Rheumatoid Arthritis: Isaacs JD

Wise EM, Isaacs JD. · No affiliation provided · Rheumatology (Oxford). · Pubmed #16105914 links to  free full text

This publication has no abstract.

Pratt AG, Isaacs JD, Mattey DL. · Musculoskeletal Research Group, Institute for Cellular Medicine, School of Clinical Medical Sciences, Newcastle University, Newcastle, UK. · Best Pract Res Clin Rheumatol. · Pubmed #19233044 links to  free full text

Abstract: Rheumatoid arthritis (RA) is a systemic inflammatory disease with a predilection for symmetrically distributed diarthroidal joints. It is clinically heterogeneous, with particular disease phenotypes defined according to a complex interplay of genes and the environment. In this chapter we first summarize current knowledge of RA genetic susceptibility, a field which has been transformed in recent years by powerful modern genotyping technologies. The importance of a recently described subclassification for the disease based upon the presence or absence of circulating autoantibodies to citrullinated peptides has further informed genetic studies, and we consider the implications for our understanding of RA pathogenesis. We then review the cellular and molecular processes that initiate and perpetuate joint destruction.

Isaacs JD. · Musculoskeletal Research Group and Wilson Horne Immunotherapy Centre, Institute of Cellular Medicine, Newcastle University, Newcastle-upon-Tyne, UK. · Rheumatology (Oxford). · Pubmed #18503092 No free full text.

Abstract: Accumulating evidence suggests that RA is a T-cell-mediated autoimmune disease. Early attempts at disease modulation using strategies such as CD4 mAbs were severely hampered by a lack of biomarkers of autoreactivity. Recently, however, co-stimulation blockade has emerged as an effective treatment for RA. Alongside a greatly improved mechanistic understanding of immune regulation, this has rekindled hopes for authentic and robust immune programming. The final pieces of the jigsaw are not yet in place for RA but, in other disciplines, emerging treatment paradigms such as non-mitogenic anti-CD3 mAbs, autoantigenic peptides and even cellular therapies are providing hope for a future in which immunopathology can be specifically and vigorously curtailed.

Anderson AE, Isaacs JD. · Musculoskeletal Research Group Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK. · Acta Reumatol Port. · Pubmed #18344919 links to  free full text

Abstract: Regulatory T cells (Tregs) are a subset of T cells which are involved in peripheral immune tolerance. Their role in autoimmune disease which occurs through a breakdown of tolerance is of particular interest in trying to ascertain the mechanism(s) of disease progression. It is hoped that by understanding the role of Tregs in autoimmunity a reliable therapy may be developed to aid in both the treatment and potentially cure of disease. This review will focus on the naturally-occurring CD4+ CD25+ regulatory T cell subset and their possible involvement in rheumatoid arthritis.

Isaacs JD. · Wilson Horne Immunotherapy Centre and Musculoskeletal Research Group, Institute of Cellular Medicine, Catherine Cookson Building, Framlington Place, Newcastle-upon-Tyne NE2 4HH, United Kingdom. · Curr Opin Pharmacol. · Pubmed #17611158 No free full text.

Abstract: The concept and practice of therapeutic tolerance has successfully been applied to animal models of autoimmunity and transplantation for more than 2 decades. Finally, there are encouraging signs of its translation to clinical practice. Short courses of anti-CD3 monoclonal antibody therapy have provided lasting benefits in recent-onset type 1 diabetes in association with evidence for the induction of immunoregulatory mechanisms. Co-stimulation blockade with abatacept (CTLA4-Ig) will soon be licensed for the treatment of rheumatoid arthritis - over the past year phase III studies have demonstrated impressive improvement in subjective and objective signs of the disease. T cell depletion is in development for several conditions, again with recent studies demonstrating evidence of immune regulation in some instances. More specific antigen-directed peptide therapies have also been applied to atopic asthma, type 1 diabetes, and adult and juvenile arthritis. The tragic sequelae of the phase I trial of TGN1412 at Northwick Park demonstrated the delicate, but unpredictable, therapeutic ratio of some T-cell-directed treatments and, in the UK, have led to new guidelines for early-phase clinical trials of immune-directed therapies.

