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Summary: The dysregulation of the Janus family tyrosine kinase‐signal transducer and activator of transcription (JAK‐STAT) is closely related to acute lymphoblastic leukaemia (ALL), whereas the clinical value of phosphorylated STAT5 (pSTAT5) remains elusive. Herein we performed a prospective study on clinical significance of flow cytometry‐based pSTAT5 in adult B‐ALL patients. A total of 184 patients were enrolled in the Precision‐Classification‐Directed‐Target‐Total‐Therapy (PDT)‐ALL‐2016 cohort between January 2018 and December 2021, and STAT5 phosphorylation was detected by flow cytometry at diagnosis. Based on flow‐pSTAT5, the population was classified into pSTAT5low (113/184, 61.1%) and pSTAT5high (71/184, 38.9%). Overall survival (OS) and event‐free survival (EFS) were inferior in pSTAT5high patients than in those with pSTAT5low (OS, 44.8% vs. 65.2%, p = 0.004; EFS, 23.5% vs. 52.1%, p < 0.001), which was further confirmed in an external validation cohort. Furthermore, pSTAT5 plus flow‐based minimal residual disease (MRD) postinduction defines a novel risk classification as being high risk (HR, pSTAT5high + MRD+), standard risk (SR, pSTAT5low + MRD−) and others as moderate‐risk group. Three identified patient subgroups are distinguishable with disparate survival curves (3‐year OS rates, 36.5%, 56.7% and 76.3%, p < 0.001), which was confirmed on multivariate analysis (hazard ratio 3.53, p = 0.003). Collectively, our study proposed a novel, simple and flow‐based risk classification by integrating pSTAT5 and MRD in favour of risk‐guided treatment for B‐ALL. [ABSTRACT FROM AUTHOR]

Summary: Acute myeloid leukaemia (AML) is a highly heterogeneous disease, exhibiting diverse subtypes according to the characteristics of tumour cells. The immunophenotype is one of the aspects acquired routinely through flow cytometry in the diagnosis of AML. Here, we characterized the antigen expression in paediatric AML cases across both morphological and molecular genetic subgroups. We discovered a subgroup of patients with unfavourable prognosis that can be immunologically characterized, irrespective of morphological FAB results or genetic aberrations. Cox regression analysis unveiled key antigens influencing the prognosis of AML patients. In terms of underlying genotypes, we observed that the antigenic profiles and outcomes of one specific group, primarily composed of CBFA2T3::GLIS2 and FUS::ERG, were analogous to the reported RAM phenotype. Overall, our data highlight the significance of immunophenotype to tailor treatment for paediatric AML. [ABSTRACT FROM AUTHOR]

Summary: Targeted therapy development for acute myeloid leukaemia (AML) requires an understanding of specific expression profiles. We collected flow cytometry data on 901 AML patients and recorded aberrant CD7 expression on leukaemic blasts. 263 (29.2%) had blasts positive for CD7. CD7+ AML was more likely to be adverse risk (64.6% vs. 55.6%, p = 0.0074) and less likely to be favourable risk (15.2% vs. 24.1%, p = 0.0074) by European LeukemiaNet 2022 criteria. Overall survival was inferior (11.9 [95% CI, 9.7–15.9] vs. 19.0 months [95% CI, 16.1–23.0], p = 0.0174). At relapse, 30.4% lost and 19.0% gained CD7, suggesting moderate instability over time. [ABSTRACT FROM AUTHOR]

Summary: Assessing minimal residual disease (MRD) in B‐cell precursor acute lymphoblastic leukaemia (BCP‐ALL) is essential for adjusting therapeutic strategies and predicting relapse. Quantitative polymerase chain reaction (qPCR) is the gold standard for MRD. Alternatively, flow cytometry is a quicker and cost‐effective method that typically uses leukaemia‐associated immunophenotype (LAIP) or different‐from‐normal (DFN) approaches for MRD assessment. This study describes an optimized 12‐colour flow cytometry antibody panel designed for BCP‐ALL diagnosis and MRD monitoring in a single tube. This method robustly differentiated hematogones and BCP‐ALL cells using two specific markers: CD43 and CD81. These and other markers (e.g. CD73, CD66c and CD49f) enhanced the specificity of BCP‐ALL cell detection. This innovative approach, based on a dual DFN/LAIP strategy with a principal component analysis method, can be used for all patients and enables MRD analysis even in the absence of a diagnostic sample. The robustness of our method for MRD monitoring was confirmed by the strong correlation (r = 0.87) with the qPCR results. Moreover, it simplifies and accelerates the preanalytical process through the use of a stain/lysis/wash method within a single tube (<2 h). Our flow cytometry‐based methodology improves the BCP‐ALL diagnosis efficiency and MRD management, offering a complementary method with considerable benefits for clinical laboratories. [ABSTRACT FROM AUTHOR]

Summary: Flow cytometry (FC) incorporating the T‐cell receptor β constant chain‐1 (TRBC1) has been recently proposed as a new standard in T‐cell clonality assessment. While early studies demonstrated high sensitivity in samples with conspicuous tumour burden, performance in real‐world samples, including those with low tumour burden and correlation with molecular methods has been limited. We evaluated TRBC1‐FC performance and correlated the results with high‐throughput TRB sequencing and a targeted next‐generation sequencing gene panel. Our cohort consisted of 90 evaluable samples from 57 patients. TRBC1‐FC confirmed T‐cell clonality in 37 out of 38 samples (97%) that were involved in a mature T‐cell neoplasm (MTCN). T‐cell clonality was also identified in nine samples from patients lacking a current or prior diagnosis of MTCN, consistent with the emerging entity T‐cell clonality of uncertain significance. TRBC‐FC was polyclonal in all samples and negative for disease involvement by standard pathology assessment. However, correlation with TRB sequencing in 17 of these samples identified two cases that harboured the known clonal sequence from index testing, indicating the presence of measurable residual disease not otherwise detected. Our study provides real‐world correlative validation of TRBC1‐FC, highlighting the strengths and limitations pertinent to its increasing implementation by general diagnostic laboratories. [ABSTRACT FROM AUTHOR]

The article discusses the use of T-cell receptor constant beta-chain 1 (TRBC1) expression as a marker for T-cell clonality in the diagnosis of T-cell lymphomas. The study compared TRBC1 testing with genetic studies and found that TRBC1 expression detected by flow cytometry showed 100% concordance with genetic testing for T-cell clonality. The authors concluded that TRBC1 testing, along with immunophenotyping, could replace genetic testing in cases where an abnormal T-cell phenotype is detected or when flow cytometry is inconclusive. The study was supported by the National Institute for Health Research and the authors declared no conflicts of interest. [Extracted from the article]

