IgG4 autoantibodies against muscle-specific kinase undergo Fab-arm exchange in myasthenia gravis patients

Abstract

Autoimmunity mediated by IgG4 subclass autoantibodies is an expanding field of research. Due to their structural characteristics a key feature of IgG4 antibodies is the ability to exchange Fab-arms with other, unrelated, IgG4 molecules, making the IgG4 molecule potentially monovalent for the specific antigen. However, whether those disease-associated antigen-specific IgG4 are mono- or divalent for their antigens is unknown. Myasthenia gravis (MG) with antibodies to muscle specific kinase (MuSK-MG) is a well-recognized disease in which the predominant pathogenic IgG4 antibody binds to extracellular epitopes on MuSK at the neuromuscular junction; this inhibits a pathway that clusters the acetylcholine (neurotransmitter) receptors and leads to failure of neuromuscular transmission. In vitro Fab-arm exchange-inducing conditions were applied to MuSK antibodies in sera, purified IgG4 and IgG1-3 sub-fractions. Solid-phase cross-linking assays were established to determine the extent of pre-existing and inducible Fab-arm exchange. Functional effects of the resulting populations of IgG4 antibodies were determined by measuring inhibition of agrin-induced AChR clustering in C2C12 cells. To confirm the results, κ/κ, λ/λ and hybrid κ/λ IgG4s were isolated and tested for MuSK antibodies. At least fifty percent of patients had IgG4, but not IgG1-3, MuSK antibodies that could undergo Fab-arm exchange in vitro under reducing conditions. Also MuSK antibodies were found in vivo that were divalent (monospecific for MuSK). Fab-arm exchange with normal human IgG4 did not prevent the inhibitory effect of serum derived MuSK antibodies on AChR clustering in C2C12 mouse myotubes. The results suggest that a considerable proportion of MuSK IgG4 could already be Fab-arm exchanged in vivo. This was confirmed by isolating endogenous IgG4 MuSK antibodies containing both κ and λ light chains, i.e. hybrid IgG4 molecules. These new findings demonstrate that Fab-arm exchanged antibodies are pathogenic.

B cells in the pathophysiology of myasthenia gravis

“…An interesting feature of secreted IgG4 antibodies is their ability to engage in “Fab‐arm exchange,” whereby a monospecific IgG4 protein may swap a heavy and light chain pair with another IgG4 to become bispecific . It has recently been shown that Fab‐arm‐exchanged antibodies are present in MuSK MG patients …”

Neuronal Surface Autoantibodies in Neuropsychiatric Disorders: Are There Implications for Depression?

“…Antibodies of the IgG1–3 subtypes are able to cross-link the antigens because of their bivalent nature, whereas the IgG4 subtype loses this ability after the fab-arm exchange with other unrelated IgG4 molecules (43). Cross-linking autoantibodies are believed to bring the antigens close together on the cell membrane and promote the degradation of the ligand–receptor complex (44).…”

“…As a result, the protein–protein interaction between the receptor and the associated protein is interrupted with the consequence that those receptors or ion channels become dysfunctional. Autoantibodies to muscle-specific kinase (anti-MuSK) are another type of autoantibodies involved in the pathogenicity of MG. Anti-MuSK (predominant IgG4) binds to an extracellular epitope on MuSK at the neuromuscular junction, inhibits the pathway involved in the clustering of the AChRs in the membrane, and leads to failure of neuromuscular transmission (43). Autoantibodies to LGI1, a VGKC complex-associated protein, play a similar role, resulting in reduced VGKC function at CNS synapses and increased cell excitability (60).…”

Utility of experimental animal models of myasthenia gravis for the elucidation of pathogenic mechanisms and development of new medications

“…64,65 Therefore, monovalent MuSK IgG4 antibodies are unlikely to disrupt MuSK action by antigenic modulation, which is required to bind divalently to adjacent MuSK molecules to cause internalization. 67 Klooster et al purified IgG4 and IgG1-3 fractions from the plasma of patients with MuSK-MG, and showed that the disease could be passively induced in mice through administration of the IgG4 fraction, but not the IgG1-3 fraction, thereby showing that IgG4 antibodies are the main pathogenic components. 66 In addition, divalent antibodies were shown to induce the activation of MuSK in the absence of agrin, whereas monovalent antibodies inhibited this activation, showing that distinct molecular mechanisms underlie the MuSK dysfunction induced by these two types of antibody.…”