IL7 genetic variation and toxicity to immune checkpoint blockade in patients with melanoma

Treatment with immune checkpoint blockade (ICB) frequently triggers immune-related adverse events (irAEs), causing considerable morbidity. In 214 patients receiving ICB for melanoma, we observed increased severe irAE risk in minor allele carriers of rs16906115, intronic to IL7. We found that rs16906115 forms a B cell-specific expression quantitative trait locus (eQTL) to IL7 in patients. Patients carrying the risk allele demonstrate increased pre-treatment B cell IL7 expression, which independently associates with irAE risk, divergent immunoglobulin expression and more B cell receptor mutations. Consistent with the role of IL-7 in T cell development, risk allele carriers have distinct ICB-induced CD8+ T cell subset responses, skewing of T cell clonality and greater proportional repertoire occupancy by large clones. Finally, analysis of TCGA data suggests that risk allele carriers independently have improved melanoma survival. These observations highlight key roles for B cells and IL-7 in both ICB response and toxicity and clinical outcomes in melanoma

Mécanismes pathogéniques des maladies inflammatoires chroniques : relier génotype, fonctions immunitaires et pathologie dans les spondylarthrites

Spondyloarthritis (SpA) is a family of related chronic inflammatory diseases (CID), and is characterised by inflammation of the spine and sacroiliac joints, the hallmarks of the axial form of this disease (AxSpA). Genome-wide association studies (GWAS) have identified genetic loci associated to SpA disease susceptibility. GWAS also have provided evidence of a key role for immune signalling pathways in the pathogenesis of SpA, as many of the identified loci map to immune genes. In particular, GWAS have suggested a role for the interleukin-23/interleukin-17 (IL-23/IL-17) axis in the pathogenesis of several CIDs. However, for most associated loci, the mechanism by which they affect pathogenesis and the immune cell populations in which they act are still not known. The goal of this project is to understand the molecular mechanisms of the disease pathogenesis by studying immune cell function and response to biologics in SpA.To understand the role of susceptibility loci in CIDs we designed an nCounter® gene expression panel (NanoString Technologies): the Autoimmune Discovery Consortium Panel. This panel was created utilising the reported loci from GWAS in 9 CIDs, with the addition of several immune genes, such as cytokine and chemokine genes. We used this panel to determine the expression pattern of “GWAS genes” in immune cell populations isolated from 50 axial SpA (AxSpA) patients. The cell-type-specific gene expression profiles were then correlated with the patient genotype to identify expression quantitative trait loci (eQTLs). Gene expression analysis showed that ~80% of genes associated with CIDs were expressed in T cell subsets (CD4+ and CD8+, under resting and activated states), implicating their role in disease. The eQTL study revealed a genomic region on chromosome 11, including SNPs that affect the expression of cathepsin W (CTSW), a lysosomal peptidase implicated in cytotoxic activity in CD8+ T cells. Furthermore, we demonstrated that transcription factor NFATC2 (Nuclear factor of activated T-cells, cytoplasmic 2) binds in an allele-specific manner to the eQTL rs12225345, supporting its role in CTSW regulation in T cells. These data illustrate how allele-specific binding of transcription factors could contribute to the regulation of disease-associated genes, and may play a role in the pathogenesis of the disease.In the second part of this project, we studied the molecular mechanism of AxSpA pathogenesis and the impact of biologics (TNF inhibitors, TNFi) on immune responses of AxSpA patients. To study the role for the IL-23/IL-17 axis in the pathogenesis of AxSpA, we have characterised the immune cells that produce IL-17A in AxSpA patients. We compared the IL-17 production capacity of cell subsets of the innate (MAIT, γδT, and neutrophils) and adaptive (CD4+ and CD8+ T-cells) arms of the immune system. We identified MAIT cells as the main producers of IL-17 in AxSpA. We also observed that both innate and adaptive lymphocytes express genes belonging to the IL-23/IL-17 pathway and genes previously associated with SpA susceptibility. To understand the mechanism of action of TNF blockade in axSpA patients, we investigated the effect of TNFi on immune responses to microbial and pathway-related stimuli. We demonstrated that anti-TNF therapy induced profound changes in patients’ innate immune responses, but has minor effects on Th1/Th17 immunity. Additionally, we observed that TNFi affect the NFκB transcriptional network – an important regulatory network for innate immune response genes. We also reported that TNFi steers