Isaacs JD. · Rheumatology and Rehabilitation Research Unit/Molecular Medicine Unit, Clinical Sciences Building, St James's University Hospital, Leeds LS9 7TF, UK. · Rheumatology (Oxford). · Pubmed #11477276 links to  free full text

Abstract: Anti-T-cell monoclonal antibodies (mAbs) form a unique class of therapeutic agent. Their precise specificity offers tremendous potential for the treatment of autoimmune and inflammatory diseases but also prevents meaningful preclinical animal studies. In particular, adverse reactions to therapy may be unanticipated, and the first administration of a novel T-cell mAb to a patient thus marks the beginning of a unique experiment. By comparing clinical parameters and laboratory measurements, small-scale pilot studies can provide detailed information about mAb biology that both predicts and suggests solutions to the complications of therapy. In this essay I illustrate this concept with reference to three specific areas: lymphocyte depletion, mAb immunogenicity and cytokine-release syndromes. In each case, systematic clinical and laboratory science has improved our understanding of the problem and suggested solutions; most of these solutions have been or are being adopted. Thus, small, open studies are an essential step in the development of novel mAbs, provide an ideal platform for the study of mAb biology, and serve as an early warning system for potential adverse effects.

Isaacs JD, Morgan AW, Strand V. · Department of Rheumatology, University of Leeds, UK. · Clin Exp Rheumatol. · Pubmed #10589370 No free full text.

Abstract: Biologic therapies refer to genetically engineered treatments such as monoclonal antibodies and receptor-immunoglobulin fusion proteins. Following many disappointments, the introduction of anti-tumor necrosis factor (anti-TNF alpha) therapies into the clinic has clearly demonstrated the exciting potential of biologic agents. Many of these have been designed to modulate a specific aspect of the underlying autoimmune process, thus avoiding generalized immunosuppression. They include products which interfere with the trimolecular complex of major histocompatibility complex II-Antigen-T cell receptor interaction; others designed to block the secondary signals for T cell activation and T cell interaction with antigen-presenting cells; and cytokine agonists as well as antagonists. Whilst reducing the degree of global immunosuppression associated with therapy, this targeted specificity may reduce the likelihood that a single therapeutic agent will provide long-term disease control. On the other hand, animal models have demonstrated synergy of combination biologic therapy, particularly for the re-induction of self-tolerance. As our knowledge of immune physiology and, particularly, immune regulation improves, it can be expected that many combination biologic therapies will be tested in the clinic. Pilot studies of combined anti-CD4 and TNF alpha blockade are underway; combination use of TNF alpha and interleukin-1 beta inhibitors are expected. This article reviews the potential for combination biologic therapy, including likely adverse effects, for the treatment of rheumatoid arthritis and other autoimmune diseases.

Verburg RJ, Flierman R, Sont JK, Ponchel F, van Dreunen L, Levarht EW, Welling MM, Toes RE, Isaacs JD, van Laar JM. · Department of Rheumatology, Division of Nuclear Medicine, Leiden University Medical Centre, PO Box 9600, 2300 RC Leiden, The Netherlands. · Ann Rheum Dis. · Pubmed #15829573 links to  free full text

