Clonally expanded CD38hi cytotoxic CD8 T cells define the T cell infiltrate in checkpoint inhibitor-associated arthritis.

Immune checkpoint inhibitor (ICI) therapies used to treat cancer, such as anti-PD-1 antibodies, can induce autoimmune conditions in some individuals. The T cell mechanisms mediating such iatrogenic autoimmunity and their overlap with spontaneous autoimmune diseases remain unclear. Here, we compared T cells from the joints of 20 patients with an inflammatory arthritis induced by ICI therapy (ICI-arthritis) with two archetypal autoimmune arthritides, rheumatoid arthritis (RA) and psoriatic arthritis (PsA). Single-cell transcriptomic and antigen receptor repertoire analyses highlighted clonal expansion of an activated effector CD8 T cell population in the joints and blood of patients with ICI-arthritis. These cells were identified as CD38hiCD127- CD8 T cells and were uniquely enriched in ICI-arthritis joints compared with RA and PsA and also displayed an elevated interferon signature. In vitro, type I interferon induced CD8 T cells to acquire the ICI-associated CD38hi phenotype and enhanced cytotoxic function. In a cohort of patients with advanced melanoma, ICI therapy markedly expanded circulating CD38hiCD127- T cells, which were frequently bound by the therapeutic anti-PD-1 drug. In patients with ICI-arthritis, drug-bound CD8 T cells in circulation showed marked clonal overlap with drug-bound CD8 T cells from synovial fluid. These results suggest that ICI therapy directly targets CD8 T cells in patients who develop ICI-arthritis and induces an autoimmune pathology that is distinct from prototypical spontaneous autoimmune arthritides.

Co-Inhibitory Molecules - Their Role in Health and Autoimmunity; Highlighted by Immune Related Adverse Events.

Immune checkpoint receptors are key players in regulating the immune response. They are responsible for both generating an immune response sufficient to kill invading pathogens, balancing the same response, and protecting against tissue destruction or the development of autoimmune events. The central role of the co-inhibitory receptors also referred to as inhibitory immune checkpoints, including PD-1 and CTLA-4 has become especially evident with the cancer treatments targeting these receptors. Blocking these pathways enhances the immune activity, resulting in both an increased chance of cancer clearance, at the same time induction of immune-related adverse events (irAE). Some of these irAE progress into actual autoimmune diseases with autoantibodies and symptoms, undistinguished from the naturally occurring diseases. This review will take advantage of the lessons learned from immune checkpoint blockade and relate this knowledge to our understanding of the same pathways in naturally occurring autoimmune diseases, mainly focusing on rheumatic diseases.

The Programmed Death-1 Pathway Counter-Regulates Inflammation-Induced Osteoclast Activity in Clinical and Experimental Settings.

Objective: The programmed death-1 (PD-1) pathway is essential for maintaining self-tolerance and plays an important role in autoimmunity, including rheumatoid arthritis (RA). Here, we investigated how membrane-bound and soluble (s)PD-1 influence bone homeostasis during chronic inflammation, exemplified in RA. Methods: Bone mineral density and bone microstructure were examined in PD-1 and PD-L1 knockout (KO) mice and compared with wild-type (WT) mice. Receptor activator of nuclear factor kappa-B ligand (RANKL) was measured in serum, and the expression examined on activated bone marrow cells. Osteoclast formation was examined in cells from murine spleen and bone marrow and from human synovial fluid cells. sPD-1 was measured in chronic and early (e)RA patients and correlated to markers of disease activity and radiographic scores. Results: PD-1 and PD-L1 KO mice showed signs of osteoporosis. This was supported by a significantly reduced trabecular bone volume fraction and deteriorated microstructure, as well as increased osteoclast formation and an increased RANKL/OPG ratio. The recombinant form of sPD-1 decreased osteoclast formation in vitro, but was closely associated with disease activity markers in eRA patients. Sustained elevated sPD-1 levels indicated ongoing inflammation and were associated with increased radiographic progression. Conclusion: The PD-1 pathway is closely associated with bone homeostasis, and lacking members of this pathway causes a deteriorated bone structure. The immunological balance in the microenvironment determines how the PD-1 pathway regulates osteoclast formation. In eRA patients, sPD-1 may serve as a biomarker, reflecting residual but clinically silent disease activity and radiographic progression.

CXCL13 predicts long-term radiographic status in early rheumatoid arthritis.

