted using a specific SOCE inhibitor. In summary, the abnormal SOCE regulation of Ca2+ flows in Mks contributes to the pathophysiology of CALR-mutated MPNs. In perspective, SOCE may represent a new therapeutic target to counteract Mk proliferation and its clinical consequences in MPNs.

The introduction of new therapeutic agents for multiple myeloma (MM), including proteasome inhibitors, immunomodulatory drugs, and monoclonal antibodies, has improved the outcomes of patients but, in parallel, has changed the frequency and epidemiology of thrombotic events. Thrombosis is now a significant cause of morbidity and mortality in MM patients, and optimal thromboprophylaxis is far from being reached. Moving from the recognition that the above issue represents an unmet clinical need, an expert panel assessed the scientific literature and composed a framework of recommendations for improving thrombosis control in patients who are candidates for active treatment for MM. The panel generated key clinical questions using the criterion of clinical relevance through a Delphi process. It explored four domains, i.e., thrombotic risk factors and risk stratification, primary thromboprophylaxis, management of acute thrombotic events, and secondary thromboprophylaxis. The recommendations issued may assist hematologists in minimizing the risk of thrombosis and guarantee adherence to treatment in patients with MM who are candidates for active treatment.

Polycythaemia vera is associated with a reduced quality of life, a high rate of vascular events, and an intrinsic risk of disease evolution. The results of several randomised trials for the treatment of this disorder are now available, and both a new ropegylated formulation of interferon alfa-2b (ropeginterferon alfa-2b; 2018) and ruxolitinib (2015) have been approved in Europe. European LeukemiaNet (ELN) investigators have therefore deemed it appropriate to provide recommendations for the use of these drugs in clinical practice. An expert panel of 14 senior haematologists from ELN centres that had actively participated in previous ELN projects or relevant randomised trials, chaired by a member of the ELN Steering Committee, developed a list of clinical questions, and a methodologist established three patient, intervention, comparator, outcome (PICO) questions and systematically reviewed the evidence. Recommendations were approved by six Delphi consensus rounds and two virtual meetings (on Jan 26, 2021, and June 24, 2021). The expert panel recommended that patients with polycythaemia vera who are younger than 60 years and have not had previous thrombotic events should start cytoreductive drug therapy if at least one of the following criteria are fulfilled: strictly defined intolerance to phlebotomy, symptomatic progressive splenomegaly, persistent leukocytosis (15 × 10

Daratumumab is approved for newly diagnosed or relapsed/refractory multiple myeloma (MM). The use of daratumumab has improved patient outcomes but has changed the frequency and epidemiology of infections. However, the optimal approach to prophylaxis and supportive therapy for daratumumab-emergent infections is unknown and represents an unmet clinical need in MM. Consequently, an expert panel convened to compose recommendations for optimal infection control in patient candidates to or under daratumumab treatment for MM. Scientific evidence on infections secondary to daratumumab was evaluated, and a consensus was developed by group discussion for key questions selected according to the clinical relevance. The following key issues were addressed: infectious risk assessment and risk stratification, infection mitigation strategies, and management of infectious complications in patients with MM treated with daratumumab.

TO THE EDITOR: The discovery of prognostically informative mutations in patients with primary myelofibrosis (PMF)[1][1] prompted the development of mutation-enhanced risk scores.[2][2][⇓][3][⇓][4][⇓][5]-[6][6] Among these mutations, those in ASXL1 are invariably associated with adverse

The introduction of new therapeutic agents in chronic lymphocytic leukemia (CLL), including the new kinase inhibitors (KIs) ibrutinib and idelalisib, has changed the therapeutic landscape of the disease. The new KIs have also changed frequency and epidemiology of infections, that represent a major cause of morbidity and mortality of the disease. Hence, the great strides in the indications and use of new KIs need parallel amelioration of prophylaxis and supportive treatment for infections. Moving from the recognition that infection control represents an unmet need, the Italian Society of Hematology (SIE) convened a panel of experts who had published and/or expressed an interest in infection complications in CLL. The goal of the project was to provide practice recommendations for the management of the infectious complications of CLL during ibrutinib or idelalisib therapy. The present publication represents the results of a series of email correspondences and meetings held during 2017 and 2018. Three domains of infectious complications during KIs therapy for CLL were explored: risk assessment, risk management and risk monitoring. We hope these recommendations will help to minimize infectious adverse events, and we believe that an optimal management of them will be rewarded by better outcomes, and better quality of life.

Three randomized controlled trials and a meta-analysis reported lenalidomide given after high-dose therapy and an autologous hemopoietic cell transplantation is associated with increase in progression-free survival (PFS) and survival in persons with plasma cell myeloma (PCM). Based on these data, posttransplant lenalidomide is considered by many a standard-of-care in this setting. However, decisions on the use of new therapies should consider not only results of such trials and meta-analyses but also other factors including quality-of-evidence, anticipated desired and undesired effects of the drug, costs and feasibility of the therapy option. In this review, we critically analyzed evidence on posttransplant lenalidomide in PCM, and we identified criteria which should be considered in designating posttransplant lenalidomide the standard-of-care. Using Grading of Evidence, Assessment, Development and Evaluation (GRADE) approach we judged posttransplant lenalidomide improves PFS with high-quality evidence. However, we identified inconsistency and imprecision as limitations in the conclusions regarding a survival benefit rating the quality-of-evidence for a survival benefit moderate. We also highlighted inconsistency in claims of an increased risk of new cancers associated with posttransplant lenalidomide. We emphasize the need for a value-based reasoning which considers PFS and survival as well as health-related quality-of-life and costs. We conclude the decision to use posttransplant lenalidomide should be individualized based on pre- and posttransplant variables such as remission state, risk category and/or posttransplant measurable residual disease (MRD)-test results. Validity of these variables in estimating benefits and risks of posttransplant lenalidomide should be tested in randomized clinical trials.

In this paper, we present a review of critical concepts, and produce recommendations on management issues in cutaneous T-cell lymphomas (CTCLs) of adults. A panel of nine experts was selected for their expertise in research and clinical practice of CTCLs. During an initial meeting, the areas of major concern in the management of CTCLs were selected by generating and rank-ordering clinical key questions using the criterion of clinical relevance, through group discussion. Recommendations were achieved by multiple-step formalized procedures to reach a consensus after a comprehensive analysis of the scientific literature. The panel produced recommendations on how to facilitate the clinical suspicion of CTCL; indication of cutaneous biopsy; optimal histological diagnosis, immunohistochemistry and genetic markers; and staging pathway and up-to-date therapeutics (with particular focus on new treatments). The critical concept of integration of the different medical expertise in the management of the patients with CTCL was thoroughly examined. These recommendations are intended for use not only by expert centers but above all by "not experienced" dermatologists and hematologists as well as general practitioners.

