Clinical efficacy of omalizumab in chronic spontaneous urticaria is associated with a reduction of FcεRI-positive cells in the skin

Background. Treatment with omalizumab, a humanized recombinant monoclonal anti-IgE antibody, results in clinical efficacy in patients with Chronic Spontaneous Urticaria (CSU). The mechanism of action of omalizumab in CSU has not been elucidated in detail. Objectives. To determine the effects of omalizumab on levels of high affinity IgE receptor-positive (FcεRI+) and IgE- positive (IgE+) dermal cells and blood basophils. Treatment efficacy and safety were also assessed. Study design. In a double-blind study, CSU patients aged 18‑75 years were randomized to receive 300 mg omalizumab (n=20) or placebo (n=10) subcutaneously every 4 weeks for 12 weeks. Changes in disease activity were assessed by use of the weekly Urticaria Activity Score (UAS7). Circulating IgE levels, basophil numbers and levels of expression of FcεRI+ and IgE+ cells in the skin and in blood basophils were determined. Results. Patients receiving omalizumab showed a significantly greater decrease in UAS7 compared with patients receiving placebo. At Week 12 the mean difference in UAS7 between treatment groups was -14.82 (p=0.0027), consistent with previous studies. Total IgE levels in serum were increased after omalizumab treatment and remained elevated up to Week 12. Free IgE levels decreased after omalizumab treatment. Mean levels of FcεRI+ skin cells in patients treated with omalizumab 300 mg were decreased at Week 12 compared with baseline in the dermis of both non-lesional and lesional skin, reaching levels comparable with those seen in healthy volunteers (HVs). There were no statistically significant changes in mean FcɛRI+ cell levels in the placebo group. Similar results were seen for changes in IgE+ cells, although the changes were not statistically significant. The level of peripheral blood basophils increased immediately after treatment start and returned to Baseline values after the follow-up period. The levels of FcεRI and IgE expression on peripheral blood basophils were rapidly reduced by omalizumab treatment up to Week 12. Conclusions. Treatment with omalizumab resulted in rapid clinical benefits in patients with CSU. Treatment with omalizumab was associated with reduction in FcɛRI+ and IgE+ basophils and intradermal cells

The Involvement of SMILE/TMTC3 in Endoplasmic Reticulum Stress Response

The state of operational tolerance has been detected sporadically in some renal transplanted patients that stopped immunosuppressive drugs, demonstrating that allograft tolerance might exist in humans. Several years ago, a study by Brouard et al. identified a molecular signature of several genes that were significantly differentially expressed in the blood of such patients compared with patients with other clinical situations. The aim of the present study is to analyze the role of one of these molecules over-expressed in the blood of operationally tolerant patients, SMILE or TMTC3, a protein whose function is still unknown.We first confirmed that SMILE mRNA is differentially expressed in the blood of operationally tolerant patients with drug-free long term graft function compared to stable and rejecting patients. Using a yeast two-hybrid approach and a colocalization study by confocal microscopy we furthermore report an interaction of SMILE with PDIA3, a molecule resident in the endoplasmic reticulum (ER). In accordance with this observation, SMILE silencing in HeLa cells correlated with the modulation of several transcripts involved in proteolysis and a decrease in proteasome activity. Finally, SMILE silencing increased HeLa cell sensitivity to the proteasome inhibitor Bortezomib, a drug that induces ER stress via protein overload, and increased transcript expression of a stress response protein, XBP-1, in HeLa cells and keratinocytes.In this study we showed that SMILE is involved in the endoplasmic reticulum stress response, by modulating proteasome activity and XBP-1 transcript expression. This function of SMILE may influence immune cell behavior in the context of transplantation, and the analysis of endoplasmic reticulum stress in transplantation may reveal new pathways of regulation in long-term graft acceptance thereby increasing our understanding of tolerance

