Search Results (70)

Background/Objectives: Juvenile myelomonocytic leukemia (JMML) is a rare, highly aggressive form of pediatric myelodysplastic/myeloproliferative neoplasm characterized by molecular heterogeneity and constitutive activation of the RAS signaling pathway. This study aimed to characterize the mutational landscape, driver genes, mutational signatures, functional pathways, and therapeutic potential of mutated epigenetic regulator genes in JMML. Methods: Tumor and matched buccal swab samples were collected from 35 JMML patients, and whole-exome sequencing was performed. Somatic variants were called with GATK-Mutect2 and annotated with ANNOVAR. maftools and OncodriveCLUST were used for mutational profiling, co-occurrence analysis, driver gene identification, and protein domain mapping. Drug–gene interactions were explored using DGIdb, mutational signatures for genes were characterized by MutationalPatterns, and functional enrichment analysis was performed by the clusterProfiler package. Results: A total of 28 variants were detected in epigenetic regulator genes, with missense mutations being the most common class of variants and a C>T nucleotide substitution pattern being the most frequent. EP300, SETD2, and DNMT3B were the genes most frequently altered, with 8.57% of cases each, followed by ASXL1, BCORL1, ATRX, KMT2A, and TET2 (5.71% each). Driver gene analysis identified ASXL1 as the top candidate driver gene, followed by BCORL1, EP300, and SETD2. Functional enrichment analysis revealed a high number of genes involved in chromatin organization, histone modification, transcriptional regulation, and oncogenic signaling pathways. The mutational signatures identified were SBS5-like, which are dominated by C>T and T>C transitions, indicating endogenous mutational processes. Analysis of drug–gene interactions revealed KMT2A, EP300, and ATRX as the most interconnected and potentially actionable therapeutic targets. Conclusions: This study provides a comprehensive characterization of epigenetic regulator gene alterations in JMML and highlights the importance of epigenetic dysregulation in disease pathogenesis. Full article

Background/Objectives: Fluorescence in situ hybridization (FISH) is an established component of the diagnostic and prognostic workup of hematologic malignancies. However, most evidence supporting FISH interpretation originates from treatment-naïve diagnostic cohorts, providing limited insight into how cytogenetic findings evolve during longitudinal disease monitoring and therapeutic exposure. We retrospectively analyzed 1357 consecutive FISH assays performed in 1092 patients with hematologic malignancies between 2019 and 2023. Methods: Multivariable logistic regression identified predictors of aberration detection, while longitudinal analyses explored temporal changes in cytogenetic profiles among serially monitored patients. Observed abnormality rates were compared with published diagnostic benchmarks. Results: The cohort had a median age of 68 years and was predominantly composed of patients with myelodysplastic/myeloproliferative neoplasms (27.2%), multiple myeloma (26.9%), and chronic lymphocytic leukemia (21.6%). Overall, 29.2% of informative assays revealed cytogenetic abnormalities. Clinical indication was the strongest predictor of aberration detection. Compared with diagnostic samples, follow-up specimens showed significantly lower odds of abnormal findings (adjusted OR 0.48, 95% CI 0.36–0.64; p < 0.001), whereas relapse samples demonstrated higher odds (adjusted OR 1.82, 95% CI 1.28–2.59; p = 0.001). Increasing age was independently associated with abnormal FISH results (adjusted OR per decade 1.24, 95% CI 1.12–1.38; p < 0.001). Several canonical abnormalities occurred less frequently than expected from historical diagnostic series, including BCR::ABL1 within the chronic myeloid leukemia testing pathway and del(13q14) in both chronic lymphocytic leukemia and multiple myeloma. In contrast, del(17p), involving the TP53 locus, remained consistently represented across disease categories and was enriched in chronic lymphocytic leukemia relative to published diagnostic cohorts. Conclusions: Longitudinal assessment revealed increasing cytogenetic heterogeneity over time, with common lesions progressively declining and rare or complex abnormalities expanding from 16.3% to 36.7% of the observed cytogenetic landscape. The cytogenetic patterns detected by FISH in contemporary hematology practice are strongly associated with clinical context, disease stage, and treatment history; however, the cross-sectional and retrospective design precludes inference of causal influence. The divergence between real-world positivity rates and historical diagnostic benchmarks highlights the need for context-specific interpretation of cytogenetic findings. The persistence of TP53 loss across disease boundaries and the progressive diversification of aberrations over time underscore the dynamic nature of clonal evolution in hematologic malignancies. Full article

