Myeloproliferative Neoplasms Subtypes Research Articles

CD56briCD38+ as a novel neutrophil-specific marker in chronic myeloid leukemia

BackgroundChronic myeloid leukemia (CML) is classified as a subtype of myeloproliferative neoplasm, and bone marrow flow cytometry does not reveal any specific immunophenotype. Interestingly, aberrant expression of cluster of differentiation (CD) markers such as CD56 and CD38 has been observed on neutrophils in CML patients. Therefore, we investigated the abnormal expression of CD56 and CD38 in CML neutrophils to explore their diagnostic value in identifying CML through flow cytometry. MethodsWe have developed a multi-parameter flow cytometry assay to identify aberrant immunophenotypes in CML neutrophils among bone marrow nucleated cells. ResultsCompared to healthy donors and patients with a reactive neutrophilia or other hematological malignancies, the percentage of CD56briCD38+ neutrophil subsets in CML patients exhibits a distinctive increase (cut-off value, 2.0 %). The specificity and sensitivity associated with the learning cohort (168 samples) were 90.8 % and 84.9 %, respectively, while in the validation cohort (194 samples), they were 90.7 % and 84.7 %. The accumulation of CD56briCD38+ neutrophil subsets, which demonstrate are abnormal characteristics, is independent of the neutrophil count, BCR/ABL1 fusions and risk stratification but associated with blast cells immunophenotype. Moreover, this increase disappears in CML patients after treatment with tyrosine kinase inhibitors when the curative effect was satisfactory. ConclusionsWe conclude that an increase in the proportion of CD56briCD38+ neutrophil subsets exceeding 2.0 % of total neutrophils serves as a highly sensitive and specific flow cytometry marker, enabling rapid and accurate identification of CML.

Association between JAK2V617F variable allele frequency and risk of thrombotic events in patients with myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPNs) are a group of chronic disorders of the bone marrow characterised by the overproduction of clonal myeloid stem cells. The most common driver mutation found in MPNs is a point mutation on exon 14 of the JAK2 gene, JAK2V617F. Various studies have suggested that measuring the variable allele frequency (VAF) of JAK2V617F may provide useful insight regarding diagnosis, treatment, risks and outcomes in MPN patients. In particular, JAK2V617F has been associated with increased risk of thrombotic events, a leading cause of mortality in MPNs. The aim of this study was to determine if JAK2V617F VAF was associated with clinical outcomes in patients with MPN. JAK2V617F VAF was determined by quantitative PCR (qPCR) in a cohort of 159 newly diagnosed MPN patients, and the association of JAK2V617F VAF and risk of thrombosis was examined in this cohort. We observed a significantly higher JAK2V617F VAF in PV and PMF versus ET. A significant association was observed between JAK2V617F VAF and risk of thrombotic events. When patients were stratified by thrombotic events prior to and post diagnosis, an association with JAK2V617F VAF was only observed with post diagnosis thrombotic events. Of note, these associations were not observed when looking at each MPN subtype in isolation. We have shown that a higher JAK2V617F VAF is associated with thrombotic events post MPN diagnosis. JAK2V617F VAF may therefore provide a valuable prognostic indicator for risk of thrombosis in MPNs.

Increased mRNA expression for serotonin receptor 1B (HTR1B) is associated with thrombosis in BCR::ABL1-negative myeloproliferative neoplasms.

BCR::ABL1-negative myeloproliferative neoplasms (MPNs) are clonal haematopoietic stem cell disorders characterized by specific driver mutations and an increased risk of both macrothrombosis and microthrombosis. Serotonin receptor type 1B (HTR1B) was found to be expressed by various solid tumours, and also primary bone marrow mononuclear cells from myelodysplastic neoplasm and acute myeloid leukaemia patients, representing a potential therapeutic target. In this study we assessed for the first time the expression levels of HTR1B mRNA in the peripheral blood mononuclear cells (PBMC) of 85 newly diagnosed MPN patients, consisting of 28 polycythemia vera, 25 essential thrombocythemia and 32 primary myelofibrosis cases. Levels of HTR1B expression between MPN subtypes and control group were not significantly different. However, at clinical data examination, it was observed that MPN patients with a recent history of major thrombosis and/or signs of impaired microcirculation exhibited significantly higher HTR1B expression levels compared to non-thrombotic MPNs and control group. Moreover, thrombotic MPN patients had significantly higher HTR1B expression than patients with recent thrombosis and absence of MPN diagnostic criteria. These findings suggest that increased levels of HTR1B expression in PBMC might be associated with thrombosis in MPN patients, but larger studies are needed for confirmation, including testing of the receptor protein expression level.

