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Although B cells are implicated in multiple sclerosis (MS) pathophysiology, a predictive or diagnostic autoantibody remains elusive. Here, the Department of Defense Serum Repository (DoDSR), a cohort of over 10 million individuals, was used to generate whole-proteome autoantibody profiles of hundreds of patients with MS (PwMS) years before and subsequently after MS onset. This analysis defines a unique cluster of PwMS that share an autoantibody signature against a common motif that has similarity with many human pathogens. These patients exhibit antibody reactivity years before developing MS symptoms and have higher levels of serum neurofilament light (sNfL) compared to other PwMS. Furthermore, this profile is preserved over time, providing molecular evidence for an immunologically active prodromal period years before clinical onset. This autoantibody reactivity was validated in samples from a separate incident MS cohort in both cerebrospinal fluid (CSF) and serum, where it is highly specific for patients eventually diagnosed with MS. This signature is a starting point for further immunological characterization of this MS patient subset and may be clinically useful as an antigen-specific biomarker for high-risk patients with clinically- or radiologically-isolated neuroinflammatory syndromes.

BACKGROUNDVaccine-elicited adaptive immunity is a prerequisite for control of SARS-CoV-2 infection. Multiple sclerosis (MS) disease-modifying therapies (DMTs) differentially target humoral and cellular immunity. A comprehensive comparison of the effects of MS DMTs on SARS-CoV-2 vaccine-specific immunity is needed, including quantitative and functional B and T cell responses.METHODSSpike-specific Ab and T cell responses were measured before and following SARS-CoV-2 vaccination in a cohort of 80 study participants, including healthy controls and patients with MS in 6 DMT groups: untreated and treated with glatiramer acetate (GA), dimethyl fumarate (DMF), natalizumab (NTZ), sphingosine-1-phosphate (S1P) receptor modulators, and anti-CD20 mAbs. Anti-spike-Ab responses were assessed by Luminex assay, VirScan, and pseudovirus neutralization. Spike-specific CD4+ and CD8+ T cell responses were characterized by activation-induced marker and cytokine expression and tetramer.RESULTSAnti-spike IgG levels were similar between healthy control participants and patients with untreated MS and those receiving GA, DMF, or NTZ but were reduced in anti-CD20 mAb- and S1P-treated patients. Anti-spike seropositivity in anti-CD20 mAb-treated patients was correlated with CD19+ B cell levels and inversely correlated with cumulative treatment duration. Spike epitope reactivity and pseudovirus neutralization were reduced in anti-CD20 mAb- and S1P-treated patients. Spike-specific CD4+ and CD8+ T cell reactivity remained robust across all groups, except in S1P-treated patients, in whom postvaccine CD4+ T cell responses were attenuated.CONCLUSIONThese findings from a large cohort of patients with MS exposed to a wide spectrum of MS immunotherapies have important implications for treatment-specific COVID-19 clinical guidelines.FUNDINGNIH grants 1K08NS107619, K08NS096117, R01AI159260, R01NS092835, R01AI131624, and R21NS108159; NMSS grants TA-1903-33713 and RG1701-26628; Westridge Foundation; Chan Zuckerberg Biohub; Maisin Foundation.

While SARS-CoV-2 infection has pleiotropic and systemic effects in some patients, many others experience milder symptoms. We sought a holistic understanding of the severe/mild distinction in COVID-19 pathology, and its origins. We performed a whole-blood preserving single-cell analysis protocol to integrate contributions from all major cell types including neutrophils, monocytes, platelets, lymphocytes and the contents of serum. Patients with mild COVID-19 disease display a coordinated pattern of interferon-stimulated gene (ISG) expression across every cell population and these cells are systemically absent in patients with severe disease. Severe COVID-19 patients also paradoxically produce very high anti-SARS-CoV-2 antibody titers and have lower viral load as compared to mild disease. Examination of the serum from severe patients demonstrates that they uniquely produce antibodies with multiple patterns of specificity against interferon-stimulated cells and that those antibodies functionally block the production of the mild disease-associated ISG-expressing cells. Overzealous and auto-directed antibody responses pit the immune system against itself in many COVID-19 patients and this defines targets for immunotherapies to allow immune systems to provide viral defense., One Sentence Summary: In severe COVID-19 patients, the immune system fails to generate cells that define mild disease; antibodies in their serum actively prevents the successful production of those cells.

One third of COVID-19 patients develop significant neurological symptoms, yet SARS-CoV-2 is rarely detected in central nervous system (CNS) tissue, suggesting a potential role for parainfectious processes, including neuroimmune responses. We therefore examined immune parameters in cerebrospinal fluid (CSF) and blood samples from a cohort of patients with COVID-19 and significant neurological complications. We found divergent immunological responses in the CNS compartment, including increased levels of IL-12 and IL-12-associated innate and adaptive immune cell activation. Moreover, we found increased proportions of B cells in the CSF relative to the periphery and evidence of clonal expansion of CSF B cells, suggesting a divergent intrathecal humoral response to SARS-CoV-2. Indeed, all COVID-19 cases examined had anti-SARS-CoV-2 IgG antibodies in the CSF whose target epitopes diverged from serum antibodies. We directly examined whether CSF resident antibodies target self-antigens and found a significant burden of CNS autoimmunity, with the CSF from most patients recognizing neural self-antigens. Finally, we produced a panel of monoclonal antibodies from patients’ CSF and show that these target both anti-viral and anti-neural antigens—including one mAb specific for the spike protein that also recognizes neural tissue. This exploratory immune survey reveals evidence of a compartmentalized and self-reactive immune response in the CNS meriting a more systematic evaluation of neurologically impaired COVID-19 patients.One Sentence SummaryA subset of COVID-19 patients with neurologic impairment show cerebrospinal fluid-specific immune alterations that point to both neuroinvasion and anti-neural autoimmunity as potential causes of impairment.

As SARS-CoV-2 continues to spread around the world, there is an urgent need for new assay formats to characterize the humoral response to infection. Convalescent serum is being used for treatment and for isolation of patient-derived antibodies. However, currently there is not a simple means to estimate serum bulk neutralizing capability. Here we present an efficient competitive serological assay that can simultaneously determine an individual’s seropositivity against the SARS-CoV-2 Spike protein and estimate the neutralizing capacity of anti-Spike antibodies to block interaction with the human angiotensin converting enzyme 2 (ACE2) required for viral entry. In this ELISA-based assay, we present natively-folded viral Spike protein receptor binding domain (RBD)-containing antigens via avidin-biotin interactions. Sera are then supplemented with soluble ACE2-Fc to compete for RBD-binding serum antibodies, and antibody binding quantified. Comparison of signal from untreated serum and ACE2-Fc-treated serum reveals the presence of antibodies that compete with ACE2 for RBD binding, as evidenced by loss of signal with ACE2-Fc treatment. In our test cohort of nine convalescent SARS-CoV-2 patients, we found all patients had developed anti-RBD antibodies targeting the epitope responsible for ACE2 engagement. This assay provides a simple and high-throughput method to screen patient sera for potentially neutralizing anti-Spike antibodies to enable identification of candidate sera for therapeutic use.