Myeloid Hyperplasia Research Articles

Chronic Myelomonocytic Leukemia and Atypical Chronic Myeloid Leukemia: A National Analysis

BackgroundMyelodysplastic/myeloproliferative overlap syndromes (MDS/MPN) are rare blood cancers characterized by concomitant myeloid hyperplasia and dysplasia. These heterogenous disorders include chronic myelomonocytic leukemia (CMML) and atypical chronic myeloid leukemia (aCML). MethodsUsing two large national cancer databases to examine a total of 15,704 CMML and 702 aCML patients, we report the largest study to date on the incidence, survival and demographic characteristics of CMML and aCML in the United States. ResultsOverall age-adjusted incidence of CMML and aCML was 0.63 per 100,000 Americans per year and 0.03 per 100,000 per year, respectively. CMML incidence in the U.S. was noted to rise steadily in the years between 2001 and 2019. Median patient age was 75 and 72 years for CMML and aCML, and the majority of CMML and aCML patients were male (62.9% and 62.0%) and White (90.1% and 86.3%). Median OS was 17.4 months for CMML, and 15.2 months for aCML. Multivariate Cox regression demonstrated features associated with reduced survival, including increasing age, comorbidities, Medicaid insurance status, and low-income residential zip code, highlighting survival disparities in underinsured and socioeconomically disadvantaged patients. In CMML, Black race was associated with inferior survival, while female sex, management at an academic center, and later calendar-year of diagnosis were associated with improved OS. ConclusionThese findings underscore the need to better understand the biological basis for such differences in survival and reflect the importance of access to specialized care for patients with these rare disorders.

Hemangiosarcoma Cells Promote Conserved Host-Derived Hematopoietic Expansion.

Hemangiosarcoma and angiosarcoma are soft-tissue sarcomas of blood vessel-forming cells in dogs and humans, respectively. These vasoformative sarcomas are aggressive and highly metastatic, with disorganized, irregular blood-filled vascular spaces. Our objective was to define molecular programs which support the niche that enables progression of canine hemangiosarcoma and human angiosarcoma. Dog-in-mouse hemangiosarcoma xenografts recapitulated the vasoformative and highly angiogenic morphology and molecular characteristics of primary tumors. Blood vessels in the tumors were complex and disorganized, and they were lined by both donor and host cells. In a series of xenografts, we observed that the transplanted hemangiosarcoma cells created exuberant myeloid hyperplasia and gave rise to lymphoproliferative tumors of mouse origin. Our functional analyses indicate that hemangiosarcoma cells generate a microenvironment that supports expansion and differentiation of hematopoietic progenitor populations. Furthermore, gene expression profiling data revealed hemangiosarcoma cells expressed a repertoire of hematopoietic cytokines capable of regulating the surrounding stromal cells. We conclude that canine hemangiosarcomas, and possibly human angiosarcomas, maintain molecular properties that provide hematopoietic support and facilitate stromal reactions, suggesting their potential involvement in promoting the growth of hematopoietic tumors.

Targeting TNF/IL-17/MAPK pathway in hE2A-PBX1 leukemia: effects of OUL35, KJ-Pyr-9, and CID44216842.

t(1;19)(q23;p13) is one of the most common translocation genes in childhood acute lymphoblastic leukemia (ALL) and is also present in acute myeloid leukemia (AML) and mixed-phenotype acute leukemia (MPAL). This translocation results in the formation of the oncogenic E2A-PBX1 fusion protein, which contains a trans-activating domain from E2A and a DNA-binding homologous domain from PBX1. Despite its clear oncogenic potential, the pathogenesis of E2A-PBX1 fusion protein is not fully understood (especially in leukemias other than ALL), and effective targeted clinical therapies have not been developed. To address this, we established a stable and heritable zebrafish line expressing human E2A-PBX1 (hE2A-PBX1) for high-throughput drug screening. Blood phenotype analysis showed that hE2A-PBX1 expression induced myeloid hyperplasia by increasing myeloid differentiation propensity of hematopoietic stem cells (HSPC) and myeloid proliferation in larvae, and progressed to AML in adults. Mechanistic studies revealed that hE2A-PBX1 activated the TNF/IL-17/MAPK signaling pathway in blood cells and induced myeloid hyperplasia by upregulating the expression of runx1. Interestingly, through high-throughput drug screening, three small molecules targeting the TNF/IL-17/MAPK signaling pathway were identified, including OUL35, KJ-Pyr-9, and CID44216842, which not only alleviated the hE2A-PBX1-induced myeloid hyperplasia in zebrafish but also inhibited the growth and oncogenicity of human pre-B ALL cells with E2A-PBX1. Overall, this study provides a novel hE2APBX1 transgenic zebrafish leukemia model and identifies potential targeted therapeutic drugs, which may offer new insights into the treatment of E2A-PBX1 leukemia.

WHIM-синдром: обзор литературы и описание двух собственных клинических наблюдений в одной семье

WHIM syndrome (warts, hypogammaglobulinemia, infections, and myelokathexis) is a rare genetic disease associated with activating germline mutations in the gene encoding chemokine receptor CXCR4. WHIM syndrome is manifested by neutropenia, lymphopenia, infections, and degenerative changes of mature neutrophils with bone marrow myeloid hyperplasia (myelokathexis). Some patients show hypogammaglobulinemia, persistent cutaneous, genital, or elsewhere localized warts. There are also cases of congenital heart defects. The present paper extensively analyzes genetic basis, pathophysiology, clinical manifestations, and diagnosis of WHIM syndrome as well as its treatment options. The paper reports two cases in one family.

