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•Ruxolitinib demonstrated clinically meaningful efficacy in pediatric patients with either treatment-naïve or steroid-refractory aGVHD.•We identified the ruxolitinib dosage across the pediatric age range treated, and no unexpected safety signals were observed in children.

•Inactivation of factor XIa by the endothelium increases vessel permeability.•FXIa–PAI-1 complex binding to very low–density lipoprotein receptor induces MAPK signaling, causing ADAM10-mediated cleavage of VE-cadherin.

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•SOX11 expression and EBV infection occur in alternative subsets of BL with different profiles of somatic mutations.•Among EBV−BL, IG∷MYCtranslocation is generated by class switch recombination in SOX11+BL and somatic hypermutation in SOX11−tumors.

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•ERCC6L2 disease causes BMF that requires timely stem cell transplantation before progressing to a high-risk myeloid malignancy.•Frequent somatic TP53mutation screening and BM examination are essential for surveillance of patients with ERCC6L2-related disease.

Myeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia, myelodysplastic syndrome with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Here, we investigate the drug sensitivity landscape of these rare malignancies. We show that acute myeloid leukemia (AML) cells with erythroid or megakaryocytic differentiation depend on the antiapoptotic protein B-cell lymphoma (BCL)-XL, rather than BCL-2, using combined ex vivo drug sensitivity testing, genetic perturbation, and transcriptomic profiling. High-throughput screening of >500 compounds identified the BCL-XL–selective inhibitor A-1331852 and navitoclax as highly effective against erythroid/megakaryoblastic leukemia cell lines. In contrast, these AML subtypes were resistant to the BCL-2 inhibitor venetoclax, which is used clinically in the treatment of AML. Consistently, genome-scale CRISPR-Cas9 and RNAi screening data demonstrated the striking essentiality of BCL-XL-encoding BCL2L1 but not BCL2 or MCL1, for the survival of erythroid/megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias identified high BCL2L1 expression compared with other subtypes of AML and other hematological malignancies, where BCL2 and MCL1 were more prominent. BCL-XL inhibition effectively killed blasts in samples from patients with AML with erythroid or megakaryocytic differentiation ex vivo and reduced tumor burden in a mouse erythroleukemia xenograft model. Combining the BCL-XL inhibitor with the JAK inhibitor ruxolitinib showed synergistic and durable responses in cell lines. Our results suggest targeting BCL-XL as a potential therapy option in erythroid/megakaryoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.

Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality following allogeneic hematopoietic transplantation. In experimental models, interleukin-22 promotes epithelial regeneration and induces innate antimicrobial molecules. We conducted a multicenter single-arm phase 2 study evaluating the safety and efficacy of a novel recombinant human interleukin-22 dimer, F-652, used in combination with systemic corticosteroids for treatment of newly diagnosed lower gastrointestinal acute GVHD. The most common adverse events were cytopenias and electrolyte abnormalities, and there were no dose-limiting toxicities. Out of 27 patients, 19 (70%; 80% confidence interval, 56%-79%) achieved a day-28 treatment response, meeting the prespecified primary endpoint. Responders exhibited a distinct fecal microbiota composition characterized by expansion of commensal anaerobes, which correlated with increased overall microbial α-diversity, suggesting improvement of GVHD-associated dysbiosis. This work demonstrates a potential approach for combining immunosuppression with tissue-supportive strategies to enhance recovery of damaged mucosa and promote microbial health in patients with gastrointestinal GVHD. This trial was registered at www.clinicaltrials.gov as NCT02406651.

