Predictors of clinical outcome in myeloproliferative neoplasm, unclassifiable: A Bone Marrow Pathology Group study.

OBJECTIVES: Myeloproliferative neoplasm, unclassifiable (MPN-U, revised to MPN, not otherwise specified in the fifth edition of the World Health Organization classification) is a heterogeneous category of primary marrow disorders with clinical, morphologic, and/or molecular features that preclude classification as a more specific MPN subtype due to stage at diagnosis, overlapping features between MPN subtypes, or the presence of coexisting disorders. Compared with other MPN subtypes, the contribution of the mutational landscape in MPN-U in conjunction with other clinical and morphologic biomarkers to prognosis has been less well investigated. METHODS: We performed a multicenter, retrospective study of MPN-U (94 cases) to better define the clinicopathologic features, genetic landscape, and clinical outcomes, including subgroups of early-stage, advanced-stage, and coexisting disorders. The Dynamic International Prognostic Scoring System (DIPSS) plus scoring system was applied to assess its relevance to MPN-U prognosis. RESULTS: Multivariate analysis demonstrated bone marrow blast count and DIPSS plus score as statistically significant in predicting overall survival. Univariate analysis identified additional potential poor prognostic markers, including abnormal karyotype and absence of JAK2 mutation. Secondary mutations were frequent in the subset analyzed by next-generation sequencing (26/37 cases, 70.3%) with a borderline association between high molecular risk mutations and overall survival. CONCLUSIONS: This study, as one of the largest of MPN-U studies incorporating both clinicopathologic and molecular data, moves toward identification of biomarkers that better predict prognosis in this heterogeneous category.

Molecular and clinical presentation of UBA1-mutated myelodysplastic syndromes.

Mutations in UBA1, which are disease-defining for VEXAS syndrome, have been reported in patients diagnosed with myelodysplastic syndromes (MDS). Here, we define the prevalence and clinical associations of UBA1 mutations in a representative cohort of patients with MDS. Digital droplet PCR profiling of a selected cohort of 375 male patients lacking MDS disease-defining mutations or established WHO disease classification identified 28 patients (7%) with UBA1 p.M41T/V/L mutations. Using targeted sequencing of UBA1 in a representative MDS cohort (n=2,027), we identified an additional 27 variants in 26 patients (1%), which we classified as likely/pathogenic (n=12) and unknown significance (n=15). Among the total 40 patients with likely/pathogenic variants (2%), all were male and 63% were classified by WHO2016 as MDS-MLD/SLD. Patients had a median of one additional myeloid gene mutation, often in TET2 (n=12), DNMT3A (n=10), ASXL1 (n=3), or SF3B1 (n=3). Retrospective clinical review where possible showed that 83% (28/34) UBA1-mutant cases had VEXAS-associated diagnoses or inflammatory clinical presentation. The prevalence of UBA1-mutations in MDS patients argues for systematic screening for UBA1 in the management of MDS.

Follicular Lymphoma Presenting With Symptomatic Bone Involvement: A Clinicopathologic and Molecular Analysis of 16 Cases.

