Abstract Details

Single antigen-directed autoantibody diseases represent a unique model to describe the pathway to disease by tracing the development and survival of pathogenic B cells. We applied this framework to CASPR2-Ab-E, an autoimmune encephalitis characterized by autoantibodies reactive against synaptic protein, CASPR2.

To investigate the mechanisms driving immune tolerance escape by autoantigen-specific B cells in CASPR2-antibody encephalitis (CASPR2-Ab-E).

Naive B cells (NBCs; CD19⁺IgD⁺CD27^-) and memory B cells (MBCs; CD19⁺IgD^-CD27⁺) from six CASPR2-Ab-E patients and six healthy controls were cultured and differentiated into plasma cells. Supernatant reactivity was screened using CASPR2-EGFP expressing HEK293T cells in a live cell-based assay (CBA). Corresponding CASPR2-reactive BCR sequences were cloned, including reversion to unmutated common ancestor sequences (UCAs). All BCRs were expressed as monoclonal antibodies (mAbs) and characterized using surface plasmon resonance, CBA, BCR structure (IMGT CDR analyses), and polyreactivity/autoreactivity determination.

CASPR2-reactive NBCs were detected at similar frequencies in health and disease (0.4% vs 0.6%; p=0.21), while MBC CASPR2-reactivity was exclusive to patients. Memory BCRs carried affinity-enhancing somatic mutations, with diverse kinetics determined by surface plasma resonance. Binding was maintained, although with reduced affinity, when expressed as UCAs. Memory mAbs showed diverse pathogenic effects in vitro and in vivo. To ask how these memory B cells escape tolerance, we compared whole proteome and CASPR2 affinities, and observed UCAs reacted with highest affinity against CASPR2 with few other proteome-wide reactivities, which may explain their successful escape from tolerance mechanisms.

Our results highlight a balance between CASPR2 and other autoreactivities is critical for effective tolerance of CASPR2-reactive NBCs. We provide a rationale for investigating B cell tolerance checkpoints for CASPR2-Ab-E and present a pathology-tracing paradigm that can be applied across autoimmunity.