DARS expression in BCR/ABL1-negative myeloproliferative neoplasms and its association with the immune microenvironment

DARS, encoding for aspartyl-tRNA synthetase, is implicated in the pathogenesis of various cancers, including renal cell carcinoma, glioblastoma, colon cancer, and gastric cancer. Its role in BCR/ABL1-negative myeloproliferative neoplasms (MPNs), however, remains unexplored. This study aimed to elucidate the expression of DARS in patients with MPNs (PV 23, ET 19, PMF 16) through immunohistochemical analysis and to examine the profiles of circulating immune cells and cytokines using flow cytometry. Our findings indicate a significant overexpression of DARS in all MPNs subtypes at the protein level compared to controls (P < 0.05). Notably, elevated DARS expression was linked to splenomegaly in MPNs patients. The expression of DARS showed a negative correlation with CD4+ T cells (R = − 0.451, P = 0.0004) and CD4+ T/CD8+ T cell ratio (R = − 0.3758, P = 0.0040), as well as with CD68+ tumor-associated macrophages (R = 0.4037, P = 0.0017). Conversely, it was positively correlated with IL-2 (R = 0.5419, P < 0.001), IL-5 (R = 0.3161, P = 0.0166), IL-6 (R = 0.2992, P = 0.0238), and IFN-γ (R = 0.3873, P = 0.0029). These findings underscore a significant association between DARS expression in MPNs patients and specific clinical characteristics, as well as immune cell composition. Further investigation into the interplay between DARS and the immune microenvironment in MPNs could shed light on the underlying mechanisms of MPNs pathogenesis and immune dysregulation.

Translation, Cultural Adaptation, and Validation into Romanian of the Myeloproliferative Neoplasm Symptom Assessment Form-Total Symptom Score (MPN-SAF TSS or MPN-10) Questionnaire.

Background: Patients with myeloproliferative neoplasms (MPNs) experience a high disease-related symptom burden. A specific instrument to evaluate quality of life (QoL), i.e., the MPN Symptom Assessment Form Total Symptom Score (MPN-SAF TSS; MPN-10), was developed. We conducted the translation, cultural adaptation, and validation into Romanian of the MPN-10. Methods: We translated the MPN-10 and tested its psychometric properties. Results: We recruited 180 MPN patients: 66 polycythemia vera (36.67%), 61 essential thrombocythemia (33.89%), 51 primary and secondary myelofibrosis (SMF) (28.33%), and 2 MPN-unclassifiable (1.11%). The mean TSS was 19.51 ± 16.51 points. Fatigue, inactivity, and concentration problems were the most cumbersome symptoms. We detected scoring differences between MPN subtypes regarding weight loss (p < 0.001), fatigue (p = 0.006), early satiety (p = 0.007), night sweats (p = 0.047), pruritus (p = 0.05), and TSS (p = 0.021). There were strong positive associations between TSS and inactivity, fatigue, and concentration problems, and moderate negative correlations between QoL scores and all MPN-10 items. Cronbach's α internal consistency coefficient was 0.855. The Kaiser-Meyer-Olkin construct validity test result was 0.870 and the Bartlett Sphericity Test was significant (p < 0.001). Symptom scores were loaded into one single factor according to the exploratory factor analysis. Conclusions: The Romanian MPN-10 version displayed excellent psychometric properties and is a reliable instrument for assessing symptom burden and QoL in Romanian MPN patients.

Patient reported outcomes among U.S. Veterans living with myeloproliferative neoplasms.

e18528 Background: Myeloproliferative Neoplasms (MPNs) profoundly affect patients' well-being, necessitating effective symptom relief. Patient-reported outcomes are pivotal for care improvement, aligning with Centers for Medicare and Medicaid Services priorities. This study aims to assess MPN symptom burden among U.S. Veterans, examining risk levels, thrombo-hemorrhagic complications, medication use, information access, care, and service-connected benefits. Methods: Conducted as a cross-sectional survey from December 1, 2023, - January 22, 2024, all Veterans in the MPN Advocacy Education &amp; International database were recruited. Symptom burden was assessed using the validated MPN Symptom Assessment Form Total Symptom Score (MPN SAF TSS), with descriptive statistics and T-tests employed for analysis. Results: Of 61 respondents, 88% were male, 92% Caucasian, 88% served during the Vietnam war. The mean age was 73±7, with MPN diagnosis at 64±9 and a mean duration of 44±10 years from service-connected exposure. Key findings included 56% Polycythemia Vera, 26% Essential Thrombocythemia, 16% Primary Myelofibrosis, 70% JAK2, 10% CALR, 3% MPL, 14% unaware of their mutation status, and 34% unaware of MPN risk status. Seventeen (28%) underwent phlebotomy, 64% used Hydroxyurea, 29% Ruxolitinib, and 3% Interferon. Fatigue and itching were the predominant and severe symptoms (Table), with 43% reporting no symptom improvement. Mean MPN SAF TSS of all respondents was higher than prior study by Emanuel et.al. which adjusted for MPN subtype (36.4 vs. 21.6, p&lt;0.0001). Twelve (20%) reported bleeding and 10% reported venous thrombosis after MPN diagnosis, while 13% reported arterial thrombosis before MPN diagnosis. Twenty-two (36%) are not seeing an MPN specialist and 61% are not seeing a hematologist at their local Veterans Affairs Medical Centers. Twenty-three (38%) did not seek a second opinion while 43% do not have a caregiver. Respondents turn to MPN organizations (75%), physician offices (74%), Google Search (49%), Webinars (41%), and Facebook (23%) for information. Five (8%) received service-connected disability benefits. Conclusions: Veterans with MPNs endure a substantial symptom burden, surpassing previous studies, signifying suboptimal management. Limited access to care persists, necessitating enhanced support. However, self-reporting bias and potential limitations in activity assessment must be acknowledged. Efforts to enhance MPN awareness, specialized care, and support mechanisms are imperative for comprehensive management. [Table: see text]

