Blocking soluble Fas Ligand ameliorates pemphigus: PC111 efficacy in ex-vivo human pemphigus models.

Pemphigus is a life-threatening, chronic, autoimmune bullous disease affecting both the skin and the mucous membranes. Based on the mainstream concept that blister formation occurs upon binding of autoantibodies to their antigen proteins (desmoglein1, DSG1 and desmoglein3, DSG3), current therapies mostly aim to suppress the immune system. To avoid the severe side effects associated with the chronic use of immunosuppressive treatments, we have developed PC111, a fully human monoclonal antibody targeting human Fas ligand (FasL). We have provided a number of in vitro and in vivo evidences showing that soluble FasL induces keratinocyte apoptosis followed by acantholysis. An anti-murine FasL prevents blister formation in the pemphigus neonatal mouse model. To confirm the mechanism of action (MoA) and the efficacy of PC111 in a human pemphigus context, we used the keratinocyte dissociation assay and two independent Human Skin Organ Cultures (HSOC) pemphigus models. PC111 reduced acantholysis in vitro, as shown by the dose-dependent reduction of fragments in the monolayer cultures. In the first HSOC model, normal human skin was subcutaneously injected with a scFv antibody fragment directed against DSG1 and DSG3, resulting in a severe acantholysis (70-100%) after 24 hours. PC111 inhibited blister formation to around 50% of control. In the second model, normal human skin was injected with a mixture of pemphigus patients' autoantibodies resulting in a less severe acantholysis (20-30%). PC111 significantly suppressed blister formation to more than 75% up to 72 hours. These results confirm PC111 MoA and demonstrates the efficacy of the anti-FasL antibody also in a pemphigus setting.

Autoimmune pre-disease.

Approximately 5% of the world-wide population is affected by autoimmune diseases. Overall, autoimmune diseases are still difficult to treat, impose a high burden on patients, and have a significant economic impact. Like other complex diseases, e.g., cancer, autoimmune diseases develop over several years. Decisive steps in the development of autoimmune diseases are (i) the development of autoantigen-specific lymphocytes and (often) autoantibodies and (ii) potentially clinical disease manifestation at a later stage. However, not all healthy individuals with autoantibodies develop disease manifestations. Identifying autoantibody-positive healthy individuals and monitoring and inhibiting their switch to inflammatory autoimmune disease conditions are currently in their infancy. The switch from harmless to inflammatory autoantigen-specific T and B-cell and autoantibody responses seems to be the hallmark for the decisive factor in inflammatory autoimmune disease conditions. Accordingly, biomarkers allowing us to predict this progression would have a significant impact. Several factors, such as genetics and the environment, especially diet, smoking, exposure to pollutants, infections, stress, and shift work, might influence the progression from harmless to inflammatory autoimmune conditions. To inspire research directed at defining and ultimately targeting autoimmune predisease, here, we review published evidence underlying the progression from health to autoimmune predisease and ultimately to clinically manifest inflammatory autoimmune disease, addressing the following 3 questions: (i) what is the current status, (ii) what is missing, (iii) and what are the future perspectives for defining and modulating autoimmune predisease.

Association of stress-related neural activity and baseline interleukin-6 plasma levels in healthy adults.

Several studies suggest a link between acute changes in inflammatory parameters due to an endotoxin or (psychological) stressor and the brain's stress response. The extent to which basal circulating levels of inflammatory markers are associated with the brain's stress response has been hardly investigated so far. In the present study, baseline plasma levels of the cytokine interleukin (IL)-6 were obtained and linked to neural markers of psychosocial stress using a modified version of the Montreal Imaging Stress Task in a sample of N = 65 healthy subjects (N = 39 female). Of three a-priori defined regions of interest - the amygdala, anterior insula, and anterior cingulate cortex - baseline IL-6 was significantly and negatively associated with stress-related neural activation in the right amygdala and left anterior insula. Our results suggest that baseline cytokines might be related to differences in the neural stress response and that this relationship could be inverse to that previously reported for induced acute changes in inflammation markers.

