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Background: Analysis of measurable residual disease (MRD) is increasingly being implemented in the clinical care of children and adults with acute myeloid leukemia (AML). However, MRD methodologies differ and discordances in results lead to difficulties in interpretation and clinical decision-making. The aim of this study was to compare results from reverse transcription quantitative polymerase chain reaction (RT-qPCR) and multiparameter flow cytometry (MFC) in childhood AML and describe the kinetics of residual leukemic burden during induction treatment. Methods: In 15 children who were treated in the NOPHO-AML 2004 trial and had fusion transcripts quantified by RT-qPCR, we compared MFC with RT-qPCR for analysis of MRD during (day 15) and after induction therapy. Eight children had RUNX1::RUNX1T1, one CBFB::MYH11 and six KMT2A::LLT3. Results: When ≥0.1% was used as cut-off for positivity, 10 of 22 samples were discordant. The majority (9/10) were MRD positive with RT-qPCR but MRD negative with MFC, and several such cases showed presence of mature myeloid cells. Fusion transcript expression was verified in mature cells as well as in CD34 expressing cells sorted from diagnostic samples. Conclusions: Measurement with RT-qPCR suggests slower response kinetics than indicated from MFC, presumably due to the presence of mature cells expressing fusion transcript. The prognostic impact of early measurements with RT-qPCR remains to be determined.

Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoma subtype, accounting for 30-40% of lymphoma diagnoses. Though aggressive, cure is achievable in approximately 60% of cases with primary chemo-immunotherapy, and in a further substantial minority by salvage therapy and autologous stem cell transplantation. Despite promising activity in early phase clinical trials, no intensified or novel treatment regimen has improved outcomes over R-CHOP21 in randomised studies. However, there remain several areas of controversy including the most appropriate prognostic markers, CNS prophylaxis and the optimal treatment for patients with high-risk disease. This position statement presents an evidence-based synthesis of the literature for application in Australasian practice. This article is protected by copyright. All rights reserved.

Von Willebrand factor (VWF) is a glycoprotein that is secreted into the circulation and controls bleeding by promoting adhesion and aggregation of blood platelets at sites of vascular injury. Substantial inter-individual variation in VWF plasma levels exists among the healthy population. Prior to secretion, VWF polymers are assembled and condensed into helical tubules, which are packaged into Weibel-Palade bodies (WPBs), a highly specialized post-Golgi storage compartment in vascular endothelial cells. In the inherited bleeding disorder Von Willebrand disease (VWD), mutations in the VWF gene can cause qualitative or quantitative defects, limiting protein function, secretion, or plasma survival. However, pathogenic VWF mutations cannot be found in all VWD cases. Although an increasing number of genetic modifiers have been identified, even more rare genetic variants that impact VWF plasma levels likely remain to be discovered. Here, we summarize recent evidence that modulation of the early secretory pathway has great impact on the biogenesis and release of WPBs. Based on these findings, we propose that rare, as yet unidentified quantitative trait loci influencing intracellular VWF transport contribute to highly variable VWF levels in the population. These may underlie the thrombotic complications linked to high VWF levels, as well as the bleeding tendency in individuals with low VWF levels.

Purpose: Few studies have compared the effects of low-concentration (5%) and high-concentration (10%) intravenous immunoglobulin (IVIG) preparations for patients with Kawasaki disease (KD) in the acute phase. The purpose of this study was to compare outcomes between low- and high-concentration IVIG preparations in children with KD, using a national inpatient database in Japan.Method: We used the Diagnostic Procedure Combination database to identify patients with KD treated with IVIG from April 2012 to March 2020. We identified those receiving high- and low-concentration IVIG preparations as an initial treatment. The outcomes included the proportions of patients with coronary artery abnormalities (CAAs) and IVIG resistance, length of stay, and medical costs. Propensity score-matched analyses were conducted to compare the outcomes between the two groups. Instrumental variable analyses were performed to confirm the results.Result: We identified 48,046 patients with KD and created 4:1 propensity score-matched pairs between the low- and high-concentration IVIG groups. There was a significant difference in the percentage with IVIG resistance between the two groups (20.6% vs 24.1%; risk difference, 3.5% [95% confidence interval, 2.3-4.7]; p < 0.001). However, there was no significant difference in CAAs (1.6% vs 1.6%; risk difference, 0.013% [95% confidence interval, −0.34 to 0.37]; p = 0.953). The instrumental variable analyses showed similar results.Conclusions: The proportion of CAAs did not differ significantly between those receiving low- and high-concentration IVIG. To confirm the results of this study, prospective studies adjusting for duration of IVIG administration and duration of observation are needed.

Background Platelet-binding Von Willebrand Factor (VWF) strings assemble upon stimulated secretion from endothelial cells. Objectives To investigate the efficiency of platelet binding to multi-molecular VWF bundles secreted from endothelial cells and to investigate the role of osteoprotegerin, a protein located in Weibel-Palade bodies that interacts with the VWF platelet binding domain. Methods The nanobody VWF/AU-a11 that specifically binds to VWF in its active platelet-binding conformation was used to investigate the conformation of VWF. Results Upon stimulated secretion from endothelial cells, VWF strings were only partially covered with platelets, while a VWD-type 2B mutation or ristocetin enhanced platelet binding by 2-3-fold. Osteoprotegrin, reduces platelet adhesion to VWF by 40±18% in perfusion assays. siRNA-mediated down-regulation of endothelial osteoprotegerin expression resulted in a 1.8-fold increase in platelet adhesion to VWF strings. Upon viral infection, there is a concordant rise in VWF and osteoprotegerin plasma levels. Unexpectedly, no such increase was observed in plasma of desmopressin-treated hemophilia A-patients. In a mouse model, osteoprotegerin expression was low in liver endothelial cells of vehicle-treated mice, and concanavalin A-treatment increased VWF and osteoprotegerin expression 4- and 40-fold, respectively. This increase was translated in a 30-fold increased osteoprotegerin/VWF ratio in plasma. Conclusions Release of VWF from endothelial cells opens the platelet-binding site, irrespective of the presence of flow. However, not all available platelet-binding sites are being occupied, suggesting some extent of regulation. Part of this regulation involves endothelial proteins that are co-secreted with VWF, like osteoprotegerin. This regulatory mechanism may be of more relevance under inflammatory conditions.

The immune microenvironment in acute myeloid leukemia (AML) is closely related to patients’ prognosis. Long noncoding RNAs (lncRNAs) are emerging as key regulators in immune systems. In this study, we established a prognostic model using an immune‐related lncRNA (IRL) signature to predict AML patients’ overall survival (OS) through Least Absolute Shrinkage and Selection Operator (LASSO) and multivariate Cox regression analysis. Kaplan‐Meier analysis, receiver operating characteristic (ROC) analysis, univariate Cox regression, and multivariate Cox regression analyses further illustrated the reliability of our prognostic model. An IRL signature‐based nomogram consisting of other clinical features efficiently predicted the OS of AML patients. The incorporation of the IRL signature improved the ELN2017 risk stratification system's prognostic accuracy. In addition, we found that monocytes and metabolism‐related pathways may play a role in AML progression. Overall, the IRL signature appears as a novel effective model for evaluating the OS of AML patients and may be implemented to contribute to the prolonged OS in AML patients.

The echinoderm microtubule associated protein‐like 4 gene (EML4) encodes the predominant anaplastic lymphoma kinase (ALK) fusion partner in non‐small‐cell lung cancer (NSCLC); however, the dynactin subunit 1 (DCTN1)‐ALK rearrangement is extremely rare. The co‐occurrence of primary epidermal growth factor receptor (EGFR) T790M mutation with EGFR exon 19 deletion (del) in patients with NSCLC is uncommon. Here we report a female lung adenocarcinoma patient with brain metastases and possible coexistence of primary EGFR T790M mutation/EGFR exon 19 del/DCTN1‐ALK translocation. The patient received multiline treatment including chemotherapy, antivascular, and targeted therapies. To overcome developed resistance to chemotherapy or targeted therapy to prolong overall survival, the patient's circulating tumor DNA (ctDNA) was dynamically monitored. The patient responded to successive osimertinib and alectinib treatment, and alectinib achieved a nearly complete response for lung and brain lesions after she acquired osimertinib resistance. Furthermore, we summarize 22 published cases of patients with lung adenocarcinoma with concurrent EGFR mutation and ALK rearrangement, including details of clinical characteristics, natural history, and pertinent therapy of this uncommon tumor subtype. This literature review shows that EGFR inhibition was an indispensable aspect of the treatment of patients with EGFR/ALK co‐alterations in the pre‐alectinib era and that ALK inhibition with crizotinib did not show more eye‐catching therapeutic results. Considering the effectiveness achieved by alectinib, this case study provides a new perspective for the treatment of lung cancer brain metastasis patients with concurrent EGFR/ALK mutations.