Primary bone lymphoma (PBL) is rare and mostly represented by diffuse large B-cell lymphomas (DLBCL). Follicular lymphoma (FL), albeit commonly disseminating to the bone marrow, rarely presents primarily as bone lesions. Here, we studied 16 patients (12 men:4 women, median age 60 years) who presented with bone pain and/or skeletal radiologic abnormalities revealing bone FL. Lesions were multifocal in 11 patients (spine ± appendicular skeleton), and unifocal in 5 patients (femoral, tibial, or vertebral). An infiltrate of centrocytes and centroblasts (CD20+ CD5- CD10+ BCL2+ BCL6+) with abundant reactive T cells and an increased reticulin fibrosis massively replaced the marrow spaces between preserved bone trabeculae. The pattern was diffuse ± nodular, often with paratrabecular reinforcement and/or peripheral paratrabecular extension. Ki-67 was usually <15%. Two cases had necrosis. BCL2 rearrangement was demonstrated in 14 of 14 evaluable cases (with concomitant BCL6 rearrangement in one). High-throughput sequencing revealed BCL2, KMT2D, and TNFRSF14 to be the most frequently mutated genes. After staging, 5 qualified for PBL (3 limited stage) and 11 had stage IV systemic FL. All patients received rituximab ± polychemotherapy as firstline treatment, and 7 received local therapy (6 radiotherapy and 2 surgery). Three patients experienced transformation to DLBCL. At the last follow-up (15/16, median 48 months), 11 patients achieved complete remission, including all cases with PBL and most patients with limited extraosseous disease (3-year progression-free survival 71%). One patient died of unrelated cause (3-year overall survival 91%). FL may manifest as a localized or polyostotic bone disease. A minority represent PBL, whereas most reveal systemic disease.

Cutaneous Manifestations of Myeloid Neoplasms Exhibit Broad and Divergent Morphologic and Immunophenotypic Features but Share Ancestral Clonal Mutations With Bone Marrow.

In this study, we performed a comprehensive molecular analysis of paired skin and peripheral blood/bone marrow (BM) samples from 17 patients with cutaneous myeloid or cutaneous histiocytic-dendritic neoplasms. The cutaneous manifestations included 10 patients with cutaneous acute myeloid leukemia (c-AML), 2 patients with full or partial Langerhans cell differentiation, 2 patients with blastic plasmacytoid dendritic cell neoplasms (BPDCN), 1 patient with both Langerhans cell differentiation and BPDCN, and 2 patients with full or partial indeterminate dendritic cell differentiation. Seven of the 10 c-AML patients (70%) exhibited concurrent or subsequent marrow involvement by acute myeloid leukemia, with all 7 cases (100%) demonstrating shared clonal mutations in both the skin and BM. However, clonal relatedness was documented in one additional case that never had any BM involvement. Nevertheless, NPM1 mutations were identified in 7 of the 10 (70%) of these c-AML cases while one had KMT2A rearrangement and one showed inv(16). All 3 patients (100%) with Langerhans cell neoplasms, 2 patients with BPDCN (100%), and one of the 2 patients (50%) with other cutaneous dendritic cell neoplasms also demonstrated shared mutations between the skin and concurrent or subsequent myeloid neoplasms. Both BM and c-AML shared identical founding drivers, with a predominance of NPM1, DNMT3A, and translocations associated with monocytic differentiation, with common cutaneous-only mutations involving genes in the signal transduction and epigenetic pathways. Cutaneous histiocytic-dendritic neoplasms shared founding drivers in ASXL1, TET2, and/or SRSF2. However, in the Langerhans cell histiocytosis or histiocytic sarcoma cases, there exist recurrent secondary RAS pathway hits, whereas cutaneous BPDCN cases exhibit copy number or structural variants. These results enrich and broaden our understanding of clonally related cutaneous manifestations of myeloid neoplasms and further illuminate the highly diverse spectrum of morphologic and immunophenotypic features they exhibit.

STAT5B mutations in myeloid neoplasms differ by disease subtypes but characterize a subset of chronic myeloid neoplasms with eosinophilia and/or basophilia.

STAT5B has been reported as a recurrent mutation in myeloid neoplasms (MNs) with eosinophilia, but the overall frequency and importance across a spectrum of MNs are largely unknown. We conducted a multicenter study on a series of 82 MNs with STAT5B mutations detected by next-generation sequencing. The estimated frequency of STAT5B mutation in MNs was low.

Morphologic Characteristics of Myelodysplastic Syndromes.