Successful Therapeutic Leukapheresis in a Patient of Lobar Pneumonia with Hyperleukocytosis: A Case Report

ABSTRACT Hyperleukocytosis is a rare and potentially life-threatening condition that can cause leukostasis and organ dysfunction. This case report describes the case of a 69-year-old female who presented with fever, drowsiness, and consolidation in the left lung lower lobe. Initial investigations showed an increased total leukocyte count (TLC) of 89 × 103/μL, which further increased to 111.9 × 103/μL despite treating with intravenous antibiotics. A bone marrow biopsy revealed myeloid hyperplasia, but the BCR-ABL fusion gene test was negative. Due to symptomatic leukostasis and rapid deterioration of the patient’s condition, therapeutic leukapheresis (TL) was performed. During the first sitting, the device parameters were optimized for maximum platelets and the leukocytes removal. The second sitting of TL was carried out on the next day and the TLC was further reduced to 22.3 × 103/μL. The patient’s general condition and mental status improved significantly and she was taken off the mechanical ventilation and discharged after 7 days with a TLC count of 14.59 × 103/μL. She was regularly followed up for the next 3 months and the counts have remained within normal limits. The case reports highlight that TL may be helpful in similar clinical conditions with symptomatic leukostasis.

First-Hit SETBP1 Mutations Cause a Myeloproliferative Disorder with Bone Marrow Fibrosis

Somatic SETBP1 mutations are found in various myeloid disorders covering both myeloproliferative neoplasms (MPN) and myelodysplastic syndromes (MDS). To characterize the early steps of SETBP1-mediated leukemogenesis, we generated a conditional mouse model expressing SETBP1 G870S mutant in the entire hematopoietic tissue through Cre-mediated recombination driven by the Vav1 promoter. In all mice signs of a hematological disease appeared between 30 and 90 days: longitudinal analysis revealed accumulation of white blood cells (WBC) in virtually all heterozygous SETBP1 G870S mice, with a marked imbalance between the lymphoid and myeloid lineages in favor of the latter, and an increase in mature myeloid cells in absence of circulating blasts or non-segmented myeloid precursors. Kaplan-Meier analysis revealed a dramatic decrease in event-free survival in SETBP1 G870S mice. BM histology showed overt myeloid hyperplasia with fibrosis and no evidence of dysplasia except for the megakaryocytic lineage. Exploration of visceral organs highlighted the presence of severe hepatosplenomegaly with massive infiltration by myeloid elements, disruption of normal tissue architecture, and signs of extramedullary hematopoiesis. Single-cell RNA-sequencing (scRNA) on BM Lin - cells identified Mecom, Setbp1 and Hoxa9 among the top upregulated genes in SETBP1 G870S precursors. Pseudotime analysis of scRNA data revealed the presence of 261 spatially autocorrelated genes. Of them 158 were also differentially expressed in SETBP1 G870Svs control mice. Spi1, encoding for the master regulator of hematopoietic differentiation PU.1, was significantly upregulated in SETBP1 G870S precursors. PU.1 promotes the maturation of bone marrow early precursors towards the granulocytic/monocytic lineages by directly impairing the transcription of Gata2 and Gata1. In line with these data, Gata2 and Gata1 expression was profoundly suppressed in the early myeloid precursors of SETBP1 G870S mice, which associated with the down-modulation of markers of the erythroid lineage, such as the Carbonic Anhydrase 1, in the MEP differentiation branch. Ourmouse model recapitulates many clinical features of primary myelofibrosis (PMF). As up to 10% of PMF are triple-negative for the classical JAK2, CALR, and MPL mutations (TN-PMF), we set out to assess SETBP1 mutations in this context. We analyzed 36 TN-PMF patients by exome sequencing. In 29 we did not find any evidence of somatic mutations; in the remaining 7 (7/36; 19.4%) high VAF SETBP1 degron mutations were identified. In two patients SETBP1 was found as a single somatic variant, while in the others it coexisted with ASXL1, NRAS TET2 SRSF2, RIT1 CBL or CSF3R mutations. Notably, SETBP1 mutations were the only shared alterations among all seven patients. A markedly reduced overall survival was observed for SETBP1 positive patients, with a median survival time of 24 months (median survival not reached at 60 months for SETBP1 negative patients). To dissect the clonal architecture of SETBP1 positive TN-PMF at single-cell resolution, we applied single-cell targeted DNA sequencing on 3 SETBP1-mutated TN-PMF samples using the Tapestri technology, showing that, as opposite to MDS/MPN, in all TN-PMF cases SETBP1 is a very early clonal event. Therefore, our study suggests a clear partition of TN-PMF into two groups, the first one characterized by oncogenic, high VAF SETBP1 mutations accompanied by other oncogenic variants and poor prognosis and the other one characterized by no evidence of an active clonal process or of driver oncogenic events and much lower aggressiveness. SETBP1 positive TN-PMF disorders lie in a gray zone comprised between the MPN and MPN/MDS boundary. In MDS/MPN, SETBP1 mutations are often found as mid or late events. In contrast, we show here that in TN-PMF SETBP1 appears to be one of the earliest hits, therefore highlighting a potentially relevant biological difference occurring in the two subsets. In this context the presence of early SETBP1 mutations seems to promote the occurrence of a myeloid disorder characterized by the triad: leukocytosis without differentiation block or dysplasia, BM fibrosis and progressive splenomegaly, hence recapitulating many clinical features of overt or prefibrotic/early PMF and resembling the SETBP1 mouse model.