Background: Juvenile myelomonocytic leukemia (JMML) is a rare RAS/MAPK-driven pediatric myelodysplastic/myeloproliferative neoplasm. The phenotype and survival relevance of NRAS/KRAS alterations remain difficult to interpret because of co-mutations, evolving sequencing, incomplete germline confirmation, and post-diagnostic HSCT. Methods: We retrospectively reviewed 34 children with JMML and classifiable NRAS and/or KRAS alterations diagnosed between November 2010 and September 2022. Patients were classified as NRAS-only, KRAS-only, or NRAS/KRAS co-mutated. Direct comparisons used single-subtype cases. OS was analyzed in evaluable patients, with HSCT modeled as a time-dependent covariate. Results: The cohort included 20 NRAS-only, 12 KRAS-only, and 2 NRAS/KRAS co-mutated cases. KRAS-only cases had higher monocyte percentage (27.2% vs. 16.7%; p = 0.023) and lymphocyte percentage (48.1% vs. 39.0%; p = 0.018), whereas absolute monocyte count was comparable (4.0 vs. 4.0 × 10⁹/L; p = 0.930). PTPN11 was the most frequent non-RAS co-mutation (7/34) and occurred in both NRAS/KRAS co-mutated cases. Methylation status was available in 11 patients and analyzed descriptively. NRAS/KRAS subtype did not significantly stratify OS (HR for KRAS-only vs. NRAS-only, 0.55; 95% CI, 0.18–1.62; p = 0.276). HbF did not significantly stratify OS, while diagnostic total hemoglobin showed only an exploratory univariable association. Exact HSCT dates were retrieved for all 11 transplanted patients; time-dependent Cox analysis showed a significant association between HSCT and better OS (HR, 0.11; 95% CI, 0.02–0.56; p = 0.007). Conclusions: NRAS/KRAS status defined a limited diagnosis-time phenotype but did not independently stratify survival. JMML survival analyses should integrate co-mutations, germline and testing-era limitations, HbF/hemoglobin risk context, and time-dependent HSCT handling. Full article

Myeloid malignancies encompass a heterogeneous group of haematological disorders, primarily including myelodysplastic syndromes (MDSs) and myeloproliferative neoplasms (MPNs). MDS is characterised by defective myeloid cell maturation, while MPNs involve the pathological overproduction of myeloid lineage cells. In the absence of timely diagnosis and effective clinical intervention, both entities carry a substantial risk of progression to acute myeloid leukaemia (AML). Although allogeneic haematopoietic stem cell transplantation remains the only potentially curative therapy, its application is frequently constrained by patient-related factors such as advanced age and comorbid conditions. While currently, hypomethylating agent therapy (azacitidine and decitabine) is mainly used in high-risk MDS patients, and ruxolitinib is primarily used in symptomatic primary myelofibrosis (PMF-MPN), their clinical efficacy remains suboptimal. More recently, focus has turned toward the role of the tumour microenvironment (TME) in disease pathogenesis and whether therapeutically targeting the TME, either alone or in combination with conventional therapy, could present a new treatment option. Emerging evidence underscores the significant influence of TME components, particularly macrophages and T cells, in modulating immune responses and shaping the leukaemic niche to either facilitate or hinder malignant progression. In response, a new generation of immune checkpoint inhibitors are being developed to target the TME, including PD-1/CTLA-4 blockers, macrophage-directed agents including CD47 inhibitors, and T cell-targeting checkpoint inhibitors such as TIM-1 and LAG-3. This review will describe the functional role of key TME constituents in the progression of myeloid malignancies and explore the current landscape and future potential of advanced cellular and molecular immunotherapies in the treatment of these disorders. Full article

