Synovial Tissue: Turning the Page to Precision Medicine in Arthritis

Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease targeting the joints. Current treatment strategies are based on clinical, biological and radiological features, yet still fail to reach the goal of early low disease activity in a significant number of cases. Hence, there is a need for refining current treatment algorithms, using accurate markers of response to therapy. Because RA induces histological and molecular alterations in the synovium even before apparition of clinical symptoms, synovial biopsies are a promising tool in the search of such new biomarkers. Histological and molecular characteristics of RA synovitis are heterogeneous. Variations in synovial lining layer hyperplasia, in cellular infiltration of the sublining by immune cells of myeloid and lymphoid lineages, and in molecular triggers of these features are currently categorized using well-defined pathotypes: myeloid, lymphoid, fibroid and pauci-immune. Here, we first bring the plasticity of RA synovitis under scrutiny, i.e., how variations in synovial characteristics are associated with relevant clinical features (disease duration, disease activity, effects of therapies, disease severity). Primary response to a specific drug could be, at least theoretically, related to the representation of the molecular pathway targeted by the drug in the synovium. Alternatively, absence of primary response to a specific agent could be due to disease severity, i.e., overrepresentation of all synovial molecular pathways driving disease activity overwhelming the capacity of any drug to block them. Using this theoretical frame, we will highlight how the findings of previous studies trying to link response to therapy with synovial changes provide promising perspectives on bridging the gap to personalized medicine in RA

Deciphering transcriptomic heterogeneity in rheumatoid arthritis synovium

Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory disease characterized by a predominant joint involvement. RA encompasses a wide spectrum of clinical disease severity spanning from limited, indolent oligoarticular involvement to severe, polyarticular destructive disease. Therapeutic armamentarium has greatly increased these last decades to involve drugs specifically targeting key inflammatory effectors. Again, response to therapy displays a wide heterogeneity across individuals, with some patients entering remission after 1st-line therapy while others suffer from “multi-resistant” RA. Importantly, accurate predictors of response to therapy are lacking. The target tissue of RA -the synovium- is the seat of alterations involving immune and resident cells. Just as clinical characteristics, features of synovial inflammation in RA are widely heterogenous from a cellular and molecular point-of-view. The development of minimally invasive biopsies techniques has enabled to study clinical correlates of synovial tissue heterogeneity in large cohorts of RA patients. While confounding factors (i.e. disease duration, ongoing treatment) may partially account for these variations, it remains unclear whether RA endotypes (commonly named “pathotypes”) can be defined based on synovial inflammatory pattern. The clinical benefit of defining such RA subgroups with different prognosis/response to therapy is evident and would represent a great step toward the grail of precision-medicine. Nevertheless, clinician scientists ought to temper their enthusiasm with a critical approach. In this thesis, I first explored the intra-patient heterogeneity (that is, across pairs of large and small joints) of synovial inflammatory features. I have shown that T cells infiltration and expression of TCR-signaling genes (and other RA-related pathways) are largely similar between pairs of joints from the same individuals. These observations have pathophysiological but also methodological implications: the location of the joint biopsy is not a confounding factor contributing to inter-patient heterogeneity in synovial signals. Then, I analyzed the cellular and molecular effects of T cell co-stimulation blockade therapy (abatacept) on RA synovitis using pre/post treatment biopsies. I took advantage of the large datasets previously acquired by our team to analyze the common transcriptomic effects of 5 drugs with different molecular targets (i.e. B cells, TNF⍺, IL6R). Interestingly, we found a large overlap between the effects of the different therapies: no matter their primary target, they all induced a correlated downregulation of a common set of genes involved in both lymphoid and myeloid cells activation. These observations argue in favor of a common pathogenic mechanism operating in RA synovitis, as opposed to differential (innate vs. adaptive) immune activation. Finally, I analyzed the synovial transcriptome in a large cohort of early, untreated RA patients using unbiased approaches. The main drivers of synovial transcriptomic heterogeneity were genes involved in B and plasma cell biology and RA-related inflammatory process. Interestingly, the synovial expression pattern of these genes strongly correlated with the systemic disease activity. Response to methotrexate therapy (despite several limitations) was mainly associated with baseline disease activity rather than with a specific gene expression pattern. In conclusion, I discuss three (non-exclusive) mechanistic models able to account for our observations on the link between synovial and clinical features in RA. Several questions are raised: Is RA a syndrome encompassing discrete, stable (sub)entities? Does RA synovitis rather represent a continuous spectrum of common mechanisms varying in intensity? Finally, could the differences in pro- and anti-inflammatory cell subtypes present in the synovium of RA patients represent a dynamic process of successive immune-regulatory mechanisms?(MED - Sciences médicales) -- UCL, 202

