DECIPHERING AND TARGETING VEXAS

Overview

VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome (1/4,000 males aged \>50) is a paradigmatic age-onset acquired, severe, autoinflammatory hematological disease due to clonal dominance of hematopoietic cells bearing a somatic mutation in the UBA1 (ubiquitin-activating enzyme 1) gene. VEXAS presents with treatment-refractory systemic myeloid-driven inflammatory manifestations and hematologic abnormalities, hence the poor prognosis. Current available therapies are poorly effective and burdened by debilitating side effects. VEXAS mutations inactivate cytoplasmic UBA1-driven ubiquitination, impairing protein clearance and triggering stress responses. By innovative base-editing (patent PCTIB2024060412), the investigators generated new in vitro and in vivo models that, along with comprehensive phenotyping of VEXAS patients, disclosed progressive inflammatory poisoning of healthy hematopoiesis and resilience to inflammation of UBA1-mutant hematopoietic stem/progenitor cells (HSPC) as key mechanisms of inflammation and clonal dominance. Deciphering these mechanisms might reveal predictive markers, actionable targets, and inform drug repurposing or novel therapies. The investigators hypothesize that cell-intrinsic and -extrinsic mechanisms triggered by altered UBA1 function are targetable drivers linking inflammation and clonal dominance. The research plan is designed to investigate, both in vitro and in BM and peripheral hematopoietic cells from VEXAS patients, the key pathogenic mechanisms sustaining inflammation and clonal dominance in VEXAS syndrome. Patients with VEXAS syndrome, defined by the presence of pathogenic mutations in the UBA1 gene identified in hematopoietic cells isolated from the peripheral blood of individuals with systemic autoinflammatory disease, will be included in the study population. The study will also enroll healthy control subjects, matched as closely as possible for age and sex, with no history of inflammatory or hematologic disorders. The dissection of mechanisms underlying the interplay between inflammation and clonal dominance will be fundamental to identify novel therapeutic strategies against disease progression that would enable the treatment of VEXAS patients at early disease stages, improving prognosis and life expectancy. Moreover, establishing robust genotype-to-phenotype correlations in patients may inform the design of personalized therapeutic options. The obtained results may inform the design of new therapeutic strategies against this cureless disease. Overall, in this observational monocentric study the investigators will use retrospective samples collected from 2020 to 2026 to conduct cytokine profiling and comprehensive omics-based characterization (transcriptomics, proteomics, metabolomics) on peripheral blood-derived monocytes/neutrophils and bone marrow (BM) hematopoietic cells.

Primary Objective. To validate, in patients affected by VEXAS syndrome, the pathogenic mechanisms associated with UBA1-mutant hematopoietic cells, with particular focus on their role in promoting systemic inflammation and clonal dominance. Primary Endpoint. Differences in the production of pro-inflammatory cytokines/chemokines by monocytes isolated from VEXAS patients compared with healthy controls, assessed by multiplex immunoassay. The primary endpoint will be assessed on blood samples collected retrospectively. Secondary Objectives. * To evaluate the role of autophagy and unfolded protein response (UPR) pathways in regulating the inflammatory response in VEXAS monocytes. * To characterize multi-omic profiles (transcriptomics, proteomics, metabolomics) of hematopoietic cells from BM and peripheral blood. * To compare molecular profiles between UBA1-mutant cells and cells from healthy donors. * To investigate genotype-phenotype correlations associated with different UBA1 mutations. * To integrate ex vivo findings with data generated from in vitro models. Secondary Endpoints. * Changes in cytokine production following ex vivo modulation of autophagy and UPR pathways. * Changes in transcriptomic profiles (RNA-seq) following ex vivo modulation of autophagy and UPR pathways. * Proteomic and metabolomic profiles of hematopoietic cells from bone marrow and peripheral blood. * Quantitative and qualitative differences across UBA1 mutation subgroups. * Correlations between molecular data and available clinical features. Secondary endpoints will be assessed on samples collected retrospectively.