Abstract: OBJECTIVE: To determine clinical and immunological correlates of high dose chemotherapy (HDC) + autologous stem cell transplantation (ASCT) in patients with severe rheumatoid arthritis (RA), refractory to conventional treatment. METHODS: Serial samples of peripheral blood and synovial tissue were obtained from seven patients with RA treated with HDC and autologous peripheral blood grafts enriched for CD34+ cells. Disease activity was assessed with the Disease Activity Score (DAS), serum concentrations of C reactive protein (CRP), and human immunoglobulin (HIg) scans, and the extent of immunoablation was determined by immunophenotyping of peripheral blood mononuclear cells, and immunohistochemistry and double immunofluorescence of synovium. RESULTS: Clinical responders (n = 5) had a larger number of cells at baseline expressing CD3, CD4, CD27, CD45RA, CD45RB, and CD45RO in synovium (p < 0.05), higher activity on HIg scans (p = 0.08), and a trend towards higher concentrations of CRP in serum than non-responders (n = 2). Subsequent remissions and relapses in responders paralleled reduction and re-expression, respectively, of T cell markers. A relatively increased expression of CD45RB and CD45RO on synovial CD3+ T cells was seen after HDC + ASCT. No correlations were found between DAS and changes in B cells or macrophage infiltration or synoviocytes. CONCLUSIONS: HDC + ASCT results in profound but incomplete immunoablation of both the memory and naïve T cell compartment, which is associated with longlasting clinical responses in most patients. The findings provide strong circumstantial evidence for a role of T cells in established RA, and demonstrate a role for the synovium in post-transplantation T cell reconstitution.

Bracewell C, Isaacs JD, Emery P, Ng WF. · Newcastle University, Musculoskeletal Research Group and Wilson Horne Immunotherapy Centre, Institute of Cellular Medicine, Newcastle-upon-Tyne, UK. · Expert Opin Biol Ther. · Pubmed #19522556 No free full text.

Abstract: BACKGROUND: Recent data suggest a key role for B cells in the pathogenesis of many autoimmune diseases including rheumatoid arthritis (RA), and biological therapies targeting B cells are promising treatments for patients with RA. Atacicept inhibits B cell maturation, differentiation and survival, and immunoglobulin production by depriving B cells of growth and development signals. Therefore, atacicept may represent an effective strategy in RA treatment. OBJECTIVE: To evaluate the potential value of atacicept in RA treatment based on preclinical and clinical studies. METHODS: Preclinical and clinical data on atacicept were identified using PubMed and systematically reviewed. RESULTS/CONCLUSION: Preclinical and clinical studies show that atacicept is well tolerated, with no increased incidence of infections. Atacicept displays non-linear pharmacokinetics, with a more than dose-proportional increase in free drug and less than dose-proportional, saturated increase in atacicept-ligand complex. Overall, the pharmacokinetic profiles of atacicept were consistent, dose-related and predictable. Dose-dependent reductions in immunoglobulins and other biomarkers, including rheumatoid factor, occurred rapidly but returned to baseline after discontinuation. There was a biphasic response in B cell number, but no effect on other leucocytes. Atacicept improved the signs and symptoms of RA, although larger studies are needed to confirm its efficacy and its optimal use.

Isaacs JD. · Wilson Horne Immunotherapy Centre and Musculoskeletal Research Group, Institute of Cellular Medicine, Newcastle University, Newcastle-Upon-Tyne NE2 4HH, UK. · Arthritis Res Ther. · Pubmed #19490600 links to  free full text

Abstract: There has been a therapeutic revolution in rheumatology over the past 15 years, characterised by a move away from oral immuno-suppressive drugs toward parenteral targeted biological therapies. The potency and relative safety of the newer agents has facilitated a more aggressive approach to treatment, with many more patients achieving disease remission. There is even a prevailing sense that disease 'cure' may be a realistic goal in the future. These developments were underpinned by an earlier revolution in molecular biology and protein engineering as well as key advances in our understanding of rheumatoid arthritis pathogenesis. This review will focus on antibody engineering as the key driver behind our current and developing range of antirheumatic treatments.

Lakey RL, Morgan TG, Rowan AD, Isaacs JD, Cawston TE, Hilkens CM. · Musculoskeletal Research Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK. · Rheumatology (Oxford). · Pubmed #19269957 No free full text.