Synovial Tissue: Turning the Page to Precision Medicine in Arthritis.

Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease targeting the joints. Current treatment strategies are based on clinical, biological and radiological features, yet still fail to reach the goal of early low disease activity in a significant number of cases. Hence, there is a need for refining current treatment algorithms, using accurate markers of response to therapy. Because RA induces histological and molecular alterations in the synovium even before apparition of clinical symptoms, synovial biopsies are a promising tool in the search of such new biomarkers. Histological and molecular characteristics of RA synovitis are heterogeneous. Variations in synovial lining layer hyperplasia, in cellular infiltration of the sublining by immune cells of myeloid and lymphoid lineages, and in molecular triggers of these features are currently categorized using well-defined pathotypes: myeloid, lymphoid, fibroid and pauci-immune. Here, we first bring the plasticity of RA synovitis under scrutiny, i.e., how variations in synovial characteristics are associated with relevant clinical features (disease duration, disease activity, effects of therapies, disease severity). Primary response to a specific drug could be, at least theoretically, related to the representation of the molecular pathway targeted by the drug in the synovium. Alternatively, absence of primary response to a specific agent could be due to disease severity, i.e., overrepresentation of all synovial molecular pathways driving disease activity overwhelming the capacity of any drug to block them. Using this theoretical frame, we will highlight how the findings of previous studies trying to link response to therapy with synovial changes provide promising perspectives on bridging the gap to personalized medicine in RA

Analyse de l’hyperplasie des synoviocytes et de l’infiltration des cellules immunitaires dans la synovie des patients atteints d’arthrite idiopathique juvénile

Analyse de l’hyperplasie des synoviocytes et de l’infiltration des cellules immunitaires dans la synovie des patients atteints d’arthrite idiopathique juvénil

Destructive juvenile idiopathic arthritis: do not overlook rare genetic skeletal disorders

We read with interest the report by Miyamae and colleagues describing severe joint damage, loss of carpal bones and foreshortened fingers in a patient diagnosed with long-standing juvenile idiopathic arthritis (JIA). [..

Comparison of transcriptomic profiles between paired joint biopsies from Rheumatoid Arthritis patients

Authors and affiliations Triaille C1,2, Vansteenkiste L1, Méric de Bellefon L1,3, Nzeusseu Toukap A1,3, Galant C1,4, Durez P1,3, Lauwerys B1,3. 1 : Pôle de pathologies rhumatismales systémiques et inflammatoires, Institut de Recherches Expérimentales Cliniques, Université catholique de Louvain, Brussels, Belgium 2 : Service d’Hématologie et Oncologie pédiatrique, Cliniques Universitaires Saint-Luc, Brussels, Belgium 3 : Service de Rhumatologie, Cliniques Universitaires Saint-Luc, Brussels, Belgium 4 : Service d’Anatomie Pathologique, Cliniques Universitaires Saint-Luc, Brussels, Belgium Background Rheumatoid Arthritis (RA) is a chronic and heterogenous condition characterized by inflammatory involvement of the synovial membrane in multiple joints. Synovial biopsies are used in research setting in order to identify diagnostic and theranostic markers. Many studies have shown a high degree of heterogeneity in histological and transcriptomic profiles between patients

Common Transcriptomic Effects of Abatacept and Other DMARDs on Rheumatoid Arthritis Synovial Tissue