Venetoclax combination therapies are becoming the standard of care in acute myeloid leukemia (AML). However, the therapeutic benefit of these drugs in older/unfit patients is limited to only a few months, highlighting the need for more effective therapies. Protein phosphatase 2A (PP2A) is a tumor suppressor phosphatase with pleiotropic functions that becomes inactivated in ∼70% of AML cases. PP2A promotes cancer cell death by modulating the phosphorylation state in a variety of proteins along the mitochondrial apoptotic pathway. We therefore hypothesized that pharmacological PP2A reactivation could increase BCL2 dependency in AML cells and, thus, potentiate venetoclax–induced cell death. Here, by using 3 structurally distinct PP2A-activating drugs, we show that PP2A reactivation synergistically enhances venetoclax activity in AML cell lines, primary cells, and xenograft models. Through the use of gene editing tools and pharmacological approaches, we demonstrate that the observed therapeutic synergy relies on PP2A complexes containing the B56α regulatory subunit, of which expression dictates response to the combination therapy. Mechanistically, PP2A reactivation enhances venetoclax-driven apoptosis through simultaneous inhibition of antiapoptotic BCL2 and extracellular signal-regulated kinase signaling, with the latter decreasing MCL1 protein stability. Finally, PP2A targeting increases the efficacy of the clinically approved venetoclax and azacitidine combination in vitro, in primary cells, and in an AML patient-derived xenograft model. These preclinical results provide a scientific rationale for testing PP2A-activating drugs with venetoclax combinations in AML.

Tγδ large granular lymphocyte leukemia (LGLL) is a rare variant of T-cell LGLL (T-LGLL) that has been less investigated as compared with the more frequent Tαβ LGLL, particularly in terms of frequency of STAT3 and STAT5b mutations. In this study, we characterized the clinical and biological features of 137 patients affected by Tγδ LGLL; data were retrospectively collected from 1997 to 2020 at 8 referral centers. Neutropenia and anemia were the most relevant clinical features, being present in 54.2% and 49.6% of cases, respectively, including severe neutropenia and anemia in ∼20% of cases each. Among the various treatments, cyclosporine A was shown to provide the best response rates. DNA samples of 97 and 94 cases were available for STAT3 and STAT5b mutation analysis, with 38.1% and 4.2% of cases being mutated, respectively. Clinical and biological features of our series of Tγδ cases were also compared with a recently published Tαβ cohort including 129 cases. Though no differences in STAT3 and STAT5b mutational frequency were found, Tγδ cases more frequently presented with neutropenia (P = .0161), anemia (P < .0001), severe anemia (P = .0065), and thrombocytopenia (P = .0187). Moreover, Vδ2− cases displayed higher frequency of symptomatic disease. Overall, Tγδ cases displayed reduced survival with respect to Tαβ cases (P = .0017). Although there was no difference in STAT3 mutation frequency, our results showed that Tγδ LGLL represents a subset of T-LGLL characterized by more frequent symptoms and reduced survival as compared with Tαβ LGLL.

•GALEis expressed during late-stage megakaryopoiesis, regulating the glycosylation and surface exposure of GPIbα and β1 integrin.•Novel GALEvariants cause syndromic macrothrombocytopenia associated with impaired platelet formation and function.

Glycosylation is recognized as a key process for proper megakaryopoiesis and platelet formation. The enzyme uridine diphosphate (UDP)-galactose-4-epimerase, encoded by GALE, is involved in galactose metabolism and protein glycosylation. Here, we studied 3 patients from 2 unrelated families who showed lifelong severe thrombocytopenia, bleeding diathesis, mental retardation, mitral valve prolapse, and jaundice. Whole-exome sequencing revealed 4 variants that affect GALE, 3 of those previously unreported (Pedigree A, p.Lys78ValfsX32 and p.Thr150Met; Pedigree B, p.Val128Met; and p.Leu223Pro). Platelet phenotype analysis showed giant and/or grey platelets, impaired platelet aggregation, and severely reduced alpha and dense granule secretion. Enzymatic activity of the UDP-galactose-4-epimerase enzyme was severely decreased in all patients. Immunoblotting of platelet lysates revealed reduced GALE protein levels, a significant decrease in N-acetyl-lactosamine (LacNAc), showing a hypoglycosylation pattern, reduced surface expression of gylcoprotein Ibα-IX-V (GPIbα-IX-V) complex and mature β1 integrin, and increased apoptosis. In vitro studies performed with patients-derived megakaryocytes showed normal ploidy and maturation but decreased proplatelet formation because of the impaired glycosylation of the GPIbα and β1 integrin, and reduced externalization to megakaryocyte and platelet membranes. Altered distribution of filamin A and actin and delocalization of the von Willebrand factor were also shown. Overall, this study expands our knowledge of GALE-related thrombocytopenia and emphasizes the critical role of GALE in the physiological glycosylation of key proteins involved in platelet production and function.