Abstract: OBJECTIVE: Dendritic cells (DCs) are enriched in RA synovium and have been implicated in the pathogenesis of RA primarily through their ability to present autoantigen and activate T cells. However, whether DCs play an effector role in cartilage destruction is unknown. The aim of this study was to investigate whether DCs can induce collagen release from cartilage and the mechanism involved. METHODS: Human monocyte-derived DCs (mDCs) were activated with CD40 ligand (CD40L) to mimic DC-T-cell interaction, and supernatants were incubated with cartilage explants. Hydroxyproline was assessed as a measure of collagen release and collagenolytic activity was measured by a bioassay using tritiated collagen. TNF-alpha in DC supernatants was measured by specific ELISA. RESULTS: Supernatants from CD40L-activated mDCs, but not unstimulated mDCs, strongly induced the destruction of cartilage collagen. mDC supernatants did not contain collagenases but did induce collagenolytic activity in cartilage explants. Neutralization of TNF-alpha in mDC supernatants completely abolished collagenolysis. CONCLUSIONS: This study shows that mDCs, upon CD40-ligation, induce cartilage collagen degradation through an indirect mechanism via the production of TNF-alpha. Our data suggest a potential important role for mDC-derived TNF-alpha in RA, which is in line with the previously reported observations that DCs are a major source of TNF-alpha in early autoimmune lesions and that anti-TNF-alpha therapeutics effectively suppress joint damage in RA patients. We propose that DCs can act as effectors in cartilage destruction, adding a new aspect to the functional role of DCs in RA pathogenesis.

Bowes JD, Potter C, Gibbons LJ, Hyrich K, Plant D, Morgan AW, Wilson AG, Isaacs JD, Worthington J, Barton A, Anonymous00030. · Arthritis Research Campaign Epidemiology Unit, University of Manchester, Manchester, UK. · Pharmacogenet Genomics. · Pubmed #19262425 No free full text.

Abstract: The introduction of anti-tumour necrosis factor (TNF) agents has greatly improved the treatment of rheumatoid arthritis; however, a significant proportion of patients fail to respond to therapy. We hypothesized that genes within the TNF receptor superfamily member 1B signalling pathway contribute towards the observed variation in patient response. This was tested by genotyping 73 single-nucleotide polymorphisms (SNPs) from six candidate genes (DUSP1, HRB, IKBKAP, MAP3K1, MAP3K14 and TANK) in a large UK cohort of rheumatoid arthritis patients (n=642). Two SNPs [rs96844 (MAP3K1) and rs4792847 (MAP3K14)] showed evidence of association (P<0.05) to treatment response and were subsequently examined in an independent cohort of patients (n=428). Replication of association to either SNP was not achieved, but combined analysis of the complete cohort (n=1070) provided nominal evidence of association to both SNPs. We conclude that analysis of the common variation in the selected candidate genes did not provide strong evidence to implicate their involvement in varying patient response to anti-TNF treatment.

Anderson AE, Swan DJ, Sayers BL, Harry RA, Patterson AM, von Delwig A, Robinson JH, Isaacs JD, Hilkens CM. · Musculoskeletal Research Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK. · J Leukoc Biol. · Pubmed #18971286 links to  free full text

Abstract: Autoimmune pathologies are caused by a breakdown in self-tolerance. Tolerogenic dendritic cells (tolDC) are a promising immunotherapeutic tool for restoring self-tolerance in an antigen-specific manner. Studies about tolDC have focused largely on generating stable maturation-resistant DC, but few have fully addressed questions about the antigen-presenting and migratory capacities of these cells, prerequisites for successful immunotherapy. Here, we investigated whether human tolDC, generated with dexamethasone and the active form of vitamin D3, maintained their tolerogenic function upon activation with LPS (LPS-tolDC), while acquiring the ability to present exogenous autoantigen and to migrate in response to the CCR7 ligand CCL19. LPS activation led to important changes in the tolDC phenotype and function. LPS-tolDC, but not tolDC, expressed the chemokine receptor CCR7 and migrated in response to CCL19. Furthermore, LPS-tolDC were superior to tolDC in their ability to present type II collagen, a candidate autoantigen in rheumatoid arthritis. tolDC and LPS-tolDC had low stimulatory capacity for allogeneic, naïve T cells and skewed T cell polarization toward an anti-inflammatory phenotype, although LPS-tolDC induced significantly higher levels of IL-10 production by T cells. Our finding that LPS activation is essential for inducing migratory and antigen-presenting activity in tolDC is important for optimizing their therapeutic potential.