OBJECTIVES: Identification of RA patients at a high risk of joint destruction remains challenging. The C-X-C motif chemokine 13 (CXCL13) has previously been suggested as a marker of disease activity in RA. Here, we investigate the potential of plasma CXCL13 as a marker of long-term radiographic status and progression. METHODS: CXCL13 was measured in plasma from treatment-naïve RA patients (n = 158) with an 11-year follow-up. At baseline, clinical and biochemical DASs were obtained; among these CRP, ESR, DAS in 28 joints with CRP (DAS28CRP), number of swollen joints (SJC28) and radiographic status, evaluated by total Sharp score (TSS). Age- and gender-matched healthy controls (HCs) were included. RESULTS: CXCL13 was significantly increased at baseline and decreased during treatment; however, it was not reduced to the level in HCs. At baseline, CXCL13 was associated with both CRP and ESR, but not with other markers of disease activity. Baseline CXCL13 was correlated with both TSS and radiographic progression (ΔTSS) at 11 years. With an 89% probability, levels of CXCL13 above 85 pg/ml predicted the risk of a TSS of 5 or above, after 11 years of treatment. Compared with CRP, DAS28CRP, SJC28 and ACPA status, CXCL13 was superior in predicting 11-year joint destruction. CONCLUSION: In early RA, one single measurement of plasma CXCL13 at baseline is superior to currently used clinical and serological disease markers in the prediction of long-term radiographic status and progression.

Programmed death ligand 2 - A link between inflammation and bone loss in rheumatoid arthritis.

OBJECTIVE: Active rheumatoid arthritis (RA) is accompanied by increased appendicular and axial bone loss, closely associated to the degree of inflammation. The programmed death-1 (PD-1) pathway is important for maintaining peripheral tolerance, and its ligand PD-L2 has recently been associated with bone morphogenetic protein activity. Here, we report that PD-L2 plays a central role in RA osteoimmunology. METHODS: Femoral bone mineral density (BMD) and trabecular bone microstructure were evaluated by micro-CT in wild type (WT) and PD-L2-/- mice. Osteoclasts were generated from RA synovial fluid mononuclear cells and peripheral blood monocytes. The effects of recombinant PD-L2, was evaluated by tartrate-resistant acid phosphatase (TRAP) activity and the development of bone erosions in the presence of anti-citrullinated protein antibodies (ACPA). Plasma soluble (s)PD-L2 levels were measured in patients with early (e)RA (n â€‹= â€‹103) treated with methotrexate alone or in combination with the TNF inhibitor Adalimumab. RESULTS: PD-L2-/- mice had a decreased BMD and deteriorated trabecular bone microstructure that was not related to the RANKL/OPG pathway. PD-L2 decreased TRAP activity in osteoclasts and decreased ACPA-induced erosions. In the RA synovial membrane PD-L2 was highly expressed especially in the lining layer and plasma sPD-L2 levels were increased in eRA patients and decreased with treatment. One-year sPD-L2 correlated inversely with erosive progression two years after treatment initiation with methotrexate and placebo. CONCLUSION: PD-L2 regulates bone homeostasis in RA. Our findings provide new insight into the relationship between the immune system and bone homeostasis, and suggest a potential therapeutic target for limiting inflammatory bone loss in RA.

Extracellular Vesicles Transfer the Receptor Programmed Death-1 in Rheumatoid Arthritis.