Objectives: Our goal was to assess for the histological and transcriptomic effects of abatacept on RA synovia, and to compare them with previously published data from four other DMARDs: tocilizumab, rituximab, methotrexate, and adalimumab. Methods: Synovial tissue was obtained using ultrasound-guided biopsy from affected joints of 14 patients, before and 16 weeks after treatment with subcutaneous abatacept 125 mg weekly. Paraffin-sections were stained and scored for CD3+, CD20+, and CD68+ cell infiltration. Transcriptional profiling was performed using GeneChip Human Genome U133 Plus 2.0 arrays (Affymetrix) and analyzed on Genespring GX (Agilent). Pathway analyses were performed on Genespring GX, Metascape, and EnrichR. Results: Gene expression analysis identified 304 transcripts modulated by abatacept in synovial tissue. Downregulated genes were significantly enriched for immune processes, strongly overlapping with our findings on other therapies. Data were pooled across these studies, revealing that genes downregulated by DMARDs are significantly enriched for both T-cell and myeloid leukocyte activation pathways. Interestingly, DMARDs seem to have coordinate effects on the two pathways, with a stronger impact in good responders to therapy as compared to moderate and non-responders. Conclusion: We provide evidence that the effects of five DMARDs on the RA synovium culminate in the same pathways. This confirms previous studies suggesting the existence of common mediators downstream of DMARDs, independent of their primary targets

2022 POS0448 SYNOVIAL TRANSCRIPTOMIC PROFILES CORRELATE WITH DISEASE ACTIVITY IN EARLY UNTREATED RHEUMATOID ARTHRITIS

Background Synovitis is the common feature across all individuals with a diagnosis of rheumatoid arthritis (RA). Yet, cellular and transcriptomic alterations occuring in RA synovium are highly variable amongst patients. So far, most data on clinical-tissue correlations either rely on hypothesis-driven approaches or are potentially biased by heterogeneous clinical characteristics (e.g. disease duration or disease-modifying antirheumatic drugs). Objectives We used transcriptomic profiling of synovial tissue from early, untreated rheumatoid arthritis patients (ERA) to 1/ identify the genes with the most variable expression amongst patients and 2/ explore the ability of unbiased (data-driven) approaches to define clinically relevant ERA subgroups. Methods Synovial biopsies were harvested from clinically involved joints of ERA patients using needle arthroscopy or ultrasound-guided biopsy. Data on disease activity were collected at inclusion. For each sample, 350ng total RNA was sent for RNAsequencing using a standardized protocol (Macrogen Europe). After quality control (Fast QC) and genome alignement (HiSat2), normalized read counts were analyzed on Qlucore Omics Explorer. To focus on inter-sample heterogeneity, genes were filtered based on variance (σ/σmax). Unbiased approaches (Principal Component Analysis, Unsupervised Clustering) were applied to define patients’ clusters. Pathway enrichment analysis were performed on Metascape. CibersortX was used to extrapolate the immune cell subsets relative composition from gene expression data. All other statistical analyses were performed on GraphPad Prism v9. Results Total RNA was obtained from synovial biopsies from 74 patients. We first applied variance filtering to identify the genes whose expression showed the greatest variation between patients (n = 894 most variable genes). PCA analysis on the level of expression of these genes did not divide samples into distinct groups, instead yielding a continuous distribution broadly associated with baseline disease activity, as measured by DAS28CRP. Consequently, we used unsupervised clustering to allow for unbiased definition of two patient clusters (PtC): PtC1 (n=52) and PtC2 (n=22) based on their expression of these 894 genes. Pathway analysis of these genes revealed significant enrichment of immune system genes, in the Inflammatory response and Rheumatoid Arthritis pathways (gene cluster 1: GC1), B cell & plasma cell-related pathways (GC2) and metabolic processes-related genes (GC3). Interestingly, PtC1 and PtC2 were characterized by very different clinical features. More specifically, patients from the group with a strong B & plasma cell signature (PtC1) displayed higher baseline indices of all disease activity score components (median DAS28CRP: 5.56 vs 4.09; p-value = 0.0003). They also had higher rates of baseline radiological erosions (erosive disease in 34.6 % vs 10%; p-value = 0.0252) but similar rates of seropositive disease. In line with our pathway analyses, we found a higher signature (inferred relative frequency) of B & plasma cells, T cells and M1-like macrophages in PtC1 compared to PtC2 synovia. PtC2 synovia instead had relatively higher M2-like macrophage and resting mast cell signatures. Conclusion In this large synovial biopsy study, we found that synovial transcriptomic profiles in ERA patients distribute continuously based on the expression of inflammatory and immune cell transcriptomic pathways. These synovial transcriptomic signatures correlate strongly with systemic disease activity