Summary: To determine the prevalence and clinical relevance of glycosylphosphatidylinositol‐anchored protein‐deficient (GPI[−]) cell populations (paroxysmal nocturnal haemoglobinuria [PNH]‐type cells) in patients with acquired aplastic anaemia (AA) or myelodysplastic syndrome (MDS), we prospectively studied peripheral blood samples of 2402 patients (1075 with AA, 900 with MDS, 144 with PNH, and 283 with other anaemia) using a high‐sensitivity flow cytometry assay in a nationwide multi‐centre observational study. PNH‐type cells were detected in 52.6% of AA and 13.7% of MDS patients. None of the 35 patients with refractory anaemia (RA) with ringed sideroblasts or the 86 patients with RA with excess of blasts carried PNH‐type cells. Among the 317 patients possessing PNH‐type granulocytes, the percentage of PNH‐type granulocytes increased by ≥10% in 47 patients (14.8%), remained unchanged in 240 patients (75.7%), and decreased by ≥10% in 30 patients (9.5%) during 3 years of follow‐up. PNH‐type granulocyte expansion occurred more frequently (27.1%) in the 144 patients who originally carried PNH‐type granulocytes ≥1% than in the 173 patients with PNH‐type granulocytes <1% (4.6%). This study confirmed that PNH‐type cells are undetectable in authentic clonal MDS patients, and the presence of ≥1% PNH‐type granulocytes predicts a higher likelihood of PNH‐type cell expansion than with <1% PNH‐type granulocytes. [ABSTRACT FROM AUTHOR]

Summary: We investigated 23 hepatitis C virus (HCV)‐infected patients with overt lymphoproliferative diseases (15 cases) or monoclonal B lymphocytosis (8 cases) treated with direct agent antiviral (DAAs) per clinical practice. DAA therapy yielded undetectable HCV‐RNA, the complete response of cryoglobulinemia vasculitis and related signs, whilst the presence of B‐cell clones (evaluated by flow cytometry, IGHV, and BCL2‐IGH rearrangements), detected in 19/23 cases at baseline, was maintained (17/19). Similarly, IGHV intraclonal diversification, supporting an antigen‐driven selection mechanism, was identified in B‐cell clones at baseline and end of follow‐up. DAA therapy alone, despite HCV eradication and good immunological responses, was less effective on the pathological B‐cell clones. [ABSTRACT FROM AUTHOR]

Summary: Undetectable measurable residual disease (MRD) is associated with favourable clinical outcomes in chronic lymphocytic leukaemia (CLL). While assessment is commonly performed using multiparameter flow cytometry (MFC), this approach is associated with limitations including user bias and expertise that may not be widely available. Implementation of unsupervised clustering algorithms in the laboratory can address these limitations and have not been previously reported in a systematic quantitative manner. We developed a computational pipeline to assess CLL MRD using FlowSOM. In the training step, a self‐organising map was generated with nodes representing the full breadth of normal immature and mature B cells along with disease immunophenotypes. This map was used to detect MRD in multiple validation cohorts containing a total of 456 samples. This included an evaluation of atypical CLL cases and samples collected from two different laboratories. Computational MRD showed high correlation with expert analysis (Pearson's r > 0.99 for typical CLL). Binary classification of typical CLL samples as either MRD positive or negative demonstrated high concordance (>98%). Interestingly, computational MRD detected disease in a small number of atypical CLL cases in which MRD was not detected by expert analysis. These results demonstrate the feasibility and value of automated MFC analysis in a diagnostic laboratory. [ABSTRACT FROM AUTHOR]

Summary: This study aimed to evaluate the concordance between minimal residual disease (MRD) results obtained by multicolour flow cytometry (MFC) and polymerase chain reaction for fusion gene transcripts (FGTs) in infants with acute lymphoblastic leukaemia (ALL) associated with rearrangement of the KMT2A gene (KMT2A‐r). A total of 942 bone marrow (BM) samples from 123 infants were studied for MFC‐MRD and FGT‐MRD. In total, 383 samples (40.7%) were concordantly MRD‐negative. MRD was detected by the two methods in 441 cases (46.8%); 99 samples (10.5%) were only FGT‐MRD‐positive and 19 (2.0%) were only MFC‐MRD‐positive. A final concordance rate of 87.4% was established. Most discordance occurred if residual leukaemia was present at levels close to the sensitivity limits. Neither the type of KMT2A fusion nor a new type of treatment hampering MFC methodology had an influence on the concordance rate. The prognostic value of MFC‐MRD and FGT‐MRD differed. MFC‐MRD was able to identify a rapid response at early time‐points, whereas FGT‐MRD was a reliable relapse predictor at later treatment stages. Additionally, the most precise risk definition was obtained when combining the two methods. Because of the high comparability in results, these two rather simple and inexpensive approaches could be good options of high clinical value. [ABSTRACT FROM AUTHOR]

Summary: A biopsy of lymphoid tissue is currently required to diagnose Kaposi sarcoma‐associated herpesvirus (KSHV)‐associated multicentric Castleman disease (KSHV–MCD). Patients showing clinical manifestations of KSHV–MCD but no pathological changes of KSHV–MCD are diagnosed as KSHV inflammatory cytokine syndrome. However, a lymph node biopsy is not always feasible to make the distinction. A pathognomonic feature of lymph nodes in KSHV–MCD is the expansion of KSHV‐infected, lambda‐restricted but polyclonal plasmablasts. To investigate whether these cells also reside in extra‐nodal sites, effusion from 11 patients with KSHV–MCD and 19 with KSHV inflammatory cytokine syndrome was analysed by multiparametric flow cytometry. A distinct, lambda‐restricted plasmablastic population (LRP) with highly consistent immunophenotype was detected in effusions in 8/11 patients with KSHV–MCD. The same population was also observed in 7/19 patients with KSHV inflammatory cytokine syndrome. The detection of LRP stratified KSHV inflammatory cytokine syndrome into two clinically distinct subgroups; those with detectable LRP closely resembled KSHV–MCD, showing similar KSHV viral load, comparable severity of thrombocytopenia and hypoalbuminaemia, and similar incidences of hepatosplenomegaly. Collectively, the detection of LRP by flow cytometry can serve as a valuable tool in diagnosing KSHV–MCD. KSHV inflammatory cytokine syndrome with LRP in effusions may represent a liquid‐form of KSHV–MCD. [ABSTRACT FROM AUTHOR]