Numerous studies have documented ultrastructural abnormalities in malignant megakaryocytes from bone marrow (BM) of myelofibrosis patients but the morphology of these cells in spleen, an important extramedullary site in this disease, was not investigated as yet. By transmission-electron microscopy, we compared the ultrastructural features of megakaryocytes from BM and spleen of myelofibrosis patients and healthy controls. The number of megakaryocytes was markedly increased in both BM and spleen. However, while most of BM megakaryocytes are immature, those from spleen appear mature with well-developed demarcation membrane systems (DMS) and platelet territories and are surrounded by platelets. In BM megakaryocytes, paucity of DMS is associated with plasma (thick with protrusions) and nuclear (dilated with large pores) membrane abnormalities and presence of numerous glycosomes, suggesting a skewed metabolism toward insoluble polyglucosan accumulation. By contrast, the membranes of the megakaryocytes from the spleen were normal but these cells show mitochondria with reduced crests, suggesting deficient aerobic energy-metabolism. These distinctive morphological features suggest that malignant megakaryocytes from BM and spleen express distinctive metabolic impairments that may play different roles in the pathogenesis of myelofibrosis.

Primary myelofibrosis (PMF) is a myeloproliferative neoplasm characterized by an excessive production of pro-inflammatory cytokines resulting in chronic inflammation and genomic instability. Besides the driver mutations in JAK2, MPL, and CALR genes, the deregulation of miRNA expression may also contribute to the pathogenesis of PMF. To this end, we recently reported the upregulation of miR-382-5p in PMF CD34+ cells. In order to unveil the mechanistic details of the role of miR-382-5p in pathogenesis of PMF, we performed gene expression profiling of CD34+ cells overexpressing miR-382-5p. Among the downregulated genes, we identified superoxide dismutase 2 (SOD2), which is a predicted target of miR-382-5p. Subsequently, we confirmed miR-382-5p/SOD2 interaction by luciferase assay and we showed that miR-382-5p overexpression in CD34+ cells causes the decrease in SOD2 activity leading to reactive oxygen species (ROS) accumulation and oxidative DNA damage. In addition, our data indicate that inhibition of miR-382-5p in PMF CD34+ cells restores SOD2 function, induces ROS disposal, and reduces DNA oxidation. Since the pro-inflammatory cytokine transforming growth factor-β1 (TGF-β1) is a key player in PMF pathogenesis, we further investigated the effect of TGF-β1 on ROS and miR-382-5p levels. Our data showed that TGF-β1 treatment enhances miR-382-5p expression and reduces SOD2 activity leading to ROS accumulation. Finally, inhibition of TGF-β1 signaling in PMF CD34+ cells by galunisertib significantly reduced miR-382-5p expression and ROS accumulation and restored SOD2 activity. As a whole, this study reports that TGF-β1/miR-382-5p/SOD2 axis deregulation in PMF cells is linked to ROS overproduction that may contribute to enhanced oxidative stress and inflammation. Our results suggest that galunisertib may represent an effective drug reducing abnormal oxidative stress induced by TGF-β1 in PMF patients. DATABASE LINKING: GEO: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE103464.

Splenic hematopoiesis is a major feature in the course of myelofibrosis (MF). In fact, the spleen of patients with MF contains malignant hematopoietic stem cells retaining a complete differentiation program, suggesting both a pivotal role of the spleen in maintaining the disease and a tight regulation of hematopoiesis by the splenic microenvironment, in particular by mesenchymal stromal cells (MSCs). Little is known about splenic MSCs (Sp-MSCs), both in normal and in pathological context. In this work, we have in vitro expanded and characterized Sp-MSCs from 25 patients with MF and 13 healthy subjects (HS). They shared similar phenotype, growth kinetics, and differentiation capacity. However, MF Sp-MSCs expressed significant lower levels of nestin, and favored megakaryocyte (Mk) differentiation in vitro at a larger extent than their normal counterpart. Moreover, they showed a significant upregulation of matrix metalloprotease 2 (MMP2) and fibronectin 1 (FN1) genes both at mRNA expression and at protein level, and, finally, developed genetic abnormalities which were never detected in HS-derived Sp-MSCs. Our data point toward the existence of a defective splenic niche in patients with MF that could be responsible of some pathological features of the disease, including the increased trafficking of CD34+ cells and the expansion of the megakaryocytic lineage.

Purpose To develop a prognostic system for transplantation-age patients with primary myelofibrosis (PMF) that integrates clinical, cytogenetic, and mutation data. Patients and Methods The study included 805 patients with PMF age ≤ 70 years recruited from multiple Italian centers and the Mayo Clinic (Rochester, MN), forming two independent learning and validation cohorts. A Cox multivariable model was used to select from among a list of 22 variables those that were predictive of overall survival (OS). Integrated clinical and genetic prognostic models with (MIPSS70-plus) or without (MIPSS70) cytogenetic information were developed. Results Multivariable analysis identified the following as significant risk factors for OS: hemoglobin < 100 g/L, leukocytes > 25 × 109/L, platelets < 100 × 109/L, circulating blasts ≥ 2%, bone marrow fibrosis grade ≥ 2, constitutional symptoms, absence of CALR type-1 mutation, presence of high–molecular risk mutation (ie, ASXL1, EZH2, SRSF2, IDH1/ 2), and presence of two or more high–molecular risk mutations. By assigning hazard ratio (HR)–weighted points to these variables, three risk categories were delineated for the MIPSS70 model; 5-year OS was 95% in low-risk, 70% in intermediate-risk, and 29% in high-risk categories, corresponding to median OS of 27.7 years (95% CI, 22 to 34 years), 7.1 years (95% CI, 6.2 to 8.1 years), and 2.3 years (95% CI, 1.9 to 2.7 years), respectively. In the MIPSS70-plus model, which included cytogenetic information, four risk categories were delineated, with 5-year OS of 91% in low-risk, 66% in intermediate-risk (HR, 3.2; 95% CI, 1.9 to 5.2), 42% in high-risk (HR, 6.4; 95% CI, 4.1 to 10.0), and 7% very high–risk categories (HR, 17.0; 95% CI, 9.8 to 29.2). Both models remained effective after inclusion of older patients in the analysis. Conclusion MIPSS70 and MIPSS70-plus provide complementary systems of risk stratification for transplantation-age patients with PMF and integrate prognostically relevant clinical, cytogenetic, and mutation data.