Metformin use and risk of myeloproliferative neoplasms: a Danish population–based case-control study

AbstractPrevious studies have suggested that metformin has beneficial effects beyond its glucose-lowering properties, particularly in terms of its potential as an antineoplastic and cancer-preventive agent. In this study, we aimed to investigate the association between metformin use and the risk of myeloproliferative neoplasms (MPN). We conducted a population-based case-control study using Danish registers. Cases with MPN diagnosed between 2010 and 2018 were identified, and metformin use before the MPN diagnosis was ascertained. We compared metformin use among cases with MPN and an age- and sex-matched control group from the Danish general population to estimate age- and sex-adjusted odds ratios (ORs) and fully adjusted ORs (aORs) for the association between metformin use and risk of MPN. The study population included 3816 cases and 19 080 controls. Overall, 7.0% of cases and 8.2% of controls were categorized as ever-users of metformin, resulting in an OR for MPN of 0.84 (95% confidence interval [CI], 0.73-0.96) and an aOR of 0.70 (95% CI, 0.61-0.81). Long-term metformin use (≥5 years) was more infrequent and comprised 1.1% of cases and 2.0% of controls, resulting in an OR of 0.57 (95% CI, 0.42-0.79) and an aOR of 0.45 (95% CI, 0.33-0.63). A dose-response relationship was observed when cumulative duration of treatment was analyzed, and this was consistent in stratified analyses of sex, age, and MPN subtypes. In conclusion, metformin use was associated with significantly lower odds of an MPN diagnosis, indicating its potential cancer-preventive effect. Given the retrospective design, causality cannot be inferred.

Trisomy 8 Defines a Distinct Subtype of Myeloproliferative Neoplasms Driven by the MYC-Alarmin Axis.

Despite advances in understanding the genetic abnormalities in myeloproliferative neoplasms (MPN) and the development of JAK2 inhibitors, there is an urgent need to devise new treatment strategies, particularly for patients with triple-negative (TN) myelofibrosis (MF) who lack mutations in the JAK2 kinase pathway and have very poor clinical outcomes. Here we report that MYC copy number gain and increased MYC expression frequently occur in TN-MF and that MYC-directed activation of S100A9, an alarmin protein that plays pivotal roles in inflammation and innate immunity, is necessary and sufficient to drive development and progression of MF. Notably, the MYC-S100A9 circuit provokes a complex network of inflammatory signaling that involves numerous hematopoietic cell types in the bone marrow microenvironment. Accordingly, genetic ablation of S100A9 or treatment with small molecules targeting the MYC-S100A9 pathway effectively ameliorates MF phenotypes, highlighting the MYC-alarmin axis as a novel therapeutic vulnerability for this subgroup of MPNs. Significance: This study establishes that MYC expression is increased in TN-MPNs via trisomy 8, that a MYC-S100A9 circuit manifest in these cases is sufficient to provoke myelofibrosis and inflammation in diverse hematopoietic cell types in the BM niche, and that the MYC-S100A9 circuit is targetable in TN-MPNs.

The personal impact of living with a myeloproliferative neoplasm.

The aim of this study is to gain insight into the physical, psychological and social impact of having a myeloproliferative neoplasm (MPN), a rare type of cancer with an often chronic course. An online survey was conducted among 455 Dutch MPN patients (62.7% female, age M 63) to explore the impact of the disease by measuring the MPN symptom burden (MPN-SAF TSS) and quality of life (QoL) (EORTC QLQ-C30) and its subscales within a hierarchical QoL model. We examined differences in MPN symptom burden and QoL in relation to sociodemographic and disease-related factors. Hierarchical regression analysis was used to explain variances in QoL. Most patients (97%) experienced MPN-related health complaints, with a significantly higher MPN symptom burden in women (M 31.50) compared to men (M 24.10). Regarding to fatigue and cognitive functioning MPN patients suffered more compared to a reference group of other cancers. MPN subtype or type of treatment did not show significant differences in MPN symptom burden or QoL. However, experiencing side effects, complications or comorbidities significantly negatively affected MPN symptom burden and QoL. 48.8% of patients reported that MPN affected their ability to work. The explained variance in overall QoL was 58%, most importantly by disease progression, comorbidities, MPN symptom burden and role, emotional and social functioning. This study revealed that having an MPN has a negative impact on several domains of QoL. Symptom assessment and support should be included in the healthcare management of MPN patients.

A Review About the Assessment of the Bleeding and Thrombosis Risk for Patients With Myeloproliferative Neoplasms Scheduled for Surgery.