Modèles de xéno et d'allo - transplantations chez le babouin

Cette thèse a eu pour but d'explorer deux stratégies des plus innovantes en transplantation : la xénotransplantation (Xt) et l'induction de tolérance en allotransplantation (At), chez le primate, entant que modèle préclinique pertinent. La partie Xt a exploré les différents mécanismes impliqués dans le rejet humoral aigu d'une xénogreffe. Notre modèle animale étudie la xénotransplantation de reins de porcs transgéniques pour les molécules régulant le complément humain (MRCh) chez le babouin. Nous avons montré que les rejets humoreaux aigus, y compris dans un contexte d'utilisation d'organes transgéniques pour les MRCh, s'accompagnaient de dépôts dans les greffons d'IgM et de complément (jusqu'à la phase terminale). Ainsi ce deuxième niveau de rejet xénogénique apparaît plutôt qu'une entité particulière, comme une forme de rejet hyperaigu retardé dans le temps par nos diverses interventions. Ce rejet est associé à une réponse majoritairement de type inné avec un infiltrat cellulaire associé à un environnement proinflammatoire et cytotoxique. Cette réponse peut être diminuée par certains protocoles d'immunosuppression. De façon surprenante, la survie du greffon n'est pas prolongée ni par une immunoadsorption unique avant greffe ni par la Mitoxantrone, nouvel agent immunosuppresseur en xénotransplantation, ciblant les cellules B. Ces données doivent être rééxaminées chez le porc " Knockout " pour l'antigène Gal, cible principale de la réponse humorale. L'objectif de la seconde partie a été d'explorer diverses façons d'induire une tolérance en allotransplantation chez le babouin, en tenant compte des acquis issus des stratégies établies chez le rongeur et in vitro, notamment l'inhibition des voies de costimulation et l'utilisation de cellules dendritiques (CD). Nous avons testé l'inhibition de la costimulation seule et en association avec la Rapamycine, immunosuppresseur facilitant potentiellement l'induction de tolérance chez le rongeur. Cette dernière stratégie s'est révélée uniquement immunosuppressive, et d'autres manœuvres immunologiques doivent être testées afin d'établir un état de tolérance. Nous avons également caractérisé pour la premier fois les CD de babouin, dérivées in vitro à partir de précurseurs de la moelle osseuse ou du sang périphérique. D'autres études in vitro sont nécessaires afin de vérifier la capacité d'induction de tolérance de ces cellules avant de les utiliser in vivo chez le babouin, soit seule, soit en association avec des inhibiteurs des voies de costimulation ou des immunosuppresseurs.The aim of this PhD was to explore the most innovative strategies in the field of transplantation: xenotransplantation (Xt) and tolerance induction in allotransplantation (At), in the non human primate, a relevant preclinical model. The objective of the Xt part was to explore the various immunological mechanisms involved in the acute humoral xenograft rejection of porcine organs transgenic for human complement regulatory molecules (hCRM) by baboons. In our studies acute humoral xenograft rejection appeared to be similar to hyperacute rejection, albeit delayed in time and less aggressive, involving intragraft IgM deposition and complement activation through to the terminal membrane attack complex, even in the case of hCRM-transgenic organs. Rejection was also associated with an innate type response to the xenograft manifest as an cellular infiltrate creating a proinflammatory and cytotoxic environment, which, in some cases, was susceptible to immunosuppression. Surprisingly, survival was not prolonged by a single pretransplant immunoadsorption or by specific B cell-targeted immunosuppression with Mitoxanotrone, a novel agent in xenotransplantation. These issues will now need to be investigated in the case of the recently developed pigs negative for the Gal antigen, the initiator of this so far insurmountable humoral response. The objective of the At part was to explore different ways of inducing transplantation tolerance in the baboon based on strategies already established in vitro and in rodents, namely costimulation blockade and dendritic cells (DC). We firstly tested costimulation blockade alone and in combination with the potentially tolerance facilitating immunosuppressor, Rapamycin, and found this to be immunosuppressive only. This approach will now have to be supplemented with other immunological manoeuvres in order for tolerance to be achieved. We also extensively characterised baboon DC for the first time, derived in vitro from bone marrow and peripheral blood precursors. Further in vitro studies are now necessary to check the tolerance inducing capacity of these DC before they can be used as part of a protocol in vivo in the baboon, either alone, or in combination with costimulation blockade or immunosuppression.NANTES-BU Médecine pharmacie (441092101) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

Étude des biomarqueurs en transplantation et plus particulièrement de Tribbles-1

Le rejet chronique humoral actif (RCHA) est une des dysfonctions chroniques qui aboutit à la perte du greffon. Malheureusement, les traitements actuels ne sont pas efficaces pour traiter ce rejet. L'identification de cette pathologie du greffon ne se fait actuellement que par l'analyse de biopsies associée à une recherche d'anticorps anti-donneur et le diagnostique n'est ainsi fait que lorsque la dysfonction est irréversible. L'identification de nouveaux biomarqueurs non invasifs de ce type de rejet serait d'un grand intérêt afin de définir des molécules pronostiques ou diagnostiques du RCHA. Ainsi, notre étude sur Tribbles-1 a montré que cette molécule était potentiellement un biomarqueur sanguin et tissulaire du RCHA. De plus, une autre étude comparant diverses dysfonctions chroniques a montrée que la molécule Granzyme B est également surexprimée au cours du RCHA dans le sang et le greffon comparé à d'autres dysfonctions chroniques. Ainsi, nous avons identifié plusieurs biomarqueurs du RCHA. En étudiant plus précisément Tribbles-1, nous avons observé que cette molécule est plus fortement exprimée dans les lymphocytes T régulateurs en comparaison des lymphocytes T CD4+CD25- chez les volontaires sains. L'étude plus poussée de Tribbles-1 dans les lymphocytes T régulateurs a montré que cette molécule interagissait avec la molécule qui confère aux Treg leurs capacités suppressives, FoxP3. Ces résultats montrent qu'il existe des biomarqueurs du RCHA et que Tribbles-1 a peut être un rôle à jouer dans cette pathologie via les lymphocytes T régulateurs.Chronic antibody-mediated rejection (CAMR) is an immune chronic dysfunction leading to graft loss. Unfortunately, current treatments are not effective in treating this rejection. The identification of this pathology of the graft is being done by the analysis of biopsies associated with anti-donor antibody and the diagnosis is made only when the dysfunction is irreversible. Thus, the identification of new non-invasive biomarkers of such rejection would be of great interest to define molecular prognostic or diagnostic of RCHA. For this purpose, our study on the Tribbles-1 molecule as shown that it was potentially a blood and graft biomarker of RCHA. In addition, another study comparing various chronic dysfunctions showed that the molecule Granzyme B is also overexpressed in the RCHA in the blood and the graft compared with other chronic dysfunctions. Thus, we identified several biomarkers of RCHA. By studying more precisely Tribbles-1, we observed that this molecule is highly expressed in regulatory T cells compared to CD4+ CD25- T cells of healthy volunteers. The advancement of Tribbles-1 in regulatory T cells has shown that this molecule interacts with the molecule that gives the suppressive capacities of Tregs, FoxP3. These results show that there are biomarkers of RCHA and one of these is Tribbles-1 which could maybe play a role in this disease through regulatory T cells.NANTES-BU Médecine pharmacie (441092101) / SudocSudocFranceF