Background and Objectives: Peripheral cytopenia occurs in approximately 2% of the population; however, in up to 0.9%, no cause is identified by conventional tests. Next-Generation Sequencing (NGS) detects somatic variants consistent with clonal hematopoiesis (CH). We aimed to determine the prevalence of CH and pre-myelodysplastic syndrome (pre-MDS) using a 51-gene panel with histopathological assessment. Materials and Methods: Bone marrow samples from 96 consecutive patients evaluated for cytopenia were retrospectively analyzed for genetic alterations. Results: In the overall cohort (n = 96), the median follow-up was 8.1 months (range, 1–20). A total of 37 (39%) out of 96 patients were diagnosed with idiopathic cytopenia of undetermined significance (ICUS), 9 (9%) with clonal cytopenia of undetermined significance (CCUS), 9 (9%) with clonal cytopenia and monocytosis of undetermined significance (CCMUS), 34 (36%) with myelodysplastic syndrome (MDS), and 7 (7%) with myelodysplastic/myeloproliferative neoplasm (MDS/MPN). Among 41 patients in whom no cytogenetic abnormalities were detected by fluorescence in situ hybridization (FISH), somatic variants were identified by NGS. In CCUS, 88% of patients had a single variant, most commonly ASXL1 (44%), followed by TET2 (22%). In CCMUS, ASXL1 and DNMT3A (each 25%) were the most frequent variants. The mean variant allele frequency (VAF) was higher in MDS (33.4%) than in CCUS/CCMUS (13.6%). In MDS patients aged 60 years and older, a higher number of variants were found compared to patients aged less than 60 years (p = 0.028). RUNX1 variants (n = 8) were associated with leukopenia (p = 0.012). Patients with SRSF2 variants (n = 4) had significantly poorer progression-free survival (p = 0.001). EZH2 and SETBP1 variants were associated with inferior overall survival (p = 0.04 and p = 0.019, respectively). In MDS patients (n = 34), thrombocytopenia (plt < 100.000) was associated with shorter PFS (p = 0.005). Conclusions: Given that pre-MDS conditions are considered predictors of hematologic malignancies, conventional diagnostic workup may be insufficient to accurately identify these entities, whereas NGS provides significant additional diagnostic value. Full article

Background: BRAF is a core component of the RAS–MAPK signaling pathway and an established oncogenic driver in several solid tumors and selected hematologic malignancies. In myeloid neoplasms, BRAF mutations are rare, and their prevalence, molecular context, and clinical significance remain incompletely defined. Available evidence is scattered across heterogeneous reports involving acute myeloid leukemia, myelodysplastic syndromes, myeloproliferative neoplasms, and overlap myelodysplastic/myeloproliferative neoplasms, with variable descriptions of mutation subtypes, co-mutational profiles, cytogenetic associations, therapeutic approaches, and clinical outcomes. To address these gaps, this review synthesizes data from the published literature up to 2025, summarizing the distribution, genetic landscape, and clinical impact of molecularly confirmed BRAF mutations across the spectrum of myeloid neoplasms. Results: Across published cohorts, BRAF mutations occurred in less than 1% of unselected myeloid neoplasms, with enrichment in chronic myelomonocytic leukemia and therapy-related or secondary acute myeloid leukemia. Both V600E and non-V600E variants were observed, typically within a complex genomic background involving ASXL1, TET2, DNMT3A, SRSF2, and RAS-pathway mutations. Acute myeloid leukemia cases showed poor prognosis, with median overall survival measured in months, whereas myelodysplastic syndromes and chronic myelomonocytic leukemia demonstrated relatively longer survival. Targeted MAPK inhibition produced hematologic responses in selected cases but rarely resulted in durable molecular clearance. Conclusions: BRAF mutations in myeloid neoplasms are rare, heterogeneous, and usually represent secondary events in clonal evolution. Although mutation clearance appears prognostically relevant, current targeted approaches provide limited durability, underscoring the need for prospective studies in this setting. Full article