Myeloproliferative neoplasms (MPNs) present a unique challenge in surgical management due to their inherent predisposition to both bleeding and thrombosis. MPNs are a heterogenous group of acquired clonal conditions. The three classic MPNs are essential thrombocythemia (ET), myelofibrosis (PMF), and polycythemia vera (PV). All subtypes of MPN are associated with both thrombotic and bleeding complications. There are four risk categories for thrombosis in MPN patients: age, thrombosis history, and JAK-2 mutation. They are further classified as very low, low, intermediate, and high risk. The genetic landscape of MPN is fascinating and complex like all myeloid disorders. Bleeding risk can be assessed through leukocytosis, thrombocytosis, acquired von Willebrand syndrome (AVWS), and a previous history of bleeding in a patient.Risk assessment and perioperative management are important aspects of improving the quality of life and preventing complications in surgeries. Preoperative management includes a risk assessment of venous thromboembolism, use of appropriate pharmacological treatment, platelet count control, and correction and cardiovascular risk factors. This review summarizes the assessment of bleeding and thrombosis risk for patients with MPNs scheduled for surgery. Furthermore, this review discusses various tools that can be used to identify MPN patients at risk of thrombosis prior to surgery.

Exploring the Molecular Aspects of Myeloproliferative Neoplasms Associated with Unusual Site Vein Thrombosis: Review of the Literature and Latest Insights.

Myeloproliferative neoplasms (MPNs) are the leading causes of unusual site thrombosis, affecting nearly 40% of individuals with conditions like Budd-Chiari syndrome or portal vein thrombosis. Diagnosing MPNs in these cases is challenging because common indicators, such as spleen enlargement and elevated blood cell counts, can be obscured by portal hypertension or bleeding issues. Recent advancements in diagnostic tools have enhanced the accuracy of MPN diagnosis and classification. While bone marrow biopsies remain significant diagnostic criteria, molecular markers now play a pivotal role in both diagnosis and prognosis assessment. Hence, it is essential to initiate the diagnostic process for splanchnic vein thrombosis with a JAK2 V617F mutation screening, but a comprehensive approach is necessary. A multidisciplinary strategy is vital to accurately determine the specific subtype of MPNs, recommend additional tests, and propose the most effective treatment plan. Establishing specialized care pathways for patients with splanchnic vein thrombosis and underlying MPNs is crucial to tailor management approaches that reduce the risk of hematological outcomes and hepatic complications.

Inherited polygenic effects on common hematological traits influence clonal selection on JAK2V617F and the development of myeloproliferative neoplasms

Myeloproliferative neoplasms (MPNs) are chronic cancers characterized by overproduction of mature blood cells. Their causative somatic mutations, for example, JAK2V617F, are common in the population, yet only a minority of carriers develop MPN. Here we show that the inherited polygenic loci that underlie common hematological traits influence JAK2V617F clonal expansion. We identify polygenic risk scores (PGSs) for monocyte count and plateletcrit as new risk factors for JAK2V617F positivity. PGSs for several hematological traits influenced the risk of different MPN subtypes, with low PGSs for two platelet traits also showing protective effects in JAK2V617F carriers, making them two to three times less likely to have essential thrombocythemia than carriers with high PGSs. We observed that extreme hematological PGSs may contribute to an MPN diagnosis in the absence of somatic driver mutations. Our study showcases how polygenic backgrounds underlying common hematological traits influence both clonal selection on somatic mutations and the subsequent phenotype of cancer.

Prevalence and predictors of pulmonary hypertension in patients with myeloproliferative neoplasms: single-center study

Background Pulmonary hypertension is a major complication of several hematologic disorders including myeloproliferative neoplasms (MPNs). Despite the potential significance of PH to the management and prognosis of MPN, accurate estimates of its prevalence in MPNs and risk factors for the condition are poorly established. Aim To determine the prevalence of pulmonary hypertension in patients with MPNs at Sohag University Hospital and to analyze data of those patients to find predictors of pulmonary hypertension in our patients. Patients and methods A total of 120 patients were diagnosed with MPNs according to the revised 2016 WHO diagnostic criteria who attended the Outpatient Hematology Clinic at Sohag University Hospital in the period from April 2021 to October 2022. Systolic pulmonary artery pressure (SPAP) by using Transthoracic echocardiography (TTE) is determined in all MPNs patients enrolled in our study. Results The overall prevalence of PH in MPN patients was found to be 26.7%. We found a significant relationship between old age, long MPN duration since diagnosis, presence of PMF, anemia, leukocytosis, hyperuricemia, and occurrence of PH. Conclusion The prevalence of PH in patients with MPNs was 26.7%. According to MPNs subtypes, the highest PH prevalence was in PMF patients. Our findings suggest that old age, long MPNs duration since diagnosis, presence of PMF, and hyperuricemia are valuable for predicting PH occurrence in patients with MPNs.

Distinct clinico-molecular arterial and venous thrombosis scores for myeloproliferative neoplasms risk stratification.