Keywords: B cell acute lymphoblastic leukaemia; B-ALL; flow cytometry; minimal residual disease; MRD; next generation flow EN B cell acute lymphoblastic leukaemia B-ALL flow cytometry minimal residual disease MRD next generation flow 381 384 4 01/23/23 20230201 NES 230201 Minimal residual disease (MRD) in acute lymphoblastic leukaemia (ALL) is a strong predictor of prognosis and its identification is incorporated into most ALL treatment protocols. Multicentric standardization of minimal/measurable residual disease in B-cell precursor acute lymphoblastic leukaemia using next-generation flow cytometry in a low/middle-level income country In an initial phase of the study, 21 laboratories submitted at least 10 Flow Cytometry Standard (FCS) files per laboratory ( I n i = 195) for centralized analyses. [Extracted from the article]

The new 2022 WHO classification has replaced the category of AML with myelodysplasia-related changes (AML-MRC) with AML-Myelodysplasia Related (AML-MR) and its diagnostic criteria are updated (see Endnote). Keywords: cytogenetics; flow cytometry; leukaemia diagnosis; molecular genetics; myeloid leukaemia EN cytogenetics flow cytometry leukaemia diagnosis molecular genetics myeloid leukaemia 150 159 10 01/16/23 20230115 NES 230115 METHODOLOGY This Good Practice Paper was compiled according to the BSH process (https://b-s-h.org.uk/media/19922/bsh-guidance-development-process-july-2021.pdf). Key changes include removal of morphology alone to make a diagnosis of AML-MR, updating the cytogenetic criteria and introducing the mutation-based diagnosis of AML-MR. INTRODUCTION Making a diagnosis of acute myeloid leukaemia (AML) requires a multi-faceted approach bringing together clinical features of patient presentation with laboratory investigations encompassing morphological, immunophenotypic and genetic evaluation of blood, bone marrow and, when appropriate, cerebrospinal fluid (CSF). [Extracted from the article]

The era of AI‐based methods to improve flow cytometry diagnostics in haematology is now at the beginning. The study by Nguyen and colleagues explored an emerging machine learning approach to assess phenotypic MRD in chronic lymphocytic leukaemia patients, showing that such AI‐driven computational analysis may represent a robust and feasible tool for advanced diagnostics of haematological malignancies. Commentary on: Nguyen et al. Computational flow cytometry provides accurate assessment of measurable residual disease in chronic lymphocytic leukaemia. Br J Haematol 2023;202:760–770. [ABSTRACT FROM AUTHOR]

On flow cytometry, HCL cells express brightly mature B-lymphoid cell antigens as CD19, CD20, CD22, CD200, are typically positive for CD11c, CD103, CD123 and CD25, while are usually negative or dim for CD5, CD23, CD10, CD79b and CD27. Three cases (3/71, 4%) - despite expressing CD25, CD11c, and CD200 on all clonal cells (which were 15%, 22% and 37% of total leukocytes, respectively) - had CD103 expressed only on a fraction of leukaemic cells (40%, 27%, and 10.8%, corresponding to 5%, 6% and 4% of total leukocytes) (Figure 1A). Keywords: CD103; flow cytometry; hairy cell leukaemia EN CD103 flow cytometry hairy cell leukaemia e28 e31 4 07/18/22 20220715 NES 220715 Classical hairy cell leukaemia (HCLc) is an uncommon mature B-cell lymphoproliferative neoplasm (MBCLN) comprising about 2% of lymphoid leukaemias. [Extracted from the article]

Multiple myeloma (MM) is a disease with known immune dysregulation. Natural killer (NK) cells have shown preclinical activity in MM. We conducted a first-in-human study of umbilical cord blood-derived (CB) NK cells for MM patients undergoing high dose chemotherapy and autologous haematopoietic stem cell transplantation (auto-HCT). Patients received lenalidomide (10 mg) on days -8 to -2, melphalan 200 mg/m2 on day -7, CB-NK cells on day -5 and auto-HCT on day 0. Twelve patients were enrolled, three on each of four CB-NK cell dose levels: 5 × 106 , 1 × 107 , 5 × 107 and 1 × 108 CB-NK cells/kg. Ten patients had either high-risk chromosomal changes or a history of relapsed/progressed disease. There were no infusional toxicities and no graft-versus-host disease. One patient failed to engraft due to poor autologous graft quality and was rescued with a back-up autologous graft. Overall, 10 patients achieved at least a very good partial response as their best response, including eight with near complete response or better. With a median follow-up of 21 months, four patients have progressed or relapsed, two of whom have died. CB-NK cells were detected in vivo in six patients, with an activated phenotype (NKG2D+ /NKp30+ ). These data warrant further development of this novel cellular therapy.

Keywords: early assessment; flow cytometry; measurable residual disease; multiple myeloma; outcome EN early assessment flow cytometry measurable residual disease multiple myeloma outcome e82 e85 4 06/14/22 20220615 NES 220615 Multiple myeloma (MM) treatment has evolved remarkably with the introduction of new agents such as proteasome inhibitors, immunomodulatory drugs (IMIDS) and therapeutic antibodies. The level of minimal residual disease in the bone marrow of patients with multiple myeloma before high-dose therapy and autologous blood stem cell transplantation is an independent predictive parameter. The 26 patients with persistent positive MRD had worse PFS compared to the negative MRD post- and pre-ASCT cases (24 months vs 45 months vs 68 months, I p i < 0.0001). [Extracted from the article]

Summary: The standardized EuroFlow protocol, including CD19 as primary B‐cell marker, enables highly sensitive and reliable minimal residual disease (MRD) assessment in B‐cell precursor acute lymphoblastic leukaemia (BCP‐ALL) patients treated with chemotherapy. We developed and validated an alternative gating strategy allowing reliable MRD analysis in BCP‐ALL patients treated with CD19‐targeting therapies. Concordant data were obtained in 92% of targeted therapy patients who remained CD19‐positive, whereas this was 81% in patients that became (partially) CD19‐negative. Nevertheless, in both groups median MRD values showed excellent correlation with the original MRD data, indicating that, despite higher interlaboratory variation, the overall MRD analysis was correct. [ABSTRACT FROM AUTHOR]

Assessment of measurable residual disease (MRD) using multiparameter flow cytometry in precursor B‐cell acute lymphoblastic leukaemia (ALL) is routine. However, studies on the harmonization of laboratory techniques as well as on the interpretation of results are limited. Here, Ikoma‐Colturato and colleagues from Brazil demonstrate multicentric standardization of B‐ALL MRD using EuroFlow protocols. Commentary on: Ikoma‐Colturato et al., Multicentric standardization of minimal/measurable residual disease in B‐cell precursor acute lymphoblastic leukaemia using next‐generation flow cytometry in a low/middle‐level income country. Br J Haematol 2023;200:381‐384. [ABSTRACT FROM AUTHOR]