Background: KMT2A partial tandem duplication (PTD) occurs in approximately 5–10% of acute myeloid leukemia (AML) cases and is associated with poor prognosis. While its cytogenetic and molecular features are well described, the immunophenotypic characteristics of AML with KMT2A-PTD remain incompletely defined. Methods: We identified 47 cases of AML with KMT2A-PTD by optical genome mapping. All cases underwent flow cytometric immunophenotypic analysis and next-generation sequencing using an 81-gene panel. Results: The cohort included 32 men and 15 women with a median age of 67 years (range, 19–87). Thirty-eight cases were de novo AML, and nine were secondary to myelodysplastic syndrome and/or myeloproliferative neoplasm. Most cases (93%) demonstrated a normal or non-complex karyotype. The most frequent mutations involved FLT3-ITD (47%), DNMT3A (43%), and RUNX1 (23%). Thirty-one cases (66%) were granulocytic, while 16 (34%) showed granulocytic and/or monocytic differentiation. Blasts uniformly expressed HLA-DR and frequently expressed CD117 (91%) and CD34 (79%). Increased expression of CD123 (74%) and CD117 (43%) and decreased expression of HLA-DR (74%) and CD38 (69%) were common. Aberrant CD25 expression was observed in 51% of cases. Increased CD123 and aberrant CD25 expression were significantly associated with FLT3-ITD mutations (both p < 0.0001) but not with other recurrent mutations. There was no correlation between FLT3-ITD mutation and expression levels of CD117, CD38 or HLA-DR (all p > 0.05). Conclusions: AML with KMT2A-PTD shows distinctive immunophenotypic features with increased CD123 and aberrant CD25 expression, both associated with FLT3-ITD. These markers may have diagnostic and therapeutic relevance in this AML subtype. Full article

Myeloproliferative neoplasms (MPNs) are a heterogeneous group of diseases originating from hematopoietic stem cell transformation, characterized by the clonal proliferation of hematopoietic progenitors. A specific subset includes myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase (TK) gene fusions, particularly involving PDGFR A or B, which are sensitive to TK inhibitor treatment. We report a case of a 21-year-old patient with a myeloproliferative/myelodysplastic neoplasm, presenting with hyperleukocytosis, anemia, thrombocytopenia, and elevated LDH. The peripheral blood smear showed hypogranular neutrophils, eosinophils, basophils, and myeloid precursors. The absence of BCR::ABL1 and mutations in JAK2, CALR, and MPL excluded common MPNs. Cytogenetic analysis revealed a rearrangement between chromosomes 5 and 14. FISH analysis confirmed an inverted insertion from chromosome 5 to chromosome 14, involving the PDGFRB gene. WGS and RNAseq identified a fusion between PDGFRB and CCDC88C, causing the constitutive activation of PDGFRB. The fusion gene was confirmed by sequencing. This allowed for targeted therapy with a tyrosine kinase inhibitor (TKI), leading to molecular remission monitored by RT-qPCR. This case highlights how a multidisciplinary approach can identify atypical transcripts in MPN, guiding targeted therapy with TK inhibitors, thus resulting in effective treatment and molecular remission. Full article

Colony-stimulating factor 3 (CSF3), additionally called granulocyte colony-stimulating factor (G-CSF), is the major cytokine regulating neutrophil production and also impacting their function. The actions of this cytokine are mediated through its unique receptor, the colony-stimulating factor 3 receptor (CSF3R). Several classes of pathogenic mutations in the CSF3R gene have been identified that have distinct biological properties and clinical impacts. This review provides an overview of CSF3R, the various pathogenic CSF3R mutations/variants and their biological effects. It also details the diseases to which they contribute, notably including chronic neutrophilic leukemia (CNL) and other myeloproliferative neoplasms (MPNs), myelodysplastic neoplasms (MDS), combined MDS/MPN disorders such as atypical chronic myeloid leukemia (aCML) and chronic myelomonocytic leukemia (CMML), as well as acute myeloid leukemia (AML) and lymphoid malignancies. Full article