Background: Until recently, ruxolitinib (RUX) was the only drug approved for treatment of intermediate- or high-risk myelofibrosis (MF). Many patients discontinue RUX due to lack of response, loss of efficacy, or intolerance. Fedratinib (INREBIC) is an oral, selective kinase inhibitor with activity against mutant and wild-type JAK2 and FLT3. Fedratinib is approved in the United States, Canada, European Union, United Kingdom, and elsewhere as front-line therapy for treatment of patients with JAK-inhibitor-naïve MF and those previously treated with RUX. In the single-arm, phase 2 JAKARTA2 trial of fedratinib 400 mg/day (starting dose) in patients with MF relapsed, refractory, or intolerant to prior RUX, 31% of pts achieved a spleen volume response and 27% achieved a symptom response with fedratinib (Harrison, Am J Hematol 2020). Gastrointestinal (GI) events were among the most common adverse events (AEs) reported during fedratinib treatment in JAKARTA2. A clinical hold was placed on fedratinib in November 2013 due to suspected cases of Wernicke encephalopathy (WE); the hold was later lifted but long-term data, including survival outcomes, are limited for fedratinib. Two phase 3 trials, FREEDOM (NCT03755518) and FREEDOM2 (NCT03952039) are ongoing to assess the safety and efficacy of fedratinib in patients with MF previously treated with RUX. Unlike earlier studies of fedratinib, these two trials prospectively include GI-directed mitigation strategies and thiamine monitoring and supplementation. Preliminary data from the single-arm FREEDOM trial show reduced incidence of GI AEs with these mitigation strategies (Gupta, CLML 2020). Here we describe the study design and methods for the FREEDOM2 trial, the first trial to compare fedratinib vs. active MF therapies. Study Design and Methods: The ongoing multicenter, open-label, randomized, phase 3 FREEDOM2 trial is enrolling patients with MF in the European Union, China, Russia, South Korea, and Australia. Key eligibility criteria include age ≥ 18 years; primary, post-polycythemia vera (post-PV), or post-essential thrombocythemia (post-ET) MF; DIPSS Intermediate-2 or High-risk disease; splenomegaly, defined as spleen volume ≥ 450 cm 3 by MRI/CT and palpable spleen measuring ≥ 5 cm below the left costal margin; ECOG PS score ≤ 2; and platelet count ≥ 50 ×10 9/L (Figure). Patients must have been relapsed or refractory to prior RUX treatment for ≥ 3 months, or intolerant to RUX after ≥ 28 days of treatment (i.e., developed RBC transfusion dependence [≥ 2 units/month for 2 months] or grade ≥ 3 thrombocytopenia, anemia, or hematoma/hemorrhage). Eligible patients are randomized 2:1 to receive fedratinib 400 mg/day or investigator-selected best available therapy (BAT), including RUX or other approved JAK inhibitors, in repeated 28-day treatment cycles (Figure). Spleen volume is assessed by MRI/CT scan at screening, end of cycles (EOC) 3, 6, 12, 18, 24, and at the end of treatment. The primary endpoint is spleen volume response rate (SVRR), the proportion of patients in each arm achieving a ≥ 35% reduction in spleen volume from baseline at EOC6, and the key secondary endpoint is MF symptom response rate, the proportion of patients achieving a ≥ 50% reduction from baseline in total symptom score (TSS) on the Myelofibrosis Symptom Assessment Form (MFSAF) at EOC6. Other secondary endpoints include durations of spleen volume and symptom responses, reductions in spleen size by palpation, overall and progression-free survival, effectiveness of the risk mitigation strategies for GI AEs and WE, patient-reported health-related quality of life (HRQoL) measures, and safety. Exploratory endpoints include relationships between fedratinib efficacy and prognostic markers (eg, mutations) and pharmacodynamic activity (eg, circulating cytokines, hematopoietic cell profiling). Patients in the BAT arm can cross-over to receive fedratinib after EOC6, or before EOC6 if the patient experiences confirmed progression of splenomegaly. As the first trial to compare fedratinib against other active therapies for MF, results from FREEDOM2 are eagerly anticipated. As of June 2021, 135 patients had been randomized, with enrollment expected to be completed by the end of 2021. Figure 1 Figure 1. Disclosures Benevolo: BMS: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Speakers Bureau. Vannucchi: Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees. Harrison: CTI BioPharma: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Abbvie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Roche: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead Sciences: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Geron: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sierra Oncology: Honoraria; Incyte Corporation: Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Promedior: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Constellation Pharmaceuticals: Research Funding; Shire: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Keros: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Galacteo: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AOP Orphan Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Loschi: CELGENE/BMS: Honoraria; AbbVie: Ended employment in the past 24 months, Honoraria; Gilead: Ended employment in the past 24 months, Honoraria; Novartis: Ended employment in the past 24 months, Honoraria; Servier: Ended employment in the past 24 months, Honoraria; MSD: Honoraria. Al-Ali: Incyte: Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding; Takeda: Consultancy; Pfizer: Consultancy; AbbVie: Consultancy, Honoraria. Bonifacio: Amgen: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees. Cervantes: Bristol Myers Squibb: Consultancy, Honoraria. Wrobel: Novartis: Honoraria, Speakers Bureau; BMS: Honoraria, Speakers Bureau; Roche: Honoraria, Research Funding, Speakers Bureau; Janssen: Honoraria, Speakers Bureau; Takeda: Honoraria, Speakers Bureau; BeiGene: Honoraria, Speakers Bureau. Kiladjian: AbbVie: Consultancy; AOP Orphan: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Verstovsek: PharmaEssentia: Research Funding; Gilead: Research Funding; Genentech: Research Funding; Ital Pharma: Research Funding; Incyte Corporation: Consultancy, Research Funding; NS Pharma: Research Funding; Blueprint Medicines Corp: Research Funding; Celgene: Consultancy, Research Funding; CTI BioPharma: Research Funding; Protagonist Therapeutics: Research Funding; Promedior: Research Funding; Roche: Research Funding; AstraZeneca: Research Funding; Sierra Oncology: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Constellation: Consultancy; Pragmatist: Consultancy. Mesa: Promedior: Research Funding; Pharma: Consultancy; Gilead: Research Funding; CTI: Research Funding; CTI: Research Funding; Novartis: Consultancy; Sierra Oncology: Consultancy, Research Funding; Incyte Corporation: Consultancy, Research Funding; Celgene: Research Funding; Constellation Pharmaceuticals: Consultancy, Research Funding; AOP: Consultancy; Abbvie: Research Funding; La Jolla Pharma: Consultancy; Samus: Research Funding; Genentech: Research Funding. Rose: Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Gharpure: Bristol Myers Squibb: Current Employment. Hernandez: Bristol Myers Squibb: Current Employment. Zhang: Bristol Myers Squibb: Current Employment. Passamonti: BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AbbVie: Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