Maxwell JR, Potter C, Hyrich KL, Anonymous00014, Barton A, Worthington J, Isaacs JD, Morgan AW, Wilson AG. · Academic Rheumatology Group, University of Sheffield, D Floor Medical School, Sheffield, UK. · Hum Mol Genet. · Pubmed #18713756 No free full text.

Abstract: Anti-tumour necrosis factor (TNF) agents have revolutionized the treatment of patients with rheumatoid arthritis (RA). These therapies are, however, expensive and 30% of patients fail to respond. In a large cohort of Caucasian RA patients treated with anti-TNF medications (total n = 1050, etanercept n = 455, infliximab n = 450), we investigated whether genotypes of eight single nucleotide polymorphisms in the region containing the TNF gene were associated with response to anti-TNF therapy. Linear regression analyses adjusted for baseline 28 joint disease activity score (DAS28), baseline health assessment questionnaire score, gender and concurrent disease modifying anti-rheumatic drug treatment were used to assess association of these polymorphisms with treatment response, defined by change in DAS28 after 6 months. Analyses were performed in the entire cohort, and also stratified by anti-TNF agent. Association between DAS28 response and TNF-308 (rs1800629) genotype (P = 0.001) was detected across the whole cohort. After stratification by anti-TNF agent, the rare TNF-308AA genotype was associated with a significantly poorer response compared with TNF-308GG in etanercept (P = 0.001, n = 7) but not infliximab (P = 0.8, n = 17) treated patients. Conversely, the GA genotype at TNF-238 (rs361525) was associated with a poorer response to infliximab (P = 0.028, n = 40), but not etanercept (P = 0.6, n = 33). Owing to the small numbers of patients in some of the genotype groups examined, our data must be regarded as preliminary and will require replication in further large cohorts of anti-TNF-treated patients. If confirmed, our findings suggest the potential for genotype at these markers to aid selection of anti-TNF agent in patients with RA.

Lorenzi AR, Morgan TA, Anderson A, Catterall J, Patterson AM, Foster HE, Isaacs JD. · Institute of Cellular Medicine, Newcastle University, Newcastle-upon-Tyne, UK. · Ann Rheum Dis. · Pubmed #18628282 links to  free full text

Abstract: OBJECTIVE: Thymic function declines exponentially with age. Impaired thymic function has been associated with autoimmune disease in adults but has never been formally assessed in childhood autoimmunity. Therefore, thymic function in children with the autoimmune disease juvenile idiopathic arthritis (JIA) was determined. METHODS: Thymic function was measured in 70 children and young adults with JIA (age range 2.1-30.8 (median 10.4)) and 110 healthy age-matched controls using four independent assays. T cell receptor excision circles (WBLogTREC/ml) and the proportion of CD4(+) CD45RA(+)CD31(+) T cells (representing recent thymic emigrants; %RTEs) were quantified and intrathymic proliferation measured by calculating the alphaTREC/SigmabetaTREC ratio. Lastly, regulatory T cells (T(Reg)) of thymic origin (CD4(+)FOXP3(+)) were quantified in peripheral blood to assess the ability of the thymus in JIA to generate this T cell subset. RESULTS: Thymic function was equivalent by all four parameters in JIA when compared with the control population. Furthermore, there was no consistent effect of JIA subtype on thymic function, although intrathymic proliferation was higher in the small rheumatoid factor (RF)(+) polyarticular group. There were no significant effects of disease-modifying antirheumatic drugs (DMARDs) or oral corticosteroids on thymic function, although those with the worst prognostic ILAR (International League of Associations for Rheumatology) subtypes were also those most likely to be on a DMARD. CONCLUSIONS: It is demonstrated that children and young adults with JIA, unlike adults with autoimmune diseases, have thymic function that is comparable with that of healthy controls. The varied pathologies represented by the term "JIA" suggest this observation may not be disease specific and raises interesting questions about the aetiology of thymic impairment in adult autoimmunity.