Background: Myelodysplastic/myeloproliferative neoplasms (MDS/MPNs) are rare hematopoietic disorders that exhibit overlapping pathological and molecular features of both MDS and MPN. This study aimed to investigate the mutational profiles and prognostic implications of MDS/MPN subtypes in Korean patients. Methods: We retrospectively reviewed 53 patients with MDS/MPN who underwent bone marrow examination and next-generation sequencing panel testing. Overall survival was analyzed with 3-year censoring. The cohort included chronic myelomonocytic leukemia (CMML; N = 30); MDS/MPN with neutrophilia (N = 6); MDS/MPN with SF3B1 mutation and thrombocytosis (N = 4); and MDS/MPN, not otherwise specified (MDS/MPN-NOS; N = 13). Results: The most frequently mutated gene was ASXL1 (52.8%), followed by TET2 (39.6%) and U2AF1 (18.9%), in total MDS/MPN. U2AF1 mutations were particularly frequent in myelodysplastic CMML (33.3%) and MDS/MPN-NOS (30.8%). In CMML, ASXL1 and TET2 mutations were associated with a trend toward better prognosis compared with wild-type (HR 0.21, p = 0.052; HR 0.25, p = 0.057, respectively), while U2AF1 mutations were associated with adverse prognosis in univariate analysis with borderline significance (HR 12.20, p = 0.050). Clinical/Molecular CMML-Specific Prognostic Scoring System and Mayo Molecular Model showed stepwise survival patterns across risk groups without statistical significance. In univariate analysis, transfusion dependency was associated with poor prognosis (HR 7.78, p = 0.013). Conclusions: This study provides the first single-institution analysis in Korean patients with MDS/MPN and identified U2AF1 mutations as a potentially significant prognostic factor, enhancing the molecular understanding of MDS/MPN. Full article

Chromosomal BCR-ABL1 fusions are the defining molecular lesions of chronic myeloid leukemia (CML) and Philadelphia-positive acute lymphoblastic leukemia, and are rarely observed in acute myeloid leukemia. Their detection have transformed treatment paradigms by enabling effective use of specific tyrosine kinase inhibitors (TKIs). Although many BCR-ABL1 rearrangements are identified by standard cytogenetics, a clinically relevant subset is cryptic and can escape detection. High-depth RNA sequencing assays have improved our capacity to detect expressed fusion transcripts. Here, we introduce two myeloid cases in which cryptic BCR-ABL1 rearrangements and precise breakpoints detection required an integrated molecular approach: we describe the initial diagnostic pitfalls, detail the downstream therapeutic and prognostic implications and offer practical recommendations for integrating targeted sequencing and cytogenetics into routine practice. In the first case, a patient initially diagnosed with a myelodysplastic/myeloproliferative neoplasm was reclassified as CML following the discovery of a cryptic e13a2 BCR-ABL1 rearrangement, enabling effective TKI treatment. In the second case, a previously undetected BCR-ABL1 fusion was identified in a relapsed AML patient, along with additional molecular lesions, underscoring the aggressive nature of the disease. Our findings support a systematic, multimodal screening strategy in patients with atypical presentations to ensure the timely detection of clinically actionable fusion events. Full article

Background: Genetic alterations in CSF3R, typically associated with chronic neutrophilic leukemia (CNL) and atypical chronic myeloid leukemia (aCML), rarely occur in other myeloid neoplasms. Methods: This study characterized the clinical, morphologic, cytogenetic, and molecular features of 13 patients with non-CNL non-aCML myeloid neoplasms with CSF3R alterations. Patients (median age, 77 years) were categorized into groups with a myelodysplastic/myeloproliferative neoplasm (MDS/MPN) (n = 5), acute leukemia (n = 4), and other myeloid neoplasms (n = 4) based on the WHO 2022 and ICC criteria. Results: The CSF3R p.Thr618Ile mutation was most frequent (11/13), with additional pathogenic variants including p.Gln743Ter and frameshift mutations affecting the cytoplasmic tail. Variant allele frequencies (VAFs) ranged from 2% to 49%, with the highest median VAF in the MDS/MPN group. Co-mutations varied by subtype; MDS/MPN, NOS, and CMML cases frequently harbored mutations in epigenetic regulators (ASXL1, TET2) and splicing factors (SF3B1, SRSF2, ZRSR2), while acute leukemia cases showed alterations in JAK3, STAT3, and NRAS. Survival analysis revealed distinct patterns across the three diagnostic groups, with MDS/MPN having the poorest prognosis. Conclusion: This study expands the recognized spectrum of CSF3R-related myeloid neoplasms and highlights the clinical and molecular heterogeneity associated with these mutations, emphasizing the need for comprehensive molecular profiling and the potential for targeted therapies. Full article