This open-label, randomized, phase 3 trial (NCT02577406) compared enasidenib, an oral IDH2 (isocitrate dehydrogenase 2) inhibitor, with conventional care regimens (CCRs) in patients aged ≥60 years with late-stage, mutant-IDH2 acute myeloid leukemia (AML) relapsed/refractory (R/R) to 2 or 3 prior AML-directed therapies. Patients were first preselected to a CCR (azacitidine, intermediate-dose cytarabine, low-dose cytarabine, or supportive care) and then randomized (1:1) to enasidenib 100 mg per day or CCR. The primary endpoint was overall survival (OS). Secondary endpoints included event-free survival (EFS), time to treatment failure (TTF), overall response rate (ORR), hematologic improvement (HI), and transfusion independence (TI). Overall, 319 patients were randomized to enasidenib (n = 158) or CCR (n = 161). The median age was 71 years, median (range) enasidenib exposure was 142 days (3 to 1270), and CCR was 36 days (1 to 1166). One enasidenib (0.6%) and 20 CCR (12%) patients received no randomized treatment, and 30% and 43%, respectively, received subsequent AML-directed therapies during follow-up. The median OS with enasidenib vs CCR was 6.5 vs 6.2 months (HR [hazard ratio], 0.86; P = .23); 1-year survival was 37.5% vs 26.1%. Enasidenib meaningfully improved EFS (median, 4.9 vs 2.6 months with CCR; HR, 0.68; P = .008), TTF (median, 4.9 vs 1.9 months; HR, 0.53; P < .001), ORR (40.5% vs 9.9%; P <.001), HI (42.4% vs 11.2%), and red blood cell (RBC)-TI (31.7% vs 9.3%). Enasidenib safety was consistent with prior reports. The primary study endpoint was not met, but OS was confounded by early dropout and subsequent AML-directed therapies. Enasidenib provided meaningful benefits in EFS, TTF, ORR, HI, and RBC-TI in this heavily pretreated older mutant-IDH2 R/R AML population.

•EFS was meaningfully improved with enasidenib vs CCR; OS was confounded by early dropout and use of subsequent AML therapies.•Enasidenib provided meaningful morphologic and hematologic responses vs CCR in this heavily pretreated older R/R mutant-IDH2AML population.

•Mortality rate in patients with hematologic malignancy with breakthrough COVID-19 is ∼9%, lower than in the prevaccination era.•Patients who received monoclonal antibodies, alone or combined with antivirals, showed a better clinical outcome.

Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of hematological cancers arising from the malignant transformation of mature T cells. In a cohort of 28 PTCL cases, we identified recurrent overexpression of MYCN, a member of the MYC family of oncogenic transcription factors. Approximately half of all PTCL cases was characterized by a MYC expression signature. Inducible expression of MYCN in lymphoid cells in a mouse model caused T-cell lymphoma that recapitulated human PTCL with an MYC expression signature. Integration of mouse and human expression data identified EZH2 as a key downstream target of MYCN. Remarkably, EZH2 was found to be an essential cofactor for the transcriptional activation of the MYCN-driven gene expression program, which was independent of methyltransferase activity but dependent on phosphorylation by CDK1. MYCN-driven T-cell lymphoma was sensitive to EZH2 degradation or CDK1 inhibition, which displayed synergy with US Food and Drug Administration–approved histone deacetylase (HDAC) inhibitors.