Current recommended risk scores to predict thrombotic events associated with myeloproliferative neoplasms (MPN) do not discriminate between arterial and venous thrombosis despite their different physiopathology. To define novel stratification systems, we delineated a comprehensive landscape of MPN associated thrombosis across a large long-term follow-up MPN cohort. Prior arterial thrombosis, age >60 years, cardiovascular risk factors and presence of TET2 or DNMT3A mutations were independently associated with arterial thrombosis in multivariable analysis. ARTS, an ARterial Thrombosis Score, based on these four factors, defined low- (0.37% patients-year) and high-risk (1.19% patients-year) patients. ARTS performance was superior to the two-tiered conventional risk stratification in our training cohort, across all MPN subtypes, as well as in two external validation cohorts. Prior venous thrombosis and presence of a JAK2V617F mutation with a variant allelic frequency ≥50% were independently associated with venous thrombosis. The discrimination potential of VETS, a VEnous Thrombosis Score based on these two factors, was poor, similar to the two-tiered conventional risk stratification. Our study pinpoints arterial and venous thrombosis clinico-molecular differences and proposes an arterial risk score for more accurate patients' stratification. Further improvement of venous risk scores, accounting for additional factors and considering venous thrombosis as a heterogeneous entity is warranted.

Polycythemia Vera: Barriers to and Strategies for Optimal Management.

Polycythemia vera (PV) is a subtype of myeloproliferative neoplasms characterized by impaired quality of life and severe complications. Despite the increasingly in-depth knowledge of this condition, it necessitates a multifaceted management approach to mitigate symptoms and prevent thrombotic and hemorrhagic events, ensuring prolonged survival. The therapeutic landscape has been revolutionized in recent years, where venesection and hydroxycarbamide associated with antiplatelet therapy have a central role and are now accompanied by other drugs, such as interferon and Janus kinase inhibitors. Ongoing research and advancements in targeted therapies hold promise for further enhancing the therapeutic choice for PV management.

A Deep Learning-Based Pathomics Methodology for Quantifying and Characterizing Nucleated Cells in the Bone Marrow Microenvironment

Introduction: The overall cellularity of the bone marrow (BM) microenvironment is a WHO criterion for differentiating BCRABL- myeloproliferative neoplasms (MPNs) like polycythemia vera (PV) and myelofibrosis (MF) [1]. However, manual assessment by hematopathologists is subjective, not scalable, and only records a single global measure, disregarding BM regional differences. Leveraging pathology slide scanners and deep learning (DL) can address these limitations and reveal valuable insights into MPN subtypes and treatment effects. Despite its potential, computational pathology (Pathomics) in this area remains understudied [2-3]. Methods: After institutional review from Stony Brook University and Weill Cornell Medicine (WCM), we identified 54 patients (pts) with a manually confirmed PV diagnosis (dx) at WCM (17% newly dx'ed, 61% treated, 17% disease progression). The dataset includes hematoxylin &amp; eosin (H&amp;E) stained whole slide images (WSIs), labs, molecular studies, and pathology report elements (reference standard for biopsy core size and cellularity). Numeric cellularity measures the ratio of hematopoietic cells to fat cells and excludes inevaluable trabecular bone. Normal cellularity ranges from (100-pt age [y])±20% and PV is characterized by hypercellularity. We evaluated three DL algorithms for nuclear segmentation: 1) Our U-Net model trained on 5B nuclei from 10 cancer types [4], 2) HistoCartography (HC): a graph neural network trained on 235K nuclei (PanNuke/MoNuSac datasets) [5], and 3) CellPose (CP): a gradient-driven U-Net model trained on 1K nuclei of varied modalities [6]. A color/morphology-based segmenter identified fat cells and evaluable BM regions, excluding vessels/tissue tears. Cellular characteristics (shape, texture, color) were computed and clustered by k-nearest neighbors (kNN) and k-means to explore differences between PV stages. A tissue-based approach for cellularity estimation derived the proportion of non-fat tissue to evaluable regions. A cell-based approach derived the proportions of nucleated cell and fat cell areas. The study calculated mean differences between biopsy core size estimates, assessed correlation (r) and intra-class correlation (ICC) for numeric cellularity estimates, and Cohen's kappa (κ) for categorical cellularity estimates. Results: Measured and estimated biopsy core sizes differed by an acceptable margin (3±4 mm). Tissue-based cellularity estimator results: r=0.2, ICC=0.0, κ=0.2. Cell-based cellularity estimator result: our approach (r=0.7, ICC=0.6, κ=0.7), HC (r=0.7, ICC=0.5, κ=0.6), and CP (r=0.2, ICC=0.2, κ=0.7). Cell size clusters aligned with known distributions in PV, with the prominent Int1 class (5.1-9μm) likely containing erythrocytes and myeloid progenitors. Overall and cell-sized cellularity heatmaps visualize regional differences (Figure 1). K-means clustering separated new and progressing PV (Figure 2). Discussion: DL and classical image analysis were employed to segment nucleated cells, fat cells, and other evaluable regions of hematopoietic tissue. Extracting shape, texture, and color features from segmented nuclei enhances BM microenvironment characterization. This pilot study resulted in a practical and interpretable Pathomics pipeline and downstream analytics as a proof of concept for hypothesis-driven research. Future directions involve applying these insights to a large cohort of diagnosis WSIs from other MPN subtypes and correlating findings with other tools, like immune cell detection models.