Multiplex allele-specific polymerase chain reaction-based prevalence study of canine trapped neutrophil syndrome in Thailand.

Trapped neutrophil syndrome (TNS) is an autosomal recessive genetic disorder found in Border Collies and is characterized by peripheral neutropenia and myeloid hyperplasia of bone marrow. The underlying cause of TNS is associated with a 4-base pair deletion mutation in the vacuolar protein sorting 13 homolog B (VPS13B) gene. In this study, we proposed and validated a novel multiplex allele specific-polymerase chain reaction (MAS-PCR) technique to assess the prevalence of TNS using VPS13B genotypes of Border Collies and Thai Ridgebacks in Thailand. We assessed the prevalence of TNS in 100 Border Collies and 30 Thai Ridgebacks using MAS-PCR-based allelic discrimination technique of the VPS13B gene. We then confirmed the VPS13B genotypes by direct DNA sequencing. A total of 130 samples were successfully genotyped using MAS-PCR assays. Of the two dog breeds examined, the VPS13B mutation was present in Border Collies, whereas Thai Ridgebacks were unaffected by this mutation. In Border Collies, 96% of dogs tested had an intact VPS13B genotype, whereas the remaining individuals had a heterozygous mutation genotype, with prevalence and mutated VPS13B allele frequencies of 4% and 2%, respectively. Using a novel MAS-PCR assay targeting the VPS13B gene, this study demonstrates for the first time that carriers of TNS exist in Border Collies in Thailand. This assay is a reliable and cost-effective tool for diagnosing TNS based on VPS13B genotypes and is suitable for routine clinical practice.

Prenatal diagnosis of Down syndrome combined with transient abnormal myelopoiesis in foetuses with a GATA1 gene variant: two case reports

BackgroundDown syndrome myeloid hyperplasia includes transient abnormal myelopoiesis (TAM) and the myeloid leukemia associated with Down syndrome (ML-DS). The mutation of GATA1 gene is essential in the development of Down syndrome combined with TAM or ML-DS. Some patients with TAM are asymptomatic and may also present with severe manifestations such as hepatosplenomegaly and hydrops.Case presentationWe report two cases of prenatally diagnosed TAM. One case was a rare placental low percentage 21 trisomy mosiacism, resulting in the occurrence of a false negative NIPT. The final diagnosis was made at 36 weeks of gestation when ultrasound revealed significant enlargement of the foetal liver and spleen and an enlarged heart; the foetus eventually died in utero. We detected a placenta with a low percentage (5–8%) of trisomy 21 mosiacism by Copy Number Variation Sequencing (CNV-seq) and Fluorescence in situ hybridization (FISH). In another case, foetal oedema was detected by ultrasound at 31 weeks of gestation. Two foetuses were diagnosed with Down syndrome by chromosomal microarray analysis via umbilical vein puncture and had significantly elevated cord blood leucocyte counts with large numbers of blasts. The GATA1 Sanger sequencing results suggested the presence of a [NM_002049.4(GATA1):c.220G > A (p. Val74Ile)] hemizygous variant and a [NM_002049.4(GATA1):c.49dupC(p. Gln17ProfsTer23)] hemizygous variant of the GATA1 gene in two cases.ConclusionIt seems highly likely that these two identified mutations are the genetic cause of prenatal TAM in foetuses with Down syndrome.

Morphological Spectrum of Bone Marrow Aspirates in Infections: A Clinico-Hematological Analysis

Abstract Context: Bone marrow examination (BME) is an invaluable tool for cases with pyrexia of unknown origin and pancytopenia. However, it is under-utilized for diagnosing infectious etiology and there is a paucity of literature regarding its role in infective pathology. Aims: This study aims to bring to light the role of BME in diagnosing infectious pathology. Settings and Design: A retrospective study was carried out on bone marrow aspirates (BMAs) sent to the hematology department over the past 4 years. Clinical details, peripheral smears and BMA were retrieved from the records and analyzed. Subjects and Methods: Leishman-stained peripheral smears and BMA were studied along with bone marrow biopsy wherever feasible. Results: A total of 52 cases were studied. The most common clinical presentation was fever, clinical finding was splenomegaly and hematological finding was anemia. Based on the morphological findings in combination with clinical history, cases were categorized into–parasitic (26.9%), viral (23.1%), tubercular (11.5%), and nonspecific infections (38.5%). Parasites such as Leishmania donovani, microfilaria, plasmodium falciparum, and vivax were reported in 14/52 (27%) cases. Associated BMA findings were plasmacytosis, eosinophilia, reactive lymphocytosis, or dyserythopoiesis. In 38% (20/52) cases, no specific cause of infection was found in the bone marrow. These patients showed histiocytosis, hemophagocytosis, maturation arrest in myeloid lineage, relative myeloid hyperplasia, dysmyelopoiesis, toxic granulation/vacuolation in myeloid cells, lymphocytosis, increased plasma cells or monocytosis in marrow. Conclusions: Increased histiocytes, hemophagocytosis, dysplastic changes, maturation arrest, relative myeloid hyperplasia or reactive plasmacytosis, lymphocytosis, and monocytosis are BMA features which must alert the pathologist towards an infectious disease process, a knowledge of these changes can help extend the scope of BME beyond hemato-lymphoid malignancies.

