Patnaik, Mrinal M.

VEXAS syndrome is a clonal hemato-inflammatory disorder impacting older, predominantly male patients, characterized by systemic inflammation and progressive bone marrow failure. It is caused by somatic mutations in UBA1, an X-linked gene, essential for initiating cellular ubiquitination. Loss-of-function mutations, most commonly M41 missense variants, lead to decreased expression of the cytoplasmic isoform, UBA1b, resulting in accumulation of misfolded proteins and activation of the endoplasmic reticulum stress pathway. Patients classically present with systemic and often refractory inflammation, requiring chronic glucocorticoid treatment and targeted immunosuppressive therapies. A central hallmark of this syndrome is progressive bone marrow failure and associated cytopenia, with a diagnosis of myelodysplastic syndromes (MDS) being reported in approximately 30-50% of patients while only 1% of unselected MDS patients have UBA1 mutations. This review examines current literature on distinct MDS features reported in VEXAS syndrome. We first summarize the defects and alterations in the hematopoietic system from UBA1 mutations, including associated peripheral blood and bone marrow findings. Next, we describe the histological and molecular characteristics of patients with VEXAS syndrome, with and without MDS, and review potential treatment strategies, including responses to hypomethylating agents. Finally, we highlight that VEXAS-associated MDS is a challenging diagnosis but one with unique clinical, laboratory, and morphological features, including prognostic implications.

ABSTRACT Hypomethylating agents (HMA) and allogeneic hematopoietic stem cell transplantation (alloHSCT) have both demonstrated remissions in VEXAS; however, comparative data is lacking. We conducted a multicenter, retrospective analysis of 66 patients diagnosed with VEXAS syndrome treated with HMA ( n = 35) or alloHSCT ( n = 31). Baseline characteristics such as genetics, co‐morbidities, and performance status were balanced between the groups, except older age in the HMA group. Median follow‐up from therapy initiation was 18 months (95% CI: 11–26), and 14 (21%) deaths were reported (alloHSCT n = 3; HMA n = 11). Among all evaluable patients within the alloHSCT cohort, all patients achieved molecular remission, and a substantial proportion of patients discontinued glucocorticoids (58%). In contrast, HMA therapy was associated with lower but meaningful rates of molecular remission (22%) and glucocorticoid discontinuation (6%). In a real‐world setting, HMA therapy was associated with a high discontinuation rate related to toxicity or lack of response. On multivariable analysis adjusted for age and Charlson Comorbidity Index, alloHSCT was associated with improved overall survival (HR = 0.20, 95% CI: 0.05–0.81; p = 0.024). This association remained consistent across multiple ancillary sensitivity analyses, including restriction to transplant‐eligible patients, patients aged ≤ 75 years, 1:1 matching, and propensity score‐based weighted analyses. Although limited by retrospective design, these findings suggest that alloHSCT remains an attractive and potentially curative strategy in selected patients with VEXAS. Prospective validation of these findings is warranted.

Abstract Germline variants in DDX41 are the most frequent genetic predisposition to adult hematologic malignancies. The most common variants are truncating, implicating loss of function in the pathogenesis. However, non-truncating variants account for 30-40% of cases, and their impact on essential DDX41 functions remains unknown. We utilized a genetic complementation assay to assess the functionality of 10 recurrent germline non-truncating variants of DDX41. All variants restored viability to Ddx41 -deficient hematopoietic progenitor cells at exogenous expression levels. In contrast, the hotspot mutant p.R525H, which is somatically acquired at disease onset in >50% of patients, failed to restore viability. CRISPR-based modeling in cell lines and mice revealed heterogeneity: some variants were non-functional at endogenous expression levels whereas others maintained complete functionality, supporting normal cell proliferation and even lifelong hematopoiesis in a homozygous setting. Notably, co-expression of p.R525H with some variants caused impaired hematopoietic progenitor cell viability, indicating a dominant-negative effect of p.R525H. In contrast, other variants, all classified as variants of unknown significance, were unaffected by the presence of p.R525H. A screen of 100 disease-associated variants confirmed that many non-truncating germline variants are susceptible to p.R525H-mediated dominant-negative effects, whereas wild-type DDX41 is not. These findings indicate that DDX41 variant curation is complicated by variable effects on functionality and variant-specific interactions with somatically-acquired DDX41 mutations. The dominant-negative effect of p.R525H provides a mechanistic basis for the conclusion of recent patient cohort analyses that co-occurrence with a somatic hotspot mutation is a reliable indicator of DDX41-driven disease in carriers of non-truncating variants.

