IL-4Rα signaling promotes barrier-altering oncostatin M and IL-6 production in aspirin-exacerbated respiratory disease

Abstract

Background: Aspirin-exacerbated respiratory disease (AERD) is a severe disease involving dysregulated type 2 inflammation. However, the role other inflammatory pathways play in AERD is poorly understood.

Objective: We sought to broadly define the inflammatory milieu of the upper respiratory tract in AERD and to determine the effects of IL-4Rα inhibition on mediators of nasal inflammation.

Methods: Twenty-two AERD patients treated with dupilumab for 3 months were followed over 3 visits and compared to 10 healthy controls. Nasal fluid was assessed for 45 cytokines and chemokines using Olink Target 48. Blood neutrophils and cultured human mast cells, monocytes/macrophages, and nasal fibroblasts were assessed for response to IL-4/13 stimulation in vitro.

Results: Of the nasal fluid cytokines measured, nearly one third were higher in AERD patients compared to healthy controls, including IL-6 and the IL-6 family-related cytokine oncostatin M (OSM), both of which correlated with nasal albumin levels, a marker of epithelial barrier dysregulation. Dupilumab significantly decreased many nasal mediators, including OSM and IL-6. IL-4 stimulation induced OSM production from mast cells and macrophages but not from neutrophils, and OSM and IL-13 stimulation induced IL-6 production from nasal fibroblasts.

Conclusion: In addition to type 2 inflammation, innate and IL-6-related cytokines are also elevated in the respiratory tract in AERD. Both OSM and IL-6 are locally produced in nasal polyps and likely promote pathology by negatively affecting epithelial barrier function. IL-4Rα blockade, although seemingly directed at type 2 inflammation, also decreases mediators of innate inflammation and epithelial dysregulation, which may contribute to dupilumab's therapeutic efficacy in AERD.

Keywords: AERD; Aspirin-exacerbated respiratory disease; dupilumab; interleukin 13; interleukin 4; interleukin 4Rα; interleukin 6; mast cells; nasal polyp; oncostatin M.

Figure 1.. Selected nasal fluid cytokine levels.

Nasal fluid levels for (A) type 2 inflammatory cytokines IL-4, IL-13, IL-33, and TSLP (B) innate cytokines OSM and IL-6, and (C) colony-stimulating factors CSF1, CSF2, and CSF3 are shown for healthy control patients (n=10) and patients with AERD before (n=23) and after treatment with dupilumab for 1 month (n=22) and 3 months (n=19). Data shown as median and interquartile range, with all p values <0.10 displayed.

Figure 2.. Sinus tissue single-cell RNA sequencing.

Uniform manifold approximation and projection (UMAP) plots of surgically excised sinus tissue cells displaying the cell clusters identified within the sinus tissues (top left) and their expression of OSM, OSMR, CSF1, CSF2, CSF3, and IL6 transcript.

Figure 3.. Effects of IL-4 on human cord blood mast cells.

(A) Normalized counts of OSM, CSF1, and CSF2 transcript levels from bulk RNA sequencing of human cord blood mast cells stimulated with or without IL-4 for 72 hours (n=2 donors). (B) OSM protein in the supernatant of human cord blood mast cells stimulated with or without IgE crosslinking, IL-33, and/or IL-4 (n=4 donors).

Figure 4.. Monocyte and neutrophil OSM production.

OSM protein levels in the supernatant of (A) human monocyte-derived macrophages (n=3–7) cultured with or without CSF1 (M-CSF) or CSF2 (GM-CSF), ±24-hour stimulation with IL-4 or IL-13; and (B) human neutrophils (n=4) cultured with or without CSF2 (GM-CSF), CSF3 (G-CSF), or IL-4 for 20 hours.

Figure 5.. Nasal fibroblast production of IL-6.

Human primary nasal fibroblasts (n=3) were cultured with or without IL-13 and were stimulated with OSM at two concentrations for 48 hours. IL-6 levels in the supernatant were measured by ELISA.

Figure 6.

Spearman’s rank correlation coefficient is shown between levels of nasal albumin and nasal OSM or IL-6 at each of three time points for patients with AERD (n=19–23).