Co-occurring mutations in ASXL1, SRSF2, and SETBP1 define a subset of myelodysplastic/ myeloproliferative neoplasm with neutrophilia

Identification of genomic signatures with consistent clinicopathological features in myelodysplastic/myeloproliferative neoplasm (MDS/MPN) is critical for improved diagnosis, elucidation of biology, inclusion in clinical trials, and development of therapies. We describe clinical and pathological features with co-existence of mutations in ASXL1 (missense or nonsense), SRSF2, and SKI homologous region of SETBP1, in 18 patients. Median age was 68 years with a male predominance (83%). Leukocytosis and neutrophilia were common at presentation. Marrow features included hypercellularity, granulocytic hyperplasia with megakaryocytic atypia, while the majority had myeloid hyperplasia and/or erythroid hypoplasia, myeloid dysplasia, and aberrant CD7 expression on blasts. Mutations in growth signaling pathways (RAS or JAK2) were noted at diagnosis or acquired during the disease course in 83% of patients. Two patients progressed upon acquisition of FLT3-TKD (acute myeloid leukemia) or KIT (aggressive systemic mastocytosis) mutations. The prognosis is poor with only two long-term survivors, thus far, who underwent blood or marrow transplantation. We propose that the presence of co-occurring ASXL1, SRSF2, and SETBP1 mutations can be diagnostic of a subtype of MDS/MPN with neutrophilia if clinical and morphological findings align. Our report underscores the association between genotype and phenotype within MDS/MPN and that genomic signatures should guide categorization of these entities.

Priapism on Chronic Myeloid Leukemia with BCR-ABL1 Fusion gene Identified by Molecular Test: A Case Report

The incidence of hematological malignancies has elevated in recent decades. Chronic Myeloid Leukemia (CML) is a hematopoietic stem cell malignant clonal disorder resulting in elevation of erythroid cells and platelets in peripheral blood and clear myeloid hyperplasia in the bone marrow. Priapism is one of a rare clinical manifestation and serious complication in Chronic Myeloid Leukemia (CML). It is cause due to hematological disorder is most likely due to venous obstruction as well as hyperviscosity due to an increased number of circulating leukocytes mature and immature forms. We report a 30-year-old male came to the emergency room, presented with priapism caused by Chronic Myeloid Leukemia (CML) with hyperleukocytosis. The rarity of this case reiterates the importance of thorough morphological, cytogenetic examination along with radiology in diagnosing, treatment and follow up of patients. Starting leukemia therapy to reduce the leukocyte count immediately, can solve the problem in hyperleukocytosis that caused priapism in Chronic Myeloid Leukemia (CML).

The Morphologic Spectrum of Myelokathexis in WHIM Syndrome and Germline CXCR4 Variants: New Insights into Cellular Changes in the Bone Marrow and Peripheral Blood