Morgan AW, Hale G, Rebello PR, Richards SJ, Gooi HC, Waldmann H, Emery P, Isaacs JD. · Leeds Institute of Molecular Medicine, Section of Musculoskeletal Disease, University of Leeds, UK. · QJM. · Pubmed #18287112 links to  free full text

Abstract: BACKGROUND: Immunological tolerance in humans using anti-T-cell monoclonal antibodies (mAbs) may be hampered by a pro-inflammatory microenvironment. All clinical trials of such therapies in rheumatoid arthritis (RA), however, have selected patients with active disease at baseline. Concurrent neutralization of inflammation with a TNFalpha antagonist should maximize the potential of anti-T-cell mAbs to induce tolerance in RA. AIM: To evaluate the safety of combining a TNFalpha antagonist and CD4 mAb in RA. DESIGN: An iterative pilot study focused on the safety of such combination therapy. METHODS: Eight poor prognosis, seropositive RA patients were treated with combined CD4 and TNFalpha blockade. Prolonged CD4 blockade was achieved with a humanized mAb, and TNFalpha blockade with a p55 TNF receptor fusion protein. RESULTS: There was a low incidence of classical first-dose reactions to the CD4 mAb, possibly reflecting concomitant TNFalpha blockade. An unusual anaphylactoid reaction was seen, however, and one patient developed a probable allergic reaction after several infusions. Skin rashes were common, as previously reported with CD4 mAb monotherapy. No serious infections were documented during follow-up, despite CD4+ lymphopenia in some patients. Most patients appeared to demonstrate improved RA disease control after the study. After 17-49 months after therapy, one patient was in remission, one remained off disease modifying anti-rheumatic drugs and five had stable disease, three on previously ineffective doses of methotrexate. CONCLUSION: We report, for the first time in man, immunotherapy with a combination of an anti-cytokine and an anti-T-cell reagent. We witnessed an unusual first-dose reaction but there were no significant infectious complications.

Lorenzi AR, Clarke AM, Wooldridge T, Waldmann H, Hale G, Symmons D, Hazleman BL, Isaacs JD. · Newcastle University, Newcastle-upon-Tyne, UK. · Arthritis Rheum. · Pubmed #18240243 links to  free full text

Abstract: OBJECTIVE: To assess immunologically relevant outcomes in a cohort of rheumatoid arthritis (RA) patients with prolonged therapy-induced lymphopenia. METHODS: Morbidity (infection or malignancy) and mortality were assessed in 53 RA patients who were treated with the lymphocytotoxic monoclonal antibody alemtuzumab between 1991 and 1994. Data were obtained by interview, medical record review, and Office for National Statistics mortality monitoring. Lymphocyte subsets were enumerated by flow cytometry. A retrospective, matched-cohort study of mortality was performed with 102 control subjects selected from the European League Against Rheumatism database of patients with rheumatic disorders. RESULTS: Lymphopenia persisted in the patients: median CD3+CD4+, CD3+CD8+, CD19+, and CD56+ lymphocyte counts measured at a median followup of 11.8 years from the first administration of alemtuzumab were 0.50 x 10(9)/liter, 0.26 x 10(9)/liter, 0.11 x 10(9)/liter, and 0.09 x 10(9)/liter, respectively. Twenty-seven of 51 cases and 46 of 101 controls with available data had died, yielding a mortality rate ratio of 1.20 (95% confidence interval 0.72-1.98). Causes of death were similar to those that would be expected in a hospital-based RA cohort. No opportunistic infections were noted, and only 3 infections were documented following 36 elective orthopedic procedures. CONCLUSION: Despite continued lymphopenia 11.8 years after therapy, our patient cohort did not exhibit excess mortality or unusual infection-related morbidity, and surgery was well tolerated. These data should be reassuring for clinicians and patients who are considering lymphocytotoxic or other immunomodulatory therapy for RA.