Melanoma is associated with an increased risk of bullous pemphigoid: a large population-based longitudinal study.

The association between bullous pemphigoid (BP) and melanoma is yet to be investigated. We aimed to assess assess the bidirectional association between BP and melanoma and to delineate the epidemiological features of patients with both diagnoses. A population-based cohort study was performed comparing BP patients (n = 3924) with age-, sex- and ethnicity-matched control subjects (n = 19,280) with regard to incident cases of melanoma. A case-control design was additionally adopted to estimate the risk of BP in individuals with a preexisting diagnosis of melanoma. The prevalence of preexisting melanoma was higher in patients with BP than in control subjects (1.5% vs. 1.0%, respectively; P = 0.004). A history of melanoma confers a 50% increase in the risk of subsequent BP (OR 1.53; 95% CI 1.14-2.06). This risk was higher among males (OR 1.66; 95% CI 1.09-2.54) and individuals older than 80 years (OR 1.63; 95% CI 1.11-2.38), and persisted after adjustment for multiple putative confounders including PD-1/PDL-1 antagonists (adjusted OR 1.53; 95% CI 1.14-2.06). Conversely, the risk of melanoma among patients with BP was slightly elevated, but did not reach the level of statistical significance (adjusted HR 1.13; 95% CI 0.73-1.74). Patients with a dual diagnosis of BP and melanoma were older at the onset of BP and had lower body mass index. A history of melanoma is associated with a 50% increase in the incidence of subsequent BP. Physicians managing patients with both conditions should be aware of this association. Further research is warranted to reveal the underlying mechanism of these findings.

Optical coherence tomography for fast bedside imaging, assessment and monitoring of autoimmune inflammatory skin diseases?

Optical coherence tomography (OCT) is a non-invasive, high-resolution imaging technique with a growing impact in dermatology. The principle of OCT is comparable to that of sonography, except that it uses infrared laser light instead of ultrasound waves. It has been clinically demonstrated that OCT is suitable for discriminating between different types of non-melanoma skin cancer at an early stage of disease. Optical coherence tomography generates two- or three-dimensional images of up to 2 mm penetration depth, a field of view of 6 mm × 6 mm, and an acquisition time of seconds. The resolution capability of OCT is more than 3 to 100 times higher than that of ultrasound imaging. It is of particular interest that the additional information on vasculature provided by OCT angiography enables the assessment and monitoring of inflammatory skin diseases. The use of OCT to locate exact blister levels was demonstrated for diagnosing autoimmune bullous diseases. It is anticipated that detection of subclinical lesions could indicate a relapse of the disease. In the future, this could enable intervention and early treatment. Furthermore, the development of high-speed OCT could allow fast scanning and bedside imaging of large body sites.

Visualization of autoantibodies and neutrophils in vivo identifies novel checkpoints in autoantibody-induced tissue injury.

In several autoimmune diseases, e.g., pemphigoid disease (PD), autoantibodies are the direct cause of pathology. Albeit key requirements for antibody-mediated diseases were identified, their interactions and exact temporal and spatial interactions remained elusive. The skin is easily accessible for imaging. Thus, we selected epidermolysis bullosa acquisita (EBA), a PD with autoantibodies to type VII collagen (COL7), to visualize interactions of autoantibodies, target tissue and effector cells (neutrophils). Following injection into mice, anti-COL7 IgG bound to the dermal-epidermal junction (DEJ) within minutes. We unexpectedly observed an inhomogeneous distribution of autoantibodies along the DEJ. Thus, we hypothesized that specific external triggers may affect autoantibody distribution. Indeed, mechanical irritation led to an increased autoantibody binding along the DEJ. Subsequently, anti-COL7 IgG was injected into mice expressing green fluorescent protein under the LysM promoter (LysM-eGFP) mice. This allows to visualize myeloid cells in vivo in these animals. Using multiphoton imaging, we observed a limited extravasation of LysM-eGFP+ cells into skin was observed within 24 hours. Intriguingly, LysM-eGFP+ cells did not immediately co-localize with autoantibodies, which was only noted at later time points. Of note, interactions of LysM-eGFP+ with the autoantibodies at the DEJ were short-lived. Collectively, our results define the following checkpoints for autoantibody-induced tissue injury: (i) autoantibody egress to target tissue influenced by mechanical trigger factors, (ii) neutrophil recruitment into the vicinity of autoantibody deposits and (iii) short-term neutrophil localization to these deposits, as well as (iv) delayed recruitment of neutrophils with subsequent autoantibody-induced inflammation.