Introduction WHIM syndrome (WHIMs) is a rare disorder caused by germline CXCR4 mutations, classically characterized by warts, hypogammaglobulinemia, infection, and neutrophil retention in the bone marrow (BM), with distinctive morphology termed myelokathexis. Demonstration of myelokathexis by BM evaluation plays a key role in disease recognition and diagnosis. However, the spectrum of BM morphologic features and correlation with specific CXCR4 mutations are not well described. Moreover, diagnostically useful peripheral blood (PB) smear findings and morphologic changes after CXCR4 antagonist therapy are largely unknown. Methods We conducted a multinational, retrospective review of clinicopathologic features of patients with established WHIMs (n=20) or germline CXCR4 variants considered likely pathogenic or of uncertain significance (n=3), including 14 BM specimens and 10 PB smears. Assessments to date were performed by 2 observers in 6 cases and a single observer in the remainder. BM morphology was compared to other neutropenia disorders (Shwachman-Diamond syndrome, n=5; GATA2 deficiency, n=5; severe congenital neutropenia with ELANE variants, n=4) and patients without hematologic disorders (n=10), together "controls." Correlations and contingency analysis (2-tailed Fisher exact test) were performed using Prism v.9.4.0 (GraphPad Software). Results Median age at WHIMs diagnosis was 18.5 years (range, 0.1-53 years), with female:male ratio of 1.1:1. Recurrent infections, warts, and hypogammaglobulinemia were present in 100%, 53%, and 40% of cases, respectively. BM aspirate smears contained neutrophils with long intranuclear chromatin strands and hypermature condensed chromatin ("myelokathectic neutrophils") in all WHIMs cases, making up 32% to 80% (median 66%) of total neutrophils (Table). In comparison, neutrophils with myelokathectic changes were seen in 0% to 4% of control BMs. Myeloid hyperplasia (myeloid:erythroid ratio >4:1) was present in 50% of WHIMs and 4% of controls (P=.0028); vacuolated neutrophils in 92% of WHIMs (≤29% of total neutrophils) and 17% of controls (P<.0001); apoptotic neutrophils in 58% of WHIMs (1%-4% of total neutrophils) and 8% of controls (P=.0025); and nonparatrabecular neutrophil clusters on core biopsy in 3/3 WHIMs and 5% of controls (P=.0017). Vacuolated eosinophils and myeloid shift toward hypermaturity were present in WHIMs. There was no correlation between percentage of BM myelokathectic neutrophils and degree of CXCR4 truncation. BM in 2 patients with CXCR4 p.S341Y showed myeloid hyperplasia, increased neutrophil apoptosis, and infrequent myelokathectic neutrophils (5%-10%). Neutropenia was present in all untreated WHIMs. Myelokathectic neutrophils were present in PB of all patients with known WHIMs and accounted for 8% to 30% (median, 18%) of total neutrophils. Percentage of PB myelokathectic neutrophils showed an inverse correlation with absolute neutrophil count (ANC) (r=-0.783; P=.0074). Neutrophil apoptosis, neutrophil vacuolization, dysmorphic (hypogranular, hyperlobated, vacuolated, or apoptotic) eosinophils, and/or dysmorphic basophils were present in 80%, 30%, 70%, and 60% of WHIMs cases, respectively (Figure). PB in 1 patient with CXCR4p.D84H showed neutrophil hypolobation without myelokathectic morphology. CXCR4 antagonist therapy with plerixafor (n=4) [Dale et al. Blood. 2011] or mavorixafor (n=1) (interval, 4-12 h) resulted in increased white blood cell (WBC) count and ANC, with a similar proportion of myelokathectic neutrophils as before therapy. Conclusions In WHIMs BM specimens, the proportion of myelokathectic neutrophils is variable and may not be prominent; the classic finding of myeloid hyperplasia is only present in a subset of cases, which may result in underrecognition of the disease and delayed diagnosis. Neutrophil vacuolization and apoptosis, eosinophil vacuolization, and prominent neutrophil clusters may also prompt disease recognition. We establish characteristic PB morphologic features which, in a patient with unexplained neutropenia, may suggest a diagnosis of WHIMs, including myelokathectic neutrophils, neutrophil vacuolization or apoptosis, and dysmorphic eosinophils and basophils. CXCR4 antagonist therapy led to an increase of WBC/ANC, while pretreatment morphologic changes persisted after a short interval. Analysis of additional cases is ongoing. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Vexas Syndrome: A Complex Necessarily Integrated Diagnosis That Can Benefit from the Help of Deep Learning

Introduction At the end of 2020, a novel entity dubbed VEXAS (for vacuoles, E1 enzyme, X-linked, auto-inflammatory, somatic) was described by Beck et al. (NEJM, 2020). This syndrome is defined molecularly by a mutation of the UBA1 gene that codes for the E1 enzyme, a major actor of ubiquitinylation. From a pathophysiological point of view, UBA1 mutations induce the transcription of a deficient enzyme. Affected cells display an altered ubiquitinylation of misfolded proteins and dysregulated autophagy. This leads in fine to an impairment of several pathways of innate immunity, overproduction of a range of pro-inflammatory cytokines and, more generally, systemic inflammation. Clinical manifestations are pleomorphic and patient-dependent, with reported cases of fever, vasculitis, ophthalmic lesions, chondritis, Sweet syndrome or pulmonary interstitial infiltration. From a hematological point of view, thromboses have also been described as well as several cytologic alterations. The latter include cytopenias of any lineage, myeloid hyperplasia with erythroblastic hypoplasia (M/E) and, notably, the presence of vacuoles essentially in granulocytes and erythroblasts precursors. Some cases are associated to myelodysplastic syndrome (MDS). Here, an in-depth morphological examination of VEXAS and non-VEXAS patients was performed, to delineate and quantify several anomalies. These data were then fed to a deep learning algorithm to evaluate the additional help provided by this artificial intelligence (AI) tool in predicting VEXAS syndrome and orienting further analyses towards the identification of the molecular anomaly. Patients, material and methods To identify diagnostic tools for VEXAS syndrome on blood and bone marrow smears, 4 patient cohorts were analyzed :i) patients with a diagnosis of VEXAS syndrome confirmed by genetic analysis (n=6), ii) MDS patients (n=4), iii) patients with an initial clinical suspicion of VEXAS invalidated by molecular analysis (UBA1 WT: n=3) and iv) 4 healthy controls. For each patient, morphological examination of blood and marrow smears was performed independently by two experts in a blind fashion. In order to guide the diagnosis of the syndrome on the blood smears, a deep learning method was trained in a multilabel classification setting to automatically evaluate the percentages of abnormal polymorphonuclears (PMN). The final dataset was therefore composed of PMN from healthy controls (n=5, 519 images) and from VEXAS syndrome (n=4, 938 images) (DI microscope, Cellavision ®). Results Among the 6 patients with VEXAS syndrome, the main clinical manifestations were: asthenia 100%; fever 83.3%; skin involvement 66.7%; thrombosis 66.7%, chondritis 33.3% and interstitial lung disease 33.3%. The most frequent cytopenia was macrocytic anemia in 66.7% of the patients. Upon morphological examination of blood smears, the following abnormalities were statistically associated with the diagnosis of VEXAS : percentage of pseudo-Pelger PMNs (p=0.003), percentage of PMNs with more than 4 nuclear projections (Barr bodies; p=0.0009), percentage of vacuolated PMNs (p=0.01). On bone marrow smears, compared to the MDS and UBA1 WT cohort, patients with VEXAS syndrome had significantly more PMNs with a pseudo-Pelger aspect (p<0.05), more vacuolization of granular (p<0.05) and erythroblastic (p<0.05) precursors. Different deep learning architectures were tested on blood smear images, with the objectives of distinguishing abnormal PMN from others. The trainings were conducted by cross-validation, and then the algorithm was evaluated on new patients. The metrics used to evaluate algorithm performance were precision, recall, F1-score and Hamming loss. These data allowed to establish a standardized computer aided diagnosis tool. Discussion/conclusion The diagnosis of VEXAS syndrome is complex. This analysis identified cytological features on blood smears leading to the development of a deep learning-based diagnosis algorithm. Promising leads are currently investigated to improve the efficiency and robustness of this deep learning algorithm such as the inclusion of other pathologies in order to train a disease agnostic anomaly detection algorithm, or leveraging data from other modalities as prior knowledge.

