Cell-binding IgM in CSF is distinctive of multiple sclerosis and targets the iron transporter SCARA5.

Intrathecal IgM production in multiple sclerosis is associated with a worse disease course. To investigate pathogenic relevance of autoreactive IgM in multiple sclerosis, CSF from two independent cohorts, including multiple sclerosis patients and controls, were screened for antibody binding to induced pluripotent stem cell-derived neurons and astrocytes, and a panel of CNS-related cell lines. IgM binding to a primitive neuro-ectodermal tumour cell line discriminated 10% of multiple sclerosis donors from controls. Transcriptomes of single IgM producing CSF B cells from patients with cell-binding IgM were sequenced and used to produce recombinant monoclonal antibodies for characterization and antigen identification. We produced five cell-binding recombinant IgM antibodies, of which one, cloned from an HLA-DR + plasma-like B cell, mediated antigen-dependent complement activation. Immunoprecipitation and mass spectrometry, and biochemical and transcriptome analysis of the target cells identified the iron transport scavenger protein SCARA5 as the antigen target of this antibody. Intrathecal injection of a SCARA5 antibody led to an increased T cell infiltration in an experimental autoimmune encephalomyelitis (EAE) model. CSF IgM might contribute to CNS inflammation in multiple sclerosis by binding to cell surface antigens like SCARA5 and activating complement, or by facilitating immune cell migration into the brain.

Potent neutralization by monoclonal human IgM against SARS-CoV-2 is impaired by class switch.

To investigate the class-dependent properties of anti-viral IgM antibodies, we use membrane antigen capture activated cell sorting to isolate spike-protein-specific B cells from donors recently infected with SARS-CoV-2, allowing production of recombinant antibodies. We isolate 20, spike-protein-specific antibodies of classes IgM, IgG, and IgA, none of which shows any antigen-independent binding to human cells. Two antibodies of class IgM mediate virus neutralization at picomolar concentrations, but this potency is lost following artificial switch to IgG. Although, as expected, the IgG versions of the antibodies appear to have lower avidity than their IgM parents, this is not sufficient to explain the loss of potency.

Receptor clustering and pathogenic complement activation in myasthenia gravis depend on synergy between antibodies with multiple subunit specificities.

Myasthenia gravis is an autoimmune disorder defined by muscle weakness and fatigability associated with antibodies against proteins of the neuromuscular junction (NMJ). The most common autoantibody target is the acetylcholine receptor (AChR). Three mechanisms have been postulated by which autoantibodies might interfere with neurotransmission: direct antagonism of the receptor, complement-mediated destruction of the postsynaptic membrane, and enhanced internalization of the receptor. It is very likely that more than one of these mechanisms act in parallel. Dissecting the mechanisms of autoantibody-mediated pathology requires patient-derived, monoclonal antibodies. Using membrane antigen capture activated cell sorting (MACACS), we isolated AChR-specific B cells from patients with myasthenia gravis, and produced six recombinant antibodies. All AChR-specific antibodies were hypermutated, including isotypes IgG1, IgG3, and IgG4, and recognized different subunits of the AChR. Despite clear binding, none of the individual antibodies showed significant antagonism of the AChR measured in an in vitro neuromuscular synapse model, or AChR-dependent complement activation, and they did not induce myasthenic signs in vivo. However, combinations of antibodies induced strong complement activation in vitro, and severe weakness in a passive transfer myasthenia gravis rat model, associated with NMJ destruction and complement activation in muscle. The strongest complement activation was mediated by combinations of antibodies targeting disparate subunits of the AChR, and such combinations also induced the formation of large clusters of AChR on the surface of live cells in vitro. We propose that synergy between antibodies of different epitope specificities is a fundamental feature of this disease, and possibly a general feature of complement-mediated autoimmune diseases. The importance of synergistic interaction between antibodies targeting different subunits of the receptor can explain the well-known discrepancy between serum anti-AChR titers and clinical severity, and has implications for therapeutic strategies currently under investigation.