Nucleophosmin-1 (NPM1) mutations in acute myeloid leukemia (AML) confer a survival advantage in the absence of FLT3-internal tandem duplication (FLT3-ITD). Here, we investigated the main predictors of outcome after allogeneic hematopoietic stem cell transplantation (allo-HCT). We identified 1572 adult (age >= 18 year) patients with NPM1-mutated AML in first complete remission (CR1:78%) or second complete remission (CR2:22%) who were transplanted from matched sibling donors (30.8%) or unrelated donors (57.4%) between 2007 and 2019 at EBMT participating centers. Median follow-up for survivors was 23.7 months. FLT3-ITD was present in 69.3% of patients and 39.2% had detectable minimal/measurable residual disease (MRD) at transplant. In multivariate analysis, relapse incidence (RI) and leukemia-free survival (LFS) were negatively affected by concomitant FLT3-ITD mutation (HR 1.66 p = 0.0001, and HR 1.53, p < 0.0001, respectively), MRD positivity at transplant (HR 2.18, p < 10(-5) and HR 1.71, p < 10(-5), respectively), and transplant in CR2 (HR 1.36, p = 0.026, and HR 1.26, p = 0.033, respectively), but positively affected by Karnofsky score >= 90 (HR 0.74, p = 0.012, and HR 0.7, p = 0.0002, respectively). Overall survival (OS) was also negatively influenced by concomitant FLT3-ITD (HR 1.6, p = 0.0001), MRD positivity at transplant (HR 1.61, p < 10(-5)), and older age (HR 1.22 per 10 years, p < 0.0001), but positively affected by matched sibling donor (unrelated donor: HR 1.35, p = 0.012; haploidentical donor: HR 1.45, p = 0.037) and Karnofsky score >= 90 (HR 0.73, p = 0.004). These results highlight the independent and significant role of FLT3-ITD, MRD status, and disease status on posttransplant outcomes in patients with NPM1-mutated AML allowing physicians to identify patients at risk of relapse who may benefit from posttransplant prophylactic interventions.

Dopamine functioning is an essential mechanism underlying addictive behaviors. This paper evaluates the association of Internet gaming disorder (IGD) with the catechol‐O‐methyltransferase (COMT) val158met polymorphism and examines the roles of impulsivity and reinforcement sensitivity in this association. Using diagnostic interviews, this study recruited 69 participants with IGD and 138 participants without. All participants underwent diagnostic interviews for IGD and an evaluation for the COMT val158met polymorphism, impulsivity, and reinforcement sensitivity. Among participants with the Val/Val genotype, the odds ratio (95% confidence interval) for IGD was 2.09 (1.15–3.80). The IGD–Val/Val genotype association was mediated by impulsivity and fun‐seeking. The Val/Val genotype is indicative of low frontal functioning and is a predictive factor of IGD, with this effect being confounded by impulsivity and fun‐seeking. Interventions targeting impulsivity and fun‐seeking might attenuate the risk of IGD, particularly among individuals with the Val/Val genotype. Additional studies are necessary to elucidate the possible role of dopamine functioning.

Aim and objectives To describe young children's experiences of valuable support in managing their fears about treatment for acute lymphoblastic leukaemia. The focus was specifically on support from parents and healthcare professionals. Design The study had a qualitative descriptive longitudinal design. Methods The study analysed 35 interviews with 13 children at three different times during their treatment period. Data were analysed using a matrix‐based method. The Consolidated criteria for reporting qualitative research (COREQ) guidelines have been followed. Results Parents and healthcare professionals provide important support to children undergoing treatment for acute lymphoblastic leukaemia, although their roles differ. Children valued their parents’ closeness and advocacy, being able to participate in their own care, and being given pain relief during procedures known to create pain. Valued support from healthcare professionals changed over time, from providing information and showing the tools that would be used in procedures, to paying attention to the child's needs and desires. It was more important for children to be able to choose between different alternatives in medical procedures than deciding on major treatment issues.

Background Most patients with methotrexate‐associated lymphoproliferative disorder (MTX‐LPD) show diffuse large B‐cell lymphoma (DLBCL) or classic Hodgkin lymphoma (CHL) types. Patients with MTX‐LPD often have spontaneous remission after MTX discontinuation, but chemotherapeutic intervention is frequently required in patients with CHL‐type MTX‐LPD. In this study, we examined whether programmed cell death‐ligand 1 (PD‐L1) expression levels were associated with the prognosis of MTX‐LPD after MTX discontinuation. Methods A total of 72 Japanese patients diagnosed with MTX‐LPD were clinicopathologically analyzed, and immunohistochemical staining of PD‐L1 was performed in 20 DLBCL‐type and 24 CHL‐type MTX‐LPD cases to compare with the clinical course. Results PD‐L1 was expressed in 5.0% (1/20) of patients with DLBCL‐type MTX‐LPD, whereas it was expressed in 66.7% (16/24) of the patients with CHL‐type MTX‐LPD in more than 51% of tumor cells. Most CHL‐type MTX‐LPD patients with high PD‐L1 expression required chemotherapy owing to exacerbations or relapses after MTX discontinuation. However, no significant differences in clinicopathologic findings at diagnosis were observed between PD‐L1 high‐ and low‐expression CHL‐type MTX‐LPD. Conclusion PD‐L1 expression was significantly higher in patients with CHL‐type than DLBCL‐type MTX‐LPD, suggesting the need for chemotherapy in addition to MTX discontinuation in CHL‐type MTX‐LPD patients to achieve complete remission. No association was observed between PD‐L1 expression levels and clinical findings at diagnosis, suggesting that PD‐L1 expression in tumor cells influences the pathogenesis of CHL‐type MTX‐LPD after MTX discontinuation., Patients with classic Hodgkin lymphoma (CHL)‐type methotrexate‐associated lymphoproliferative disorder (MTX‐LPD) with high programmed cell death‐ligand 1 (PD‐L1) expression tended to have exacerbations and relapses after MTX discontinuation. Our study suggests that the PD‐1/PD‐L1 pathway may be involved in refractoriness to MTX discontinuation in CHL‐type MTX‐LPD.

How Epstein–Barr virus (EBV)‐positive diffuse large B‐cell lymphoma (DLBCL) occasionally occurs following chronic inflammation remains to be elucidated. The case of a 57‐year‐old man who developed pulmonary EBV‐positive DLBCL from underlying silicosis lesions is presented. Immunohistochemical examination of the resected silicosis lesions showed predominant helper T cells and M1/M2 macrophages, with a lack of B cells, regulatory T cells, and resident memory T cells. Two years later, EBV‐positive DLBCL emerged unexpectedly from the silicosis. The imbalance of the immune cells in the microenvironment, at least in part, may help explain how chronic inflammation contributes to EBV‐positive DLBCL., The underlying silicosis lesions showed predominant Th cells and M1/M2 macrophages, with a lack of B cells, Treg cells, and TRM cells. The imbalance of the immune cells in the microenvironment, at least in part, may help to explain how chronic inflammation contributes to EBV‐positive diffuse large B‐cell lymphoma.

Background Anemia is a common comorbidity in patients with atrial fibrillation (AF). Reports on the association of anemia and adverse events in patients with AF, especially from Asia, are limited. Methods and Results Based on data from the Chinese Atrial Fibrillation Registry Study (CAFR), a total of 18,106 AF patients enrolled between August 2011 and December 2018 had hemoglobin (Hb) values recorded at baseline. Patients were classified into three groups according to Hb levels: 15,606 patients (86.2%) into the no anemia group (male Hb≥130 g/L; female Hb≥120 g/L), 1800 (9.9%) with mild anemia (male 110≤Hb

AML can be associated with autoimmune or inflammatory phenomena (AIP) occurring prior, concomitantly, or after its diagnosis. Trisomy 8 is one of the most common cytogenetic abnormalities associated with AML. We describe three patients with AML, trisomy 8, and associated AIP and review the known literature on this association. All of our patients had major symptomatic relief when treated with leukemia-directed therapy and corticosteroids. AIP in AML may be an underdiagnosed phenomenon, particularly in patients with trisomy 8.