monocytes/macrophages towards an M2-like profile, which may be an important factor in the regulation of inflammatory responses.La spondylarthrite (SpA) est une famille de maladies inflammatoires chroniques apparentées (CID), et se caractérise par une inflammation de la colonne vertébrale et des articulations sacro-iliaques, caractéristiques de la forme axiale de cette maladie (AxSpA). Des études d'association pangénomique (GWAS) ont identifié des locus génétiques associés à la susceptibilité à la maladie de la SpA. GWAS a également fourni la preuve d'un rôle clé pour les voies de signalisation immunitaire dans la pathogenèse de la SpA, car de nombreux loci identifiés sont associés à des gènes immunitaires. En particulier, les GWAS ont suggéré un rôle pour l'axe interleukine-23 / interleukine-17 (IL-23 / IL-17) dans la pathogenèse de plusieurs CID. Cependant, pour la plupart des loci associés, le mécanisme par lequel ils affectent la pathogenèse et les populations de cellules immunitaires dans lesquelles ils agissent ne sont toujours pas connus. L’objectif de ce projet est de comprendre les mécanismes moléculaires de la pathogenèse de la maladie en étudiant la fonction des cellules immunitaires et la réponse aux traitement biologique dans la SpA.Pour comprendre le rôle des loci de susceptibilité dans les CID, nous avons conçu un panel d'expression génique nCounter® (NanoString Technologies): le panel Autoimmune Discovery Consortium. Ce panel a été créé en utilisant les loci rapportés de GWAS dans 9 CID, avec l'ajout de plusieurs gènes immunitaires, tels que les gènes de cytokine et de chimiokine. Nous avons utilisé ce panel pour déterminer le modèle d'expression des «gènes GWAS» dans les populations de cellules immunitaires isolées de 50 patients SpA axiaux (AxSpA). Les profils d'expression génique spécifiques au type de cellule ont ensuite été corrélés avec le génotype du patient pour identifier les locus de caractères quantitatifs d'expression (eQTL). L'analyse de l'expression génique a montré qu'environ 80% des gènes associés aux CID étaient exprimés dans des sous-ensembles de cellules T (CD4+ et CD8+, à l'état de repos et activé), ce qui implique leur rôle dans la maladie. L'étude eQTL a révélé un cluster sur le chromosome 11, y compris des SNP qui affectent l'expression de la CTSW (cathepsin W), une peptidase lysosomale impliquée dans l'activité cytotoxique des cellules T CD8+. En outre, nous avons démontré que le facteur de transcription NFATC2 (Nuclear factor of activated T-cells, cytoplasmic 2) se lie de manière allélique spécifique à l'eQTL rs12225345, soutenant son rôle dans la régulation CTSW dans les lymphocytes T. Ces données illustrent comment la liaison allélique spécifique des facteurs de transcription pourrait contribuer à la régulation des gènes associés à la maladie et jouer un rôle dans la pathogenèse de la maladie.Dans la deuxième partie de ce projet, nous avons étudié le mécanisme moléculaire de la pathogenèse AxSpA et l'impact des traitement biologiques (inhibiteurs du TNF, TNFi) sur les réponses immunitaires des patients axSpA. Pour étudier le rôle de l'axe IL-23 / IL-17 dans la pathogenèse de AxSpA, nous avons caractérisé les cellules immunitaires qui produisent l'IL-17A chez les patients AxSpA. Nous avons comparé la capacité de production d'IL-17 de sous-ensembles cellulaires des bras innés (MAIT, γδT et polynucléaires neutrophiles) et adaptatifs (cellules T CD4+ et CD8+) du système immunitaire. Nous avons identifié les cellules MAIT comme les principaux producteurs d'IL-17 dans AxSpA. Nous avons également observé que les lymphocytes innés et adaptatifs expriment des gènes appartenant à la voie IL-23 / IL-17 et des gènes précédemment associés à la sensibilité SpA

T cells in the pathogenesis of axial spondyloarthritis

International audienceAxial spondyloarthritis (axSpA) is the prototype of the spondyloarthritis spectrum. The involvement of T cells in its pathogenesis has long been suspected on the basis of the association with the major histocompatibility complex I molecule HLA-B27 and the pivotal role of interleukin 17 in the inflammatory mechanisms associated with the disease. Moreover, the presence of unconventional or "innate-like" T cells within the axial enthesis suggests an important role for these cells in the pathophysiology of the disease. In this review, we describe the characteristics and the interleukin 17 secretion capacity of the T-cell subsets identified in axSpA. We discuss the genetic and epigenetic mechanisms that support the alteration of T-cell functions and promote their activation in axSpA. We also discuss recent data on T cells that could explain the extra-articular manifestations of the SpA spectrum

Characterization of Blood Mucosal Associated Invariant T ( MAIT ) cells in Axial Spondyloarthritis and of resident MAITs from control axial enthesis