Abstract While missense somatic mutations in the serine/arginine rich splicing factor 2 (SRSF2) occur early in the clonal development of myeloid neoplasms, they are uniquely enriched in chronic myelomonocytic leukemia (CMML), an MDS/MPN overlap neoplasm, where their estimated prevalence is ∼50%. Short-read RNA-sequencing experiments have demonstrated that missense alterations of the proline 95 residue hotspot (SRSF2P95) globally dysregulate exon inclusion, though the full transcript structures induced by such alternative splicing events remain uncharacterized. Given the striking enrichment of SRSF2 mutations in CMML, we hypothesized that there exist unique splicing alterations contributing to the biology of this disease, and sought to profile the landscape of alternative transcription induced by SRSF2 mutations in CMML using long-read RNA-Seq. We conducted bulk long-read RNA-Seq on bone marrow mononuclear cells from 5 CMML patients with SRSF2P95H (n=3) and SRSF2P95L (n=2) mutations, with VAFs ranging 20-48%, along with 5 CMML splicing factor wildtype patients. Libraries were prepared with the Pacbio Kinnex full-length RNA kit and sequenced on a Revio SPRQ sequencer. Reads were processed and aligned using the IsoSeq pipeline, and isoform collapsing, quality control, reference comparison, and quantification were performed with IsoQuant and SQANTI3. Differential transcript and gene expression was assessed with DESeq2, while differential transcript usage was assessed with the DRIM-Seq and stageR packages. After long-read collapsing and quality control, we identified 255,505 unique transcript isoforms, of which 14,845 (∼5.8%) were novel to the reference transcriptome annotation (GENCODE v39). We identified 23 transcripts differentially expressed in SRSF2-mutant samples, including 9 novel transcripts, the majority of which (n=7) were upregulated in SRSF2-mutant cases. A higher number of transcripts (n=80) and genes (n=65) demonstrated significant changes in proportional transcript usage. Among transcripts with increased expression and usage in SRSF2-mutant cases, we identified a novel transcript of the gene YBX1, which encodes an RNA- and DNA-binding transcription factor required for cell survival and self-renewal in myeloid neoplasms. This novel transcript, which we termed YBX1ΔEx4, has an in-frame loss of exon 4, and is predicted to encode a full-length protein missing a 30 amino acid region of the cold-shock (DNA-binding) domain. Using short read RNA-Seq data from an additional 29 CMML patients, as well as an independent dataset (GSE251806) comprising 43 blast-phase CMML patients, we confirmed increased exclusion of YBX1 exon 4 in SRSF2-mutant cases, with an even greater loss of exon 4 in blast-phase SRSF2-mutant cases. We hypothesize an important role of this isoform in CMML disease progression and are currently performing functional studies to characterize the effect of YBX1ΔEx4. Citation Format: Nickolas Steinauer, Terra Lasho, Christy Finke, Pankaj Pradeep, Jenna Fernandez, Alejandro Ferrer, Moritz Binder, Abhishek Mangaonkar, Mrinal M. Patnaik, . Long-read RNA-Seq reveals a novel isoform landscape of SRSF2-mutant chronic myelomonocytic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1497.

Abstract Recent updates to monocyte count thresholds recognize oligomonocytic chronic myelomonocytic leukemia (OM-CMML) as an early form of CMML. However, the clinical validity of these changes remains uncertain without incorporating biological and genomic factors. In this study, we analyzed a cohort of 911 patients (249 with OM-CMML, 359 with overt CMML, and 303 with myelodysplastic syndromes) using unsupervised clustering to evaluate the role of genomic determinants in refining CMML diagnosis. Our findings show that CMML molecular signatures (biallelic TET2 mutations or SRSF2-TET2 comutations) are linked to a distinct transcriptome, monocytic bias, classical monocytosis, and a higher risk of progression to overt CMML in OM-CMML cases. We developed a weighted genomic model and diagnostic workflow showing that combining genomic signatures with bone marrow monocyte frequencies in OM-CMML more accurately predicts progression to overt CMML. These findings support integrating genomic determinants and our clinic-ready diagnostic workflow into the CMML diagnostic framework to improve accuracy. Significance: Through comprehensive clinical and genomic profiling of a large patient cohort, alongside immunophenotypic and transcriptional cellular analyses, this study provides evidence that incorporating genomic determinants into the diagnostic criteria for OM-CMML improves diagnostic accuracy and refines the identification of early-stage CMML, thereby preventing misclassification.