First Reported Case Series of Combined Treatment with CLAG and Venetoclax for Acute Myeloid Leukemia (AML)

Background: Since the 1970s, the standard of care for patients with AML has been a "7+3" induction regimen consisting of 7 days of cytarabine and 3 days of anthracycline. CLAG has been demonstrated to be effective in patients with newly diagnosed, high-risk AML. Venetoclax is a Bcl2 inhibitor FDA approved to treat AML in elderly patients in combination with hypomethylating agents. Here we describe two patients who received both CLAG and venetoclax to treat R/R AML. Both patients were heavily treated (unsuccessfully), and this novel combination therapy was administered near the end of their treatment. Patient #1 was a 54 year old female with AML. Genetic analysis revealed a FLT3-ITD mutation. Patient was treated with cytarabine 3mg/m2 x 5 and mitoxantrone 60 mg/m2 on day 1 of cycle 1. She went into morphologic remission and then received 2 cycles of HIDAC. Unfortunately, patient relapsed 5 months after starting chemotherapy, thus was not a candidate for allo-SCT. At relapse, the patient had mutations of FLT3-ITD, DNMT3A, NPM1, and WT1. The patient was then treated with CLAG -Idarubicin. Venetoclax was added on day 3 of the cycle, in combination to the CLAG regimen, and she received the venetoclax for 7 days (titrated on day 1 at 50 mg, then 100 mg on day 2, and 200 mg for the next 5 days). She also received midostaurin starting day 6 of the CLAG cycle at a dose of 50 mg and received it for 14 days. Patient #2 was a 71 year old female with AML. Patient Karyotype was 45, XX, add(5)(q11), der(6;7)(p10;q10)[20]. Analysis did not detect a targetable mutation, but an NRAS mutation was detected. The patient began cycle 1 of CLAG-Ida, and on day 23 bone marrow biopsy showed myeloid hyperplasia without increased blasts. The patient then received two more cycles of CLAG-Ida at approximate 1 month intervals. 4 months after diagnosis, the patient underwent an allo-SCT. Patient developed skin lesions and on day +96 biopsy revealed leukemia cutis. Bone marrow biopsy showed no morphologic evidence of AML but abnormal karyotype was noted 45,XX,del(5)(q13q33), dic(6;7)(q11;p11.2)[4]/46,XX[16]. On day +103 the patient began decitabine-venetoclax 200 mg. She received 2 more cycles of decitabine-venetoclax. A right axillary lymph node biopsy was done day on day +238 and showed an abnormal immature population expressing CD34, CD117, MPO, CD4, and CD56. Patient then started on GO 4.5mg and then HIDAC. Patient did not respond to these regiments and received CLAG and Venetoclax starting day +307. She completed this cycle but progressed further. She then received hydrea. This was followed by vyxeos on day+357. Patient did not respond and elected to go to hospice; she died day+381 post SCT. Discussion: To our knowledge, this is the first reported case series of concomitant treatment with CLAG and Venetoclax. During the literature review, we looked at FLAG, which is similar to CLAG but uses a different purine analogue fludarabine instead of cladribine. We also saw that FLAG and venetoclax have been combined to treat AML patients as they transition into SCT. When compared to FLAG, CLAG has been shown to be more effective in treating patients with high risk AML. Although in both cases the patients ultimately succumbed to AML, this was a proof of concept, and we suspect the CLAG venetoclax combination will be more efficacious than FLAG-venetoclax in treating high risk patients. This case series is intended to show that these two therapies can be combined, and we propose a further, more complete investigation into the concomitant use of CLAG and venetoclax to treat patients with AML.

