Autoantibodies neutralizing type I interferons (IFN-Is; IFNα or IFNω) exacerbate severe viral disease, but specific treatments are unavailable. With footprint profiling, we delineate two dominant IFN-I faces commonly recognized by neutralizing IFN-I autoantibody-containing plasmas from aged individuals with HIV-1 and from individuals with severe COVID-19. These faces overlap with IFN-I regions independently essential for engaging the IFNAR1/IFNAR2 heterodimer, and neutralizing plasmas efficiently block the interaction of IFN-I with both receptor subunits in vitro. In contrast, non-neutralizing autoantibody-containing plasmas limit the interaction of IFN-I with only one receptor subunit and display relatively low IFN-I-binding avidities, thus likely hindering neutralizing function. Iterative engineering of signaling-inert mutant IFN-Is (simIFN-Is) retaining dominant autoantibody targets created potent decoys that prevent IFN-I neutralization by autoantibody-containing plasmas and that restore IFN-I-mediated antiviral activity. Additionally, microparticle-coupled simIFN-Is were effective at depleting IFN-I autoantibodies from plasmas, leaving antiviral antibodies unaffected. Our study reveals mechanisms of action for IFN-I autoantibodies and demonstrates a proof-of-concept strategy to alleviate pathogenic effects.
© 2025 Groen et al.

Autoantibodies (autoAbs) that neutralize type 1 interferons (T1IFNs) are a major risk factor associated with developing critical COVID-19 disease and are most commonly found in individuals over age 70 and in patients with genetic or acquired thymic defects. Swift identification of autoAb-positive individuals may allow targeted interventions to prevent critical COVID-19 disease. Herein, we provide a workflow and protocols aimed at rapidly identifying individuals who are autoAb positive from a large cohort. Basic Protocol 1 describes a multiplex particle-based assay to screen large cohorts of individuals for binding levels of anti-T1IFN autoAbs, and Basic Protocol 2 describes a functional assay to test if autoAbs in patient plasma can block T1IFN-induced JAK/STAT signaling. © Published 2022. This article is a U.S. Government work and is in the public domain in the USA. Basic Protocol 1: Multiplex particle-based bead assay to screen for binding levels of anti-type 1 interferon autoantibodies Alternate Protocol: Multiplex particle-based bead assay to screen for binding levels of anti-type 1 interferon immunoglobulin subtypes and isotypes Support Protocol: Coupling type 1 interferons (IFN-α, IFN-β, and IFN-ω) to magnetic beads Basic Protocol 2: pSTAT1 functional assay to test for neutralization activity of anti-type 1 interferon autoantibodies.
© Published 2022. This article is a U.S. Government work and is in the public domain in the USA.

Dimethyl fumarate (DMF) is a prescribed treatment for multiple sclerosis and has also been used to treat psoriasis. The electrophilicity of DMF suggests that its immunosuppressive activity is related to the covalent modification of cysteine residues in the human proteome. Nonetheless, our understanding of the proteins modified by DMF in human immune cells and the functional consequences of these reactions remains incomplete. In this study, we report that DMF inhibits human plasmacytoid dendritic cell function through a mechanism of action that is independent of the major electrophile sensor NRF2. Using chemical proteomics, we instead identify cysteine 13 of the innate immune kinase IRAK4 as a principal cellular target of DMF. We show that DMF blocks IRAK4-MyD88 interactions and IRAK4-mediated cytokine production in a cysteine 13-dependent manner. Our studies thus identify a proteomic hotspot for DMF action that constitutes a druggable protein-protein interface crucial for initiating innate immune responses.
Copyright © 2019 by The American Association of Immunologists, Inc.