International audienceObjectivesThe importance of IL-17A in the pathogenesis of Axial Spondyloarthritis (AxSpA) has been demonstrated by the success of IL-17A blockade. However, the nature of the cell populations producing this important pro-inflammatory cytokine remains poorly defined. Accordingly, we characterized major IL-17A-producing blood cell populations in AxSpA, with reference to entheseal biology, with a focus on mucosal associated invariant T-cells, (MAITs) a population known to be capable of producing IL-17.MethodsIL-17A production from 5 sorted peripheral blood cell populations, namely MAITs, γδ T-cells, CD4+T-cells, CD8+T-cells and neutrophils was evaluated pre- and post-stimulation with PMA, the calcium ionophore A23187 and β1,3 glucan. IL-17A transcript and protein expression were determined using nCounter Technology and ultra-sensitive SimoA technology, respectively. MAITs from control human axial entheses (n=5) were further characterized by flow cytometric immunophenotyping, qPCR and IL-17 production assessed following stimulation.ResultsIn blood, on a per cell basis, MAIT cells produced the highest amount of IL-17A compared to CD4+ (p <0.01), CD8+ (p <0.0001) and γδ T-cells (p <0.0001). IL-17A was not produced by neutrophils. Gene expression analysis also showed significantly higher IL17A and IL23R expression in MAIT cells. Stimulation of blood MAIT cells with αCD3/CD28 and IL-7 and/or IL-18 induced strong expression of IL17F. MAITs were present in the normal enthesis and showed elevated AHR, JAK1, STAT4 and TGFB1 transcript expression with inducible IL-17A protein. IL-18 protein expression was evident in spinal enthesis digests.ConclusionBoth blood MAITs and resident MAITs in axial entheses contribute to IL-17 production and may play important roles in AxSpA pathogenesis

Natural Killer cells demonstrate distinct eQTL and transcriptome-wide disease associations, highlighting their role in autoimmunity

Natural Killer cells are innate lymphocytes with central roles in immunosurveillance and are implicated in autoimmune pathogenesis. The degree to which regulatory variants affect Natural Killer cell gene expression is poorly understood. Here we perform expression quantitative trait locus mapping of negatively selected Natural Killer cells from a population of healthy Europeans (n = 245). We find a significant subset of genes demonstrate expression quantitative trait loci specific to Natural Killer cells and these are highly informative of human disease, in particular autoimmunity. A Natural Killer cell transcriptome-wide association study across five common autoimmune diseases identifies further novel associations at 27 genes. In addition to these cis observations, we find novel master-regulatory regions impacting expression of trans gene networks at regions including 19q13.4, the Killer cell Immunoglobulin-like Receptor region, GNLY, MC1R and UVSSA. Our findings provide new insights into the unique biology of Natural Killer cells, demonstrating markedly different expression quantitative trait loci from other immune cells, with implications for disease mechanisms

Immune response profiling of patients with spondyloarthritis reveals signalling networks mediating TNF-blocker function in vivo

International audienceObjectives Antitumour necrosis factor (TNF) therapy has revolutionised treatment of several chronic inflammatory diseases, including spondyloarthritis (SpA). However, TNF inhibitors (TNFi) are not effective in all patients and the biological basis for treatment failure remains unknown. We have analysed induced immune responses to define the mechanism of action of TNF blockers in SpA and to identify immunological correlates of responsiveness to TNFi. Methods Immune responses to microbial and pathwayspecific stimuli were analysed in peripheral blood samples from 80 patients with axial SpA before and after TNFi treatment, using highly standardised wholeblood stimulation assays. Cytokines and chemokines were measured in a Clinical Laboratory Improvement Amendments (CLIA)-certified laboratory, and gene expression was monitored using nCounter assays. Results Anti-TNF therapy induced profound changes in patients' innate immune responses. TNFi action was selective, and had only minor effects on Th1/Th17 immunity. Modular transcriptional repertoire analysis identified prostaglandin E 2 synthesis and signalling, leucocyte recirculation, macrophage polarisation, dectin and interleukin (IL)-1 signalling, as well as the nuclear factor kappa B (NF-kB) transcription factor family as key pathways targeted by TNF blockers in vivo. Analysis of induced immune responses before treatment initiation revealed that expression of molecules associated with leucocyte adhesion and invasion, chemotaxis and IL-1 signalling are correlated with therapeutic responses to anti-TNF. Conclusions We show that TNFi target multiple immune cell pathways that cooperate to resolve inflammation. We propose that immune response profiling provides new insight into the biology of TNFblocker action in patients and can identify signalling pathways associated with therapeutic responses to biological therapies