Successful Management of D816V Mutated c-KIT Systemic Mastocytosis with Chronic Imatinib: A Case Report

First characterized in France in 1936, systemic mastocytosis (SM) is a rare disorder by an excessive proliferation and accumulation of abnormal mast cells in extracutaneous tissues (Nettleship et al., 1869). The release of large quantities of vasoactive mediators by these cells results in allergic- and anaphylactic-type reactions in one or more organ systems. Given the similarity of symptoms between allergic reactions and SM and the rarity of the condition, SM is often missed and misdiagnosed (Schwaab et al., 2020). Once considered a subtype of myeloproliferative neoplasms, mastocytosis was reclassified in the 2016 revision to the World Health Organization (WHO) due to advances in understanding of the disorder (Arber, et al., 2016). Presented below is a case of a severe systemic mastocytosis with an initially positive D816V mutated c-kit successfully managed with long-term imatinib use alongside intermittent bone marrow biopsies (BMB) to monitor what is now an indolent systemic mastocytosis. 69 year-old female with a past medical history of paroxysmal atrial fibrilation, hashimoto's thyroiditis, metabolic syndrome, and gastro-esophageal reflux (GERD) presents to clinic for routine management of what was previously severe systemic mastocytosis (SSM) and now is indolent systemic mastocytosis with the help of imatinib. To help control her ISM her medications include a daily histamine-1 blocker (ranitidine), a daily histamine-2 blocker (diphenhydramine), and a twice-daily histamine-1 antagonist (cetirizine). Originally evaluated in 2005 with a bone marrow biopsy (BMB) and found to have a Asp816Val (D816V) c-kit mutation, she was placed on imatinib with significant clinical improvement. This seemed to support that the D816V mutation may cause constitutional activation of the c-kit transmembrane protein and thus sensitivity to imatinib therapy. This despite the FDA labeling for imatinib as only to be used in SM patients without the common D816V mutation or for whom the mutational status is unknown (Reference ID: 4957210, 2001). Interestingly, after being placed on imatinib her repeat bone marrow biopsies in 2011 and 2015 indicated that the D816V mutation was no longer present in her CD117 positive mast cells but continued to benefit from imatinib therapy. This finding is suggestive of two unique mast cell populations as imatinib continued to help maintain a low tryptase level and decrease the severity of reactions. When reactions did occur, she treated them at one point with a cocktail of an histamine-2 blocker (ranitidine), histamine-1 blocker (diphenhydramine), and a mast cell stabilizer (cromolyn) and without requiring the use of epinephrine. As a part of her longitudinal observation, she received an additional BMB in 2022 that revealed a minimal residual monocytosis that persisted with less than 10% nucleated cells, an age-appropriate trilinear hematopoiesis, decreased iron stores without ringed sideroblast formation. Flow cytometry was indicative of a typical female karyotype with the population of aberrant mast cells that is consistent with the diagnosis of ISM. Importantly, no abnormalities were detected in the FLT3, IDH1, IDH2, NPM1 genes which, if abnormal, could obfuscate the apparent efficacy of imatinib treatment in the management of SM. Given the dearth of mastocytosis in the BMB and the absence of ongoing end-organ damage, the patient meets the definition of ISM. After the most recent BMB the patient altered her daily regimen due to gastric upset and found she could take of her histamine-1 blocker (diphenhydramine) once daily along with a twice daily regimen of her imatinib (split into two smaller daily doses), histamine-1 antagonist (cetirizine), her histamine-2 antagonist (famotidine), and citalopram (selective serotonin reuptake inhibitor or SSRI) with improved symptomatic control to this day. This case serves to illustrate the diagnostic and therapeutic challenges associated with SM, as well as the potential therapeutic use of imatinib in patients with D816V outside of its original FDA indication. Further characterization of the effect of imatinib on D816V mutated SM may warrant the consideration of expanding the FDA indications for imatinib as its role and efficacy outside of solely D816V negative c-kit MS.

The Utility of a Coagulation Laboratory Profile in Addition to Clinical Risk Factors for Thrombosis Risk Evaluation in Patients with Myeloproliferative Neoplasms: A Pilot Study

