This study suggests that EPIT using Viaskin® is efficient only when applied on intact skin.
In the present study, the immune response generated by Viaskin® appears to be strongly influenced by the condition of the skin. When Viaskin® has been applied on intact skin, the profile of the immune response generated by the treatment is predominantly Th1/Treg whereas in case of Viaskin® applied on stripped skin, it is clearly Th2 oriented. The current work strongly suggests that during EPIT, skin preparation, ie. removing the stratum corneum layer and damaging epidermis, dramatically alters the outcome of treatment and immune reaction.
Skin stripping has never been investigated during the course of EPIT, but has long been considered an enhancing factor of sensitization. Tape-stripping in mice was shown to act as a natural adjuvant. Indeed, according to Strid et al. [10–12] and Spergel et al. , the application of antigen wihout adjuvant on previously stripped skin in naive mice switches the antigen-specific T helper cell responses from Th1-type to Th2-type: epicutaneous immunization on stripped skin converts an established Th1 response (induced by previous subcutaneous injection with adjuvant) into a Th2 response, with a specific reduction of IFN-γ and IgG2a and the enhancement of IL-4 and IgE. In model of food allergy in which mice were sensitized by epicutaneous application of ovalbumin on skin abraded by tape-stripping , mice developed diarrhea and accumulated mast cells in the small intestine, while vast amounts of MMCP-1were released from these cells into the circulation.
In vaccination models, a strong immune reaction was induced by application of antigen on the skin together with an adjuvant [6, 7, 15]. In these models, antigen and adjuvant were applied on the skin previously prepared by gentle removal of the superficial layer of the stratum corneum in order to enhance the transcutaneous passage of both the antigen and adjuvant. The immune responses were clearly Th2 oriented and results consistent with those reported here.
The importance of the state of skin in the profile of immune response on contact with antigen has also been illustrated in humans by Lack et al. , showing that the exposure of skin to peanut proteins may facilitate the sensitization process in very young children, when the normal architecture of the skin is altered by local or generalized eczema [16–18].
In our experiments, EPIT induced on intact skin a major decrease in specific IgE together with a huge increase in specific IgG2a whereas on stripped skin it reinforced specific IgE and did not modify specific IgG2a. The opposite modulation of humoral response was illustrated by the IgG1/IgG2a ratio that slightly decreased with EPIT on intact skin and significantly increased with EPIT on stripped skin. Moreover, at a systemic level, after oral challenge, histamine release was lower when mice were treated by EPIT on intact skin than on stripped skin. At the cellular level, splenocytes of EPIT group secreted lower levels of Th2 cytokines than sensitized and untreated mice.
However, in clinical situations, stripping of the skin appears not to be playing the same “clear-cut” role. Indeed, mouse skin is more sensitive to tape-stripping than human skin. In a recent paper, Senti et al. [8, 19] treated patients allergic to pollen by repeated applications of pollen extract on a previously stripped skin with encouraging results. Despite no improvement in the provocation test, the primary outcome, in the active group versus the control group always showed a significant improvement of the seasonal symptoms (hay fever). The patch was applied for 48 hours on the skin prepared by 6 times tape-stripping.
In this study, we showed that EPIT on stripped skin leads to a free passage of allergens (ie. Ara h 1) into the bloodstream whereas no detectable level is measured when EPIT was applied on intact skin. The kinetics of allergen delivery is different: application on stripped skin induces a passive passage of allergens through the skin into the lymphatics, with high counts in the dermis and numerous allergen-positive cells in draining LNs as early as 2h after application  (paper in preparation). Dendritic cells targeted by the two modes of application of EPIT (intact vs stripped skin) exhibit different phenotypes in term of activation and maturation  (paper in preparation).Taken together - allergen specific capture by DCs through LNs and no detectable level of allergen in bloodstream - these results underline the safety of application only on intact skin. Importantly, these data are consistent with clinical observations. In the human trial, some local adverse events (33% of patients) and systemic allergic reactions required intervention (8% of patients) during the dose-effect study . In children severely allergic to cow’s milk treated by EPIT on intact skin, no severe adverse event was reported .
The application on stripped skin clearly allows the passive and massive passage of allergen through the skin into the lymphatics  completely modifying the biodistribution of allergen and the targeted cells, i.e. less activated Langerhans cells. Tape-stripping also triggers mechanical injuries which activate keratinocytes and upregulate thymic stromal lymphopoietin (TSLP) expression by keratinocytes and mRNA expression of inflammatory cytokines, all of them involved in the polarization of skin DCs to elicit a Th2 response seeing that a link between TSLP expression and the pathogenesis of AD has been shown in several mouse models [22–24]. By contrast, in a recent paper, Li et al. (2012) describe an epicutaneous treatment on intact skin to prevent oral food sensitization in a mouse model . More specifically, the authors showed that high-dose PPE cutaneous application on intact skin is capable of promoting skin local regulatory T-cell responses. At a systemic level, their results showed that the defined exposure of food allergens to intact skin suppresses the subsequent food allergy oral sensitization with suppression of multiple Ig isotypes (IgE, IgG1, IgG2a). Altogether, this greatly suggests that for treatment of Th2 disease, such as immunotherapy of food allergy, avoiding tape-stripping could be of importance, leading to efficacy of EPIT and to maintain the safety of the treatment. The mechanisms involved with the epicutaneous allergen application for the treatment of food allergy are being explored (data submitted for publication) and is likely due to a specific targeting to Langherans cells responsible to antigen presentation to T cells in lymph nodes and Treg expansion .
The model of peanut-sensitized mice exposed to sustained peanut oral regimen in order to induce esophageal and jejunum injuries was recently published . The digestive tract is one of the main organs targeted during food allergies. Based on our previous model developed for the evaluation of new therapeutics in the field of food allergies , we compared the eosinophilic infiltration in mice treated by EPIT on intact or stripped skin. As already published, the decrease of the digestive eosinophil infiltration induced by the ingestion of peanut in sensitized mice treated by EPIT on intact skin was accompanied by a significant decrease of mRNA expression of Th2 cytokines, eotaxin and GATA-3 as well as an increase of FoxP3, underlining the involvement of Tregs in down-regulation of the Th2 pathway. EPIT on stripped skin maintain the high infiltration of eosinophil in jejunum mucosa as well as the villus sub-atrophia, did not induce any increase in mRNA expression of FoxP3 and maintain the high mRNA expression of Th2 cytokines and GATA-3.