INTRODUCTION: Ruxolitinib (RUX) has demonstrated efficacy in patients (pts) with MF, but many pts discontinue (D/C) RUX due to loss of response or treatment (Tx)-related cytopenias. Fedratinib (FEDR) is an oral, selective JAK2 inhibitor approved for Tx of pts with MF, including those previously treated with RUX. In the single-arm, phase 2 JAKARTA2 trial of FEDR 400 mg/day (d) in pts with MF resistant/intolerant to prior RUX, 31% of pts achieved a spleen volume response and 27% achieved a symptom response with FEDR. The most common adverse events (AEs) in JAKARTA2 were gastrointestinal (GI) events, including diarrhea in 62% of pts, nausea in 56%, and vomiting in 41%. A temporary clinical hold was placed on FEDR in 2013 due to suspected cases of Wernicke's encephalopathy (WE). The ongoing, single-arm, phase 3b FREEDOM trial (NCT03755518) further evaluates the safety and efficacy of FEDR in pts with MF previously treated with RUX. Unlike JAKARTA2 and other early FEDR trials, FREEDOM includes prospective strategies for preventing or mitigating GI AEs, thiamine level decreases, and potential WE. We investigated the safety of FEDR 400 mg/d in FREEDOM, including the effectiveness of these strategies. METHODS: Eligible pts are aged ≥ 18 years with DIPSS-defined intermediate- or high-risk, primary or post-PV/ET MF, ECOG PS ≤ 2, platelet count ≥ 50 ×10 9/L, and spleen volume ≥ 450 cm 3 by MRI/CT or palpable spleen ≥ 5 cm below the left costal margin (LCM). Pts must have previously received RUX for ≥ 3 months (mo), or for ≥ 28d with development of RBC transfusion requirement (≥ 2 units/mo for 2 mo) or grade ≥ 3 thrombocytopenia, anemia, hematoma, or hemorrhage. All pts received FEDR 400 mg QD in continuous 28d cycles. AE mitigation strategies include prophylactic and symptomatic use of anti-nausea/vomiting and anti-diarrheal Tx, thiamine supplementation, FEDR dosing modifications, and administration of FEDR with food. All pts who received ≥ 1 FEDR dose were evaluated for safety. AEs were coded using MedDRA v24.0 and graded (G) by CTCAE v5.0. RESULTS: At data cutoff (April 9, 2021), 34 pts had been enrolled and 16 pts continued to receive FEDR. Reasons for FEDR Tx D/C in > 1 pt were lack of efficacy (n = 5) AEs (n=4; 1 was Tx-related [G3 thrombocytopenia]), disease progression (n = 2), pt decision (n = 2), and to undergo transplant (n = 2). Median FEDR Tx duration was 28.3 weeks (range 1.6-101.3); 20 pts (59%) had received > 6 Tx cycles and 14 pts (41%) completed > 12 cycles. Median average FEDR dose was 400 mg/d (range 298-400).At BL, median (range) age was 68.5 years (49-82), time from MF diagnosis was 3.4 years (0.1-17.4), and spleen size was 15 cm (3-31). Most pts (62%) had primary MF, and all pts had received ≥ 3 mo of prior RUX Tx; the most common reason for prior RUX D/C was loss of response/Tx failure (41%). During FEDR Tx, 22 pts (65%) received ondansetron and 11 (32%) received loperamide. GI AEs reported in > 10% of pts were constipation (47%), diarrhea (35%), nausea (26%), abdominal pain (24%), and vomiting (18%). The frequency of nausea, vomiting, and diarrhea decreased as Tx continued (Figure). Most GI AEs were G1/2 (including all events of nausea, vomiting and diarrhea) and there were no Tx-related G3/4 GI AEs. No pt required FEDR dose-reduction, interruption, or D/C due to a Tx-related GI AE Overall, Tx-related G3/4 AEs were reported in 11 pts (32%), including anemia in 7 pts (21%), and neutropenia, thrombocytopenia, and hyperkalemia in 2 pts each (6%). The cases of hyperkalemia occurred in the setting of acute renal failure or of relevant concomitant conditions. At BL, 1 pt had a thiamine level below the lower limit of normal (LLN; 70 nmol/L); thiamine level was normalized before the pt received FEDR Tx. Thiamine levels dropped below the LLN during FEDR Tx for 4 pts between cycles 2-3 (and for 1 pt at end of Tx); levels returned to normal for these pts with thiamine supplementation and did not require FEDR interruption or reduction. Five other pts received prophylactic thiamine supplementation. There were no reported cases of WE. CONCLUSIONS: FEDR was generally well tolerated. These data suggest the frequency and severity of GI AEs may be reduced via early implementation of GI prophylaxis. Use of GI-directed therapies may have increased the incidence of low-grade constipation. Monitoring thiamine levels before FEDR initiation and periodically during FEDR therapy is recommended. Figure 1 Figure 1. Disclosures Gupta: Pfizer: Consultancy; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Constellation Pharma: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; BMS-Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy; Incyte: Honoraria, Research Funding; Sierra Oncology: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Yacoub: Agios: Membership on an entity's Board of Directors or advisory committees; Acceleron Pharma: Membership on an entity's Board of Directors or advisory committees; CTI Biopharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Incyte: Speakers Bureau. Verstovsek: Gilead: Research Funding; Genentech: Research Funding; Ital Pharma: Research Funding; Promedior: Research Funding; NS Pharma: Research Funding; Celgene: Consultancy, Research Funding; CTI BioPharma: Research Funding; PharmaEssentia: Research Funding; Protagonist Therapeutics: Research Funding; Incyte Corporation: Consultancy, Research Funding; Blueprint Medicines Corp: Research Funding; Roche: Research Funding; Sierra Oncology: Consultancy, Research Funding; AstraZeneca: Research Funding; Novartis: Consultancy, Research Funding; Constellation: Consultancy; Pragmatist: Consultancy. Mesa: Novartis: Consultancy; CTI: Research Funding; Promedior: Research Funding; AOP: Consultancy; Sierra Oncology: Consultancy, Research Funding; La Jolla Pharma: Consultancy; Genentech: Research Funding; CTI: Research Funding; Celgene: Research Funding; Constellation Pharmaceuticals: Consultancy, Research Funding; Pharma: Consultancy; Incyte Corporation: Consultancy, Research Funding; Samus: Research Funding; Gilead: Research Funding; Abbvie: Research Funding. Harrison: BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Shire: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead Sciences: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Roche: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AOP Orphan Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Promedior: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Keros: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Geron: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Constellation Pharmaceuticals: Research Funding; Galacteo: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Abbvie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte Corporation: Speakers Bureau; Sierra Oncology: Honoraria; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; CTI BioPharma: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Kiladjian: Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; PharmaEssentia: Other: Personal fees; Taiho Oncology, Inc.: Research Funding; Incyte Corporation: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; AOP Orphan: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees. Fazal: Agios: Consultancy, Honoraria, Speakers Bureau; Amgen: Consultancy, Honoraria, Speakers Bureau; BMS: Consultancy, Honoraria, Speakers Bureau; Celgene: Consultancy, Honoraria, Speakers Bureau; Gilead: Consultancy, Honoraria, Speakers Bureau; GlaxoSmithKline: Consultancy, Honoraria, Speakers Bureau; Incyte: Consultancy, Honoraria, Speakers Bureau; Janssen: Consultancy, Honoraria, Speakers Bureau; Jazz Pharma: Consultancy, Honoraria, Speakers Bureau; Karyopharm: Consultancy, Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Sanofi Genzyme: Consultancy, Honoraria, Speakers Bureau; Stemline: Consultancy, Honoraria, Speakers Bureau; Taiho: Consultancy, Honoraria, Speakers Bureau; Takeda: Consultancy, Honoraria, Speakers Bureau; Tolero: Consultancy. Foltz: Incyte: Research Funding; CTI Biopharma: Research Funding; Constellation: Research Funding; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sierra: Research Funding; Amgen: Other: Spouse employment and equity ownership.. Miller: CTI: Consultancy, Research Funding; Incyte: Consultancy, Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria; Takeda: Honoraria, Speakers Bureau; Celgene: Speakers Bureau. Gharpure: Bristol Myers Squibb: Current Employment. Hernandez: Bristol Myers Squibb: Current Employment. Wang: Bristol Myers Squibb: Current equity holder in publicly-traded company, Ended employment in the past 24 months; VIR Biotechnology: Current equity holder in publicly-traded company. Talpaz: Imago: Consultancy; Celgene: Consultancy; Constellation: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Other: Grant/research support .