Background: The bone marrow blast count is central to the diagnosis and monitoring of myelodysplastic syndromes (MDS). It is an independent risk factor for worse prognosis whether based on the morphology blast count or the flow cytometry (FC) myeloid progenitor (MyP) count. It is a principal population in FC MDS analysis also because once defined; it provides significant contributions to the overall FC MDS score. Methods: We elected to investigate inter-analyst agreement for the most fundamental parameter of the FC MDS diagnostic score: the MyP count. A common gating strategy was agreed and used by seven cytometrists for blind analysis of 34 routine bone marrows sent for MDS work-up. Additionally, we compared the results with a computational approach. Results: Concordance was excellent: Intraclass correlation was 0.993 whether measuring %MyP of total cells or CD45+ cells, and no significant difference was observed between files from different centers or for samples with abnormal MyP phenotypes. Computational and manual results were similar. Applying the common strategy to individual laboratories' control cohorts produced similar MyP reference ranges across centers. Conclusion: The FC MyP count offers a reliable diagnostic and prognostic measurement in MDS. The use of manual and computational approaches side by side may allow for optimizing both strategies. Considering its known prognostic power, the MyP count could be considered a useful and reliable addition to existing prognostic scoring systems.

Aims It is unknown whether Epstein-Barr virus (EBV) infection can occur in high-grade B-cell lymphoma with MYC and BCL2 and/or BCL6 rearrangements, also known as double-hit or triple-hit lymphoma (DHL/THL). Methods and results Here we report 16 cases of EBV+ DHL/THL from screening 846 cases of DHL/THL and obtaining additional EBV+ cases through multi-institutional collaboration: 8 MYC/BCL2 DHL, 6 MYC/BCL6 DHL, and 2 THL. There were 8 men and 8 women with a median age of 65 years (range, 32-86). Two patients had a history of follicular lymphoma and one had AIDS. Nine of 14 patients had an International Prognostic Index of ≥3. Half of the cases showed high-grade/Burkitt-like morphology and the other half diffuse large B-cell lymphoma morphology. By immunohistochemistry, the lymphoma cells were positive for MYC (n=14/16), BCL2 (n=12/16), BCL6 (n=14/16), CD10 (n=13/16), and MUM1 (n=6/14). By Hans algorithm, 13 cases were classified as GCB and 3 as non-GCB. The lymphomas frequently showed an EBV latency type I with a median EBV-encoded small RNAs of 80% positive cells (range, 20-100%). After a median follow-up of 36.3 months (range, 2.0-41.6), 7 patients died with a median survival of 15.4 months (range, 3.4-47.3) after diagnosis of EBV+ DHL/THL. Five of 6 patients with MYC/BCL6 DHL were alive including 4 in complete remission. In contrast, only 4/10 patients with MYC/BCL2 DHL or THL were alive including 2 in complete remission. The median survival in patients with MYC/BCL6 DHL was unreached and was 21.6 months in patients with MYC/BCL2 DHL or THL. Conclusions EBV infection in DHL/THL is rare (~1.5%). Cases of EBV+ DHL/THL are largely similar to their EBV-negative counterparts clinicopathologically. Our findings expand the spectrum of EBV+ B-cell lymphomas currently recognized in the WHO classification and suggest differences between EBV+ MYC/BCL2 and MYC/BCL6 DHL that may have therapeutic implications.

The BCR-ABL1 inhibitor ponatinib is approved for the treatment of adults with chronic myeloid leukemia or Philadelphia chromosome-positive acute lymphoblastic leukemia, including those with the T315I mutation. We report a population pharmacokinetic (PK) model-based analysis for ponatinib and its application to inform dose selection for pediatric development. Plasma concentration-time data were collected from 260 participants (86 healthy volunteers; 174 patients with hematologic malignancies) enrolled across 7 clinical trials. Data were analyzed using nonlinear mixed-effects modelling. Ponatinib PK was described by a 2-compartment model with first-order elimination from the central compartment. The final model included body weight and age as covariates on the apparent central volume of distribution; however, exposure variability explained by these covariates was small compared with overall variability in the population. None of the covariates evaluated, including sex, age (19-85 years), race, body weight (40.7-152.0 kg), total bilirubin (0.1-3.16 mg/dL), alanine aminotransferase (6-188 U/L), albumin (23.0-52.5 g/L), and creatinine clearance (≥28 mL/min) had clinically meaningful effects on apparent oral clearance. Simulations based on the final model predicted that daily doses of 15-45 mg result in steady-state average concentrations that are in the pharmacological range for BCR-ABL1 inhibition and approximate or exceed concentrations associated with suppression of T315I mutant clones. The final model was adapted using allometric scaling to inform dose selection for pediatric development. Clinicaltrials.gov identifier NCT00660920; NCT01667133; NCT01650805 This article is protected by copyright. All rights reserved.

Several lines of research suggest that BCL-XL-mediated anti-apoptotic effects may contribute to pathogenesis of myeloproliferative neoplasms driven by JAK2V617F and serve as therapeutic target. Here, we used a knock-in JAK2V617F mouse model and confirmed that Bcl-xl was overexpressed in erythroid progenitors. MPN-induced phenotype in the peripheral blood by conditional knockin of JAK2V617F was abrogated by conditional knockout of Bcl2l1, which presented anemia and thrombocytopenia independently of JAK2 mutation status. The Mx1-Cre Jak2V617W/VF /Bcl2l1f/f mice presented persistent splenomegaly as a result of extramedullary hematopoiesis and pro-apoptotic stimuli in terminally differentiated erythroid progenitors. The pan-BH3 mimetic inhibitor obatoclax showed superior cytotoxicity in JAK2V617F cell models, and reduced clonogenic capacity in ex vivo assay using Vav-Cre Jak2V617F bone marrow cells. Both ruxolitinib and obatoclax significantly reduced spleen weights in a murine Jak2V617F MPN model but did not show additive effect. The tumor burden reduction was observed with either ruxolitinib or obatoclax in terminal differentiation stage neoplastic cells but not in myeloid-erythroid precursors. Therefore, disrupting BCL2 balance is not sufficient to treat MPN at the stem cell level, but it is certainly an additional option for controlling the critical myeloid expansion of the disease.

INTRODUCTION This study aimed to explore the application value of two laboratory tests in the diagnosis of Epstein-Barr virus-associated infectious mononucleosis (EBV-IM) in children. METHODS From January 2018 to December 2020, 166 patients with EBV-IM were included in this study. Two methods were used in the analysis. The results of both tests were compared and analyzed. RESULTS The age of onset of EBV-IM is mainly distributed in the range of 0-6 years, and no difference by gender is observed. The sensitivity and specificity of EBV testing by PCR were 49.4% and 89.8%, respectively, and the area under the receiver operating characteristic curve (AUC) was 0.714 (0.662-0.762). When using the immunofluorescence method to detect EBV antibodies, the two indices with the highest diagnostic efficacy were low-affinity EBV-CA IgG and EBV-CA IgM, and their AUC values were 0.798 (0.751-0.840) and 0.663 (0.609-0.713), respectively. When combining the two indices for testing, the AUC values of EBV-CA IgM + low-affinity EBV-CA IgG, EBV-DNA + EBV-CA IgM, and EBV-DNA + low-affinity EBV-CA IgG were 0.904(0.867-0.933), 0.768 (0.719-0.812) and 0.963 (0.937-0.981), respectively. The diagnostic efficacy of the combined EBV-DNA + EBV-CA IgM + low-affinity EBV-CA IgG test was optimal compared with that of a single index or the combination of two indices, with an AUC of 0.999 (0.986-1.000) (P < 0.05), sensitivity of 100%, and specificity of 89.8%. CONCLUSIONS The combined immunofluorescence and real-time PCR method has high sensitivity and specificity and good application value in the clinical diagnosis of EBV-IM. This article is protected by copyright. All rights reserved.

Penttinen type of premature aging syndrome is an autosomal-dominant disorder that can be caused by the c.1994T>A pVal665Ala pathogenic variant in platelet-derived growth factor receptor-B (PDGFRB). Imatinib, a receptor tyrosine kinase (RTK) inhibitor, has been used in Penttinen syndrome (PS) patients with good results. A 21-year-old male presented shortly after birth with a prematurely aged appearance with distinctive facial features and cutaneous atrophy with hypertrophic scar-like lesions. Generalized brachydactyly with acro-osteolysis was observed. Flexion contractures limited his daily activities. Cognitive impairment was not present. Genetic testing found a heterozygous variant c.1994T>A pVal665Ala in exon 14 of PDGFRB. A diagnosis of PS was made and imatinib treatment was started with partial response. After lack of further improvement, in vitro molecular studies with imatinib and dasatinib showed that the Val665Ala variant had greater sensitivity to dasatinib than imatinib. This was seen examining levels of P-PDGFRB directly and on downstream ligands P-AKT and P-STAT. Improved clinical response was observed after treatment with dasatinib. We report a new case of PS with clinical and molecular response to dasatinib after incomplete response to imatinib. Our work provides further molecular and clinical evidence of RTK inhibitors' efficacy in this rare disorder.