Background: Sanger sequencing remains the gold standard for characterizing genetic variants in short DNA fragments (<700 bp). However, the increasing demand for short TATs and high sensitivities in variant detection, particularly in oncohematology, is driving the need for more efficient methods. Next-generation sequencing (NGS) has improved sensitivity and allows for the simultaneous analysis of multiple genes, but it is still costly and time-consuming. Consequently, Sanger sequencing continues to be widely used. In this study, we have compared Sanger sequencing with Oxford Nanopore technology (ONT), which offers enhanced sensitivity and faster sequencing, delivering diagnostic results within 24 h. Methods: This study involves 164 samples (for a total of 174 analyzed regions of interest) previously characterized using either Sanger sequencing or a next-generation sequencing (NGS) panel, categorized by their genetic alterations. Validation was conducted on 15 genes crucial for the diagnosis, prognosis, or identification of drug resistance in myeloproliferative neoplasms (MPN), myelodysplastic syndromes (MDS), acute myeloid leukemia (AML), and chronic myeloid leukemia (CML). The primary objective was to assess whether MinION could identify the same variants previously detected in these patients. Results and Conclusions: With a 99.43% concordance observed in our comparison, our results support the implementation of MinION technology in routine variant detection in MPN, MDS, AML, and CML cases due to its significant advantages over Sanger sequencing. Full article

Background and Objective: The primary objective of this study is to evaluate the added value of optical genome mapping (OGM) when integrated into the standard cytogenetic workup (SCGW) for hematological malignancies. Methods: The study cohort comprised 519 cases with different types of hematological malignancies. OGM and SCGW (including G-banded karyotyping and fluorescence in situ hybridization) were performed on blood and/or bone marrow. The analytical sensitivity of OGM, defined as the detection of all additional cytogenomic aberrations, and its clinical utility, referring to aberrations with diagnostic, prognostic, or therapeutic significance, were assessed. Results: OGM led to increased analytical sensitivity and clinical utility in 58% and 15% of the cases, respectively. The clinical utility varied across different malignancies, with the highest utility in T-lymphoblast leukemia (52%), followed by mixed phenotype acute leukemia (43%), B-lymphoblastic leukemia (37%), other B-cell lymphomas (22%), mature T-cell leukemia/lymphoma (20%), chronic lymphocytic leukemia (14%), acute myeloid leukemia (13%), multiple myeloma (13%), mantle cell lymphoma (8%), myelodysplastic/myeloproliferative neoplasms (6%), myelodysplastic syndrome (5%), and myeloproliferative neoplasms (0%). Conclusion: Compared to SCGW, OGM detects additional cytogenomic aberrations in approximately 58% of cases. OGM provides clinical utility at varying rates across different types of hematological malignancies. Given these differences, strategic triaging can help maximize the clinical value of OGM by focusing on diseases where it offers the most significant benefit. Full article

Myeloid sarcoma (MS), or extramedullary acute myeloid leukaemia tumour (eAML), is a rare hematopoietic neoplasm. Recognised as a distinct entity within acute myeloid leukaemia (AML), MS presents significant diagnostic challenges due to its rarity, clinical heterogeneity, and variable immunophenotypic and genetic characteristics. The mechanisms by which leukaemic stem cells (LSCs) migrate to form solid tumours in extramedullary (EM) sites remain unclear. MS can occur de novo, precede AML, and manifest alongside AML relapse. It can also develop with myelodysplastic syndromes (MDSs) or myeloproliferative neoplasms (MPNs). MS frequently presents in organs such as the skin, lymph nodes, gastrointestinal (GI) tract, and central nervous system (CNS), often resulting in diverse clinical manifestations. Diagnosis relies on a comprehensive approach, including tissue biopsy, bone marrow (BM) evaluation, and advanced imaging modalities. Accurate diagnosis is crucial for risk stratification and treatment selection. Prognosis is influenced by several factors: MS’s anatomical location, timing of MS diagnosis, genetic profile, and possible treatment. This review emphasises the need for comprehensive diagnostic methods to better define individual MS characteristics and prognosis. It explores the role of novel targeted therapies in improving patient outcomes and further highlights the critical need for future multicentre data collection to optimise diagnostic and therapeutic approaches. Full article