The prognostic value of minimal residual disease (MRD) assessed by multi-parameter flow cytometry (MFC) was investigated among 340 adult patients with B-cell acute lymphoblastic leukaemia (B-ALL) treated between 2004 and 2014 using regimens including the hyperCVAD (hyperfractionated cyclophosphamide, vincristine, doxorubicin, dexamethasone, methotrexate, cytarabine) backbone. Among them, 323 (95%) achieved complete remission (CR) and were included in this study. Median age was 52 years (range, 15-84). Median white blood cell count (WBC) was 9·35 × 10(9) /l (range, 0·4-658·1 ×1 0(9) /l). MRD by MFC was initially assessed with a sensitivity of 0·01%, using a 15-marker, 4-colour panel and subsequently a 6-colour panel on bone marrow specimens obtained at CR achievement and at approximately 3 month intervals thereafter. MRD negative status at CR was associated with improved disease-free survival (DFS) and overall survival (OS) (P = 0·004 and P = 0·03, respectively). Similarly, achieving MRD negative status at approximately 3 and 6 months was associated with improved DFS (P = 0·004 and P

Summary: Monoclonal gammopathy of unknown significance (MGUS), smouldering multiple myeloma (SMM), and multiple myeloma (MM) are very common neoplasms. However, it is often difficult to distinguish between these entities. In the present study, we aimed to classify the most powerful markers that could improve diagnosis by multiparametric flow cytometry (MFC). The present study included 348 patients based on two independent cohorts. We first assessed how representative the data were in the discovery cohort (123 MM, 97 MGUS) and then analysed their respective plasma cell (PC) phenotype in order to obtain a set of correlations with a hypersphere visualisation. Cluster of differentiation (CD)27 and CD38 were differentially expressed in MGUS and MM (P < 0·001). We found by a gradient boosting machine method that the percentage of abnormal PCs and the ratio PC/CD117 positive precursors were the most influential parameters at diagnosis to distinguish MGUS and MM. Finally, we designed a decisional algorithm allowing a predictive classification ≥95% when PC dyscrasias were suspected, without any misclassification between MGUS and SMM. We validated this algorithm in an independent cohort of PC dyscrasias (n = 87 MM, n = 41 MGUS). This artificial intelligence model is freely available online as a diagnostic tool application website for all MFC centers worldwide (https://aihematology.shinyapps.io/PCdyscrasiasToolDg/). [ABSTRACT FROM AUTHOR]

Summary: High‐sensitivity multicolour flow cytometry (MFC)‐based B‐lymphoblastic leukaemia (B‐ALL) measurable residual disease (BMRD) assay is increasingly being used in clinical practice. Herein, we describe six consistently present low‐level populations immunophenotypically mimicking abnormal B‐ALL blasts in 441 BMRD samples from 301 children. These included CD19+CD123+ plasmacytoid dendritic cells differentiating from lymphoid precursors, CD10+ transitional B cells with CD10+/CD38dim‐to‐negative/CD20bright/CD45bright phenotype, CD19+ natural killer (NK) cells, CD73bright/CD10+ mesenchymal stromal/stem cells, CD73bright/CD34+ endothelial cells, and a CD34+CD38dim‐to‐negative/CD10−/CD20bright/CD45bright subset of mature B cells. We provide the proportions, comprehensive immunophenotype, and practical clues for proper identification of these low‐level populations. Knowledge regarding the presence and immunophenotype of these mimics is essential for accurate interpretation in high‐sensitivity MFC‐BMRD analysis. [ABSTRACT FROM AUTHOR]

Serial evaluation of CD19 surface expression in pediatric B-cell malignancies following CD19-targeted therapy. In contrast to the CD19-negative precursors, the level of CD19-positives increased significantly during the first three months of maintenance therapy, although the continued presence of B-lineage regeneration remained more or less stable (Fig. Keywords: CD19 targeting; minimal residual disease; ALL; flow cytometry; CD19-negative precursors EN CD19 targeting minimal residual disease ALL flow cytometry CD19-negative precursors e6 e9 4 12/27/21 20220101 NES 220101 Targeting the pan-B-lymphocyte antigen CD19 with the bispecific T-cell engager blinatumomab is one of the modern ways of treating acute lymphocytic B-cell precursor leukaemia (BCP-ALL).1,2 This drug has been very successful in treating patients with relapsed/refractory (R/R) disease,3,4 but it has recently been shown that blinatumomab is also effective in eradicating minimal residual disease (MRD) in patients with relapsed or primary BCP-ALL.5,6 Loss of the target molecule is one of the key mechanisms for leukaemia cells to escape the selective pressure of blinatumomab.7,8 The resulting possible CD19 negativity interferes with MRD monitoring by multicolour flow cytometry (MFC), which is usually based on analysis of the CD19-positive compartment.9 On the other hand, the use of other early B-line antigens [CD22, CD24, CD10, intracellular (i) CD79a] instead of CD19 for primary B-cell gating, such as to improve the MFC-MRD technique in the case of CD19-targeted treatment,7,10,11 could lead to errors in the interpretation of the immunophenotype of normal haematopoietic cells. CD19-negative B-cell precursors were gated as described previously.12 All BM samples obtained after completion of blinatumomab were MFC-MRD-negative, including those from patients who were MFC-MRD-positive at EOI ( I n i = 7, median MFC-MRD 0-030%, range 0-001-0-166%). [Extracted from the article]

The clinical problem of a non‐healing fistula in ano in a child affected with poikiloderma with neutropenia (PN) was the stimulus for an innovative study by Parajuli et al. that sheds light on the pathological mechanisms in this disease. Multiparametric analyses of the patient's blood mononuclear cells by cell culture, flow cytometry and multiplex cytokine assay suggested a block of monocyte differentiation. Monocyte transcriptome profiling revealed a signature consistent with the haematological picture and the clinical presentation. Commentary on: Parajuli et al. Defective monocyte plasticity and altered cAMP pathway characterize USB1‐mutated poikiloderma with neutropenia Clericuzio type. Br J Haematol 2024;204:683‐693. [ABSTRACT FROM AUTHOR]

