Thrombosis is a major cause of morbidity and mortality in Philadelphia chromosome–negative myeloproliferative neoplasms (MPNs), clonal disorders of hematopoiesis characterized by activated Janus kinase (JAK)–signal transducer and activator of transcription (STAT) signaling. Neutrophil extracellular trap (NET) formation, a component of innate immunity, has been linked to thrombosis. We demonstrate that neutrophils from patients with MPNs are primed for NET formation, an effect blunted by pharmacological inhibition of JAK signaling. Mice with conditional knock-in of Jak2^V617F, the most common molecular driver of MPN, have an increased propensity for NET formation and thrombosis. Inhibition of JAK-STAT signaling with the clinically available JAK2 inhibitor ruxolitinib abrogated NET formation and reduced thrombosis in a deep vein stenosis murine model. We further show that expression of PAD4, a protein required for NET formation, is increased in JAK2^V617F-expressing neutrophils and that PAD4 is required for Jak2^V617F-driven NET formation and thrombosis in vivo. Finally, in a population study of more than 10,000 individuals without a known myeloid disorder, JAK2^V617F-positive clonal hematopoiesis was associated with an increased incidence of thrombosis. In aggregate, our results link JAK2^V617F expression to NET formation and thrombosis and suggest that JAK2 inhibition may reduce thrombosis in MPNs through cell-intrinsic effects on neutrophil function.

Somatic mutations in calreticulin (CALR) are present in approximately 40% of patients with myeloproliferative neoplasms (MPN), but the mechanism by which mutant CALR is oncogenic remains unclear. Here, we demonstrate that expression of mutant CALR alone is sufficient to engender MPN in mice and recapitulates the disease phenotype of patients with CALR-mutant MPN. We further show that the thrombopoietin receptor MPL is required for mutant CALR-driven transformation through JAK–STAT pathway activation, thus rendering mutant CALR-transformed hematopoietic cells sensitive to JAK2 inhibition. Finally, we demonstrate that the oncogenicity of mutant CALR is dependent on the positive electrostatic charge of the C-terminus of the mutant protein, which is necessary for physical interaction between mutant CALR and MPL. Together, our findings elucidate a novel paradigm of cancer pathogenesis and reveal how CALR mutations induce MPN. Significance: The mechanism by which CALR mutations induce MPN remains unknown. In this report, we show that the positive charge of the CALR mutant C-terminus is necessary to transform hematopoietic cells by enabling binding between mutant CALR and the thrombopoietin receptor MPL. Cancer Discov; 6(4); 368–81. ©2016 AACR. See related commentary by Stanley and Steidl, p. 344. This article is highlighted in the In This Issue feature, p. 331

Mixed-phenotype acute leukemia (MPAL) encompasses a heterogeneous group of rare leukemias in which assigning a single lineage of origin is not possible. A variety of different terms and classification systems have been used historically to describe this entity. MPAL is currently defined by a limited set of lineage-specific markers proposed in the 2008 World Health Organization monograph on classification of tumors of hematopoietic and lymphoid tissues. In adult patients, MPAL is characterized by relative therapeutic resistance that may be attributed in part to the high proportion of patients with adverse cytogenetic abnormalities. No prospective, controlled trials exist to guide therapy. The limited available data suggest that an “acute lymphoblastic leukemia–like” regimen followed by allogeneic stem-cell transplant may be advisable; addition of a tyrosine kinase inhibitor in patients with t(9;22) translocation is recommended. The role of immunophenotypic and genetic markers in guiding chemotherapy choice and postremission strategy, as well as the utility of targeted therapies in non–Ph-positive MPALs is unknown.