Simple Summary Disease progression and prognosis in PMF are usually associated with worsening of anemia, increase of circulating blasts, and, more recently, with the presence, in addition to the “classical” driver mutations, of JAK2, MPL, and CALR genes, as well as of cytogenetic and molecular abnormalities that have been incorporated into new genetically based prognostic scoring systems. We have recently focused our attention on new biological markers of the disease, namely sIL-2Rα, eNAMPT, and CXCR4 expression on circulating CD34+ cells and CCL2 and VEGF-A polymorphisms, which have turned out to be associated with disease evolution and patient survival. Here, we discuss the role that recently described biological markers of the disease play in PMF progression and prognosis. Abstract Primary myelofibrosis (PMF) is a myeloproliferative neoplasm due to the clonal proliferation of a hematopoietic stem cell. The vast majority of patients harbor a somatic gain of function mutation either of JAK2 or MPL or CALR genes in their hematopoietic cells, resulting in the activation of the JAK/STAT pathway. Patients display variable clinical and laboratoristic features, including anemia, thrombocytopenia, splenomegaly, thrombotic complications, systemic symptoms, and curtailed survival due to infections, thrombo-hemorrhagic events, or progression to leukemic transformation. New drugs have been developed in the last decade for the treatment of PMF-associated symptoms; however, the only curative option is currently represented by allogeneic hematopoietic cell transplantation, which can only be offered to a small percentage of patients. Disease prognosis is based at diagnosis on the classical International Prognostic Scoring System (IPSS) and Dynamic-IPSS (during disease course), which comprehend clinical parameters; recently, new prognostic scoring systems, including genetic and molecular parameters, have been proposed as meaningful tools for a better patient stratification. Moreover, new biological markers predicting clinical evolution and patient survival have been associated with the disease. This review summarizes basic concepts of PMF pathogenesis, clinics, and therapy, focusing on classical prognostic scoring systems and new biological markers of the disease.

We measured plasma levels of high-sensitivity C-reactive protein (hs-CRP) in 526 subjects with primary myelofibrosis (PMF). Thirty-eight percent had an elevated hs-CRP level (≥0.3mg/dL). Elevated hs-CRP levels were associated with a progressive disease phenotype, including anemia, high white blood cell count, low platelet count, increased splenomegaly, increased risk of blast transformation, and worse survival. Age≥52years, but no other demographic characteristics, was associated with an elevated hs-CRP level in multivariable logistic regression (odds ratio [OR], 4.29; 95% CI, 2.73-6.77; P

Splanchnic vein thrombosis (SVT) is one of the vascular complications of myeloproliferative neoplasms (MPN). We designed a phase 2 clinical trial to evaluate safety and efficacy of ruxolitinib in reducing splenomegaly and improving disease-related symptoms in patients with MPN-associated SVT. Patients diagnosed with myelofibrosis (12 cases), polycythemia vera (5 cases) and essential thrombocythemia (4 cases) received ruxolitinib for 24 weeks in the core study period. Spleen volume was assessed by magnetic resonance imaging (MRI) and splanchnic vein circulation by echo-Doppler analysis. Nineteen patients carried JAK2V617F, one had MPLW515L, and one CALRL367fs*46 mutation. Eighteen patients had spleno-portal-mesenteric thrombosis, two had Budd-Chiari syndrome, and one had both sites involved; 16 patients had esophageal varices. Ruxolitinib was well tolerated with hematological toxicities consistent with those of patients without SVT and no hemorrhagic adverse events were recorded. After 24 weeks of treatment, spleen volume reduction ≥35% by MRI was achieved by 6/21 (29%) patients, and a ≥50% spleen length reduction by palpation at any time up to week 24 was obtained by 13/21 (62%) patients. At week 72, 8 of the 13 (62%) patients maintained the spleen response by palpation. No significant effect of treatment on esophageal varices or in splanchnic circulation was observed. MPN-related symptoms, evaluated by MPN-symptom assessment form (SAF) TSS questionnaire, improved significantly during the first 4 weeks and remained stable up to week 24. In conclusion, this trial shows that ruxolitinib is safe in patients with MPN-associated SVT, and effective in reducing spleen size and disease-related symptoms.