The transformation of chronic lymphocytic leukemia (CLL) to high-grade B-cell lymphoma is known as Richter syndrome (RS), a rare event with dismal prognosis. In this study, we conducted whole-genome sequencing (WGS) of paired circulating CLL (PB-CLL) and RS biopsies (tissue-RS) from 17 patients recruited into a clinical trial (CHOP-O). We found that tissue-RS was enriched for mutations in poor-risk CLL drivers and genes in the DNA damage response (DDR) pathway. In addition, we identified genomic aberrations not previously implicated in RS, including the protein tyrosine phosphatase receptor (PTPRD) and tumor necrosis factor receptor–associated factor 3 (TRAF3). In the noncoding genome, we discovered activation-induced cytidine deaminase–related and unrelated kataegis in tissue-RS affecting regulatory regions of key immune-regulatory genes. These include BTG2, CXCR4, NFATC1, PAX5, NOTCH-1, SLC44A5, FCRL3, SELL, TNIP2, and TRIM13. Furthermore, differences between the global mutation signatures of pairs of PB-CLL and tissue-RS samples implicate DDR as the dominant mechanism driving transformation. Pathway-based clonal deconvolution analysis showed that genes in the MAPK and DDR pathways demonstrate high clonal-expansion probability. Direct comparison of nodal-CLL and tissue-RS pairs from an independent cohort confirmed differential expression of the same pathways by RNA expression profiling. Our integrated analysis of WGS and RNA expression data significantly extends previous targeted approaches, which were limited by the lack of germline samples, and it facilitates the identification of novel genomic correlates implicated in RS transformation, which could be targeted therapeutically. Our results inform the future selection of investigative agents for a UK clinical platform study. This trial was registered at www.clinicaltrials.gov as #NCT03899337.

Sickle cell disease (SCD) is characterized by frequent and unpredictable vaso-occlusive crises (VOCs). Sickle erythrocytes (SSRBCs) contribute to VOCs by participating in a series of adhesive events with blood cells and the vascular endothelium. Adhesion assays have been used to evaluate the relationship between SSRBC adhesion and SCD severity. We developed a standardized, clinical flow adhesion assay of whole blood to vascular cell adhesion molecule (FA-WB-VCAM). The objective of this study was to assess the variability and clinical predictive value of FA-WB-VCAM in a six-month longitudinal, observational study (ELIPSIS) in SCD subjects during at-home, steady-state and self-reported VOCs, and following VOC resolution. We observed a strong relationship between FA-WB-VCAM and SCD severity. Adhesion indices were significantly lower in SCD subjects on hydroxycarbamide and increased during VOCs; at-home VOCs had significantly higher FA-WB-VCAM than steady-state and contact VOCs. SCD subjects with a high frequency of self-reported VOCs had a pro-adhesive phenotype at steady state and were stratified into a high-adhesive phenotype cohort; two years prospectively we observed a higher frequency of VOCs in the high-adhesion cohort. This study supports stratifying SCD subjects based on steady-state FA-WB-VCAM and suggests that FA-WB-VCAM may be a plausible surrogate end-point for SCD severity.

Myelodysplastic syndromes (MDS) are a group of malignant hematological disorders characterized by the abnormal development of hematopoietic stem cells and increased risk of acute myelogenous leukemia. Although the pathogenesis of MDS has not been fully understood, various alterations of microRNAs (miRNAs) have been reported in MDS. This study aimed to explore the molecular mechanisms of MDS by integrative bioinformatics analysis of miRNAs expression profile. The GSE81372 expression profile dataset was downloaded from Gene Expression Omnibus database. The differentially expressed miRNAs (DEMs) between MDS and normal controls were identified and targets of miRNAs were predicted. Subsequently, gene ontology (GO) functional and pathway enrichment analyses of target genes were performed. Finally, pathway relation network and miRNA–GO regulatory network were constructed and analyzed. A total of six upregulated and 35 downregulated DEMs were identified. The results showed that target genes of DEMs mainly participated in the process of signal transduction, blood coagulation, apoptotic process, cell proliferation, transmembrane transport, and angiogenesis. The significantly enriched pathways included MAPK signaling pathway, PI3K‐Akt signaling pathway, TGF‐beta signaling pathway, Hippo signaling pathway, and P53 signaling pathway. Moreover, miR‐195‐5p, miR‐4505, miR‐22‐3p, and miR‐148a‐3p were selected as hub miRNAs in miRNA–GO regulatory network and their aberrant expression might be closely associated with MDS pathogenesis. Our discovery provides a registry of miRNAs and pathways that are disrupted in MDS, which has the potential to be used in clinic for diagnosis and target therapy of MDS in future.

The association of previous hepatitis B virus (HBV) exposure [hepatitis B surface antigen (HBsAg) negative, hepatitis B core antibody (anti-HBc/HBcAb) positive] with disease severity and decision on treatment option in primary immune thrombocytopenia (ITP) patients remains unclear. Data from 725 patients diagnosed with ITP were analyzed to elucidate the association between anti-HBc serological status and disease severity. Data from a published prospective study [high-dose dexamethasone (HD-DXM), HD-DXM plus recombinant human thrombopoietin, NCT01734044] and two retrospective studies (standard-dose and low-dose rituximab) were rearranged to evaluate the impact of anti-HBc serological status on the response and outcome to ITP-specific treatments and the risk of HBV reactivation related to these treatments. The prevalence of HBsAg- HBcAb+ and HBsAg- HBcAb- in ITP patients was 51·03% and 48·97% respectively. Compared to the HBsAg- HBcAb- group, patients in the HBsAg- HBcAb+ group had lower platelet count, higher bleeding score, and longer hospitalization (P = 0·002, 0·033, and 0·008 respectively). Moreover, the initial complete response rate of HBsAg- HBcAb+ patients was lower than that of HBsAg- HBcAb- patients (45·2% vs 59·8%, P = 0·027). In conclusion, previous HBV exposure was correlated with disease severity and hospitalization in ITP patients. Anti-HBc positivity may be considered as a predictor for poor response to ITP-specific treatments.

Epigenetic alterations of chromatin structure affect chromatin accessibility and collaborate with genetic alterations in the development of cancer. Lysine demethylase 4B (KDM4B) has been identified as a JmjC domain‐containing epigenetic modifier that possesses histone demethylase activity. Although recent studies have demonstrated that KDM4B positively regulates the pathogenesis of multiple types of solid tumors, the tissue specificity and context dependency have not been fully elucidated. In this study, we investigated gene expression profiles established from clinical samples and found that KDM4B is elevated specifically in acute myeloid leukemia (AML) associated with chromosomal translocation 8;21 [t(8;21)], which results in a fusion of the AML1 and the eight‐twenty‐one (ETO) genes to generate a leukemia oncogene, AML1‐ETO fusion transcription factor. Short hairpin RNA‐mediated KDM4B silencing significantly reduced cell proliferation in t(8;21)‐positive AML cell lines. Meanwhile, KDM4B silencing suppressed the expression of AML1‐ETO‐inducible genes, and consistently perturbed chromatin accessibility of AML1‐ETO‐binding sites involving altered active enhancer marks and functional cis‐regulatory elements. Notably, transduction of murine KDM4B orthologue mutants followed by KDM4B silencing demonstrated a requirement of methylated‐histone binding modules for a proliferative surge. To address the role of KDM4B in leukemia development, we further generated and analyzed Kdm4b conditional knockout mice. As a result, Kdm4b deficiency attenuated clonogenic potential mediated by AML1‐ETO and delayed leukemia progression in vivo. Thus, our results highlight a tumor‐promoting role of KDM4B in AML associated with t(8;21).

The definitive efficacy of anaplastic lymphoma kinase (ALK) inhibitors in ALK positive lung squamous cell carcinoma (sqCC) patients remain unclear. Here, we report a case in which brigatinib had a therapeutic effect on ALK‐positive lung squamous cell carcinoma. The patient in this report was diagnosed with ALK‐positive lung squamous cell carcinoma with brain metastases, and received brigatinib after failure of first‐line chemotherapy. Response duration was approximately 11 months, with tolerable side effects. In conclusion, a good clinical effect was achieved in a patient with ALK positive lung squamous cell carcinoma who received treatment with an ALK inhibitor., Our case was first report to determine the excellent antitumor effect of Brigatinib on ALK positive lung SqCC. PFS was more than 11 months, and the side effects were tolerable.

