Periodic Reporting for period 3 - RELENT (RELapses prevENTion in chronic autoimmune disease: common mechanisms and co-morbidities) Période du rapport: 2018-11-01 au 2020-04-30 Résumé du contexte et des objectifs généraux du projet RELENT (RELapse PrevENTion in autoimmune disease: mechanisms and co-morbidities) is a multidisciplinary consortium that was formed to meet the objectives of Horizon 2020 PHC 3 – 2015, namely to identify mechanisms that were common to a number of different diseases and had important effects on clinical outcome. RELENT’s work focussed on severe autoimmune and inflammatory diseases, including some of the most disabling chronic diseases that can affect individuals of all ages and both sex and can cause severe injury to many organs; like rheumatoid arthritis, inflammatory bowel disease and various forms of vasculitis. Management of these diseases is difficult and typically includes long-term treatment with powerful immunosuppressive and anti-inflammatory drugs and therapeutic monoclonal antibodies. These drugs can be associated with severe side effects and despite recent advances the outlook for many patients with these diseases is worse than for many cancers, many patients become severely disabled or die prematurely either from the diseases themselves or from complications of the drugs. These chronic diseases present a severe challenge for patients throughout Europe as well as for the physicians who care for them and healthcare delivery systems that provide the resources to do so. Currently there is an urgent need for better strategies to tailor drug treatment to the individual needs of patients but this requires deeper knowledge of the immune and inflammatory pathways that cause the injury: this is what the RELENT Consortium was assembled to provide. The RELENT investigators have used a group of related diseases that cause inflammation of blood vessels (vasculitis) to model chronic immune mediated inflammatory diseases more generally. In particular, much of the work focussed on one of the commonest forms of vasculitis, ANCA-associated vasculitis (AAV), so-called because it is caused by autoimmunity to one of two proteins expressed in the cytoplasm of neutrophils (white blood cells), namely myeloperoxidase (MPO) and proteinase-3 (PR3). AAV was chosen because it is relatively homogeneous and sufficiently well understood to provide the foundation for deeper analysis of inflammatory pathways, and because members of the consortium had previously shown it could be used to identify pathogenic pathways that were shared with other diseases. The consortium consisted of 13 Partners (9 from academia and 4 from industry) including geneticists, biochemists, cell biologists and clinical investigators with specific expertise in AAV and related disorders, as well as expert patients with these diseases. The work combined genetic and proteomic analysis of patient cohorts with cellular and animal models of disease separated into four interrelated Work Packages. Travail effectué depuis le début du projet jusqu’à la fin de la période considérée dans le rapport et principaux résultats atteints jusqu’à présent Work Package 1 combined subset analysis of genome wide association studies (GWAS) with classical cell biology to uncover pathways that influence the outcome of AAV and its response to treatment. New genetic variants that confer susceptibility to AAV were identified in a prognosis-oriented GWAS of 2500 cases. This has produced several suggestive signals, although none yet with genome-wide levels of significance, and so the analysis is now being extended to an additional 700 genotyped DNA samples. Inflammatory pathways activated in inflamed tissue have been examined in unprecedented detail by digital spatial profiling of patients’ renal biopsies. Comparison of transcriptomic and proteomic patterns in different glomeruli in the same (or different) kidney biopsies revealed distinct gene expression signatures identified transcripts of genes not previously known to be involved in glomerular inflammation. Cellular pathways have been analysed in induced pluripotent stem cells (iPSC) generated from patients with vasculitis and other individuals with defined genetic background that have been differentiated into subsets of white blood cells and into glomerular epithelial cells. These are used to dissect mechanism of autophagy with specific inhibitory molecules developed during the project. Work Package 2 examined the proteome of patients’ blood to identify protein signatures that predict organ distribution and outcome in vasculitis. Systematic autoantibody profiling using multiplexed antigen arrays identified novel autoantibodies that are associated with renal disease in AAV and involvement of the eye in giant cell arteritis (GCA). A more generalised biomarker search identified a previously unrecognised interferon signature in anti-MPO AAV with implications for pathogenesis and disease monitoring. Proteomic analysis of the MPO revealed profound and unexpected post translational modification that may imply a novel repurposing of the lysosome trafficking pathway in neutrophils with implications for antigenicity. Work Package 3 characterised T and B cell abnormalities that develop in the ageing immune system that could underlie the increased susceptibility to autoimmune disease and their complications in the elderly. We identified age-associated distortions in the frequencies of T cells expressing activation markers and checkpoint molecules that inhibit immune responses. Analogous changes were found in aged mice, as well as fewer regulatory T cells. Induction of autoimmunity to MPO in such mice was associated with greater expression of co-stimulatory molecules on dendritic cells and more severe injury, albeit with lower concentrations of autoantibodies. This analysis of differential expression of checkpoint molecules in immune cells suggests that age-related changes in their expression and function may lead to an unstable immune system, which is prone to autoimmunity. Work Package 4 analysed pathogenic effector T cells and their control by macrophages and dendritic cells in normal and transgenic mice that express relevant human proteins. The development and characterisation of a new murine model of anti-MPO AAV that better reflects the relapsing nature of the human disease allowed to characterise renal and lung injury in AAV. The studies revealed unexpected cross-reactivity between autoantibodies which provides a potential explanation for the interferon signal identified in the patients with anti-MPO AAV. They also provided important insights into which cells and pathways are potential targets for therapy and those that are not. It defined consequences of elevated levels of the protease cathepsin S in increasing vascular permeability and endothelial injury providing important insights into clinical vasculitis and its role as novel potential therapeutic targets. Progrès au-delà de l’état des connaissances et impact potentiel prévu (y compris l’impact socio-économique et les conséquences sociétales plus larges du projet jusqu’à présent) The comprehensive approach of RELENT research of dissecting the role of the individual components of the immune system has led to better understand how they interact in concert to mediate the autoimmune response in young and elderly people and in those with autoimmune diseases. The pathway discovery has led to measures for better disease monitoring and the development of tools and biomarkers by RELENT’s SME partners that allowed translation into clinically applicable assays. The results have been communicated through RELENT’s web page, a periodic newsletter, social media, in 66 international conferences, 58 scientific papers and to the broader public, most notably in a comprehensive review on AAV. Thus, the research of RELENT will change our current understanding of autoimmunity and broaden our tools to investigate and monitor them.