Central nervous system (CNS) relapse of mantle cell lymphoma (MCL) is a rare phenomenon with dismal prognosis, where no standard therapy exists. Since the covalent Bruton tyrosine kinase (BTK) inhibitor ibrutinib is effective in relapsed/refractory MCL and penetrates the blood–brain barrier (BBB), on behalf of Fondazione Italiana Linfomi and European Mantle Cell Lymphoma Network we performed a multicenter retrospective international study to investigate the outcomes of patients treated with ibrutinib or chemoimmunotherapy. In this observational study, we recruited patients with MCL with CNS involvement at relapse who received CNS-directed therapy between 2000 and 2019. The primary objective was to compare the overall survival (OS) of patients treated with ibrutinib or BBB crossing chemotherapy. A propensity score based on a multivariable binary regression model was applied to balance treatment cohorts. Eighty-eight patients were included. The median age at study entry was 65 years (range, 39-87), 76% were males, and the median time from lymphoma diagnosis to CNS relapse was 16 months (range, 1-122). Patients were treated with ibrutinib (n = 29, ibrutinib cohort), BBB crossing chemotherapy (ie, high-dose methotrexate ± cytarabine; n = 29, BBB cohort), or miscellaneous treatments (n = 30, other therapy cohort). Both median OS (16.8 vs 4.4 months; P = .007) and median progression-free survival (PFS) (13.1 vs 3.0 months; P = .009) were superior in the ibrutinib cohort compared with the BBB cohort. Multivariable Cox regression model revealed that ibrutinib therapeutic choice was the strongest independent favorable predictive factor for both OS (hazard ratio [HR], 6.8; 95% confidence interval [CI], 2.2-21.3; P < .001) and PFS (HR, 4.6; 95% CI, 1.7-12.5; P = .002), followed by CNS progression of disease (POD) >24 months from first MCL diagnosis (HR for death, 2.4; 95% CI, 1.1-5.3; P = .026; HR for death or progression, 2.3; 95% CI, 1.1-4.6; P = .023). The addition of intrathecal (IT) chemotherapy to systemic CNS-directed therapy was not associated with superior OS (P = .502) as the morphological variant (classical vs others, P = .118). Ibrutinib was associated with superior survival compared with BBB-penetrating chemotherapy in patients with CNS relapse of MCL and should be considered as a therapeutic option.

Allogeneic hematopoietic stem cell transplantation (HSCT) is the gold standard curative therapy for infants and children with many inborn errors of immunity (IEI), but adolescents and adults with IEI are rarely referred for transplant. Lack of published HSCT outcome data outside small, single-center studies and perceived high risk of transplant-related mortality have delayed the adoption of HSCT for IEI patients presenting or developing significant organ damage later in life. This large retrospective, multicenter HSCT outcome study reports on 329 IEI patients (age range, 15-62.5 years at HSCT). Patients underwent first HSCT between 2000 and 2019. Primary endpoints were overall survival (OS) and event-free survival (EFS). We also evaluated the influence of IEI-subgroup and IEI-specific risk factors at HSCT, including infections, bronchiectasis, colitis, malignancy, inflammatory lung disease, splenectomy, hepatic dysfunction, and systemic immunosuppression. At a median follow-up of 44.3 months, the estimated OS at 1 and 5 years post-HSCT for all patients was 78% and 71%, and EFS was 65% and 62%, respectively, with low rates of severe acute (8%) or extensive chronic (7%) graft-versus-host disease. On univariate analysis, OS and EFS were inferior in patients with primary antibody deficiency, bronchiectasis, prior splenectomy, hepatic comorbidity, and higher hematopoietic cell transplant comorbidity index scores. On multivariable analysis, EFS was inferior in those with a higher number of IEI-associated complications. Neither age nor donor had a significant effect on OS or EFS. We have identified age-independent risk factors for adverse outcome, providing much needed evidence to identify which patients are most likely to benefit from HSCT.

Persons with mild hemophilia A (HA) may use intranasal desmopressin prior to sports participation. Desmopressin is expensive and can cause vomiting, headache, palpitation, and occasionally seizures. Our group has previously documented a 2.3-fold increase in factor VIII activity (FVIII:C) in adolescents with mild HA after moderate-intensity aerobic exercise. Herein, we report principal findings of a randomized trial of intranasal desmopressin vs a standardized, moderate-intensity aerobic exercise regimen in adolescents with mild HA. Our primary objective was to compare the change in FVIII:C associated with these 2 interventions. We also examined changes in hemostatic parameters arising from their sequential administration. The study was conducted simultaneously at the Hospital for Sick Children, Canada, and Nationwide Children’s Hospital, USA. Thirty-two eligible male adolescents (mean age ± standard deviation: 16.1 ± 2.6 years) with mild HA (mean baseline FVIII:C: 27.9% ± 18.4%) were randomized to 1 of 4 study arms (desmopressin followed by exercise, desmopressin alone, exercise followed by desmopressin, and exercise alone). Blood work was obtained at baseline and at 3 subsequent time-points. Participants randomized to exercise cycled on an ergometer for approximately 12 minutes, with the final 3 minutes at 85% of their predicted maximum heart rate. Standard weight-based dosing of desmopressin was used. Mean immediate increase in FVIII:C was 1.7-fold with exercise compared with 1.9-fold with desmopressin (noninferiority, P = .04). Exercise-induced improvement in hemostatic parameters including FVIII:C was brief compared with more sustained improvements seen with desmopressin. More than 60% of participants randomized to receive both exercise and desmopressin achieved normal (>50%) FVIII:C, 75 and 135 minutes into the study protocol.