Background Glucose-6-phosphate dehydrogenase (G6PD) deficiency, hereditary spherocytosis (HS), and alpha thalassemia (α-thal) are frequent erythrocyte pathologies with different geographic distributions worldwide. Our aim is to report hematological and molecular findings of G6PD deficient Mexican patients in coinheritance with suggestive hereditary spherocytosis (sHS) and α-thal. Methods We studied 78 G6PD deficiency patients. Hematological parameters, acidified glycerol lysis test, erythrocyte morphology, electrophoresis, and hemoglobin quantification were obtained. G6PD and HBA2/HBA1 variants were identified using ARMS-PCR, Gap-PCR, or Sanger sequencing. Results Nine G6PD variants were identified; A-202A/376G , A-376G/968C , and A+376G as the most frequent. G6PD Santiago de Cuba1339A and Kamiube1387T were detected in Mexicans for first time. Hematological analysis revealed additional erythrocyte pathologies in 52 patients, 32 with positive osmotic fragility test and spherocytes in blood smear (suggestive hereditary spherocytosis, sHS), 12 with microcytosis and 8 with all three defects who had the most severe phenotype, with significantly lower hematological parameters (Hb, PCV, MCV, and MCH). α-thal variants (αHph α, α-59C>T α and -α3.7 ) were observed in 65% of patients with microcytosis. Conclusion Additional erythrocyte defects were observed in 69.3% of G6PD deficiency patients. We stress the importance of searching for the presence of additional erythrocyte hereditary diseases in patients with G6PD deficiency.

Due to the considerable success of cancer immunotherapy for leukemia, the tumor immune environment has become a focus of intense research; however, there are few reports on the dynamics of the tumor immune environment in leukemia. Here, we analyzed the tumor immune environment in pediatric B cell precursor acute lymphoblastic leukemia by analyzing serial bone marrow samples from nine patients with primary and recurrent disease by mass cytometry using 39 immunophenotype markers, and transcriptome analysis. High‐dimensional single‐cell mass cytometry analysis elucidated a dynamic shift of T cells from naïve to effector subsets, and clarified that, during relapse, the tumor immune environment comprised a T helper 1‐polarized immune profile, together with an increased number of effector regulatory T cells. These results were confirmed in a validation cohort using conventional flow cytometry. Furthermore, RNA transcriptome analysis identified the upregulation of immune‐related pathways in B cell precursor acute lymphoblastic leukemia cells during relapse, suggesting interaction with the surrounding environment. In conclusion, a tumor immune environment characterized by a T helper 1‐polarized immune profile, with an increased number of effector regulatory T cells, could contribute to the pathophysiology of recurrent B cell precursor acute lymphoblastic leukemia. This information could contribute to the development of effective immunotherapeutic approaches against B cell precursor acute lymphoblastic leukemia relapse., Tumor immune environment characterized by a T helper 1‐polarized immune profile was observed in recurrent B cell precursor acute lymphoblastic leukemia. Regulatory T cells were activated in recurrent B cell precursor acute lymphoblastic leukemia.

Pan-proteasome inhibitors (pPIs) significantly improve outcomes in patients with multiple myeloma; however, their indiscriminate inhibition of multiple proteasome and immunoproteasome subunits causes diverse toxicities, including thrombocytopenia. We investigated the mechanisms underlying the platelet depletion induced by the pPIs bortezomib, carfilzomib, and ixazomib. An established thrombocytopenia model was adapted for each compound (bortezomib, ixazomib, and carfilzomib) to compare the following two pharmacodynamic mechanisms: a reversible inhibition of new progenitor cell formation (the myelosuppression model) and a reversible effect on the function of megakaryocytes to bud new platelets (platelet formation model). Bortezomib, ixazomib, and carfilzomib plasma concentration profiles and platelet counts were extracted from the literature. Pharmacokinetic (PK) and thrombocytopenia models were developed to predict the PK of these drugs and to describe their effects on proliferating cells and platelet budding. The PK models reproduced the exposure of the three compounds at steady state well compared with those reported in the literature. Both the platelet formation and myelosuppression models seemed able to describe the platelet depletion caused by bortezomib, ixazomib, and carfilzomib. Estimated structural parameters in the myelosuppression model were in the range of the values reported in the literature, whereas the mean transit time estimated with the platelet formation model was 3-fold to 10-fold higher than the highest reported value. The model of drug-induced myelosuppression yielded estimates of structural parameters in the range of those previously reported. The platelet formation model captured the temporal variation reported in clinical studies.

Aims Imatinib is considered the standard first line treatment in newly diagnosed patients with chronic-phase myeloid leukaemia (CML). Several imatinib population pharmacokinetic (popPK) models have been developed. However, their predictive performance has not been well established when extrapolated to different populations. Therefore, this study aimed to perform an external evaluation of available imatinib popPK models developed mainly in adult patients, and to evaluate the improvement in individual model-based predictions through Bayesian forecasting computed by each model at different treatment occasions. Methods A literature review was conducted through PubMed and Scopus to identify popPK models. Therapeutic drug monitoring data collected in adult CML patients treated with imatinib was used for external evaluation including prediction- and simulated-based diagnostics together with Bayesian forecasting analysis. Results Fourteen imatinib popPK studies were included for model-performance evaluation. A total of 99 imatinib samples were collected from 48 adult CML patients undergoing imatinib treatment with a minimum of one plasma concentrations measured at steady-state between January 2016 and December 2020. The model proposed by Petain et al. showed the best performance concerning prediction-based diagnostics in the studied population. Bayesian forecasting demonstrated a significant improvement in predictive performance at the second visit. Inter-occasion variability contributed to reducing bias and improving individual model-based predictions. Conclusions Imatinib popPK studies developed in Caucasian subjects including α1-acid glycoprotein showed the best model performance in terms of overall bias and precision. Moreover, two imatinib samples from different visits appear sufficient to reach an adequate model-based individual prediction performance trough Bayesian forecasting.

Immune thrombocytopenia (ITP) is an autoimmune haemorrhagic disease, in which the overactivation of T cells is crucial in the pathogenesis. Atorvastatin (AT), a lipid-lowering medicine, has shown promising immunomodulatory effects in certain inflammatory conditions. However, the immunoregulatory role of AT in ITP remains elusive. To investigate the effect of AT in the treatment of ITP, cluster of differentiation 4 (CD4)+ T cells were isolated from patients with ITP and cultured with different dosages of AT. We found that AT significantly inhibited cell proliferation, led to cell cycle arrest, induced apoptosis, and repressed the activation of CD4+ T cells in vitro. ITP murine models were then established, and results showed that AT treatment led to faster recovery of the platelet count to normal and exhibited comparable immunomodulatory function. Furthermore, we found the phosphorylation of mammalian target of rapamycin (mTOR), protein kinase B (AKT) and extracellular signal-regulated kinase (ERK), as well as activation of rat sarcoma virus (RAS) were all reduced dramatically after AT treatment in vitro. In conclusion, our present study demonstrated that AT could reinstate the functions of CD4+ T cells by inhibiting the excessive activation, proliferation, and survival of CD4+ T cells in ITP via the RAS/mitogen-activated protein kinase kinase (MEK)/ERK and the mTOR/phosphatidylinositol-3 kinase (PI3K)/AKT pathway. Therefore, we propose that AT could be used as a potential therapeutic option for ITP by restoring the over-activated cellular immunity.

DISEASE OVERVIEW Chronic lymphocytic leukemia (CLL) is one of the most frequent types of leukemia. It typically occurs in elderly patients and has a highly variable clinical course. Leukemic transformation is initiated by specific genomic alterations that interfere with the regulation of proliferation and of apoptosis in clonal B-cells. DIAGNOSIS The diagnosis is established by blood counts, blood smears, and immunophenotyping of circulating B-lymphocytes, which identify a clonal B-cell population carrying the CD5 antigen as well as typical B-cell markers. PROGNOSIS AND STAGING The clinical staging systems provide prognostic information by using the results of physical examination and blood counts. Various biological and genetic markers provide additional prognostic information. Deletions of the short arm of chromosome 17 (del[17p]) and/or mutations of the TP53 gene predict resistance to chemoimmunotherapy and a shorter time to progression with most targeted therapies. The CLL international prognostic index integrates genetic, biological, and clinical variables to identify distinct risk groups of patients with CLL. THERAPY Only patients with active or symptomatic disease or with advanced Binet or Rai stages require therapy. When treatment is indicated, several therapeutic options exist: a combination of the B-cell lymphoma 2 (BCL2) inhibitor venetoclax with obinutuzumab, monotherapy with inhibitors of Bruton tyrosine kinase (BTK) such as ibrutinib and acalabrutinib, or chemoimmunotherapy. At relapse, the initial treatment may be repeated, if the treatment-free interval exceeds 3 years. If the disease relapses earlier, therapy should be changed using an alternative regimen. Patients with a del(17p) or TP53 mutation are usually resistant to chemotherapy and should, therefore, be treated with targeted agents. FUTURE CHALLENGES Combinations of targeted agents are now being investigated to create efficient, potentially curative therapies of CLL with fixed duration. One of the most relevant questions currently addressed in clinical trials is the comparison of monotherapies with BTK inhibitors with fixed duration combination therapies. Moreover, the optimal sequencing of targeted therapies remains to be determined. Alternative therapies are needed for patients with BTK and BCL2 inhibitor double-refractory disease.