Morphologic characterization remains a cornerstone in the diagnosis and classification of myelodysplastic syndromes (MDS) in the updated International Consensus Classification (ICC) and 5th edition World Health Organization Classification of Myeloid Neoplasms (Arber, Orazi, & Hasserjian, 2022; Khoury & Solary, 2022). The presence of dysplasia is one of the key diagnostic criteria required for establishing a diagnosis of MDS, and the percentage of myeloblasts in the blood and bone marrow impacts both disease classification and prognostication. Morphologic features also aid in distinguishing MDS from a myriad of other myeloid neoplasms and non-neoplastic mimics. Additional key morphologic features that should be recorded in any MDS case are the bone marrow cellularity and the degree of reticulin fibrosis. In this review, the morphologic assessment of the bone marrow biopsy, bone marrow aspirate, and peripheral blood smear as it pertains to the diagnosis and up-to-date classification of MDS will be described. The implications of the findings on classification and prognosis will also be discussed.

Diagnosis and classification of myelodysplastic syndromes.

ABSTRACT: Myelodysplastic syndromes (MDSs) are neoplastic myeloid proliferations characterized by ineffective hematopoiesis resulting in peripheral blood cytopenias. MDS is distinguished from nonneoplastic clonal myeloid proliferations by the presence of morphologic dysplasia and from acute myeloid leukemia by a blast threshold of 20%. The diagnosis of MDS can be challenging because of the myriad other causes of cytopenias: accurate diagnosis requires the integration of clinical features with bone marrow and peripheral blood morphology, immunophenotyping, and genetic testing. MDS has historically been subdivided into several subtypes by classification schemes, the most recent of which are the International Consensus Classification and World Health Organization Classification (fifth edition), both published in 2022. The aim of MDS classification is to identify entities with shared genetic underpinnings and molecular pathogenesis, and the specific subtype can inform clinical decision-making alongside prognostic risk categorization. The current MDS classification schemes incorporate morphologic features (bone marrow and blood blast percentage, degree of dysplasia, ring sideroblasts, bone marrow fibrosis, and bone marrow hypocellularity) and also recognize 3 entities defined by genetics: isolated del(5q) cytogenetic abnormality, SF3B1 mutation, and TP53 mutation. It is anticipated that with advancing understanding of the genetic basis of MDS pathogenesis, future MDS classification will be based increasingly on genetic classes. Nevertheless, morphologic features in MDS reflect the phenotypic expression of the underlying abnormal genetic pathways and will undoubtedly retain importance to inform prognosis and guide treatment.

Classification, risk stratification and response assessment in myelodysplastic syndromes/neoplasms (MDS): A state-of-the-art report on behalf of the International Consortium for MDS (icMDS).

The guidelines for classification, prognostication, and response assessment of myelodysplastic syndromes/neoplasms (MDS) have all recently been updated. In this report on behalf of the International Consortium for MDS (icMDS) we summarize these developments. We first critically examine the updated World Health Organization (WHO) classification and the International Consensus Classification (ICC) of MDS. We then compare traditional and molecularly based risk MDS risk assessment tools. Lastly, we discuss limitations of criteria in measuring therapeutic benefit and highlight how the International Working Group (IWG) 2018 and 2023 response criteria addressed these deficiencies and are endorsed by the icMDS. We also address the importance of patient centered care by discussing the value of quality-of-life assessment. We hope that the reader of this review will have a better understanding of how to classify MDS, predict clinical outcomes and evaluate therapeutic outcomes.

Presence but not number of secondary type mutations influences outcome in de novo AML without MDS-associated or recurring cytogenetic abnormalities.

A group of gene mutations has been identified to be strongly associated with secondary acute myeloid leukemias (AML) arising from prior myeloid neoplasms. The International Consensus Classification (ICC) and proposed 5th edition of the World Health Organization (WHO) classification differ by inclusion of RUNX1. A recent study suggested that having two or more secondary mutations is associated with a particularly poor prognosis. In a study of 294 de novo AML patients, we found that patients with at least one ICC-defined secondary mutation had shorter survival when compared to those without secondary mutations, and ICC/WHO groups of two or more mutations did not predict for worse outcomes.