Les xénogreffes finiront-elles par être acceptées ?

La pénurie d’organes disponibles est responsable d’un nombre croissant de décès de patients en attente de greffe. Dans ces conditions, des solutions alternatives sont envisagées, notamment l’utilisation d’organes porcins. Les xénogreffes sont cependant soumises à différents types de rejet (hyperaigu, vasculaire aigu et cellulaire) dont les mécanismes commencent à être mieux connus. Les stratégies visant à faire de la xénotransplantation une réalité clinique existent et, bien que le nombre de laboratoires engagés dans ce domaine soit encore modeste, des progrès substantiels ont été récemment obtenus. Elles incluent de nouvelles modifications génétiques des animaux donneurs combinées à la mise au point de traitements immunosuppresseurs adaptés à la situation clinique, ainsi que les tentatives d’induction d’une tolérance immunitaire et la gestion du risque sanitaire. L’acceptation des xénogreffes par les receveurs potentiels et son impact sur la société sont également pris en considération.Transplantation represents a major advance in modern medicine with a major impact on the interactions between individuals and society. The numbers of patients undergoing organ transplantation increased steadily over the years and around 250,000 individuals are living nowadays in Europe with a transplanted organ. On the other hand, the numbers of cadaveric (brain-dead) donors used for organ transplantation remains stable, at around 5,000 each year, and the numbers of transplantation from living donors only slowly increase in Europe. Therefore, a gap is growing between the numbers of patients in need of a transplant and the numbers of organs available for transplantation. About 45,000 patients are currently on renal transplant waiting lists in Europe and, depending on the countries considered, 15 to 30 % of candidates for liver or heart transplantation die before a life-saving transplant becomes available to them. There is therefore an urgent need to implement innovative research and to take full advantage of recent biotechnological advances to explore new avenues in xenotransplantation, and to simultaneously address the ethical, societal and public health issues related to organ replacement. Much progresses have been accomplished in the understanding of xenograft rejection processes that include hyperacute, acute vascular and cellular rejection mechanisms. Strategies to promote xenograft survival that are currently under evaluation include genetic engineering of donor pigs, adapted immunosuppressive treatments and tolerance induction. Also, the psychological acceptance has been evaluated

An original approach was used to better evaluate the capacity of a prognostic marker using published survival curves

International audienceObjectives: Predicting chronic disease evolution from a prognostic marker is a key field of research in clinical epidemiology. However, the prognostic capacity of a marker is not systematically evaluated using the appropriate methodology. We proposed the use of simple equations to calculate time-dependent sensitivity and specificity based on published survival curves and other time-dependent indicators as pre-dictive values, likelihood ratios, and posttest probability ratios to reappraise prognostic marker accuracy.Study Design and Setting: The methodology is illustrated by back calculating time-dependent indicators from published articles presenting a marker as highly correlated with the time to event, concluding on the high prognostic capacity of the marker, and presenting the KaplaneMeier survival curves. The tools necessary to run these direct and simple computations are available online at http://www.divat.fr/ en/online-calculators/evalbiom.Results: Our examples illustrate that published conclusions about prognostic marker accuracy may be overoptimistic, thus giving potential for major mistakes in therapeutic decisions.Conclusion: Our approach should help readers better evaluate clinical articles reporting on prognostic markers. Time-dependent sensitivity and specificity inform on the inherent prognostic capacity of a marker for a defined prognostic time. Time-dependent predictive values, likelihood ratios, and posttest probability ratios may additionally contribute to interpret the marker's prognostic capacity