The aim of the present study was to evaluate the significance of low-level minimal/measurable residual disease (MRD) during early consolidation treatment in adult BCR-ABL1-negative acute lymphoblastic leukaemia. The MRD load was monitored by immunoglobulin/T-cell receptor rearrangements and assessed as negative [complete MRD response (CMR)], positive non-quantifiable (MRDnq) and positive quantifiable (MRDq). MRDnq before the first and second consolidation blocks had a comparable negative effect on survival as MRDq. The 5-year overall survival for CMR, MRDnq and MRDq at week 11 was 74·0%, 42·3% and 35·0% respectively. No central nervous system infiltration and MRD at week 11 were independent prognostic factors for survival on multivariate analysis (hazard ratios 0·32 and 2·25).

Aim In order to further understand the feature of natural killer cell (NK) dysfunction in acute myeloid leukemia (AML), The distribution of NK cell subset the expression of the inhibitory receptors immunoglobulin and ITIM domain (TIGIT), killer cell lectin-like receptor (KLRG1), and the expression of maturation marker CD57 in NK cell subsets and their correlation with patient outcomes were analyzed in this study. Methods We collected peripheral blood (PB) and bone marrow (BM) samples from de novo AML (AML-DN) patients, patients who achieved complete remission after chemotherapy (AML-CR), and healthy individuals. An eight-color flow cytometry panel was used to identify different NK subsets and their expression of TIGIT, CD57 and KLRG1. Results Decreased percentage of CD56dim CD16+ NK cells was found only in the PB of AML-DN and AML-CR patients but not in the BM. The expression frequency of TIGIT and KLRG1 was elevated on NK cells from the PB of AML-DN patients, while it was recovered in AML-CR patients. Moreover, a higher percentage of CD57+ CD56dim CD16+ NK cells, representing a terminally differentiated NK subset with strong cytotoxic capacity but defective replication potential, was detected in the BM of AML-DN patients and predicted sub-optimal survival for patients. Conclusion The results indicated that the NK cell subsets in the PB of AML patients had an exhaustion phenotype, while the BM NK cells had a terminally differentiated phenotype, which correlated with short survival for AML patients.

CD49d, the α4 chain of the VLA-4 integrin, is a negative prognosticator in chronic lymphocytic leukemia (CLL) with a key role in CLL cell-microenvironment interactions mainly occurring via its ligands VCAM-1 and fibronectin. In the present study, we focused on EMILIN-1 (Elastin-MIcrofibriL-INterfacer-1), an alternative VLA-4 ligand whose role has been so far reported only in non-hematological settings, by investigating: i) the distribution of EMILIN-1 in CLL-involved tissues; ii) the capability of EMILIN-1 to operate, via its globular C1q (gC1q) domain, as additional adhesion ligand in CLL; iii) the functional meaning of EMILIN-1 gC1q/VLA-4 interactions in CLL. EMILIN-1 is widely present in the CLL-involved areas of bone marrow biopsies (BMBs) without difference between CD49d negative and positive cases, displaying at least three different expression patterns: "fibrillar", "dot-like" and "mixed". The lack in CLL-BMB of neutrophil elastase, whose proteolytic activity degrades EMILIN-1 and impairs EMILIN-1 function, suggests full functional EMILIN-1 in CLL independently of its expression pattern. Functionally, EMILIN-1 gC1q domain promotes adhesion of CLL cells through specific interaction with VLA-4, and releases pro-survival signals for CLL cells, as demonstrated by enhanced ERK and AKT phosphorylation and impairment of in-vitro-induced apoptosis. EMILIN-1/VLA-4 interaction can efficiently contribute to the maintenance of the neoplastic clone in CLL. This article is protected by copyright. All rights reserved.

BACKGROUND Up to 15% of people aged 60 and over are anemic, and the prevalence of anemia increases with age. In older men and women, anemia is associated with increases in the risk of death and all-cause hospitalization, poor functional capacity, quality of life, and depression. METHODS AND RESULTS We reviewed the literature describing anemia in aging populations, focusing on the specific diagnostic criteria of anemia and potential causes in older men and women. Even after extensive etiologic workup that involves careful medical history, physical examination, laboratory measurements, and additional studies such as bone marrow biopsy, anemia of aging is unexplained in up to 40% of older patients with anemia. As a result, treatment options remain limited. CONCLUSIONS The prevalence of unexplained anemia of aging (UAA; also called unexplained anemia of the elderly, UAE), its deleterious impacts on health, physical function, and quality of life, and the lack of effective treatment or therapy guidelines represent a compelling unmet clinical need. In this review and consensus document, we discuss the scope of the problem, possible causes of UAA, diagnostic criteria, and potential treatment options. Because even mild anemia is strongly linked to poor clinical outcomes, it should receive clinical attention rather than simply being considered a normal part of aging.

INTRODUCTION The combination of cytology and multiparametric flow cytometry (MFC) may be useful in the diagnosis of breast implant-associated anaplastic large cell lymphoma (BIA-ALCL) and may be a practical way to differentiate lymphoma from benign and reactive seromas. Although the Brazilian breast implant market is the second largest in the world, with several manufacturers and the almost exclusive use of textured implants, the occurrence of BIA-ALCL in Brazil is underreported. METHODS One hundred seventeen sequential collections of suspicious periprosthetic fluid (PF) from 105 Brazilian patients registered between March/2018 and March/2021 were evaluated by routine cytomorphology and flow cytometry. The combination of CD30, HLA-DR, and CD25 was used together with T and B lymphocyte and monocyte evaluation. The PF samples were divided into positive, acute reactive (neutrophilic exudate), or chronic reactive (macrophage or lymphocyte rich), and unavailable samples. RESULTS Nine BIA-ALCL positive cases (7.7%) were identified, with typical morphology and increased FSC/SSC dispersion, bright expression of CD30, CD25 and HLA-DR, and absence or weakness of T-cell antigens (CD3, CD8, CD4, CD5, and CD7). Reactive samples were acute (n = 18, 15.4%) and chronic (n = 70, 59.8%). Twenty samples were excluded. The mean age of BIA-ALCL patients was 50 years (31-57 years) and 35 years in reactive patients (20-69 years). CONCLUSION Use of MFC with a comprehensive antibody panel consisting of CD30 in conjunction with CD25 and HLA-DR can discriminate anaplastic cells of BIA-ALCL from lymphoid or neutrophilic reactive cells and should be considered in the initial evaluation of seroma.

Background Since azathioprine is associated with lymphoproliferative disorders in Epstein-Barr virus (EBV)-naive patients with inflammatory bowel disease, guidelines advise avoidance. No recommendations exist for autoimmune liver disease (AILD). Aims To evaluate EBV status and EBV-related complications in paediatric AILD. Methods Single-centre, retrospective, observational study of paediatric AILD. Results In 245 paediatric patients with AILD, azathioprine was used in 168 (68.6%) and mycophenolate mofetil in 69 (28.2%). EBV status was assessed in 18 (10.7%) prior to azathioprine and 6 (8.7%) MMF. Acute EBV infection was diagnosed in five patients while on immunosuppression, resulting in one transient hepatitis and one persistent hepatitis. There were no cases of lymphoproliferative disorder in native livers. Liver transplantation (LT) was performed in 39 (15.9%) patients, with 8 EBV IgG-negative at LT. Post-LT EBV viraemia developed in 29 (74.4%), first detected at median 26 days (IQR, 13-86). EBV IgG-negative recipients had higher peak viraemia (266 984 IU/mL [IQR, 41108-2429050] v 5333 [IQR, 2036-38770], P = .004) and longer time to peak viraemia (375 days [IQR, 251-884] v 70 [IQR, 21-604], P = .04). Early EBV-associated post-transplant lymphoproliferative disorder (PTLD) was diagnosed in two patients, both EBV-IgG negative with prior azathioprine. Conclusions Real-world data demonstrate that EBV serostatus is not routinely checked before immunosuppression for paediatric AILD. Lymphoproliferative disorder was not diagnosed in those with native livers; however, EBV IgG-negative LT recipients receiving EBV IgG-positive donor organs are at risk of early PTLD. Large multicentre studies with longer follow-up are required to further evaluate the risk.

