Multiplexed autoantigen microarrays identify HLA as a key driver of anti-desmoglein and -non-desmoglein reactivities in pemphigus

Abstract

Patients with pemphigus vulgaris (PV) harbor antibodies reactive against self-antigens expressed at the surface of keratinocytes, primarily desmoglein (Dsg) 3 and, to a lesser extent, Dsg1. Conventionally, only antibodies targeting these molecules have been thought to contribute to disease pathogenesis. This notion has been challenged by a growing pool of evidence that suggests that antibodies toward additional targets may play a role in disease. The aims of this study were to (i) establish high-throughput protein microarray technology as a method to investigate traditional and putative autoantibodies (autoAbs) in PV and (ii) use multiplexed protein array technology to define the scope and specificity of the autoAb response in PV. Our analysis demonstrated significant IgG reactivity in patients with PV toward the muscarinic acetylcholine receptor subtypes 3, 4, and 5 as well as thyroid peroxidase. Furthermore, we found that healthy first-and second-degree relatives of patients with PV express autoAbs toward desmoglein and non-Dsg targets. Our analysis also identified genetic elements, particularly HLA, as key drivers of autoAb expression. Finally, we show that patients with PV exhibit significantly reduced IgM reactivity toward disease-associated antigens relative to controls. The use of protein microarrays to profile the autoAb response in PV advanced the current understanding of disease and provided insight into the complex relationship between genetics and disease development.pemphigus | autoantibody | protein microarray P emphigus vulgaris (PV) is a blistering autoimmune skin disease characterized by the presence of autoantibodies (autoAbs) directed against keratinocyte surface antigens (1, 2). Although early immunofluorescence studies demonstrated the presence of autoAbs in patient sera that bound to the surface of keratinocytes, a direct role of autoAbs in disease pathogenesis was not established until purified patient IgG (PVIgG) was shown to elicit blister formation upon passive transfer in mice (3). The main targets of these autoAbs were identified as Desmoglein (Dsg) 3 and 1, cadherin proteins that constitute key components of desmosomes, protein complexes responsible for maintaining cell-cell adhesion (4-7). Later experiments in which patient sera depleted of anti-Dsg3 Abs failed to produce blisters when passively transferred to mice (8) seemingly cemented the notion that these autoAbs alone were responsible for blister formation. As a result, subsequent research in the field has focused primarily on autoAbs directed against Dsgs.The assertion that anti-Dsg autoAbs alone are pathogenic was first challenged when PVIgG, lacking any anti-Dsg1 autoAbs, produced blisters when passively transferred into Dsg3-null mice (9). Additionally, several studies have shown that anti-Dsg Ab titers do not necessarily correlate with disease activity, and a subset of patients with PV do not harbor any detectable anti-Dsg Abs (10-14). The presence of pathogenic autoAbs directed against non-Dsg targets could account for th...

Pemphigus

“…Moreover, a pemphigus vulgaris-associated functional risk variant residing within the ST18 promoter region may be linked to the secretion of key inflammatory molecules by keratinocytes after autoantibody binding (tumour necrosis factor (TNF), IL-1α and IL-6) 30 . In addition to regulating autoantibody expression, the HLA status is also an important driver of oxidative stress in pemphigus vulgaris 31,32 .…”

“…However, in very rare cases of pemphigus vulgaris, IgG autoantibodies against desmocollin 3 (another desmosomal cadherin) can cause the disease phenotype without coexisting anti-desmoglein autoantibodies 41,42 . In addition to desmoglein and desmocollin, other autoimmune targets have also been found with various approaches 31,43–45 . Although their potential role in the pathogenesis of pemphigus is still under investigation, these non-desmoglein autoantibodies might have some supporting roles and contribute the complex phenotype of pemphigus or might have pathogenic roles on their own that are sufficient for blister formation in rare cases.…”

“…Anti-desmoglein 1 antibody titres tend to show a closer correlation with the course of the disease activity compared with anti-desmoglein 3 antibody titres 120,121 . However, active lesions might occur, albeit rarely, even in the absence of detectable anti-desmoglein 3 and desmoglein 1 IgG autoantibodies 122 , whereas 20–40% of patients in clinical remission still have detectable levels of circulating autoantibodies 123,124 , and low levels of anti-desmoglein IgG autoantibodies are also occasionally detected among healthy individuals 25,31,125,126 .…”

A reciprocal HLA-Disease Association in Rheumatoid Arthritis and Pemphigus Vulgaris

“…PV, the most common type of pemphigus, is a potentially lethal blistering disease that affects the mucosal membranes and the skin, and is associated with antibodies against the desmosomal cadherins desmoglein 3 (Dsg3) and Dsg1 (17-20). These antibodies are believed to attack the desmosomes, thereby disrupting the adhesion between the keratinocytes in the epidermis, which is followed by accumulation of transudate fluid.…”

“…It was recently shown that the PV associated HLA alleles, including HLA-DRB1*0402 , are important drivers for the reactivity against autoantigen targets in PV (20). However, little is currently known about the mechanistic basis of the association of HLA-DRB1*04:02 with PV.…”