Background/Purpose: VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome is a severe adult-onset inflammatory disease caused by somatic mutations of the ubiquitin-like modifier activating enzyme 1 (UBA1) gene. Expansion of UBA1 mutated hematopoietic stem cells in the bone marrow of patients produces skewing towards the myeloid compartment. Neutrophils are enriched at sites of inflammation, yet prior functional studies in VEXAS have not focused on neutrophils. The aim of this study was to examine the transcriptomes and maturation of neutrophils in peripheral blood and marrow from patients with VEXAS.

Methods: We collected paired bone marrow and peripheral blood samples from 26 VEXAS patients with somatic pathogenic variants in UBA1 at p.Met41 for single-cell RNA sequencing (scRNA-seq). Negatively isolated neutrophils were captured using 10X Genomics Chromium 3’ Kits. Libraries were sequenced on the Illumina Novaseq 6000/X plus. Reads were aligned and quantified using Cell Ranger v8.0.0; dimensional reduction, clustering, and gene expression analysis was conducted using R package Seurat v5.2.1. Trajectory analysis and lineage differential expression tests were performed by R packages slingshot v3.20 and tradeSeq v1.23.0. Longitudinal analyses were performed in patients who were treated with hypomethylating agents, where one patient achieved molecular remission (i.e. eradication of the mutant clone).

Results: Canonical mutation types were proportional among the cohort (p.Met41Thr (n=10), p.Met41Leu (n=8), and p.Met41Val (n=8)) with median variant allele fraction 74% (range 24-89%). Circulating neutrophil counts were preserved to elevated in patients with VEXAS, while lymphocyte, monocyte, and natural killer cell counts were typically reduced. Immature granulocytes were commonly detected in peripheral blood and confirmed by scRNA-seq. Upregulation of genes related to neutrophil survival and inflammation were seen in neutrophils from patients with VEXAS. In particular, interferon response genes were highly upregulated in VEXAS neutrophils. Trajectory analysis identified two pathways of neutrophil differentiation in VEXAS: traditional development and an atypical pathway where myeloid-committed stem cells rapidly differentiated into mature neutrophils. Over-expression of genes associated with emergency granulopoiesis and immune metabolism defined the atypical pathway. Mutational burden in blood and bone marrow was eradicated in one patients with VEXAS following treatment with azacytidine. Comparing neutrophils before and after molecular remission revealed with significant reduction in interferon response gene expression, elimination of the atypical trajectory in peripheral blood, and normalization of neutrophil maturity subsets relative to healthy controls in both blood and bone marrow.

Conclusion: Neutrophils are critical effector cells in VEXAS syndrome, with distinct inflammatory transcriptomic profiles that normalize in molecular remission. These results are informative about the heterogeneity of neutrophils in VEXAS syndrome and may lead to discovery of novel myeloid-directed therapeutic targets.

« Back to ACR Convergence 2025

ACR Meeting Abstracts - https://acrabstracts.org/abstract/neutrophil-transcriptomics-and-maturation-pathways-in-vexas-syndrome/