Background: Patients (pts) with myeloproliferative neoplasms (MPN) have a significantly increased risk of arterial and venous thrombotic events (TEs), while some pts may paradoxically suffer from bleeding manifestations. Thrombosis risk evaluation in polycythemia vera (PV) and essential thrombocythemia (ET) has important implications for patient management, however, the currently used risk stratification models are extremely limited. Methods: We designed a pilot study to assess the utility of an expanded coagulation laboratory profile in addition to other clinical variables for thrombosis and bleeding risk evaluation in a heterogenous MPN patient population. Our laboratory profile included PT, aPTT, fibrinogen, D-dimer, von-Willebrand Factor (vWF) antigen and activity testing, platelet (PLT) aggregation studies and thrombin generation assay (TGA, ST-Genesia, Stago, France), along with CBC and comprehensive biochemistry studies. Blood was drawn from pts visiting the hematology clinics or day care at Hadassah Medical Center. Clinical and demographic data were collected retrospectively from electronic medical records. The study was approved by the local IRB committee. Results:74 pts were included in the study; the mean age was 59.7 years and 51% were women. Sixty percent had a diagnosis of PV, 20% had ET, 17.6% had myelofibrosis (MF) and 2.7% had MPN unclassifiable. 78% of the pts harbored a JAK2V617F mutation, 8% and 2.7% harbored CALR and MPL mutations, respectively, and 11% were negative to all three MPN driver mutations. 63% of the pts had at least one cardiovascular risk factor, most commonly hypertension (HTN) and dyslipidemia. Most of the pts were treated with antiplatelet agents (62%), anticoagulants (16%) and/or cytoreductive agents (58%). Of 74 pts, 31% had a history of TEs (20% arterial, 11% venous), 22% reported suffering from microvascular symptoms, and 20% had experienced bleeding. The high rate of bleeding manifestations in our study population suggests an underestimation of the bleeding burden in MPN, as proposed by our group previously [Rottenstreich, Eur J Intern Med 2017]. Other patient characteristics were overall comparable to MPN patient populations in other studies (Table 1). In multivariate analyses, history of TEs was significantly associated with older age, type 2 diabetes mellitus (DM2), HTN and dyslipidemia, but not with gender, MPN subtype, driver mutation status or smoking. Pts with history of TEs had significantly lower hematocrit and prolonged PT/INR, associated with a higher rate of treatment with cytoreduction and anticoagulation, respectively, in these pts. Interestingly, pts with TEs had significantly higher vWF antigen levels (p=0.007), which were independent of age, blood counts and treatments, a finding not previously reported in MPN. No association was observed between TEs and PLT or WBC counts, D-dimer levels and vWF activity (Table 2), however, vWF activity was decreased in pts with higher PLT counts. TGA was of key interest in this study. In 62 pts not receiving anticoagulation, we observed an association between prolonged lag time (suggesting decreased coagulability) and older age, male gender and (near significant trend) JAK2V617 negative status, but did not find a correlation to TEs, blood counts and other variables. PLT aggregation indices were not associated with TEs, bleeding history or blood counts. In multivariate logistic regression analyses, history of DM2 and HTN, and vWF antigen levels, showed the strongest association to TEs, which was independent of age, blood counts and all other variables. A risk-model incorporating these variables showed a sensitivity of 77% and specificity of 92% for history of TEs in our study population. Conclusions: Despite being limited by a small and heterogenous study population and retrospective data collection; these findings support the utility of a coagulation laboratory profile in the evaluation of thrombosis and bleeding risk in MPN. The potential of TGA in risk evaluation in MPN is intriguing but merits further research. The multivariate risk-model incorporating history of DM2 and HTN along with vWF antigen levels merits validation in larger studies. Based on this pilot, a prospective study following over 250 pts with PV and ET (excluding overt MF) was initiated. This study will include NGS mutational profiling and JAK2V617F allele burden testing and omit the use of PLT aggregation studies.

Cardiovascular Outcomes Among Myeloproliferative Neoplasm Patients with or without Agent Orange Exposure Using a Large Veteran Database