Introduction: MF is a clonal stem cell disorder characterized by progressive bone marrow failure, extramedullary hematopoiesis, and debilitating constitutional symptoms. Approximately 60% of pts develop anemia within a year of diagnosis; pts who are transfusion dependent have worse survival and quality-of-life outcomes (Tefferi A, et al. Mayo Clin Proc 2012;87:25-33). Treatment for MF-associated anemia remains critically unaddressed, with no approved therapies available. Luspatercept is a first-in-class erythroid maturation agent that has been shown to increase hemoglobin and reduce transfusion burden in pts with myelodysplastic syndromes (Fenaux P, et al. N Engl J Med 2020;382:140-51) and β-thalassemia (Cappellini MD, et al. N Engl J Med 2020;382:1219-31). Pts have also achieved multiple response episodes with luspatercept (Fenaux P, et al. Blood 2019;134:841). We previously reported findings from the ongoing open-label, phase 2 ACE-536-MF-001 trial evaluating luspatercept in pts with MF-associated anemia (Gerds AT, et al. Blood 2019;134:557). Here we report updated study results, focusing on response in pts requiring RBC transfusions. Methods: 79 pts with MF and anemia were enrolled (Figure). Of these, 43 were requiring RBC transfusions, defined as 6-12 U/84 d prior to treatment, or 4-12 U/84 d for the 3 expansion cohort pts. Pts in Cohort 3B received a stable ruxolitinib (RUX) dose for ≥ 16 wks (40 wks for expansion cohort pts), whereas Cohort 2 pts did not (Figure). Pts in Cohort 3B received a median RUX dose of 20 mg/d (range 5-40) for a median of 73 wks (range 29-359) prior to treatment. Pts received luspatercept every 21 d at a starting dose of 1.0 mg/kg (1.33 mg/kg for 3 expansion cohort pts in Cohort 3B), with doses escalating up to 1.75 mg/kg; 81% and 50% of pts in Cohorts 2 and 3B escalated to 1.75 mg/kg. Primary endpoint for pts receiving transfusions was RBC transfusion-independence (RBC-TI) for ≥ 12 consecutive wks within the first 24 wks on study. However, as responses were observed that did not fall within the 24-wk response window, additional assessments were conducted for the entire treatment duration for RBC-TI and ≥ 50% decrease in RBC transfusions (minimum 4 RBC U decrease) from baseline. Intent-to-treat data were analyzed as of March 29, 2020. Results: Pts in Cohorts 2 and 3B received a median of 8 cycles of luspatercept (range 1-39), for a median (mean) duration of 24 (31) and 23 (39) wks, respectively. Pts in Cohorts 2 and 3B received a median of 2.7 RBC U/28 d (range 1-5) and 2.7 RBC U/28 d (range 2-4), respectively. For the primary endpoint, 2/21 (10%) and 6/22 (27%) pts receiving transfusions in Cohorts 2 and 3B achieved RBC-TI over any consecutive 12 wks during the first 24 wks. Median time to start of first RBC-TI response period in Cohorts 2 and 3B was 1.5 days and 37 days; median duration of response was 49 wks (range 16-82) and 42 wks (range 12-111), respectively. When assessing the entire treatment period, 4/21 (19%) and 8/22 (36%) pts in Cohorts 2 and 3B achieved RBC-TI ≥ 12 wks, respectively. 25% of the RBC-TI responders in both cohorts experienced 2 separate episodes of RBC-TI ≥ 12 wks. Median cumulative duration of all individual ≥ 12-wk response episodes of RBC-TI was 59 wks (range 24-82) in Cohort 2 and 55 wks (range 12-116) in Cohort 3B. 8/21 (38%) and 10/22 (46%) pts in Cohorts 2 and 3B achieved a ≥ 50% reduction in RBC transfusion burden (minimum ≥ 4 U reduction) over 12 wks; of these, 38% and 20% of responders in Cohorts 2 and 3B experienced 2 separate ≥ 12-wk response episodes, and 1 pt (13%) in Cohort 2 experienced 3 separate responses. 4/15 (27%) and 8/14 (57%) pts in Cohorts 2 and 3B who reached 24 wks of treatment achieved clinical benefit and therefore continued luspatercept on an ongoing basis. Safety is reported for all 79 pts on study. 5 (6%) pts had grade 3-4 treatment-related adverse events (AEs); these AEs were diarrhea (n = 2), dehydration (n = 1), and hypertension (n = 3). There were no treatment-related deaths. Treatment-related AEs (any grade) occurring in ≥ 5% of pts were hypertension (13%), bone pain (9%), and diarrhea (5%). 8 (10%) pts had ≥ 1 AE leading to discontinuation. 16 (20%) pts remain on treatment. Conclusions: These longer-term findings suggest durable activity of luspatercept in pts with MF-associated anemia. In addition, a quarter of pts receiving transfusions achieved more than one ≥ 12-wk episode of RBC-TI response with luspatercept. The incidence of grade 3-4 AEs with luspatercept was low. Disclosures Gerds: Roche/Genentech: Research Funding; Celgene: Consultancy, Research Funding; Sierra Oncology: Research Funding; AstraZeneca/MedImmune: Consultancy; Incyte Corporation: Consultancy, Research Funding; Pfizer: Research Funding; Apexx Oncology: Consultancy; CTI Biopharma: Consultancy, Research Funding; Gilead Sciences: Research Funding; Imago Biosciences: Research Funding. Vannucchi:Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Blueprint: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene/BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees. Passamonti:Novartis: Speakers Bureau; BMS: Speakers Bureau; Roche: Other: Support of parent study and funding of editorial support. Kremyanskaya:Constellation Pharmaceuticals: Research Funding; Incyte Corporation: Research Funding; Bristol Myers Squibb: Research Funding; Protagonist Therapeutics: Consultancy, Research Funding; Astex Pharmaceuticals: Research Funding. Gotlib:CTI Biopharma: Research Funding; Kartos: Consultancy, Honoraria, Research Funding; Incyte: Consultancy, Honoraria, Research Funding; Roche: Honoraria, Research Funding; BMS: Research Funding. McCaul:BMS: Speakers Bureau; Seattle Genetics: Speakers Bureau; Karyopharm: Speakers Bureau; Incyte: Speakers Bureau. Ribrag:argenX: Research Funding; pharmamar: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; servier: Consultancy; nanostring: Honoraria, Membership on an entity's Board of Directors or advisory committees; gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; AZD: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other; Infinity: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; MSD: Honoraria, Membership on an entity's Board of Directors or advisory committees; gustave roussy comprehensive cancer center: Current Employment; EPZ: Honoraria, Membership on an entity's Board of Directors or advisory committees; epizyme (EPZ): Research Funding. Mead:Novartis: Consultancy, Honoraria, Other: travel, accommodations, expenses, Research Funding, Speakers Bureau; Celgene/BMS: Consultancy, Honoraria, Other: travel, accommodations, expenses, Research Funding; Abbvie: Consultancy; CTI: Consultancy; Gilead: Consultancy. Harrison:Roche: Honoraria; Gilead Sciences: Honoraria, Speakers Bureau; Incyte Corporation: Speakers Bureau; Janssen: Speakers Bureau; CTI Biopharma Corp: Honoraria, Speakers Bureau; Sierra Oncology: Honoraria; Celgene: Honoraria, Research Funding, Speakers Bureau; Shire: Honoraria, Speakers Bureau; Novartis: Honoraria, Research Funding, Speakers Bureau; AOP Orphan Pharmaceuticals: Honoraria; Promedior: Honoraria. Mesa:Sierra Oncology: Consultancy; LaJolla Pharmaceutical Company: Consultancy; Novartis: Consultancy; Incyte: Research Funding; CTI BioPharma: Research Funding; Genentech: Research Funding; AbbVie: Research Funding; Promedior: Research Funding; Bristol Myers Squibb: Research Funding; Samus Therapeutics: Research Funding. Kiladjian:AbbVie: Membership on an entity's Board of Directors or advisory committees; AOP Orphan: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Barosi:Bristol Myers Squibb: Consultancy. Gerike:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company. Shetty:BMS: Current Employment, Current equity holder in publicly-traded company. Pariseau:Bristol Myers Squibb: Current Employment. Miranda:Bristol Myers Squibb: Current Employment. Schwickart:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company. Giuseppi:BMS: Current Employment, Current equity holder in publicly-traded company. Zhang:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company. Backstrom:Acceleron Pharma: Current Employment, Current equity holder in publicly-traded company; BMS: Current equity holder in publicly-traded company. Verstovsek:AstraZeneca: Research Funding; Roche: Research Funding; CTI Biopharma Corp: Research Funding; Genentech: Research Funding; Incyte Corporation: Consultancy, Research Funding; NS Pharma: Research Funding; Novartis: Consultancy, Research Funding; PharmaEssentia: Research Funding; ItalPharma: Research Funding; Sierra Oncology: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Blueprint Medicines Corp: Research Funding; Promedior: Research Funding; Gilead: Research Funding; Protagonist Therapeutics: Research Funding.