Myelodysplastic Syndrome/Myeloproliferative Neoplasm Overlap Syndromes: A National Analysis of Overall Survival and Demographic Features

Introduction Myelodysplastic/myeloproliferative (MDS/MPN) overlap syndromes (MDS/MPN) are rare disorders characterized by concomitant myeloid hyperplasia and dysplasia. Only recognized as distinct disorders since 2001, this heterogeneous group includes chronic myelomonocytic leukemia (CMML), MDS/MPN with ringed sideroblasts and thrombocytosis (MDS/MPN-RS-T), atypical chronic myeloid leukemia (aCML), juvenile myelomonocytic leukemia (JMML) and MDS/MPN-unclassifiable (MDS/MPN-U). Timely and accurate diagnosis can be difficult outside academic medical centers, and epidemiological data are lacking. This study aims to evaluate national survival outcomes, population and disease characteristics, and socioeconomic and demographic features associated with disparate survival in these rare and complex disorders. Methods Utilizing the National Cancer Database from 2004-2019, MDS/MPN was identified by histology codes for aCML (9876), CMML (9945), MDS/MPN-U (9975), and MDS/MPN-RS-T (9982). Only adult (age >18) patients (pts) were included, excluding JMML. Demographic, socioeconomic, and first line treatment characteristics were identified for all MDS/MPN and by histology. Kaplan-Meier survival analysis was used to compare overall survival (OS), with multivariable Cox regression, adjusted for age, sex, race, ethnicity, Charlson-Deyo comorbidity score, treatment facility type, and insurance to evaluate features associated with disparate survival outcomes. Results Of the 34,245 MDS/MPN pts identified, 42% (N=14,338) had CMML, 24% (N=8,055) had MDS/MPN-RS-T, 2% (N=634) had aCML, and 33% (N=11,318) were diagnosed with MDS/MPN-U. The median age at diagnosis was 75 years (yrs) (interquartile range 66-82). MDS/MPN-RS-T and aCML pts presented at the oldest and youngest ages (median 76 yrs vs 71 yrs). The majority of pts were male (58%), and White (88%; 8% Black and 2% Asian). Notably, aCML and MDS/MPN-U pts were more likely to be a race other than White (both 15% non-White), compared to CMML (11%) and MDS/MPN-RS-T (9%), p<0.001. While molecular data is limited in the NCDB, of the 10,583 cases with JAK2 testing reported, 49% had a JAK2 mutation. This was greatest in the MDS/MPN-U group, at 65% of pts, followed by 39% of MDS/MPN-RS-T pts, and15% for both aCML and CMML pts. Approximately half of MDS/MPN pts (51%) received disease directed therapy at diagnosis, with 22% managed with active surveillance, and 27% not receiving treatment, for reasons including early death, contraindication, or hospice care. aCML pts were the most likely to be placed on disease directed therapy (78%), followed by CMML (56%), MDS/MPN-U (55%), and MDS/MPN-RS-T (28%). The median OS for all MDS/MPN was 31.4 months (mos) (95% confidence interval [CI] 30.7-32.1), with a 1-, 5-, and 10-year OS of 71%, 33%, and 16%, respectively. MDS/MPN-RS-T pts had a median OS of 52.5 mos (95% CI 50.6-54.3), similar to MDS/MPN-U with a median OS of 48.2 mos (95% CI 46.1-50.4). CMML and aCML had significantly worse survival with a median OS of 16.8 mos (95% CI 16.3-17.4) and 14.5 mos (95% CI 12.5-16.5), respectively, p<0.001. On multivariable Cox regression, MDS/MPN-RS-T and MDS/MPN-U retained a significant survival advantage over CMML (Hazard ratio [HR] 0.45 [95% CI 0.43-0.46], p<0.001 and HR 0.51 [95% CI 0.49-0.53], p<0.001, respectively). However, aCML had a significantly worse survival, with HR 1.27 (95% CI 1.15-1.40, p<0.001). Age, increasing comorbidity burden, and Black race were associated with reduced survival outcomes, while female sex, private insurance, and treatment at academic centers was associated with significantly improved OS. Conclusion We present the largest study to date on the real-world survival and demographic characteristics of MDS/MPN. Compared to prior data, which was generally limited in size, we found a lower median OS than previously reported, though MDS/MPN-U had a greater OS than prior estimates. We further identified a higher portion of MDS/MPN pts identified as MDS/MPN-U than previously estimated, and significantly higher rates of JAK2 mutations than previously reported. This study also adds clarity to ages of diagnosis, sex ratios, and racial and ethnic characteristics. We also note survival disparities in Black pts, and those who are underinsured or treated at non-academic centers, indicating that access to treatment remains a significant barrier to survival in these pts. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Peripheral smear reckons the nature of IgM cold agglutinins: A case‐based study asserting the role of thermal amplitude and clonality

Peripheral smear reckons the nature of IgM cold agglutinins: A case‐based study asserting the role of thermal amplitude and clonality

A Rare PTPN11 Mutation in a Patient with Juvenile Myelomonocytic Leukemia: A Case Report