Patients with acute myeloid leukemia (AML) often achieve remission after allogeneic hematopoietic cell transplantation (allo-HCT) but subsequently die of relapse driven by leukemia cells resistant to elimination by allogeneic T cells based on decreased major histocompatibility complex II (MHC-II) expression and apoptosis resistance. Here we demonstrate that mouse-double-minute-2 (MDM2) inhibition can counteract immune evasion of AML. MDM2 inhibition induced MHC class I and II expression in murine and human AML cells. Using xenografts of human AML and syngeneic mouse models of leukemia, we show that MDM2 inhibition enhanced cytotoxicity against leukemia cells and improved survival. MDM2 inhibition also led to increases in tumor necrosis factor-related apoptosis-inducing ligand receptor-1 and -2 (TRAIL-R1/2) on leukemia cells and higher frequencies of CD8+CD27lowPD-1lowTIM-3low T cells, with features of cytotoxicity (perforin+CD107a+TRAIL+) and longevity (bcl-2+IL-7R+). CD8+ T cells isolated from leukemia-bearing MDM2 inhibitor-treated allo-HCT recipients exhibited higher glycolytic activity and enrichment for nucleotides and their precursors compared with vehicle control subjects. T cells isolated from MDM2 inhibitor-treated AML-bearing mice eradicated leukemia in secondary AML-bearing recipients. Mechanistically, the MDM2 inhibitor-mediated effects were p53-dependent because p53 knockdown abolished TRAIL-R1/2 and MHC-II upregulation, whereas p53 binding to TRAILR1/2 promotors increased upon MDM2 inhibition. The observations in the mouse models were complemented by data from human individuals. Patient-derived AML cells exhibited increased TRAIL-R1/2 and MHC-II expression on MDM2 inhibition. In summary, we identified a targetable vulnerability of AML cells to allogeneic T-cell–mediated cytotoxicity through the restoration of p53-dependent TRAIL-R1/2 and MHC-II production via MDM2 inhibition.

Activation of coagulation factor (F) XI promotes multiorgan failure in rodent models of sepsis and in a baboon model of lethal systemic inflammation induced by infusion of heat-inactivated Staphylococcus aureus. Here we used the anticoagulant FXII-neutralizing antibody 5C12 to verify the mechanistic role of FXII in this baboon model. Compared with untreated control animals, repeated 5C12 administration before and at 8 and 24 hours after bacterial challenge prevented the dramatic increase in circulating complexes of contact system enzymes FXIIa, FXIa, and kallikrein with antithrombin or C1 inhibitor, and prevented cleavage and consumption of high-molecular-weight kininogen. Activation of several coagulation factors and fibrinolytic enzymes was also prevented. D-dimer levels exhibited a profound increase in the untreated animals but not in the treated animals. The antibody also blocked the increase in plasma biomarkers of inflammation and cell damage, including tumor necrosis factor, interleukin (IL)-1β, IL-6, IL-8, IL-10, granulocyte-macrophage colony-stimulating factor, nucleosomes, and myeloperoxidase. Based on clinical presentation and circulating biomarkers, inhibition of FXII prevented fever, terminal hypotension, respiratory distress, and multiorgan failure. All animals receiving 5C12 had milder and transient clinical symptoms and were asymptomatic at day 7, whereas untreated control animals suffered irreversible multiorgan failure and had to be euthanized within 2 days after the bacterial challenge. This study confirms and extends our previous finding that at least 2 enzymes of the contact activation complex, FXIa and FXIIa, play critical roles in the development of an acute and terminal inflammatory response in baboons challenged with heat-inactivated S aureus.