Prognostic impact of secondary versus de novo ontogeny in acute myeloid leukemia is accounted for by the European LeukemiaNet 2022 risk classification.

Secondary AML (sAML), defined by either history of antecedent hematologic disease (AHD) or prior genotoxic therapy (tAML), is classically regarded as having worse prognosis than de novo disease (dnAML). Clinicians may infer a new AML diagnosis is secondary based on a history of antecedent blood count (ABC) abnormalities in the absence of known prior AHD, but whether abnormal ABCs are associated with worse outcomes is unclear. Secondary-type mutations have recently been incorporated into the European LeukemiaNet (ELN) 2022 guidelines as adverse-risk features, raising the question of whether clinical descriptors of ontogeny (i.e., de novo or secondary) are prognostically significant when accounting for genetic risk by ELN 2022. In a large multicenter cohort of patients (n = 734), we found that abnormal ABCs are not independently prognostic after adjusting for genetic characteristics in dnAML patients. Furthermore, history of AHD and tAML do not confer increased risk of death compared to dnAML on multivariate analysis, suggesting the prognostic impact of ontogeny is accounted for by disease genetics as stratified by ELN 2022 risk and TP53 mutation status. These findings emphasize the importance that disease genetics should play in risk stratification and clinical trial eligibility in AML.

Guide to the Diagnosis of Myeloid Neoplasms: A Bone Marrow Pathology Group Approach.

OBJECTIVES: The practicing pathologist is challenged by the ever-increasing diagnostic complexity of myeloid neoplasms. This guide is intended to provide a general roadmap from initial case detection, often triggered by complete blood count results with subsequent blood smear review, to final diagnosis. METHODS: The integration of hematologic, morphologic, immunophenotypic, and genetic features into routine practice is standard of care. The requirement for molecular genetic testing has increased along with the complexity of test types, the utility of different testing modalities in identifying key gene mutations, and the sensitivity and turnaround time for various assays. RESULTS: Classification systems for myeloid neoplasms have evolved to achieve the goal of providing a pathology diagnosis that enhances patient care, outcome prediction, and treatment options for individual patients and is formulated, endorsed, and adopted by hematologists/oncologists. CONCLUSIONS: This guide provides diagnostic strategies for all myeloid neoplasm subtypes. Special considerations are provided for each category of testing and neoplasm category, along with classification information, genetic testing requirements, interpretation information, and case reporting recommendations based on the experience of 11 Bone Marrow Pathology Group members.

The international consensus classification of eosinophilic disorders and systemic mastocytosis.

Based on new data and increased understanding of disease molecular genetics, the international consensus classification (ICC) has made several changes in the diagnosis and classification of eosinophilic disorders and systemic mastocytosis. Myeloid/lymphoid neoplasms with eosinophilia (M/LN-eo) and gene rearrangements have been renamed as M/LN-eo with tyrosine kinase gene fusions (M/LN-eo-TK). The category has been expanded to include ETV6::ABL1 and FLT3 fusions, and to accept PCM1::JAK2 and its genetic variants as formal members. The overlaps and differences between M/LN-eo-TK and BCR::ABL1-like B-lymphoblastic leukemia (ALL)/de novo T-ALL sharing the same genetic lesions are addressed. Besides genetics, ICC for the first time has introduced bone marrow morphologic criteria in distinguishing idiopathic hypereosinophilia/hypereosinophilic syndrome from chronic eosinophilic leukemia, not otherwise specified. The major diagnostic criteria for systemic mastocytosis (SM) in the ICC remain largely based on morphology, but several minor modifications/refinements have been made in criteria related to diagnosis, subclassification, and assessment of disease burden (B- and C-findings). This review is to focus on the ICC updates related to these disease entities, illustrated through changes related to morphology, molecular genetics, clinical features, prognosis, and treatment. Two practical algorithms are provided in navigating through the diagnosis and classification systems of hypereosinophilia and SM.