Background: Juvenile myelomonocytic leukemia (JMML) is a rare childhood disease characterized by hepatosplenomegaly, monocytosis, anemia, increased white blood cell count, thrombocytopenia, skin infiltration, and elevated fetal hemoglobin. Mutation in specific genes, including KRAS, NRAS, PTPN11, and NF1, can lead to the development of JMML. Case Presentation: A two-year-old boy with a history of inguinal abscess at the age of 12 months and surgery due to infectious lymphadenitis was referred to the hospital. His parents also reported a history of oral candidiasis, recurrent otitis media, and lymphadenopathy in the patient. The physical examination showed splenomegaly, macular rash, lymphadenopathy in the neck region, and rashes in the inguinal region and on the hands and feet. Laboratory and flow cytometry data showed lymphocytosis, low hemoglobin, thrombocytopenia, monocytosis, eosinophilia, and a shift to the left in the peripheral blood. The bone marrow aspiration showed a cellular marrow with myeloid hyperplasia. Whole-exome sequencing revealed a rare heterozygous ENST00000351677.2:c.1508G&gt;C, p.Gly503Ala variant in PTPN11. The patient was diagnosed with JMML but, unfortunately, passed away. Conclusion: We report a rare heterozygous mutation in the PTPN11 gene in a two-year-old boy diagnosed with JMML. This uncommon mutation should be considered in the mutational screening protocol of JMML. Management of JMML with RAS pathway targeted therapy may also have promising results and needs further investigations.

Spontaneous scapular region hematoma extending upto anterior chest wall in a patient of chronic myeloid leukaemia: an unusual site presentation

Spontaneous haemorrhage and thrombosis should always raise the suspicion of CML as one of the differential as it is associated with qualitative platelet dysfunction. We here report a rare case of CML with spontaneous hematoma at an unusual site. A 26 years male presented with complaints of sudden onset non traumatic spontaneous swelling over the left scapular region and progressively extending up-to the left anterior chest wall which was associated with dull aching moderate intensity pain. Swelling was warm, soft in consistency and tender. He also had hepato-splenomegaly. General blood picture showed marked leucocytosis with myeloid hyperplasia. CT scan thorax reported a large hypodense lesion in the scapular and anterior chest wall region which on diagnostic aspiration revealed blood thereby confirming the diagnosis of spontaneous hematoma with CML. Imatinib therapy was started and hematoma was managed conservatively with antibiotics. The swelling resolved spontaneously without any surgical intervention.

Circulating CD34+ cells of primary myelofibrosis patients contribute to myeloid-dominant hematopoiesis and bone marrow fibrosis in immunodeficient mice.

Primary myelofibrosis (PMF) is a clonal stem cell disorder characterized by myeloid dominant hematopoiesis and dysregulated proliferation of fibroblasts in the bone marrow. However, how these aberrant myeloid cells and fibroblasts are produced remains unclear. In this study, we examined in vivo engraftment kinetics of PMF patient-derived CD34+ cells in immunecompromised NOD/SCID/IL2rgKO (NSG) mice. Engrafted human cells were analyzed with flow cytometry, and proliferation of fibroblastic cells and bone marrow fibrosis were assessed with the histo-pathological examination. Transplantation of PMF patient-derived circulating CD34+ fractions into NSG newborns recapitulates clinical features of human PMF. Engraftment of human CD45+ leukocytes resulted in anemia and myeloid hyperplasia accompanied by bone marrow fibrosis by six months post-transplantation. Fibrotic bone marrow contained CD45-vimentin+ cells of both human and mouse origin, suggesting that circulating malignant CD34+ subsets contribute to myelofibrotic changes in PMF through direct and indirect mechanisms. A patient-derived xenotransplantation (PDX) model of PMF allows in vivo examination of disease onset and propagation originating from immature CD34+ cells and will support the investigation of pathogenesis and development of therapeutic modalities for the disorder.

AML-214: Case Report: A Rare Case Presentation and Detection of Acute Promyelocytic Leukemia

Acute promyelocytic leukemia (APL) is an uncommon subtype of acute myeloid leukemia (AML) characterized by the translocation t (15;17) (q22; q21) with a favorable prognosis upon immediate recognition and treatment. It is very rare when the insertion into PML could not be demonstrated by initial FISH and demonstrated by molecular analysis. We present a case of a 44-year-old female who presented in October 2020 to the Emergency Room with severe fatigue and blood tests showing Hb=6 g/dL, thrombocytopenia, and leukopenia (WBC 2.5 x10⁹/L). The patient was admitted to the hospital, where bone marrow biopsy and flow cytometry showed a negative result. Initial diagnosis was mainly related to a viral infection, when the patient had blood transfusion. On December 15, 2020, the patient presented with leukocytosis (55.31 x10⁹/L), anemia (Hb 7.6 g/dL), thrombocytopenia PLT (67 x10⁹/L), and swelling in her right knee. Repeated flow cytometry showed signs of dysplasia with abnormal myeloid lineage. Total body scans showed findings mostly related to underlying myeloproliferative or lymphoproliferative disease with hepatosplenomegaly, as well as retroperitoneal soft tissue thickening encasing the abdominal aorta, with aortic wall mostly related to associated para-neuroplastic aortitis. Stills disease was diagnosed. Initiation of steroids 1 mg/kg was given, leading to amelioration of the patient and less of blood transfusion. After 1 week, a repeated cytogenetics and molecular analysis showed PML-RARA transcripts and t (15;17), negative for acute leukemia, hypercellular aspirate showing left-shifted myeloid hyperplasia, and <1% blasts. She was diagnosed with promyelocytic leukemia and was started on treatment with ATRA + arsenic per guidelines and is now in complete remission.