Context: Fedratinib is a selective JAK2/FLT3 inhibitor shown to induce spleen responses and symptom improvement in patients with myelofibrosis. Gastrointestinal (GI) events including nausea, vomiting, and diarrhea, were the most common adverse events (AEs) in fedratinib clinical trials, and were most frequent during early treatment. The JAK1/2 inhibitor, ruxolitinib, has demonstrated efficacy in patients with myelofibrosis; however, many patients discontinue ruxolitinib due to loss of response or drug-related cytopenias. The ongoing, single-arm, phase IIIb FREEDOM study evaluates fedratinib in myelofibrosis patients previously treated with ruxolitinib. FREEDOM includes prospective mitigation strategies for managing GI events. Objective: Review the frequency, severity, and mitigation of GI events during early fedratinib treatment in FREEDOM. Methods: Eligible patients are aged ≥18 years, with intermediate- or high-risk MF, ECOG-PS score 0-2, platelet count ≥50 × 109/L, and splenomegaly ≥ 450cm3 or palpable spleen > 5 cm below LCM; and had previously received ruxolitinib for ≥3 months (or ≥28 days with intolerance). Patients receive fedratinib 400 mg/day in continuous 28-day cycles. Study design includes mitigation strategies (prophylaxis, symptomatic treatment, dosing modifications, dosing with food) for managing GI toxicities. AEs are evaluated in all treated patients. Results: At data cutoff (26-Mar-2020), 22 patients had enrolled; 4 patients discontinued (none due to GI-related events). Median age was 68.5 years (range, 49-80). All patients had received ≥3 months of prior ruxolitinib treatment; the most common reason for ruxolitinib discontinuation was relapse (27%). Median fedratinib treatment duration was 18.6 weeks (1.6-48); 10 patients (46%) received >6 fedratinib cycles. Diarrhea, vomiting, and nausea occurred in 32%, 18%, and 14% of patients, respectively; none of these events were grade 3-4 or required dose modification or discontinuation. 12 patients (55%) received ondansetron and 6 (27%) received loperamide. Conclusions: Fedratinib was generally well tolerated. In similar patients receiving fedratinib 400 mg/day (starting dose) in the phase II JAKARTA2 study, which did not prespecify mitigation for GI events, rates of diarrhea, nausea, and vomiting through end-of-cycle 6, ranged from 41% to 62%. Early data from the FREEDOM study suggest frequency and severity of GI events may be reduced via mitigation strategies. Study enrollment is ongoing.

Primary Myelofibrosis (PMF) is a myeloproliferative disorder associated with JAK2V617F, Calreticulin (CALR) indels, and MPLW515L/K mutations activating the tyrosine kinase JAK2 and its downstream signaling pathway. The nature of signaling abnormalities in primary cells from PMF patients is poorly understood, since most of the work has been performed in cell lines or animal models. By flow cytometry we measured constitutive and cytokine induced phosphorylation of STAT5, STAT3, and ERK1/2 in circulating CD34+ cells from 57 patients with PMF (20 with prefibrotic-PMF) and 13 healthy controls (CTRLs). Levels of constitutive and TPO induced p-STAT5, and IL6 induced p-STAT3 were higher in patients than in CTRLs. Constitutive p-STAT5 values were lower in CALR than homozygous JAK2V617F mutated CD34+ cells from PMF patients. Moreover, constitutive p-STAT5 and IL6 induced p-STAT3 values correlated directly with circulating CD34+ cell number/L, and inversely with the frequency of circulating CD34+ cells expressing CXCR4. Constitutive p-STAT5 values of CD34+ cells were also inversely correlated with hemoglobin levels. When the patients were divided according with presence/absence of JAK2V617F mutation, all the correlations described characterized the JAK2V617F+ patients with prefibrotic-PMF (P-PMF). In conclusion, increased constitutive p-STAT5 and IL6 induced p-STAT3 values in circulating CD34+ cells characterize patients with PMF. Constitutive p-STAT5 and IL6 induced p-STAT3 values correlate with circulating CD34+ cell number/L, the frequency of circulating CD34+ cells expressing CXCR4 and hemoglobin levels within the prefibrotic JAK2V617F+ patient population. Our data point toward a complex activation of STAT5-dependent pathways in the stem/progenitor cell compartment, that characterize the phenotypic diversity of PMF.

The prognostic significance of bone marrow (BM) fibrosis grade in patients with primary myelofibrosis (PMF) is still debated. A fibrosis grade greater than 1 was shown to associate with higher risk of death, and addition of fibrosis grade to IPSS score resulted in a more accurate prediction of survival. The aim of this study was to analyze the prognostic impact of BM fibrosis in 490 patients with PMF, evaluated at diagnosis, molecularly annotated and with extensive follow-up information. We found that fibrosis grade 2 and greater on a 0-3 scale was associated with clinical characteristics indicative of a more advanced disease, such as anemia, leukopenia, thrombocytopenia, constitutional symptoms, larger splenomegaly and a higher IPSS risk category. Patients with higher grade of fibrosis were also more likely to have additional somatic mutations in ASXL1 and EZH2, that are prognostically adverse. Median survival was significantly reduced in patients with grade 2 and 3 fibrosis as compared with grade 1; this effect was maintained when analysis was restricted to younger patients. In multivariate analysis, fibrosis grade independently predicted for survival regardless of IPSS variables and mutational status; the adverse impact of fibrosis was noticeable especially in lower IPSS risk categories. Overall, results indicate that higher grades of fibrosis correlate with unique clinical and molecular aspects and represent an independent adverse variable in patients with PMF; these observations deserve confirmation in prospectively designed series of patients. Am. J. Hematol. 91:918-922, 2016. © 2016 Wiley Periodicals, Inc.