Introduction Turoctocog alfa (NovoEight® ) is a B-domain-truncated recombinant factor VIII (FVIII) approved for patients with haemophilia A. Aim To investigate the long-term safety and efficacy of turoctocog alfa in routine clinical practice. Methods Guardian 5 was a prospective, multinational, non-interventional, post-authorisation safety study. Male previously treated patients (> 150 exposure days [EDs]) of any age with severe/moderately severe haemophilia A (FVIII ≤ 2%) and a negative inhibitor test prior to first dosing (independent of FVIII-inhibitor history) were included to receive prophylaxis or on-demand treatment. The primary endpoint was the proportion of patients developing FVIII inhibitors (≥.6 Bethesda Units [BU]) after baseline visit, measured as per routine practice of each study site during clinic visits. Secondary endpoints included haemostatic effect, annualised bleeding rate (ABR), and adverse reactions assessment. The study concluded when 50 patients reached 100 EDs/patient minimum. Results Seventy patients were screened and 68 exposed to turoctocog alfa; 63 (92.6%) were on prophylaxis and five received on-demand treatment. Six (8.8%) patients reported a history of positive inhibitors. During the study, patients were exposed to turoctocog alfa for a mean (standard deviation) of 131.9 (99.0) days/patient. Fifty-five of 58 patients who completed the study were tested for FVIII inhibitors; no positive tests were reported. Overall success rate of turoctocog alfa for treatment of bleeds was 87.3%. Among patients receiving prophylaxis, median (range) ABR was 1.97 (.0-25.5) bleeds/year; estimated ABR (negative binomial model) was 3.65 (95% confidence interval: 2.53-5.25). Conclusion Turoctocog alfa was safe and efficacious for haemophilia A treatment in routine clinical practice.

Natural killer (NK) cells exert anti-viral effects after haematopoietic stem cell transplantation (HSCT). The balance between inhibition and activation of NK cells determined by the inherited repertoire of killer cell immunoglobulin-like receptors (KIR) genes may influence Epstein-Barr virus (EBV) reactivation after transplantation. To evaluate the relative contributions of KIR genotypes to EBV reactivation, we prospectively enrolled 300 patients with malignant haematological disease who were suitable for haploidentical HSCT. Univariate analysis showed that donors with KIR2DS1, KIR2DS3 or KIR3DS1 genes were associated with an increased risk of EBV reactivation [hazard ratio (HR) 1·86, 95% confidence interval (CI) 1·19-2·9, P = 0·0067; HR 1·78, 95% CI 1·07-2·97, P = 0·027; HR 1·86, 95% CI 1·19-2·91, P = 0·0065 respectively]. Multivariate analysis revealed that the presence of KIR2DS1, KIR2DS3 or KIR3DS1 genes was associated with increased EBV reactivation after HSCT. This effect was more evident in the absence of the cognate ligands for the corresponding activating receptors. Our present data firstly showed that donors with activating KIR genes, specifically activating KIR2DS1, KIR2DS3 and KIR3DS1, had an increased risk of EBV reactivation. Precaution for patients whose donors carry activating genes will help prevent EBV reactivation and improve patient prognosis after HSCT.

Acute myeloid leukaemia (AML) is an aggressive form of blood cancer that carries a dismal prognosis. Several studies suggest that the poor outcome is due to a small fraction of leukaemic cells that elude treatment and survive in specialised, oxygen (O2)‐deprived niches of the bone marrow. Although several AML drug targets such as FLT3, IDH1/2 and CD33 have been established in recent years, survival rates remain unsatisfactory, which indicates that other, yet unrecognized, mechanisms influence the ability of AML cells to escape cell death and to proliferate in hypoxic environments. Our data illustrates that Carbonic Anhydrases IX and XII (CA IX/XII) are critical for leukaemic cell survival in the O2‐deprived milieu. CA IX and XII function as transmembrane proteins that mediate intracellular pH under low O2 conditions. Because maintaining a neutral pH represents a key survival mechanism for tumour cells in O2‐deprived settings, we sought to elucidate the role of dual CA IX/XII inhibition as a novel strategy to eliminate AML cells under hypoxic conditions. Our findings demonstrate that the dual CA IX/XII inhibitor FC531 may prove to be of value as an adjunct to chemotherapy for the treatment of AML.

High‐dose methylprednisolone plus rituximab (R‐HDMP) is a useful treatment in chronic lymphocytic leukaemia/small lymphocytic lymphoma (CLL/SLL) patients unfit for chemo‐immunotherapy and has proven its utility on the treatment of CLL/SLL complicated by auto‐immune cytopenias. We performed a retrospective, single‐centre study, of CLL/SLL patients treated with R‐HDMP for 9 years. Thirty‐nine patients were included, median age at time of treatment was 77 years. Most patients had stage Rai III/IV and Binet C disease. Twenty‐eight patients had relapsed/refractory disease at time of treatment with a median of 1 previous line of therapy; 53.8% had prior exposure to fludarabine and 25% to rituximab. Grade 3–4 neutropenia and thrombocytopenia were recorded in 10.2% and 17.9% patients, respectively. While on treatment, 51.3% had documented infectious complications, but no other non‐haematological toxicities grades 3–4 were identified. Overall response rate was 64%. Median overall survival and progression‐free survival were 24 and 13 months, respectively. Twenty four patients relapsed and 16 received another line of treatment after R‐HDMP, with median time to next treatment of 13.5 months. Thirteen out of the 24 patients improved performance status and were subsequently considered fit for chemo‐immunotherapy. R‐HDMP is a valuable option for elderly and frail patients, with low risk of severe myelotoxicity and other severe adverse events. It was shown to work as a bridge to other lines of treatment, including chemo‐immunotherapy., High dose methylprednisolone plus rituximab (R‐HDMP) is a useful treatment in chronic lymphocytic leukaemia/small lymphocytic lymphoma patients unfit for chemo‐immunotherapy. We present a cohort of patients with median age of 77 years, most with stage Rai III/IV and Binet C disease. Overall response rate was 64% and median overall survival and progression‐free survival were 24 and 13 months, respectively, with an acceptable toxicity profile, highlighting R‐HDMP utility for elderly and frail patients.

Acquired thrombotic thrombocytopenic purpura (aTTP) is a rare disease and has occasionally been described after vaccination, especially against viral agents. We present a case of a patient who presents with the classic pentad of TTP a few days after receiving the first dose of the mRNA Pfizer COVID‐19 vaccine. To our knowledge, this is the second report of a de novo TTP following mRNA Pfizer COVID‐19 vaccination.

With the advent of targeted therapies for chronic lymphocytic leukemia (CLL), treatment choice has expanded and patients are living longer. Careful consideration is needed regarding treatment duration and sequence, how best to meet patients' needs, balancing toxicities while improving long-term survival and maximizing depth of response. This review addresses these considerations and discusses current targeted treatment dilemmas. Targeted therapies have dramatically transformed the CLL treatment landscape. Two treatment paradigms have emerged using B-cell lymphoma 2 inhibitors (BCL2i) and Bruton's tyrosine kinase (BTK): (i) fixed duration and (ii) continuous treatment. The BCL2i venetoclax can attain deep remissions with a fixed-duration approach, resulting in high rates of undetectable minimal residual disease (uMRD) in treatment-naïve and relapsed/refractory (R/R) patients with CLL. BTKis such as ibrutinib and acalabrutinib achieve high objective response rates and long-term disease control, although they rarely attain complete response or uMRD status as monotherapy. Numerous studies are evaluating the clinical utility of BTKi and BCL2i as combination therapies, where deep remissions have been found to occur. MRD status may also be a useful marker for deciding when to stop continuous therapy, and randomized trials on MRD-guided treatment strategies are currently ongoing. The current treatment choice between continuous or fixed-duration therapy should be based on comorbidities, risks, preferences, and treatment goals, whilst areas of emerging clinical interest include the potential utility of BTKi-BCL2i combination therapies, as well as an MRD-guided treatment strategies in the future.

