Printed Poster | Session 2

032 – Mechanisms of IgG4 autoantibodies to neurofascins in autoimmune demyelinating neuropathies

Kristina Patterson (1) – Xiaolin He (2) – Elisabeth Burnor (1) – Leah Zuroff (1) – Amit Bar-Or (1) – Eric Lancaster (1)
Hospital of the University of Pennsylvania, Neurology, Philadelphia, United States (1) – Northwestern University, Molecular Pharmacology and Biological Chemistry, Chicago, United States (2)

Background: Autoantibodies to Neurofascin 155 (NF155) define a subset of patients with severe acquired demyelinating neuropathy and are usually of the IgG4 subclass. IgG4, unlike other IgG subtypes, undergoes hemi-antibody exchange making each antibody bi-specific (functionally monovalent), such that crosslinking and internalization of the target antigen is unlikely. At the paranode, NF155 interacts with contactin/Caspr complex, an association necessary for anchoring myelin to the axon. NF155 is known to dimerize in vitro, though the functional implications of this interaction are not well understood. We hypothesize that IgG4 autoantibodies disrupt important heterophilic and homophilic interactions of NF155 which, in turn, may disrupt the function of the protein.
Methods: Using cell-based assay screening, we confirmed the presence NF155 autoantibodies in a subset of patients with demyelinating neuropathy. A solid phase binding assay was used to test the effect of NF155 autoantibodies on the binding of NF155 to contactin, Caspr, and neurofascin in vitro. The effects of NF155 autoantibodies on the binding of NF155 multimers to contactin was evaluated using a clustering assay in which a NF155 Fc fusion protein was pre-clustered with an Alexa 488-conjugated antibody against human IgG then incubated with HEK293 cells transiently transfected with contactin.

Results: We identified 4 cases of acquired demyelinating neuropathy harboring serum autoantibodies to NF155. IgG4 autoantibodies to NF155 were found in all cases and typically comprised the predominant subtype. In binding assays, NF155 bound contactin and neurofascin with nanomolar affinity while NF155 and Caspr did not appear to directly interact. Formation of NF155 multimers was necessary for contactin binding. Though antibodies to neurofascin had very modest direct effects on the interaction of NF155 with contactin, these neurofascin autoantibodies modulated homophilic interactions of NF155 which, in turn, indirectly inhibited binding of NF155 to contactin.
Conclusions: Our results suggest that inhibition of important protein-protein interactions is a possible mechanism by which NF155 IgG4 autoantibodies may disrupt the paranode. This work demonstrates that NF155 self-association increases affinity of contactin for NF155 and that interference with the formation of NF155 multimers indirectly inhibits the association of NF155 with contactin. Future work will focus on further characterizing homophilic NF155 interactions and their role in myelination.