INTRODUCTION: Extracellular vesicles (EVs) have been recognized as route of communication in the microenvironment. They transfer proteins and microRNAs (miRNAs) between cells, and possess immunoregulatory properties. However, their role in immune-mediated diseases remains to be elucidated. We hypothesized a role for EVs in the rheumatoid arthritis (RA) joint, potentially involving the development of T cell exhaustion and transfer of the co-inhibitory receptor programmed death 1 (PD-1). METHODS: Synovial fluid mononuclear cells (SFMCs) and peripheral blood mononuclear cells (PBMCs) from RA patients were investigated for PD-1 and other markers of T cell inhibition. EVs were isolated from RA plasma and synovial fluid. In addition, healthy control (HC) and RA PBMCs and SFMCs were cultured to produce EVs. These were isolated and investigated by immunogold electron microscopy (EM) and also co-cultured with lymphocytes and PD-1 negative cells to investigate their functions. Finally, the miRNA expression profiles were assessed in EVs isolated from RA and HC cell cultures. RESULTS: Cells from the RA joint expressed several T cell co-inhibitory receptors, including PD-1, TIM-3, and Tigit. ELISA demonstrated the presence of PD-1 in EVs from RA plasma and synovial fluid. Immunogold EM visualized PD-1 expression by EVs. Co-culturing lymphocytes and the PD-1 negative cell line, U937 with EVs resulted in an induction of PD-1 on these cells. Moreover, EVs from RA PBMCs increased proliferation in lymphocytes when co-cultured with these. All EVs contained miRNAs associated with PD-1 and other markers of T cell inhibition and the content was significantly lower in EVs from RA PBMCs than HC PBMCs. Stimulation of the cells increased the miRNA expression. However, EVs isolated from stimulated RA SFMCs did not change their miRNA expression profile to the same extend. CONCLUSION: EVs carrying both the PD-1 receptor and miRNAs associated with T cell inhibition were present in RA cell cultures. Upon stimulation, these miRNAs failed to be upregulated in EVs from RA SFMCs. This was in line with increased expression of T cell co-inhibitory markers on SFMCs. In conclusion, we suggest EVs to play a significant role in the RA microenvironment, potentially favoring the progression of T cell exhaustion.

Soluble programmed death-1 levels are associated with disease activity and treatment response in patients with autoimmune hepatitis.

PURPOSE: Autoimmune hepatitis (AIH) is a chronic liver disease caused by impaired immune regulation. Programmed death-1 (PD-1) is an inhibitory receptor mainly expressed by T cells and with its ligands, PD-L1 and PD-L2 present on antigen-presenting cells. We hypothesised the PD-1 axis to be impaired in AIH and investigated systemic levels of soluble(s) PD-1 and T cells ability to up-regulate PD-1 following in vitro activation in AIH patients. MATERIALS AND METHODS: We included 67 AIH patients; 9 with active disease, 31 responders and 27 incomplete-responders to standard therapy. Forty-seven healthy controls (HC) were included for comparison. Soluble PD-1 was measured by enzyme-linked immunosorbent assay. The PD-1 expression on T cells was measured using flow cytometry before and after 48-h stimulation in vitro with CD3/CD28 in 13 AIH patients and 10 HC. RESULTS: Soluble PD-1 was significantly elevated in AIH patients with active disease [0.24 ng/mL (range 0.16-0.28)] and in incomplete responders to standard therapy [0.17 (0.11-0.22)] compared with responders [0.11 (0.08-0.16), p = .008 and p = .01, respectively] and HC [0.12 (0.05-0.16), p = .02, both]. Following in vitro activation, PD-1 was significantly up-regulated (3.3-fold) on CD4+ T cells from AIH patients compared with HC (1.5-fold) (p = .0006). CONCLUSIONS: AIH patients with active disease and incomplete response to standard treatment have similarly increased sPD-1 levels. Further, AIH patients have increased ability to up-regulate PD-1 following in vitro activation. Together these data suggests an impaired PD-1 axis in AIH.

Spontaneous generation of functional osteoclasts from synovial fluid mononuclear cells as a model of inflammatory osteoclastogenesis.

In osteoimmunology, osteoclastogenesis is understood in the context of the immune system. Today, the in vitro model for osteoclastogenesis necessitates the addition of recombinant human receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). The peripheral joints of patients with rheumatoid arthritis (RA) and spondyloarthritis (SpA) are characterized by an immune-mediated inflammation that can lead to bone destruction. Here, we evaluate spontaneous in vitro osteoclastogenesis in cultures of synovial fluid mononuclear cells (SFMCs) activated only in vivo. SFMCs were isolated and cultured for 21 days at 0.5-1.0 × 10(6) cells/mL in culture medium. SFMCs and healthy control peripheral blood monocytes were cultured with RANKL and M-CSF as controls. Tartrate-resistant acid phosphatase (TRAP) positive multinucleated cells were found in the SFMC cultures after 21 days. These cells expressed the osteoclast genes calcitonin receptor, cathepsin K, and integrin ß3, formed lacunae on dentin plates and secreted matrix metalloproteinase 9 (MMP9) and TRAP. Adding RANKL and M-CSF potentiated this secretion. In conclusion, we show that SFMCs from inflamed peripheral joints can spontaneously develop into functionally active osteoclasts ex vivo. Our study provides a simple in vitro model for studying inflammatory osteoclastogenesis.