Rituximab is a chimeric monoclonal antibody against CD20 that is used mainly for the treatment of CD20-positive lymphoma. Recently, its use has been expanded to include treatment of other nonmalignant diseases such as rheumatologic diseases and autoimmune cytopenia. Correlating with the increased use of rituximab has been an increased number of reports of its late adverse effects. One of these is late-onset neutropenia (LON). Most investigators define LON as grade III-IV neutropenia occurring 3-4 weeks after the last treatment with rituximab, in the absence of an alternative explanation for the neutropenia. We report 6 cases of LON identified in our institution. Four patients were treated for diffuse large B-cell lymphoma, and 2 patients for follicular lymphoma. Median patient age was 68 years (range, 33-83 yr); LON appeared after a median interval of 77 days (range, 42-153 d) and lasted for a median of 5 days (range, 1-45 d). Five of the 6 patients presented with infectious complications, and 4 patients experienced recurrent episodes of neutropenia. One patient presented with LON and concomitant subacute pulmonary disease that was attributed to rituximab therapy. In addition to our own case series we present a systematic review of the literature, which we performed to compile data to describe better the syndrome of LON. Systematic studies, case series, and case reports were extracted. Most studies dealing with LON are retrospective by design and are limited by the heterogeneous populations included in the analysis. The incidence of LON is generally reported to be in the range of 3%-27%. Data regarding populations at risk are not consistent, and in some instances are conflicting. Patients considered at increased risk of LON include patients after autologous stem cell transplantation, patients treated for acquired immunodeficiency syndrome (AIDS)-related lymphoma, and patients treated with purine analogues. Patients who received previous cytotoxic treatment as well as those treated with more intensive chemotherapy or with chemotherapy in combination with radiotherapy are also considered to be at risk of LON. In addition, advanced stages of disease and having received multiple doses of rituximab are risk factors for LON. The mechanism of LON is poorly understood. Direct toxicity is very unlikely. Some speculate that there may be an infectious etiology involved, as well as an antibody-mediated process, but these ideas have not been substantiated. The concept of a lymphocyte subpopulation imbalance leading to LON has been presented based on the demonstration of T-LGL in peripheral blood and bone marrow of patients with LON. Perturbations in stromal-derived factor-1 and in the BAFF cytokine have also been discussed as potential players in the pathogenesis of LON. A recent study correlated specific polymorphism in the immunoglobulin G Fc receptor FCγRIIIa 158 V/F with increased rates of LON. The clinical significance of LON is important because it may affect treatment strategies. Of note, infectious complications are not very frequent and not very severe. Pooling data from the major retrospective studies reveals an infection rate of 16.9%. Most infections were mild and resolved promptly. One death occurred from infection during neutropenia. Repeated episodes of LON are not uncommon, but it is so far impossible to identify those patients at risk of these relapsing episodes of LON. Re-treatment with rituximab after LON may result in recurrent episodes, but the implications and risks are uncertain at the present time. The role of growth factors once LON appears is ill defined, and the decision to use them should be made on a case-by-case basis.

Survival of adults with acute lymphoblastic leukemia (ALL) is inferior to that of pediatric patients. Strategies to improve the outcome of adult population are warranted. This study aims to evaluate the efficacy and safety of pediatric‐inspired regimens given to adolescents and young adults (AYA), usually defined as 16–39 years, with ALL. Systematic review and meta‐analysis of comparative trials of AYA patients with ALL given induction chemotherapy with either pediatric‐inspired regimens or conventional‐adult chemotherapy was conducted. Relative risks (RR) with 95% confidence intervals (CIs) were estimated and pooled. Our search yielded 11 trials, including 2,489 patients. AYA patients given pediatric‐inspired regimens had a statistically significant lower all cause mortality rate at 3 years (RR 0.58; 95% CI 0.51–0.67). Complete remission rate after induction chemotherapy and event free survival were superior in the pediatric‐inspired regimens arm (RR 1.05; 95% CI 1.01–1.10 and RR 1.66; 95% CI 1.39–1.99, respectively). Relapse rate was also lower in patients given pediatric‐inspired regimens (RR 0.51; 95% CI 0.39–0.66) with comparable nonrelapse mortality between the two groups (RR 0.53, 95% CI 0.19–1.48). Pediatric‐inspired regimens are superior to conventional‐adult chemotherapy in AYA ALL patients. Further randomized controlled studies to investigate this approach in adult ALL patients are warranted. Am. J. Hematol. 87:472–478, 2012.