The transmembrane protein LRIG2 increases tumor progression in skin carcinogenesis.

Over the last few decades, the number of cases of non-melanoma skin cancer (NMSC) has risen to over 3 million cases every year worldwide. Members of the ERBB receptor family are important regulators of skin development and homeostasis and, when dysregulated, contribute to skin pathogenesis. In this study, we investigated leucine-rich repeats and immunoglobulin-like domains 2 (LRIG2), a transmembrane protein involved in feedback loop regulation of the ERBB receptor family during NMSC. LRIG2 was identified to be up-regulated in various types of squamous cell carcinoma (SCC), but little is known about LRIG2 in cutaneous SCC (cSCC). To investigate the function of LRIG2 in cSCC in vivo, we generated a skin-specific LRIG2 overexpressing transgenic mouse line (LRIG2-TG) using the Tet-Off system. We employed the 7,12-dimethylbenz(a)anthracene/12-O-tetra-decanoylphorbol-13-acetate (DMBA/TPA) two-stage chemical carcinogenesis model and analyzed the skin during homeostasis and tumorigenesis. LRIG2-TG mice did not exhibit alterations in skin development or homeostasis but showed an interaction between LRIG2 and thrombospondin-1, which is often involved in angiogenesis and tumorigenesis. However, during carcinogenesis, transgenic animals showed significantly increased tumor progression and a more rapid development of cSCC. This was accompanied by changes in the ERBB system. After a single TPA application, inflammation of the epidermis was enhanced during LRIG2 overexpression. In human skin samples, LRIG2 expression was identified in the basal layer of the epidermis and in hair follicles of normal skin, but also in cSCC samples. In conclusion, epidermal LRIG2 excess is associated with activated EGFR/ERBB4-MAPK signaling and accelerated tumor progression in experimentally induced NMSC, suggesting LRIG2 as a potential oncoprotein in skin.

Reduced skin blistering in experimental epidermolysis bullosa acquisita after anti-TNF treatment.

Epidermolysis bullosa acquisita (EBA) is a difficult-to-treat subepidermal autoimmune blistering skin disease (AIBD) with circulating and tissue-bound anti-type VII collagen antibodies. Different reports have indicated an increased concentration of tumor necrosis factor alpha (TNF) in the serum and blister fluid of patients with subepidermal AIBDs. Furthermore, successful anti-TNF treatment has been reported for individual patients with AIBDs. Here, we show that in mice, induction of experimental EBA by repeated injections of rabbit-anti mouse type VII collagen antibodies led to increased expression of TNF in skin, as determined by real-time PCR and immunohistochemistry. To investigate if the increased TNF expression is of functional relevance in experimental EBA, we inhibited TNF function using the soluble TNF receptor fusion protein etanercept (Enbrel®) or a monoclonal antibody to murine TNF. Interestingly, mice receiving either of these two treatments showed significantly milder disease progression than controls. In addition, immunohistochemical staining demonstrated reduced numbers of macrophages in lesional skin in mice treated with TNF inhibitors compared to controls. Furthermore, etanercept treatment significantly reduced the disease progression in immunization-induced EBA. In conclusion, the increased expression of TNF in experimental EBA is of functional relevance, as both the prophylactic blockade of TNF and the therapeutic use of etanercept impaired the induction and progression of experimental EBA. Thus, TNF is likely to serve as a new therapeutic target for EBA and AIBDs with a similar pathogenesis.