Paired Rheumatoid Arthritis Synovial Biopsies From Small and Large Joints Show Similar Global Transcriptomic Patterns With Enrichment of Private Specificity and TCR Signaling Pathways.

We explored histological and transcriptomic profiles of paired synovial biopsies from rheumatoid arthritis (RA) patients, in order to assess homogeneity in synovial tissue at the individual level. Synovial biopsies were performed simultaneously in one small and one large joint per patient using needle-arthroscopy for the knee and ultrasound-guided biopsy for the hand or wrist. Synovium from individuals with osteoarthritis was used as controls. Paraffin-embedded samples were stained for CD3, CD20, and CD68. Total RNA was hybridized on high-density microarrays. variable sequences were obtained from synovial and blood RNA samples. Twenty paired biopsies from 10 RA patients with active disease were analyzed. Semi-quantification of histological markers showed a positive correlation for synovial hyperplasia, inflammatory infiltrates and CD3-positive T cells between pairs. Pairwise comparison of transcriptomic profiles showed similar expression of RA-related molecular pathways (TCR signaling, T cell costimulation and response to TNFα). T cells clonotypes were enriched in all but one joints compared to blood, regardless of the magnitude of T cell infiltration. Enriched clonotypes were shared between pairs (23-100%), but this was less the case in pairs of joints displaying weaker T cell signatures and more pronounced germinal center-like transcriptomic profiles. Cellular and molecular alterations in RA synovitis are similar between small and large joints from the same patient. Interindividual differences in magnitude of T cell infiltrates and distribution of enriched T cell clonotypes support the concept of distinct synovial pathotypes in RA that are associated with systemic versus local antigen-driven activation of T cells

Disease activity drives transcriptomic heterogeneity in early untreated rheumatoid synovitis.

Transcriptomic profiling of synovial tissue from patients with early, untreated rheumatoid arthritis (RA) was used to explore the ability of unbiased, data-driven approaches to define clinically relevant subgroups. RNASeq was performed on 74 samples, with disease activity data collected at inclusion. Principal components analysis (PCA) and unsupervised clustering were used to define patient clusters based on expression of the most variable genes, followed by pathway analysis and inference of relative abundance of immune cell subsets. Histological assessment and multiplex immunofluorescence (for CD45, CD68, CD206) were performed on paraffin sections. PCA on expression of the (n=894) most variable genes across this series did not divide samples into distinct groups, instead yielding a continuum correlated with baseline disease activity. Two patient clusters (PtC1, n=52; PtC2, n=22) were defined based on expression of these genes. PtC1, with significantly higher disease activity and probability of response to methotrexate therapy, showed upregulation of immune system genes; PtC2 showed upregulation of lipid metabolism genes, described to characterise tissue resident or M2-like macrophages. In keeping with these data, M2-like:M1-like macrophage ratios were inversely correlated with disease activity scores and were associated with lower synovial immune infiltration and the presence of thinner, M2-like macrophage-rich synovial lining layers. In this large series of early, untreated RA, we show that the synovial transcriptome closely mirrors clinical disease activity and correlates with synovial inflammation. Intriguingly, lower inflammation and disease activity are associated with higher ratios of M2:M1 macrophages, particularly striking in the synovial lining layer. This may point to a protective role for tissue resident macrophages in RA