Summary: Infants with acute lymphoblastic leukaemia (ALL) have a high frequency of central nervous system (CNS) involvement. Flow cytometric analysis of cerebrospinal fluid (CSF) was recently demonstrated to be a sensitive method for detecting CNS involvement in childhood ALL. In the present study, CSF from 14 infants was collected at routine lumbar punctures and analysed by multicolour flow cytometry. At initial diagnosis, leukaemic blasts were detected in CSF by flow cytometry in 11 patients (78·6%) compared to seven patients (50%) by cytospin. Larger studies are needed to determine if CSF flow cytometry has prognostic value in infant ALL. [ABSTRACT FROM AUTHOR]

Summary: Plasma cell leukaemia (PCL) is a rare and very aggressive plasma cell disorder. Preventing a dismal outcome of PCL requires early diagnosis with appropriate analytical tools. Therefore, the investigation of 33 patients with primary and secondary PCL was done when the quantity of circulating plasma cells (PCs) using flow cytometry (FC) and morphology assessment was evaluated. The phenotypic profile of the PCs was also analysed to determine if there is an association with clinical outcomes and to evaluate the prognostic value of analysed markers. Our results revealed that FC is an excellent method for identifying circulating PCs as a significantly higher number was identified by FC than by morphology (26·7% vs. 13·5%, P = 0·02). None of secondary PCL cases expressed CD19 or CD20. A low level of expression with similar positivity of CD27, CD28, CD81 and CD117 was found in both PCL groups. A decrease of CD44 expression was detected only in secondary PCL. Expression of CD56 was present in more than half of PCL cases as well as cytoplasmic nestin. A decreased level of platelets, Eastern Cooperative Oncology Group score of 2–3 and lack of CD20+ PC were associated with a higher risk of death. FC could be incorporated in PCL diagnostics not only to determine the number of circulating PCs, but also to assess their phenotype profile and this information should be useful in patients' diagnosis and possible prognosis. [ABSTRACT FROM AUTHOR]

Increased platelet activation in SARS-CoV-2 infected non-hospitalised children and adults, and their household contacts Platelet activation and platelet-monocyte aggregates formation trigger tissue factor expression in severe COVID-19 patients. Keywords: platelet activation; Paediatrics; virology; flow cytometry; platelet function EN platelet activation Paediatrics virology flow cytometry platelet function 90 94 5 09/30/21 20211001 NES 211001 COVID-19 is associated with haemostatic dysregulation,1 with thromboembolism occurring in 25% of hospitalised COVID-19 patients and microvascular thrombi reported at autopsy.2 Platelets are activated in COVID-19 patients requiring intensive care,3 while limited data in mild COVID-19 shows no changes in platelet phenotype.4 Children have low-risk of severe COVID-19 and thrombosis.5 If haemostasis is fundamental to COVID-19 pathogenesis, then age-related haemostatic differences may protect children from COVID-19. Platelet activation, Paediatrics, virology, flow cytometry, platelet function. [Extracted from the article]

Summary: Hydroxyurea (hydroxycarbamide) is an effective treatment for sickle cell anaemia (SCA), but clinical responses depend primarily upon the degree of fetal haemoglobin (HbF) induction and the heterogeneity of HbF expression across erythrocytes. The number and characteristics of HbF‐containing cells (F‐cells) are not assessed by traditional HbF measurements. Conventional hydroxyurea dosing (e.g. fixed doses or low starting doses with stepwise escalation) produces a moderate heterocellular HbF induction, but haemolysis and clinical complications continue. Robust, pancellular HbF induction is needed to minimise or fully inhibit polymerisation of sickle haemoglobin. We treated children with hydroxyurea using an individualised, pharmacokinetics‐guided regimen starting at predicted maximum tolerated dose (MTD). We observed sustained HbF induction (mean >30%) for up to 6 years, which was not dependent on genetic determinants of HbF expression. Nearly 70% of patients had ≥80% F‐cells (near‐pancellular), and almost half had ≥90% F‐cells (pancellular). The mean HbF/F‐cell content was ~12 pg. Earlier age of initiation and better medication adherence were associated with high F‐cell responses. In summary, early initiation of hydroxyurea using pharmacokinetics‐guided starting doses at predicted MTD can achieve sustained near‐pancellular or pancellular HbF expression and should be considered an achievable goal for children with SCA treated with hydroxyurea at optimal doses. Clinical trial registration number: NCT02286154 (clinicaltrials.gov). [ABSTRACT FROM AUTHOR]

Summary: Minimal residual disease (MRD) is an important independent prognostic factor for relapse and survival in acute lymphoblastic leukaemia (ALL). Compared with adult B‐cell ALL, reports of adult T‐cell ALL (T‐ALL) MRD have been scarce and mostly based on molecular methods. We evaluated the prognostic value of multiparameter flow cytometry (FCM)‐based MRD at the end of induction (EOI‐MRD). The present retrospective study included 94 adult patients with T‐ALL. MRD was detected by six‐ to eight‐colour FCM. Patients who were EOI‐MRD positive had a higher cumulative incidence of relapse (CIR) (87·6% vs. 38·8%, P = 0·0020), and a lower relapse‐free survival (RFS) (5·4% vs. 61·0%, P = 0·0005) and overall survival (OS) (32·7% vs. 69·7%, P < 0·0001) than those who were EOI‐MRD negative. Moreover, for patients who received allogeneic haematopoietic stem cell transplantation (allo‐HSCT) at their first remission, EOI‐MRD positivity was predictive of post‐transplant relapse (2‐year CIR: 68·2% vs. 4·0%, P = 0·0003). Multivariate analysis showed that EOI‐MRD was an independent prognostic factor for CIR [hazard ratio (HR) 2·139, P = 0·046], RFS (HR 2·125, P = 0·048) and OS (HR 2·987, P = 0·017). In conclusion, EOI‐MRD based on FCM was an independent prognostic factor for relapse and survival in adult T‐ALL. For patients who underwent HSCT, EOI‐MRD could be used to identify patients with a high risk of relapse after allo‐HSCT. [ABSTRACT FROM AUTHOR]

Summary: Little data are available for the expression of immune checkpoint (IC) molecules within myelodysplastic syndrome (MDS). Here, we report increased PD‐L1+CD34+CD38− and PD‐L1+CD34+CD38+ stem cell frequencies within MDS patients compared to stem cell recipients in remission. Additionally, we observed exceedingly similar PD1+ and Tim‐3+ T‐cell frequencies between acute myeloid leukaemia (AML) and MDS samples that were elevated compared to patients in remission. Furthermore, we found highly dynamic Tim‐3+ and PD1+ T‐cell frequencies within serial samples of relapsing MDS with excess blasts (MDS‐EB II) patients, correlating with further disease markers. These findings support the idea of a potential successful implementation of IC inhibitor treatment in suitable MDS patients. [ABSTRACT FROM AUTHOR]