Cardiovascular Disease (CVD) and its complications contribute significantly to the morbidity and mortality of individuals with Myeloproliferative Neoplasms (MPNs), apart from the increased incidence of untoward thrombosis and bleeding. The prevalence of CVD, such as Heart Failure (HF) and Pulmonary Hypertension (PH) in individuals with MPNs and its association with exposure to Agent Orange (AO), a dioxin-containing carcinogen, remain undefined. These findings will be essential in risk stratification and multidisciplinary management of MPNs to prevent progression of CVD. Utilizing the Veterans Affairs Informatics and Computing Infrastructure (VINCI) database, a case-control study was performed from January 1, 2006 - January 26, 2023 to characterize the association of CVD in Veterans with MPNs, arterial thrombosis (AT), venous thrombosis (VT), bleeding, and AO exposure. ICD-9 and -10 codes identified Veterans with MPN, AT, VT, bleeding, CV risk factors, such as hypertension (HTN), hyperlipidemia (HLD), diabetes (DM), smoking, and CVD sequelae, such as HF and PH. Age-, sex-, and race-matched controls (1:1) were selected from VINCI, who are Veterans without MPNs from Illinois, the state most representative of the U.S. population. Composite CVD outcome includes any Veteran with either HTN, HLD, DM, HF, or PH. Exposure to AO was self-reported and verified through Veterans' duration and location of service. Qualitative data were compared by chi-square tests. Among 93,269 Veterans with MPN, there were higher rates of composite CVD, CV risk factors, HF, PH, AT, VT, and bleeding (Figure 1) compared to matched controls. All MPN subtypes, Essential Thrombocythemia (ET), Polycythemia Vera (PV), and Primary Myelofibrosis (PMF) were each associated with increased prevalence of HTN, HLD, DM, HF, and PH (p&amp;lt;0.0001). Among Veterans with MPNs with AT, VT, or bleeding, there was a significant association with each CV risk factors, HF, and PH (p&amp;lt;.001). Among Veterans with MPN with AO exposure, there were higher rates of composite CVD, CV risk factors, HF, PH, AT, and bleeding (Figure 1) when compared to those without AO exposure. Among matched controls with AO, there were higher rates of HLD and AT compared to matched controls without AO. Veterans born before 1960 with AO exposure had younger mean age at MPN diagnosis compared to those without AO, 66 ± 7 (SD) years vs. 69 ± 10 (SD) years, p&amp;lt;0.0001, respectively. Among the 24,385 (26.1%) Veterans with MPNs with HF vs. 1,749 (1.9%) Veterans with MPNs with PH: 46.1% vs. 38.9% had ET; 48.8% vs. 56% had PV; and 5.1% vs. 5.1% had PMF; 92.7% vs. 91.2% were born before 1960; 96.8% vs. 95.3% were male; 75.8% vs. 72.1% White; 14.7% vs. 19.3% Black; 7.5% vs. 6.9% unknown race, respectively. Among MPNs with PH, there is a higher rate of pulmonary embolism (4.87% vs. 1.1%, p&amp;lt;0.0001) when compared to matched controls. There is decreased overall survival among Veterans with MPNs with HF and PH (Figure 2) and Veterans with MPNs compared to matched controls. In summary, the CV risk factors, HF and PH, are increased in Veterans with MPNs and are associated with increased risk for AT, VT, and bleeding complications. Not only is AO exposure associated with development of MPN, as previously reported (J Clin Oncology 2023 41:16_suppl, 7011), but AO exposure appears to increase CVD complications among Veterans with MPNs. Further studies evaluating contributions of driver mutation and CHIPs to cardiac and vascular remodeling are needed and will elucidate the implications of AO and its imputed toxic effects on DNA to the accelerated development of MPNs and its CVD complications. Future targeted therapies and strategies should be developed to mitigate the consequences of progressive CVD, and prevention of AT, VT, and bleeding complications in individuals with MPNs.

Signaling Pathway Gene Set Enrichment Analysis in Whole Exome Sequencing Data from Discordant Identical Twins Unveils Relevant Pathways As a Clue to Understand Somatic Mutation Acquisition and Clonal Evolution in Myeloproliferative Neoplasms

Although the role of classic driver mutations (i.e., JAK2, MPL, and CALR) in myeloproliferative neoplasm (MPN) development is well characterized, additional somatic mutations likely modulate the phenotype. Recurrent somatic mutations, such as those in TET2, DNMT3A, ASXL1 genes are commonly identified in MPN using targeted myeloid panels, however identification of somatic mutations using an unbiased approach may unveil novel pathways involved in MPN pathogenesis. We performed whole exome sequencing of blood samples of three sets of discordant identical twins in which one twin has MPN and the other twin is unaffected. The twin sets included all MPN subtypes (1 polycythemia vera, 1 essential thrombocythemia, 1 primary myelofibrosis). We also performed whole exome sequencing on additional family members, including the mother and father of the PV twin and the brother of the ET twin. Sequencing information from the discordant identical twins is a unique opportunity to identify somatic mutations in the affected twin that may contribute to MPN pathogenesis, and also to potentially identify clonal hematopoiesis in the unaffected twin. We mapped all the identified genetic variants to the affected genes, and we filtered all the genes mapped to those variants depending on whether they were present either in the MPN-affected twin or in the unaffected counterpart for further gene set enrichment pathway analysis. The gene set enriched signaling pathways from identified genetic variants for each condition-exclusive (either MPN or unaffected twin) allows us to identify and prioritize key pathways in MPN development in terms of somatic mutations acquisition. Furthermore, besides performing the analysis within each single MPN set, we analyzed the gene sets resulting from combining the different families allowing us not only to identify pathways at each single MPN type level but also among the different MPN diseases (PV, ET, and MF). We divided all the enrichment analyses into coding and silent variants to obtain a higher degree of pathway refinement. In single discordant twin sets we identified several pathways that may play an important role within MPN pathogenesis such as defective mismatch DNA repair, facilitated mitotic progression, extracellular matrix degradation, MAPK signaling, FGFR signaling, O-glycosylation of proteins, collagen metabolism and laminin interactions, neutrophil degranulation, interleukin-33 signaling, Rho GTPases and cGMP regulation, and nuclear pore complex (NPC) assembly. Interestingly, only when we analyze all the MPN-types together all the statistically significant pathways resulting from coding variants correspond only to DNA repair, specifically defective mismatch repair (PMS2, MLH1, MutSα, MutSβ, and TP53). Our findings reveal that DNA repair genes might occupy a central role within MPN acquisition and development, whereas at the same time each single MPN clinical entity might possess its own distinct mechanisms differentiating each one from the others.