Serum Proteomics Distinguish Subtypes of NMO Spectrum Disorder and MOG Antibody-Associated Disease and Highlight Effects of B-Cell Depletion

Abstract

Background and objectives: AQP4 antibody-positive NMOSD (AQP4-NMOSD), MOG antibody-associated disease (MOGAD), and seronegative NMOSD (SN-NMOSD) are neuroautoimmune conditions that have overlapping clinical manifestations. Yet, important differences exist in these diseases, particularly in B-cell depletion (BCD) efficacy. Yet, the biology driving these differences remains unclear. Our study aims to clarify biological pathways distinguishing these diseases beyond autoantibodies and investigate variable BCD effects through proteomic comparisons.

Methods: In a retrospective study, 1,463 serum proteins were measured in 53 AQP4-NMOSD, 25 MOGAD, 18 SN-NMOSD, and 49 healthy individuals. To identify disease subtype-associated signatures, we examined serum proteins in patients without anti-CD20 B-cell depletion (NoBCD). We then assessed the effect of BCD treatment within each subtype by comparing proteins between BCD-treated and NoBCD-treated patients.

Results: In NoBCD-treated patients, serum profiles distinguished the 3 diseases. AQP4-NMOSD showed elevated type I interferon-induced chemokines (CXCL9 and CXCL10) and TFH chemokine (CXCL13). MOGAD exhibited increased cytotoxic T-cell proteases (granzyme B and granzyme H), while SN-NMOSD displayed elevated Wnt inhibitory factor 1, a marker for nerve injury. Across all subtypes, BCD-treated patients showed reduction of B-cell-associated proteins. In AQP4-NMOSD, BCD led to a decrease in several inflammatory pathways, including IL-17 signaling, cytokine storm, and macrophage activation. By contrast, BCD elevated these pathways in patients with MOGAD. BCD had no effect on these pathways in SN-NMOSD.

Discussion: Proteomic profiles show unique biological pathways that distinguish AQP4-NMOSD, MOGAD, or SN-NMOSD. Furthermore, BCD uniquely affects inflammatory pathways in each disease type, providing an explanation for the disparate therapeutic response in AQP4-NMOSD and MOGAD.

Figure 1. Serum Protein Profiles Distinguish NMOSD Subtypes and MOGAD That Are Not Undergoing B-Cell Depletion From Healthy Controls

Comparison of serum protein profiles between (A) noBCD AQP4-NMOSD and healthy controls, (B) noBCD MOGAD and healthy controls, and (C) noBCD SN-NMOSD and healthy controls. Significant proteins were determined using unpaired t tests with multiple comparisons setting a p-value of 0.05. (D) Heatmap of hierarchical clustering of serum proteins significantly different between diseases and healthy controls. BCD = B-cell depletion; MOGAD = myelin oligodendrocyte glycoprotein antibody-associated disease; NMOSD = neuromyelitis optica spectrum disorder.

Figure 2. Serum Protein Profiles Distinguish AQP4-NMOSD, MOGAD, and SN-NMOSD Patients Who Are Not Undergoing B-Cell Depletion

Comparison of serum protein profiles between (A) noBCD AQP4-NMOSD and noBCD SN-NMOSD, (B) noBCD AQP4-NMOSD and noBCD MOGAD, and (C) noBCD MOGAD and noBCD SN-NMOSD. Significant proteins were determined using unpaired t tests with multiple comparisons setting a p-value of 0.05. (D) Principal component analysis (PCA) of significantly different proteins between diseases. BCD = B-cell depletion; MOGAD = myelin oligodendrocyte glycoprotein antibody-associated disease; NMOSD = neuromyelitis optica spectrum disorder.

Figure 3. Effect of B-Cell Depletion on Serum Proteomic Signatures in AQP4-NMOSD, MOGAD, and SN-NMOSD Patients

Heatmap depicts supervised clustering of the proteins significantly altered in BCD vs NoBCD in (A) AQP4-NMOSD, (B) MOGAD, and (C) SN-NMOSD. The significant proteins used to generate the heatmaps were determined using unpaired t-tests with multiple comparisons setting a p-value of 0.05. (D) Ingenuity Pathway Analysis (IPA) of pathways altered by BCD vs NoBCD in each disease. Z-scores and -log10p values were determined using IPA. BCD = B-cell depletion; MOGAD = myelin oligodendrocyte glycoprotein antibody-associated disease; NMOSD = neuromyelitis optica spectrum disorder.

Figure 4. Effect of B-Cell Depletion on Individual Serum Proteins

Comparison of serum levels of (A) CD22, (B) CXCL13, (C) CXCL10, and (D) MMP10 between untreated (UnTx), non-BCD (nonBCD), and BCD-treated patients for each disease type. One-way ANOVA test with a Benjamini, Krieger, and Yekutieli correction for multiple comparisons was used.