Summary: Given its myeloid‐restricted expression, myeloperoxidase (MPO) is typically used for lineage assignment (myeloid vs. lymphoid) during acute leukaemia (AL) diagnostics. In the present study, a robust flow cytometric definition for MPO positivity was established based on the standardised EuroFlow protocols, the standardised Acute Leukaemia Orientation Tube and 1734 multicentre AL cases (with confirmed assay stability). The best diagnostic performance was achieved by defining MPO positivity as ≥20% of the AL cells exceeding a lymphocyte‐based threshold. The methodology employed should be applicable to any form of standardised flow cytometry. [ABSTRACT FROM AUTHOR]

Summary: Splicing factor 3B subunit 1 (SF3B1) mutations define a distinct myelodysplastic syndromes (MDS) patient group with a relatively favourable disease course and high response rates to luspatercept. Few data are available on bone marrow phenotype beyond ring sideroblasts in this subgroup of patients with MDS. In the present study, we identified immunophenotypic erythroid, myelomonocyte and progenitor features associated with SF3B1 mutations. In addition, we illustrate that SF3B1‐mutation type is associated with distinct immunophenotypic features, and show the impact of co‐occurrence of a SF3B1 mutation and a deletion of chromosome 5q on bone marrow immunophenotype. These genotype–phenotype associations and phenotypic subtypes within SF3B1‐MDS provide leads that may further refine prognostication and therapeutic strategies for this particular MDS subgroup. [ABSTRACT FROM AUTHOR]

Summary: CD19‐directed treatment in B‐cell precursor acute lymphoblastic leukaemia (BCP‐ALL) frequently leads to the downmodulation of targeted antigens. As multicolour flow cytometry (MFC) application for minimal/measurable residual disease (MRD) assessment in BCP‐ALL is based on B‐cell compartment study, CD19 loss could hamper MFC‐MRD monitoring after blinatumomab or chimeric antigen receptor T‐cell (CAR‐T) therapy. The use of other antigens (CD22, CD10, CD79a, etc.) as B‐lineage gating markers allows the identification of CD19‐negative leukaemia, but it could also lead to misidentification of normal very‐early CD19‐negative BCPs as tumour blasts. In the current study, we summarized the results of the investigation of CD19‐negative normal BCPs in 106 children with BCP‐ALL who underwent CD19 targeting (blinatumomab, n = 64; CAR‐T, n = 25; or both, n = 17). It was found that normal CD19‐negative BCPs could be found in bone marrow after CD19‐directed treatment more frequently than in healthy donors and children with BCP‐ALL during chemotherapy or after stem cell transplantation. Analysis of the antigen expression profile revealed that normal CD19‐negative BCPs could be mixed up with residual leukaemic blasts, even in bioinformatic analyses of MFC data. The results of our study should help to investigate MFC‐MRD more accurately in patients who have undergone CD19‐targeted therapy, even in cases with normal CD19‐negative BCP expansion. [ABSTRACT FROM AUTHOR]

Summary: Eltrombopag has been added to first‐line treatment of immune aplastic anaemia (AA), resulting in higher responses. We analysed marrow samples of AA patients who responded to immunosuppressive therapy (IST) alone or in combination with eltrombopag for the composition of the haematopoietic stem and progenitor cell (HSPC) compartment. The number of CD34+ cells and multipotent progenitors was higher in patients treated with eltrombopag (P < 0·005; P < 0·05; respectively), but not the number of stem cells. No aberrant phenotype was observed. These results indicate that eltrombopag augments CD34+ cells in vivo and preferentially expands multipotent progenitors, but not stem cells. [ABSTRACT FROM AUTHOR]

Summary: Next‐generation sequencing (NGS) increasingly influences diagnosis, prognosis and management of myelodysplastic syndrome (MDS). In addition to marrow morphology and flow cytometry, our institution performs cytogenetics (CG) and NGS‐based testing routinely in patients with suspected MDS. We evaluated the relative value of NGS in the assessment of patients with suspected MDS. We initially compared the diagnostic and prognostic information derived from CG and NGS in 134 patients. NGS enhanced the diagnostic yield compared to CG for clonal myeloid disorders (sensitivity 77% vs. 42·2%; specificity 90·2% vs. 78%; positive predictive value 92·8% vs. 76%; and negative predictive value 70·8% vs. 45·5%). The identification of poor prognosis mutations by NGS altered risk category in 27/39 (69·2%) patients with MDS with good/intermediate risk CG. Subsequently, we prospectively evaluated 70 patients with suspected MDS using an 'NGS‐first approach' with CG restricted to samples with morphological abnormalities. We rarely identified mutations or CG abnormalities in patients without dysplastic features. NGS has a superior diagnostic performance compared to CG in patients with suspected MDS. We estimate that by using an 'NGS‐first approach' we could reduce karyotyping by approximately 30%. [ABSTRACT FROM AUTHOR]

Summary: Thrombotic events are common in patients with multiple myeloma (MM), smouldering myeloma (SM) and monoclonal gammopathy of undetermined significance (MGUS). Previous studies have indicated platelet hyperactivation as a feature of thrombotic risk in MM, but there is a dearth of data in MGUS. In the present study, multiparameter analysis of platelet activation and responsiveness was investigated by flow cytometry in patients with MGUS, SM/MM and healthy controls (HCs). The median platelet surface CD63 levels, annexin V and PAC‐1 antibody (specific for activated integrin αIIbβ3) binding were significantly elevated in patients with MGUS versus the HCs. These markers were also elevated in SM/MM, but not significantly. In all, 74% of MGUS and 38% of SM/MM patients had one or more elevated marker of platelet activation, compared to 19% of the HCs. Marker‐specific hyporesponsiveness of platelets to agonist [adenosine diphosphate (ADP), thrombin receptor‐activating peptide 6] stimulation in vitro was observed, with significantly reduced surface levels of P‐selectin in response to ADP in patients with MGUS. Platelet‐leucocyte aggregates were not altered in patients, while platelet‐associated immunoglobulins were elevated in a subset of patients. Overall, we found that platelet hyperactivation is prevalent in both MGUS and SM/MM patients and is potentially related to hyporesponsiveness. These observations suggest that further investigation of the predictive and prognostic value of platelet hyperactivation in such patients is warranted. [ABSTRACT FROM AUTHOR]