Computer vision quantitation of erythrocyte shape abnormalities provides diagnostic, prognostic, and mechanistic insight.

Examination of red blood cell (RBC) morphology in peripheral blood smears can help diagnose hematologic diseases, even in resource-limited settings, but this analysis remains subjective and semiquantitative with low throughput. Prior attempts to develop automated tools have been hampered by their poor reproducibility and limited clinical validation. Here, we present a novel, open-source machine-learning approach (denoted as RBC-diff) to quantify abnormal RBCs in peripheral smear images and generate an RBC morphology differential. RBC-diff cell counts showed high accuracy for single-cell classification (mean AUC, 0.93) and quantitation across smears (mean R2, 0.76 compared with experts, interexperts R2, 0.75). RBC-diff counts were concordant with the clinical morphology grading for 300 000+ images and recovered the expected pathophysiologic signals in diverse clinical cohorts. Criteria using RBC-diff counts distinguished thrombotic thrombocytopenic purpura and hemolytic uremic syndrome from other thrombotic microangiopathies, providing greater specificity than clinical morphology grading (72% vs 41%; P < .001) while maintaining high sensitivity (94% to 100%). Elevated RBC-diff schistocyte counts were associated with increased 6-month all-cause mortality in a cohort of 58 950 inpatients (9.5% mortality for schist. >1%, vs 4.7% for schist; <0.5%; P < .001) after controlling for comorbidities, demographics, clinical morphology grading, and blood count indices. RBC-diff also enabled the estimation of single-cell volume-morphology distributions, providing insight into the influence of morphology on routine blood count measures. Our codebase and expert-annotated images are included here to spur further advancement. These results illustrate that computer vision can enable rapid and accurate quantitation of RBC morphology, which may provide value in both clinical and research contexts.

Updates in Classification of Myelodysplastic Syndrome.

ABSTRACT: Myelodysplastic syndrome includes a broad range of myeloid neoplasms characterized by cytopenia and morphologic dysplasia. Recently, 2 new classification systems emerged to further define how these diseases are diagnosed and risk stratified. This review compares these models, provides detailed approaches, and reveals practical ways to move forward in clinical practice of myelodysplastic syndrome diagnosis.

Acute leukemias with complex karyotype show a similarly poor outcome independent of mixed, myeloid or lymphoblastic immunophenotype: A study from the Bone Marrow Pathology Group.

Mixed phenotype acute leukemia (MPAL) is a heterogenous group of acute leukemias characterized by leukemic blasts that express markers of multiple lineages. The revised 4th edition WHO classification of MPAL excludes AML with myelodysplasia related changes (AML-MRC), including those with complex karyotype (CK), from a diagnosis of MPAL. Abnormal karyotype is frequent in MPAL with the reported rate of CK in MPAL ranging from 19% to 32%. Due its rarity, the clinical and genetic features of MPAL with CK remain poorly characterized. This study aims to further characterize the genetic features of MPAL with CK in comparison to cases of AML and ALL with CK. Cases of de novo MPAL, AML, and B- and T-ALL patients with CK were collected from 8 member institutions of the Bone Marrow Pathology Group. We found no significant difference in overall survival between MPAL with CK compared to AML and ALL with CK. AML with CK was more strongly associated with TP53 mutations, however the presence of TP53 mutations conferred a worse prognosis regardless of lineage. ALL with CK seems to show increased IKZF1 mutation rates which is known to confer a worse prognosis in ALL. Additionally, MPAL with CK showed similarly poor outcomes regardless of whether a lymphoid or myeloid chemotherapy regimen is chosen. Our results suggest that acute leukemias with complex karyotype show a similarly poor outcome regardless of lineage differentiation and that mutation in TP53 confers a poor prognosis in all lineages. Our results support the exclusion of immunophenotypic MPAL with CK from MPAL and appear to confirm the approach proposed in the revised 4th edition WHO to include them as AML with myelodysplasia-related changes and similar myelodysplasia-related AML categories of newer classifications.

