Dysregulation of B Cell Repertoire Formation in Myasthenia Gravis Patients Revealed through Deep Sequencing

Abstract

Myasthenia gravis (MG) is a prototypical B cell-mediated autoimmune disease affecting 20–50 per 100,000 people. The majority of patients fall into two clinically distinguishable types based on whether they produce autoantibodies targeting the acetylcholine receptor (AChR-MG) or muscle specific kinase (MuSK-MG). The autoantibodies are pathogenic, but whether their generation is associated with broader defects in the B cell repertoire is unknown. To address this question, we performed deep sequencing of the B cell receptor repertoire of AChR-MG, MuSK-MG and healthy subjects to generate approximately 518,000 unique VH and VL sequences from sorted naïve and memory B cell populations. AChR-MG and MuSK-MG subjects displayed distinct gene segment usage biases in both VH and VL sequences within the naïve and memory compartments. The memory compartment of AChR-MG was further characterized by reduced positive selection of somatic mutations in the H-CDR regions and altered H-CDR3 physicochemical properties. The VL repertoire of MuSK-MG was specifically characterized by reduced V/J segment distance in recombined sequences, suggesting diminished VL receptor editing during B cell development. Our results identify large-scale abnormalities in both the naïve and memory B cell repertoires. Particular abnormalities were unique to either AChR-MG or MuSK-MG indicating that the repertoires reflect the distinct properties of the subtypes. These repertoire abnormalities are consistent with previously observed defects in B cell tolerance checkpoints in MG, thereby offering additional insight regarding the impact of tolerance defects on peripheral autoimmune repertoires. These collective findings point toward a deformed B cell repertoire as a fundamental component of MG.

Mechanisms underlying B cell immune dysregulation and autoantibody production in MuSK myasthenia gravis

“…The most prominent repertoire abnormality involved the biased use of variable (V) region gene segments. We observed significant shifts in the relative abundance and variability of each V gene family between MuSK MG and HDs . In the naive compartment of MuSK MG subjects, V‐family biases were observed in the form of decreased usage of IGHV3 as well as an increase in IGHV1 and IGHV4 usage.…”

“…We recently applied this approach to compare the broad repertoire features of both the naive and memory B cell compartments in MuSK MG patients with those of healthy donors (HDs) . We reasoned that a disease driven by autoantibodies may have conspicuous abnormalities present in the circulating B cell repertoire.…”

Impaired B‐cell tolerance checkpoints promote the development of autoimmune diseases and pathogenic autoantibodies

“…82 In contrast, naive BCR repertoires from MG patients were highly diverse among individuals and may reflect dysregulated B-cell development in these patients. 81 ities. An AIRR-seq investigation revealed that differential expression of IGHV genes in the repertoires from SSc cohorts relative to a healthy cohort are present in the IgD/IgM compartment.…”

“…80 We more specifically applied AIRR-seq to analyze the BCR repertoire of naive B cells from AChR and MuSK MG patients that are enriched in autoreactive clones due to impaired central and peripheral B-cell tolerance checkpoints in MG and compared them to those of healthy donors (HD) to potentially identify naive BCR repertoire features associated with autoimmunity. 81 Our initial study focused on determining whether the naive BCR repertoires in MG included both skewed immunoglobulin gene usage and physiochemical properties. 81 In agreement with a previous study, the naive B-cell repertoires obtained from multiple healthy control subjects displayed highly consistent IGH variable (IGHV) gene usage with limited variability.…”

Autoantibody-producing plasmablasts after B cell depletion identified in muscle-specific kinase myasthenia gravis

“…Although broad immune dysregulation (13)(14)(15) in MuSK MG is recognized, the mechanistic details of autoantibody production remain undefined. Given that MuSK MG autoantibodies are required for pathogenesis (2) and that they are, consequently, the fundamental target of immunotherapy, it is important to establish how they are generated.…”