Thalassaemia is caused by genetic globin defects leading to anaemia, transfusion-dependence and comorbidities. Reduced survival and systemic organ disease affect transfusion-dependent thalassaemia major and thalassaemia intermedia. Recent improvements in clinical management have reduced thalassaemia mortality. The therapeutic landscape of thalassaemia may soon include gene therapies as functional cures. An analysis of the adult US thalassaemia population has not been performed since the Thalassemia Clinical Research Network cohort study from 2000 to 2006. The Centers for Disease Control and Prevention supported US thalassaemia treatment centres (TTCs) to compile longitudinal information on individuals with thalassaemia. This dataset provided an opportunity to evaluate iron balance, chelation, comorbidities and demographics of adults with thalassaemia receiving care at TTCs. Two adult cohorts were compared: those over 40 years old (n = 75) and younger adults ages 18-39 (n = 201). The older adult cohort was characterized by higher numbers of iron-related comorbidities and transfusion-related complications. By contrast, younger adults had excess hepatic and cardiac iron and were receiving combination chelation therapy. The ethnic composition of the younger cohort was predominantly of Asian origin, reflecting the demographics of immigration. These findings demonstrate that comprehensive care and periodic surveys are needed to ensure optimal health and access to emerging therapies.

BACKGROUND Myelodysplastic syndromes (MDS) are a heterogenous collection of clonal bone marrow diseases characterized by cytopenias, abnormal karyotypes, molecular abnormalities, and dysplasia by flow cytometry and/or morphology. The progression of MDS to severe cytopenias and/or overt leukemia is associated with the accumulation of additional cytogenetic abnormalities, suggesting clonal evolution. The impact of these accumulated abnormalities on myeloid maturation and the severity of the disease is poorly understood. METHODS Bone marrow specimens from 16 patients with cytogenetic abnormalities were flow cytometrically sorted into three myeloid populations: progenitors, immature myeloid cells, and mature myeloid cells. Fluorescence in situ hybridization analysis was performed on each to determine the distribution of chromosomal abnormalities during myeloid maturation. RESULTS Our findings revealed three distinct distributions of cytogenetic abnormalities across myeloid maturation, each of which corresponded to specific cytogenetic abnormalities. Group 1 had continuous distribution across all maturational stages and contained patients with a single cytogenetic aberration associated with good-to-intermediate prognosis; Group 2 had accumulation of abnormalities in immature cells and contained patients with high-risk monosomy 7; and Group 3 had abnormalities defining the founding clone equally distributed across maturational stages while subclonal abnormalities were enriched in progenitor cells and contained patients with multiple, non-monosomy 7, abnormalities with evidence of clonal evolution. CONCLUSIONS Our findings demonstrate that low-risk abnormalities (e.g., del(20q) and trisomy 8) occurring in the founding clone display a markedly different disease etiology, with respect to myeloid maturation, than monosomy 7 or abnormalities acquired in subclones, which result in a disruption of myeloid cell maturation in MDS.

Chronic-lymphocytic-leukemia (CLL) is the most prevalent leukemia in developed countries, caused by monoclonal proliferation of CD5+ B-cells and accumulation of mature-appearing-neoplastic lymphocytes in blood, bone marrow, and secondary lymphoid organs. Oral manifestations of CLL are infrequent and rarely reported in literature. We report a new case of a 67-year old man who presented with the complaints of tooth mobility and gingival swelling. Extra-oral examination was remarkable for cutaneous pallor and bilateral cervical lymphadenopathy involving the submandibular, and deep cervical lymph nodes on both sides of the neck. Complete blood count revealed normal red blood cell count (4.25 × 106/μl), normal platelet count (136 × 103/μl) and increased white blood cell count (25.3 × 103/μl). Differential white blood cell count showed marked lymphocytosis (88%), and blood film revealed the presence of leukocytosis, with small mature-looking lymphocytes, and mild thrombocytopenia. A flow cytometry immune-phenotyping revealed that 55% of peripheral blood cells were monoclonal B-lymphocytes expressing CD19, CD20, CD23, CD200, CD22, CD5, CD38, CD11c, sIgD and Kappa light chain confirming the diagnosis of CLL. Oral healthcare professionals should consider systemic causes, such as CLL, in the differential diagnosis of generalized tooth mobility and gingival swelling, particularly in patients with associated symptoms such as lymphadenopathy, fever, weight loss, and general fatigue.

Multiple molecular features, such as activation of specific oncogenes (e.g., MYC, BCL2) or a variety of gene expression signatures, have been associated with disease course in diffuse large B-cell lymphoma (DLBCL), although their relationships and implications for targeted therapy remain to be fully unraveled. We report that MYC activity is closely correlated with-and most likely a driver of-gene signatures related to oxidative phosphorylation (OxPhos) in DLBCL, pointing to OxPhos enzymes, in particular mitochondrial electron transport chain (ETC) complexes, as possible therapeutic targets in high-grade MYC-associated lymphomas. In our experiments, indeed, MYC sensitized B cells to the ETC complex I inhibitor IACS-010759. Mechanistically, IACS-010759 triggered the integrated stress response (ISR) pathway, driven by the transcription factors ATF4 and CHOP, which engaged the intrinsic apoptosis pathway and lowered the apoptotic threshold in MYC-overexpressing cells. In line with these findings, the BCL2-inhibitory compound venetoclax synergized with IACS-010759 against double-hit lymphoma (DHL), a high-grade malignancy with concurrent activation of MYC and BCL2. In BCL2-negative lymphoma cells, instead, killing by IACS-010759 was potentiated by the Mcl-1 inhibitor S63845. Thus, combining an OxPhos inhibitor with select BH3-mimetic drugs provides a novel therapeutic principle against aggressive, MYC-associated DLBCL variants.

BACKGROUND Human papillomavirus-related oropharyngeal squamous cell carcinoma (HPV-OPSCC) presents frequently as metastasis in a neck lymph node that may be cystic or necrotic. Fine-needle aspiration (FNA) biopsies are often first-line diagnostic procedures. p16 immunohistochemistry (IHC) is a surrogate marker for high-risk HPV (hrHPV) infection but can be challenging to interpret. This study evaluated the use of hrHPV in situ hybridization (ISH) in cytology cell blocks of cystic neck lesions. METHODS Twenty-four FNA cases with cell blocks and surgical correlates were evaluated. p16 IHC and hrHPV ISH were assessed on cell blocks (C-p16 and C-hrHPV ISH), and hrHPV ISH on surgical samples (S-hrHPV ISH). All results were classified as negative, positive, or equivocal. RESULTS Two cases were excluded because of insufficient tissue on recut. On the basis of C-hrHPV ISH cases, 12 were positive, 5 were negative, and 5 were equivocal. All 12 positive C-hrHPV ISH cases had concordant S-hrHPV ISH with no false positives. Of the 5 negative C-hrHPV ISH cases, 4 had concordant S-hrHPV ISH, and 1 had a discordant S-hrHPV ISH. Of the 5 equivocal C-hrHPV ISH cases, S-hrHPV ISH were both positive and negative. Fourteen cases were equivocal by C-p16; 9 cases were reliably classified by C-hrHPV ISH (5 positive, 4 negative; 64%). CONCLUSIONS C-hrHPV ISH can be reliably used, especially when positive. A negative or equivocal interpretation of C-hrHPV ISH may warrant repeat testing. Compared to C-p16, C-hrHPV ISH is more frequently diagnostic and could be helpful for HPV-OSCC diagnosis and management.

We analysed long-term outcome of patients receiving haematopoietic allogeneic stem cell transplantation (allo-HSCT) as a first transplant for high-risk Hodgkin lymphoma (HL). One hundred and ninety patients were included in this study, 63% of them had previously received brentuximab vedotin and/or checkpoint inhibitors. Seventy patients (37%) received an unrelated donor allo-HSCT, 99 (51%) had myeloablative conditioning (MAC) and 60% had in vivo T-cell/depleted grafts (TCD). The 100-day cumulative incidence (CI) of grade II-IV acute graft-versus-host disease (GVHD) was 25% and the 3-year CI of chronic GVHD was 38%. The 3-year CI of non-relapse mortality (NRM) and relapse rate were 21% and 38% respectively. After a median follow-up of 58 months, 3-year overall survival (OS) and progression-free survival (PFS) were 58% and 41% respectively. Multivariate analysis showed that, in comparison to reduced-intensity conditioning regimens with or without TCD, MAC using TCD had similar NRM and a lower risk of relapse leading to significantly better OS and PFS. MAC without TCD was associated with higher NRM and worse survival outcomes. These results suggest that in patients with high-risk HL and candidates of allo-HSCT, a MAC strategy with TCD might be the best option.