Myeloid sarcoma with NPM1 mutation may be clinically and genetically distinct from AML with NPM1 mutation: a study from the Bone Marrow Pathology Group.

Myeloid sarcoma (MS) is currently considered equivalent to de novo acute myeloid leukemia (AML); however, the relationship between these entities is poorly understood. This retrospective multi-institutional cohort study compared 43 MS with NPM1 mutation to 106 AML with NPM1 mutation. Compared to AML, MS had more frequent cytogenetic abnormalities including complex karyotype (p = .009 and p = .007, respectively) and was enriched in mutations of genes involved in histone modification, including ASXL1 (p = .007 and p = .008, respectively). AML harbored a higher average number of gene mutations (p = .002) including more frequent PTPN11 mutations (p < .001) and mutations of DNA-methylating genes including DNMT3A and IDH1 (both p < .001). MS had significantly shorter overall survival (OS) than AML (median OS: 44.9 vs. 93.2 months, respectively, p = .037). MS with NPM1 mutation has a unique genetic landscape, and poorer OS, compared to AML with NPM1 mutation.

First study comparing genetic profiles of MS and AML with a common disease-defining lesion.NPM1^Mut MS may be genetically distinct from NPM1^Mut AML.NPM1^Mut MS may have inferior overall survival compared to NPM1^Mut AML.

Current landscape of translational and clinical research in myelodysplastic syndromes/neoplasms (MDS): Proceedings from the 1st International Workshop on MDS (iwMDS) Of the International Consortium for MDS (icMDS).

Biological events that contribute to the pathogenesis of myelodysplastic syndromes/neoplasms (MDS) are becoming increasingly characterized and are being translated into rationally designed therapeutic strategies. Herein, we provide updates from the first International Workshop on MDS (iwMDS) of the International Consortium for MDS (icMDS) detailing recent advances in understanding the genetic landscape of MDS, including germline predisposition, epigenetic and immune dysregulation, the complexities of clonal hematopoiesis progression to MDS, as well as novel animal models of the disease. Connected to this progress is the development of novel therapies targeting specific molecular alterations, the innate immune system, and immune checkpoint inhibitors. While some of these agents have entered clinical trials (e.g., splicing modulators, IRAK1/4 inhibitors, anti-CD47 and anti-TIM3 antibodies, and cellular therapies), none have been approved for MDS. Additional preclinical and clinical work is needed to develop a truly individualized approach to the care of MDS patients.

Triple-Negative Primary Myelofibrosis: A Bone Marrow Pathology Group Study.

Primary myelofibrosis (PMF) is a clonal myeloproliferative neoplasm driven by canonical gene mutations in JAK2, CALR, or MPL in >80% of the cases. PMF that lacks these canonical alterations is termed triple-negative PMF (TN-PMF). The pathologic and genetic characteristics of TN-PMF compared with those of conventional PMF with canonical driver mutations (DM-PMF) have not been well studied. We aimed to identify clinicopathologic and molecular genetic differences between patients with TN-PMF (n = 56) and DM-PMF (n = 89), all of whom fulfilled the 2016 World Health Organization diagnostic criteria for PMF. Compared with the control group, patients in the TN-PMF group were more likely to have thrombocytopenia and less likely to have organomegaly. The bone marrow in patients with TN-PMF showed fewer granulocytic elements and more frequent dyserythropoiesis. Cytogenetic analysis showed a higher incidence of trisomy 8. Targeted next-generation sequencing revealed a lower frequency of ASXL1 mutations but enrichment of ASXL1/SRSF2 comutations. Our findings demonstrated several clinicopathologic and molecular differences between TN-PMF and DM-PMF. These findings, particularly the observed mutation profile characterized by a higher frequency of ASXL1 and SRSF2 comutation, suggest that at least a subset of TN-PMF may be pathogenetically different from DM-PMF, with potential prognostic implications.