Burgoyne CH, Field SL, Brown AK, Hensor EM, English A, Bingham SL, Verburg R, Fearon U, Lawson CA, Hamlin PJ, Straszynski L, Veale D, Conaghan P, Hull MA, van Laar JM, Tennant A, Emery P, Isaacs JD, Ponchel F. · Academic Unit of Musculoskeletal Disease, University of Leeds, Leeds, UK. · Ann Rheum Dis. · Pubmed #17644540 No free full text.

Abstract: OBJECTIVES: An abnormal CD4+ T cell subset related to inflammation exposure (inflammation-related cells, IRC) has been identified in rheumatoid arthritis (RA). Patients with inflammatory and non-inflammatory diseases were used to examine the relationship between inflammation and this T cell subset in vivo. METHODS: Blood was collected from healthy controls and patients with RA (active disease or in clinical remission), Crohn's disease and osteoarthritis. IRC and chemokine receptors were quantified by flow cytometry. Thymic activity and apoptotic factors were measured by real-time polymerase chain reaction. Circulating cytokines were measured by enzyme-linked immunosorbent assay. CXCR4 and SDF1 in synovial biopsies were measured using immunohistochemistry. RESULTS: IRC were identified in patients with RA (p<0.0001) and Crohn's disease (p = 0.005), but not in those with osteoarthritis. In RA in remission, IRC persisted (p<0.001). In remission, hyperproliferation of IRC was lost, chemokine receptor expression was significantly lowered (p<0.007), Bax expression dropped significantly (p<0.001) and was inversely correlated with IRC (rho = -0.755, p = 0.03). High IRC frequency in remission was associated with relapse within 18 months (OR = 6.4, p<0.001) and a regression model predicted 72% of relapse. CONCLUSIONS: These results suggest a model in which, despite the lack of systemic inflammation, IRC persist in remission, indicating that IRC are an acquired feature of RA. They have, however, lost their hyper-responsiveness, acquired a potential for survival, and no longer express chemokine receptors. IRC persistence in remission confirms their important role in chronic inflammation as circulating precursors of pathogenic cells. This was further demonstrated by much higher incidence of relapse in patients with high IRC frequency in remission.

von Delwig A, Hilkens CM, Altmann DM, Holmdahl R, Isaacs JD, Harding CV, Robertson H, McKie N, Robinson JH. · Musculoskeletal Research Group, Clinical Medical Sciences, University of Newcastle upon Tyne, Framlington Place, Newcastle upon Tyne, UK. · Arthritis Res Ther. · Pubmed #16704744 links to  free full text

Abstract: Professional antigen-presenting cells, such as dendritic cells, macrophages and B cells have been implicated in the pathogenesis of rheumatoid arthritis, constituting a possible target for antigen-specific immunotherapy. We addressed the possibility of blocking antigen presentation of the type II collagen (CII)-derived immunodominant arthritogenic epitope CII259-273 to specific CD4 T cells by inhibition of antigen uptake in HLA-DR1-transgenic mice in vitro and in vivo. Electron microscopy, confocal microscopy, subcellular fractionation and antigen presentation assays were used to establish the mechanisms of uptake, intracellular localization and antigen presentation of CII by dendritic cells and macrophages. We show that CII accumulated in membrane fractions of intermediate density corresponding to late endosomes. Treatment of dendritic cells and macrophages with cytochalasin D or amiloride prevented the intracellular appearance of CII and blocked antigen presentation of CII259-273 to HLA-DR1-restricted T cell hybridomas. The data suggest that CII was taken up by dendritic cells and macrophages predominantly via macropinocytosis. Administration of amiloride in vivo prevented activation of CII-specific polyclonal T cells in the draining popliteal lymph nodes. This study suggests that selective targeting of CII internalization in professional antigen-presenting cells prevents activation of autoimmune T cells, constituting a novel therapeutic strategy for the immunotherapy of rheumatoid arthritis.

Lawson CA, Brown AK, Bejarano V, