Background: Bullous pemphigoid (BP) and mucous membrane pemphigoid (MMP) are rare autoimmune blistering disorders characterized by autoantibodies (autoAbs) targeting dermo-epidermal junction components such as BP180 and BP230. The differential diagnosis, based on both the time of appearance and the extension of cutaneous and/or mucosal lesions, is crucial to distinguish these diseases for improving therapy outcomes and delineating the correct prognosis; however, in some cases, it can be challenging. In addition, negative results obtained by commercially available enzyme-linked immunosorbent assays (ELISAs) with BP and MMP sera, especially from patients with ocular involvement, often delay diagnosis and treatment, leading to a greater risk of poor outcomes.Objectives: Our aim was to find potentially different reactivity profiles in BP and MMP and improve available approaches for diagnosis with focus on ocular MMP.Methods: Two cohorts of 90 BP and 90 MMP, recruited from different Italian clinical centers, were characterized also employing a novel ELISA based on the BP180 extracellular domain (ECD-BP180).Results: Immunoglobulin G (IgG) reactivity to BP180 and BP230 in MMP sera was significantly reduced in comparison with BP, mostly affecting BP230 and E-1080 (53% and 36% in BP vs. 11% and 3% in MMP, respectively, p < 0.0001). The combined sensitivity of BP180-NC16A and ECD-BP180 ELISAs was greater compared to BP180-NC16A and BP230 ELISAs both in BP (97% and 92%, respectively) and in MMP (42% and 31%, respectively). The present study shows that MMP patients with ocular involvement rarely reacted to BP180 by IgG in contrast with patients with oral and/or cutaneous involvement (p = 0.0245 and p = 0.0377, respectively), suggesting that an oral and/or cutaneous MMP positive to BP180 hardly evolves to ocular MMP. Of note, one-third of ocular MMP showed immunoglobulin A (IgA) reactivity to ECD-BP180 by immunoblotting.Conclusions: The present study provides several hints to perform a correct and timely diagnosis in BP and MMP, which is crucial for improving therapy outcomes and delineating the correct prognosis.

Pemphigus vulgaris (PV) is an autoimmune bullous disease of the skin and mucous membranes characterized by the presence of circulating and tissue-bound autoantibodies against keratinocyte cell surface antigens, specifically desmoglein (Dsg) 1 and 3. The pathogenic role of anti-Dsg antibodies is well-established, while the mechanism of blister formation is only partly defined. We have applied a previously developed method for the efficient immortalization of IgG+ memory B cells to identify novel target antigens in PV. A human monoclonal antibody reactive with a hitherto unreported non-Dsg antigen was isolated. Immunoprecipitation and immunoblotting studies with keratinocyte extracts indicated α-catenin as the putative antigen, then confirmed by immunoblotting on the recombinant protein. Four of ten PV sera reacted with recombinant α-catenin. Although the isolated human monoclonal antibody was per se unable to dissociate keratinocyte monolayers and also to synergize with a pathogenic antibody in vitro, further studies are warranted to assess its possible in vivo contribution in the multifactorial pathogenesis and heterogeneous manifestations of PV disease.

Growing evidence has linked bullous pemphigoid (BP) to immune checkpoint inhibitor (ICI) therapy in cancer treatment. However, the immunological features of ICI-associated BP (ICI-BP) are not yet fully elucidated. In order to characterize the humoral response in ICI BP patients and investigate whether their epitope profile differs from idiopathic BP (IBP), 53 ICI-BP patients were enrolled, immunologically characterized and compared with 59 IBP patients. ICI-BP had a distinctive IgG humoral profile, with reduced reactivity toward BP230 and recognition of multiple BP180 epitopes beyond the immunodominant extracellular noncollagenous 16A domain (NC16A). Specifically, reactivity to BP180 ectodomain was present in 94% of ICI-BP and 78% of IBP (p=0.044). Moreover, BP180 C-terminal epitope was more frequently targeted in ICI-BP than IBP (72% vs 41%, p=0.002). Notably, the combined use of an in-house BP180 ectodomain ELISA and the commercial BP180 test increased diagnostic sensitivity from 83% to 100%. Enhanced IgG reactivity toward nonimmunodominant epitopes, and especially C-terminal epitope recognition, characterize the humoral immune response in ICI-BP. Our data suggest that combining NC16A and fulllength BP180 ectodomain ELISAs may help reduce diagnostic delay in ICI-BP patients, in whom a timely diagnosis is crucial to appropriately manage the disease and ultimately avoid discontinuation ofcancer therapy.