The diagnosis of juvenile myelomonocytic leukaemia (JMML) is based on clinical, laboratory and molecular features but immunophenotyping [multiparametric flow cytometry (MFC)] has not been used routinely. In the present study, we describe the flow cytometric features at diagnosis with special attention to the distribution of monocytic subsets and the relation between MFC and molecular subgroups. MFC was performed with an eight‐colour platform based on Euroflow. We studied 33 JMML cases. CD34+/CD117+/CD13+ cells >2% was found in 25 cases, and 51·5% presented an aberrant expression of CD7. A decrease of CD34+/CD19+/CD10+ cells was seen in eight cases and in four they were absent. The granulocytic population had a decreased side scatter in 29 cases. Bone marrow monocytic precursors were increased in 28 patients, with a decrease in classical monocytes (median 80·7%) and increase in CD16+ (intermediate and non‐classical). A more pronounced increase in myeloid CD34+ cells was seen in patients with Neurofibromatosis type 1 (NF1) and tyrosine‐protein phosphatase non‐receptor type 11 (PTPN11), with aberrant CD7 expression in four of six and 10/12 patients respectively. Thus, JMML shows an immunophenotypic profile similar to myelodysplastic syndromes, and a different monocyte subset distribution when compared with chronic MML. MFC proved to be an important diagnostic tool that can help in differential diagnosis with other clonal diseases with monocytosis. [ABSTRACT FROM AUTHOR]

Induction therapy followed by CD34 + cell mobilisation and autologous transplantation represents standard of care for multiple myeloma (MM). However, the anti-CD38 monoclonal antibodies daratumumab and isatuximab have been associated with mobilisation impairment, yet the mechanism remains unclear. In this study, we investigated the effect of three different regimens (dara-VCd, isa-KRd and VTd) on CD34 + cells using flow cytometry and transcriptomics. Decreased CD34 + cell peak concentration and yields, longer collection and delayed engraftment were reproduced after dara-VCd/isa-KRd versus VTd induction in 34 patients in total. Using flow cytometry, we detected major changes in the proportion of apheresis product and bone marrow CD34 + subsets in patients treated with regimens containing anti-CD38 therapy; however, without any decrease in CD38high B-lymphoid progenitors in both materials. RNA-seq of mobilised CD34 + cells from 21 patients showed that adhesion genes are overexpressed in CD34 + cells after dara-VCd/isa-KRd and JCAD, NRP2, MDK, ITGA3 and CLEC3B were identified as potential target genes. Finally, direct in vitro effect of isatuximab in upregulating JCAD and CLEC3B was confirmed by quantitative PCR. These findings suggest that upregulated adhesion-related interactions, rather than killing of CD34 + cells by effector mechanisms, could be leading causes of decreased mobilisation efficacy in MM patients treated with anti-CD38 therapy., (© 2023 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Immune aplastic anaemia (AA) is caused by cytotoxic T lymphocytes (CTLs) that destroy haematopoietic stem and progenitor cells. Enhanced type 1 T helper (Th1) responses and reduced regulatory T cells (Tregs) are involved in the immune pathophysiology. CD24hiCD38hi regulatory B cells (Bregs) suppress CTLs and Th1 responses, and induce Tregs via interleukin 10 (IL‐10). We investigated circulating B‐cell subpopulations, including CD24hiCD38hi Bregs, as well as total B cells, CD4+ T cells, CD8+ T cells and natural killer cells in 104 untreated patients with severe and very severe AA, aged ≥18 years. All patients were treated with standard immunosuppressive therapy (IST) plus eltrombopag. CD24hiCD38hi Bregs were markedly reduced in patients with AA compared to healthy individuals, especially in very severe AA, but residual Bregs remained functional, capable of producing IL‐10; total B‐cell counts and the other B‐cell subpopulations were similar to those of healthy individuals. CD24hiCD38hi Bregs did not correlate with responses to IST, and they recovered to levels present in healthy individuals after therapy. Mature naïve B‐cell counts were unexpectedly associated with IST response. Markedly reduced CD24hiCD38hi Bregs, especially in very severe AA, with recovery after IST suggest Breg deficits may contribute to the pathophysiology of immune AA. [ABSTRACT FROM AUTHOR]

Summary: Imatinib mesylate (IM) resistance has become a major clinical problem for chronic myeloid leukaemia (CML). It is known that Bcl‐x splicing is deregulated and is involved in multiple malignant cancer initiation and chemotherapy resistance, including CML. The aim of the present study was to correct the abnormal splicing of Bcl‐x in CML and investigate the subsequent malignant phenotype changes, especially response to IM. The aberrant Bcl‐x splicing in CML cells was effectively restored using vivo‐Morpholino Antisense Oligomer (vMO). CCK‐8 cell viability assay and flow cytometry showed that restoring of Bcl‐x splicing increases IM‐induced growth inhibition and apoptosis of K562 cells. Moreover, a more significant similar phenomenon was observed in imatinib‐resistant CML cell lines K562/G01. Finally, establishment of CML xenograft model had also proved that correcting Bcl‐x splicing in vivo can also enhance the anti‐tumor effect of IM. Our findings suggest that vMO co‐operating with IM can effectively increase the sensitivity of CML cells to IM both in vitro and in vivo, and Bcl‐x splicing could become good candidates for chemotherapy‐sensitized target in IM‐resistant CML. [ABSTRACT FROM AUTHOR]

Summary: Matrix‐assisted laser desorption ionisation time‐of‐flight mass spectrometry (MALDI‐TOF MS) may soon replace routine electrophoretic methods for monitoring monoclonal proteins in patients with multiple myeloma. To further evaluate the clinical utility of this assay, we compared the performance of MALDI‐TOF‐MS head‐to‐head with an established bone marrow‐based measurable residual disease assay by flow cytometry (Flow‐BM‐MRD), using Memorial Sloan Kettering Cancer Center's 10‐color, single‐tube method. Our results suggest that MALDI‐TOF‐MS adds value to bone marrow‐based MRD testing and may be most useful for early detection of relapse in peripheral blood compared to current electrophoretic methods. [ABSTRACT FROM AUTHOR]

Summary: A retrospective analysis of presentation clinical, laboratory and immunophenotypic features of 1 081 patients with paroxysmal nocturnal haemoglobinuria (PNH) clones [glycosylphosphatidylinositol (GPI)‐deficient blood cells] identified at our hospital by flow cytometry over the past 25 years was undertaken. Three distinct clusters of patients were identified and significant correlations between presentation disease type and PNH clone sizes were evident. Smaller PNH clones predominate in cytopenic and myelodysplastic subtypes; large PNH clones were associated with haemolytic, thrombotic and haemolytic/thrombotic subtypes. Rare cases with an associated chronic myeloproliferative disorder had either large or small PNH clones. Cytopenia was a frequent finding, highlighting bone marrow failure as the major underlying feature associated with the detection of PNH clones in the peripheral blood. Red cell PNH clones showed significant correlations between the presence of type II (partial GPI deficiency) red cells and thrombotic disease. Haemolytic PNH was associated with type III (complete GPI deficiency) red cell populations of >20%. Those with both haemolytic and thrombotic features had major type II and type III red cell populations. Distinct patterns of presentation age decade were evident for clinical subtypes with a peak incidence of haemolytic PNH in the 30–49 year age group and a biphasic age distribution for the cytopenia group. [ABSTRACT FROM AUTHOR]