The clinical benefit of extended prophylaxis for venous thromboembolism (VTE) after laparoscopic surgery for cancer is unclear. The efficacy and safety of direct oral anticoagulants for this indication are unexplored. PROphylaxis of venous thromboembolism after LAParoscopic Surgery for colorectal cancer Study II (PROLAPS II) was a randomized, double-blind, placebo-controlled, investigator-initiated, superiority study aimed at assessing the efficacy and safety of extended prophylaxis with rivaroxaban after laparoscopic surgery for colorectal cancer. Consecutive patients who had laparoscopic surgery for colorectal cancer were randomized to receive rivaroxaban (10 mg once daily) or a placebo to be started at 7 ± 2 days after surgery and given for the subsequent 3 weeks. All patients received antithrombotic prophylaxis with low-molecular-weight heparin from surgery to randomization. The primary study outcome was the composite of symptomatic objectively confirmed VTE, asymptomatic ultrasonography-detected deep vein thrombosis (DVT), or VTE-related death at 28 ± 2 days after surgery. The primary safety outcome was major bleeding. Patient recruitment was prematurely closed due to study drug expiry after the inclusion of 582 of the 646 planned patients. A primary study outcome event occurred in 11 of 282 patients in the placebo group compared with 3 of 287 in the rivaroxaban group (3.9 vs 1.0%; odds ratio, 0.26; 95% confidence interval [CI], 0.07-0.94; log-rank P = .032). Major bleeding occurred in none of the patients in the placebo group and 2 patients in the rivaroxaban group (incidence rate 0.7%; 95% CI, 0-1.0). Oral rivaroxaban was more effective than placebo for extended prevention of VTE after laparoscopic surgery for colorectal cancer without an increase in major bleeding. This trial was registered at www.clinicaltrials.gov as #NCT03055026.

Cytogenetics has long represented a critical component in the clinical evaluation of hematologic malignancies. Chromosome banding studies provide a simultaneous snapshot of genome-wide copy number and structural variation, which have been shown to drive tumorigenesis, define diseases, and guide treatment. Technological innovations in sequencing have ushered in our present-day clinical genomics era. With recent publications highlighting novel sequencing technologies as alternatives to conventional cytogenetic approaches, we, an international consortium of laboratory geneticists, pathologists, and oncologists, describe herein the advantages and limitations of both conventional chromosome banding and novel sequencing technologies and share our considerations on crucial next steps to implement these novel technologies in the global clinical setting for a more accurate cytogenetic evaluation, which may provide improved diagnosis and treatment management. Considering the clinical, logistic, technical, and financial implications, we provide points to consider for the global evolution of cytogenetic testing.

Disseminated intravascular coagulation (DIC) is a syndrome triggered by infectious and noninfectious pathologies characterized by excessive generation of thrombin within the vasculature and widespread proteolytic conversion of fibrinogen. Despite diverse clinical manifestations ranging from thrombo-occlusive damage to bleeding diathesis, DIC etiology commonly involves excessive activation of blood coagulation and overlapping dysregulation of anticoagulants and fibrinolysis. Initiation of blood coagulation follows intravascular expression of tissue factor or activation of the contact pathway in response to pathogen-associated or host-derived, damage-associated molecular patterns. The process is further amplified through inflammatory and immunothrombotic mechanisms. Consumption of anticoagulants and disruption of endothelial homeostasis lower the regulatory control and disseminate microvascular thrombosis. Clinical DIC development in patients is associated with worsening morbidities and increased mortality, regardless of the underlying pathology; therefore, timely recognition of DIC is critical for reducing the pathologic burden. Due to the diversity of triggers and pathogenic mechanisms leading to DIC, diagnosis is based on algorithms that quantify hemostatic imbalance, thrombocytopenia, and fibrinogen conversion. Because current diagnosis primarily assesses overt consumptive coagulopathies, there is a critical need for better recognition of nonovert DIC and/or pre-DIC states. Therapeutic strategies for patients with DIC involve resolution of the eliciting triggers and supportive care for the hemostatic imbalance. Despite medical care, mortality in patients with DIC remains high, and new strategies, tailored to the underlying pathologic mechanisms, are needed.