Myeloproliferative neoplasm (MPN)-associated myelofibrosis is a clonal, neoplastic disorder of the hematopoietic stem cells, in which inflammation and immune dysregulation play an important role. Extracellular nicotinamide phosphoribosyltransferase (eNAMPT), also known as visfatin, is a cytokine implicated in a number of inflammatory and neoplastic diseases. Here plasma levels of eNAMPT in patients with MPN-associated myelofibrosis and their effects on disease phenotype and outcomes were examined. The concordance of eNAMPT levels with the marker of general inflammation high-sensitivity C-reactive protein (hs-CRP) was also studied. A total of 333 MPN-associated myelofibrosis patients (187 males and 146 females) and 31 age- and gender-matched normal-weight healthy subjects were enrolled in the study main body. Levels of eNAMPT and hs-CRP were simultaneously assayed in 209 MPN-associated myelofibrosis patients. Twenty-four polycythemia vera or essential thrombocythemia patients were used as controls. eNAMPT was over expressed in MPN-associated myelofibrosis, and eNAMPT expression was correlated with higher white blood cell count, higher hemoglobin, and higher platelet count, suggesting that eNAMPT is an indispensable permissive agent for myeloproliferation of MPN-associated myelofibrosis. The lack of correlation between eNAMPT and hs-CRP revealed that eNAMPT in MPN-associated myelofibrosis does not behave as a canonical inflammatory cytokine. In addition, higher levels of eNAMPT predicted longer time to blast transformation, and protected against progression toward thrombocytopenia and large splenomegaly. In conclusion, in MPN-associated myelofibrosis high levels of eNAMPT mark the myeloproliferative potential and, at variance with a high number of cancers, are protective against disease progression. Am. J. Hematol. 91:709-713, 2016. © 2016 Wiley Periodicals, Inc.

JAK2 V617F allele burden 50% is associated with response to ruxolitinib in persons with MPN-associated myelofibrosis and splenomegaly requiring therapy

This is a retrospective analysis of 95 patients with myelofibrosis who were allografted between 2001 and 2014. The aims of the study were to assess whether the outcome of alternative donor grafts has improved with time and how this compares with the outcome of identical sibling grafts. Patients were studied in 2 time intervals: 2000 to 2010 (n = 58) and 2011 to 2014 (n = 37). The Dynamic International Prognostic Scoring System score was comparable in the 2 time periods, but differences in the most recent group included older age (58 versus 53 years, P = .004), more family haploidentical donors (54% versus 5%, P.0001), and the introduction of the thiotepa-fludarabine-busulfan conditioning regimen (70% of patients versus 2%, P.0001). Acute and chronic graft-versus-host disease were comparable in the 2 time periods. The 3-year transplantation-related mortality (TRM) in the 2011 to 2014 period versus the 2000 to 2010 period is 16% versus 32% (P = .10), the relapse rate 16% versus 40% (P = .06), and actuarial survival 70% versus 39% (P = .08). Improved survival was most pronounced in alternative donor grafts (69% versus 21%, P = .02), compared with matched sibling grafts (72% versus 45%, P = .40). In conclusion, the outcome of allografts in patients with myelofibrosis has improved in recent years because of a reduction of both TRM and relapse. Improvement is most significant in alternative donor transplantations, with modifications in donor type and conditioning regimen.

Extracellular matrix (ECM) components initiate crucial biochemical and biomechanical cues that are required for bone marrow homeostasis. In our research, we prove that a peri-cellular matrix composed primarily of type III and type IV collagens, and fibronectin surrounds human megakaryocytes in the bone marrow. The data we collected support the hypothesis that bone marrow megakaryocytes possess a complete mechanism to synthesize the ECM components, and that thrombopoietin is a pivotal regulator of this new function inducing transforming growth factor-β1 (TGF-β1) release and consequent activation of the downstream pathways, both in vitro and in vivo. This activation results in a dose dependent increase of ECM component synthesis by megakaryocytes, which is reverted upon incubation with JAK and TGF-β1 receptor specific inhibitors. These data are pivotal for understanding the central role of megakaryocytes in creating their own regulatory niche within the bone marrow environment.

Patients with high-risk myeloproliferative neoplasms (MPNs), and in particular myelofibrosis (MF), can be cured only with allogeneic hematopoietic stem cell transplantation (HSCT). Because MPNs and JAK2V617F-mutated cells show genomic instability, stalled replication forks, and baseline DNA double-strand breaks, DNA repair inhibition with poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors represents a potential novel therapy. Because the alkylating agent busulfan is integral in conditioning regimens for HSCT and leads to stalled replication forks through DNA strand cross-linking, we hypothesized that PARP inhibition with veliparib in combination with busulfan may lead to synergistic cytotoxicity in MPN cells. We first treated 2 MPN cell lines harboring the JAK2V617F mutation (SET2 and HEL) with veliparib at increasing concentrations and measured cell proliferation. SET2 and HEL cells were relatively sensitive to veliparib (IC50 of 11.3 μM and 74.2 μM, respectively). We next treated cells with increasing doses of busulfan in combination with 4 μM veliparib and found that the busulfan IC50 decreased from 27 μM to 4 μM in SET2 cells and from 45.1 μM to 28.1 μM in HEL cells. The mean combination index was .55 for SET2 cells and .40 for HEL cells. Combination treatment of SET2 cells caused G2M arrest in 53% of cells, compared with 30% with veliparib alone and 35% with busulfan alone. G2M arrest was associated with activation of the ATR-Chk1 pathway, as shown by an immunofluorescence assay for phosphorylated Chk1 (p-Chk1). We then tested in vivo the effect of combined low doses of busulfan and veliparib in a JAK2V617F MPN-AML xenotransplant model. Vehicle- and veliparib-treated mice had similar median survival of 39 and 40 days, respectively. Combination treatment increased median survival from 47 days (busulfan alone) to 50 days (P = .02). Finally, we tested the combined effect of busulfan and veliparib on CD34+ cells obtained from the bone marrow or peripheral blood of 5 patients with JAK2V617F-mutated and 2 patients with CALR-mutated MF. MF cells treated with the combination of veliparib and busulfan showed reduced colony formation compared with busulfan alone (87% versus 68%; P = .001). In contrast, treatment of normal CD34+ cells with veliparib did not affect colony growth. Here we show that in vivo confirmation that treatment with the PARP-1 inhibitor veliparib and busulfan results in synergistic cytotoxicity in MPN cells. Our data provide the rationale for testing novel pretransplantation conditioning regimens with combinations of PARP-1 inhibition and reduced doses of alkylators, such as busulfan and melphalan, for high-risk MPNs or MPN-derived acute myelogenous leukemia.

The introduction of new therapeutic agents in multiple myeloma (MM), including proteasome inhibitors, immunoregulatory drugs and monoclonal antibodies, has improved the outcomes of patients, but in parallel has changed the frequency and epidemiology of infections. Hence, the great strides in the indications and use of new active treatments for MM need parallel progresses on the best approach to prophylaxis and supportive therapy for infections. Moving from the recognition that the above issue represents an unmet clinical need in MM, an expert panel assessed the scientific literature and composed a framework of recommendations for optimal infection control in patients candidate to active treatment for MM. The present publication represents a consensus document from questionnaires and consensus meetings held during 2017. The issues tackled in the project dealt with: infectious risk assessment, risk management and prophylaxis, intravenous immunoglobulin replacement therapy, antiviral and antibacterial vaccination. Considering the lack of conclusive and/or enough large studies for certain topics several recommendations derived from the personal experience of the experts.