Summary: Platelet function of immune thrombocytopenia (ITP) has been controversial because of methodological problems associated with low platelet counts. In this study, we evaluated platelet function in 21 patients with chronic ITP (cITP) using the recently developed flow cytometry (FCM)‐based platelet aggregation assay (FCA) along with a PAC1/CD62P assay. Since ITP platelets are larger than controls, whole platelets (whole gating method) and size‐adjusted platelets (size‐adjusted method) were analysed in the PAC1/CD62P via FCM. We found that: (i) aggregation was equivalent [phorbol myristate acetate (PMA) or adenosine diphosphate (ADP)‐induced] or enhanced [protease‐activated receptor 1‐activating peptide (PAR1AP)‐induced] in cITP compared with control by FCA; (ii) PAC1 or CD62P was also equivalent or enhanced in cITP in the whole gating method; and (iii) in sharp contrast, the size‐adjusted method revealed that ADP‐, PAR1AP‐, and collagen synthetic liquid reactive peptide (SRP)‐induced PAC1 and ADP‐induced CD62P were impaired in cITP. These data suggested that an increase in the number of larger‐sized platelets may compensate for the impaired platelet function of cITP, leading to non‐inferiority of overall platelet function in cITP. Furthermore, we revealed that ADP‐induced aggregation was impaired in the patients with thrombopoietin receptor agonists (TPO‐RAs) or platelet‐associated anti‐αIIbβ3 antibodies compared with the control, suggesting that the presence of anti‐αIIbβ3 autoantibodies and/or administration of TPO‐RAs may have a negative impact on platelet function. [ABSTRACT FROM AUTHOR]

Summary: Mesenchymal stromal cells (MSCs) demonstrate the properties of self‐renewal, multipotentiality, and immunosuppression, which are responsible for their widespread clinical applications in tissue repair and regeneration. MSCs have been isolated from bone marrow, adipose tissue, and cord blood using culture in specialised media. Their presence in peripheral blood (PB) is debatable. We studied the presence of MSCs at baseline (PB) and following mobilisation with growth factors [PB and apheresis product (AP)] in patients undergoing autologous stem cell transplantation and healthy donors using flow cytometry. We conclude that both mobilised PB and AP are potential sources of MSCs. Given their small numbers in PB/AP, clinical use is feasible following ex‐vivo expansion. Variables affecting the presence of MSCs in PB and AP are also discussed. [ABSTRACT FROM AUTHOR]

Keywords: haematopoietic stem cell transplantation; granulocyte colony-stimulating factor; stem cell mobilization; mesenchymal stromal cells; blood circulation; peripheral blood; apheresis product; flow cytometry EN haematopoietic stem cell transplantation granulocyte colony-stimulating factor stem cell mobilization mesenchymal stromal cells blood circulation peripheral blood apheresis product flow cytometry 593 596 4 05/19/20 20200515 NES 200515 In this issue of the I British Journal of Haematology i , Jain I et al. i identified circulating mesenchymal stromal cells (MSCs) in peripheral blood (PB) and apheresis product (AP) collected in the context of haematopoietic stem cell (HSC) mobilization for HSC transplantation (HSCT). [Colour figure can be viewed at wileyonlinelibrary.com] gl Intriguingly, Larsen I et al. i noted in a baboon model that MSC mobilization and colony-forming unit fibroblast (CFU-F) in PB in response to G-CSF did only occur when adding stem cell factor (To I et al. i , [25]). Haematopoietic stem cell transplantation, granulocyte colony-stimulating factor, stem cell mobilization, mesenchymal stromal cells, blood circulation, peripheral blood, apheresis product, flow cytometry. [Extracted from the article]

Summary: This study was conducted to evaluate the expression of fibrinogen receptors on platelets of Philadelphia‐negative chronic myeloproliferative neoplasm (MPN) patients. We collected blood samples from 40 consecutive MPN patients and healthy volunteers. We performed flow cytometry analysis of P‐selectin expression and integrin beta‐3, activation of glycoprotein (GP) IIb/IIIa and fibrinogen receptor exposure (PAC‐1 binding). Surprisingly, we found a very low PAC‐1 binding capacity in MPN patients; however, the expression of PAC‐1 was almost completely recovered with aspirin intake. We hypothesize that the hypercoagulable states observed in MPN patients could depend on a primarily plasma‐driven impairment of fibrin turnover and thrombin generation. [ABSTRACT FROM AUTHOR]

Deficiency in regulatory B cells has been suggested in immune thrombocytopenia. In this study, Stimpson et al. emphasize the importance of considering the treatments received for immunological analyses. Commentary on: Stimpson et al. Systemic immunosuppression depletes peripheral blood regulatory B cells in patients with immune thrombocytopenia. Br J Haematol 2024;204:644-648., (© 2023 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Keywords: platelet function; flow cytometry; immune thrombocytopenia; bleeding risk; romiplostim Immune thrombocytopenia (ITP) is an autoimmune disease caused by auto-antibodies against platelets. Prior to treatment, 9 patients had bleeding complications and 22 patients were asymptomatic. In contrast, platelet activation responses to potent stimulation with thrombin plus collagen receptor agonists were not essentially affected in ITP, and did not differ between the bleeding and non-bleeding groups (Fig I; Figure S1F-I). [Extracted from the article]

Summary: Treatment response assessment in multiple myeloma (MM) relies on the detection of paraprotein in serum and/or urine, bone marrow morphology and immunohistochemistry. With remarkable advances in therapy, particularly in the newly diagnosed setting, achievement of complete remission became frequent, creating the need to identify smaller amounts of residual disease and understand their prognostic and therapeutic implications. Measurable residual disease (MRD) can be assessed primarily by flow cytometry and next generation sequencing and state‐of‐the‐art assays have sensitivity approaching 1 in 106 cells. This review discusses the existing challenges in utilizing MRD to inform management of MM and highlights open research questions and opportunities as MRD is more routinely incorporated into clinical practice for patients with MM. [ABSTRACT FROM AUTHOR]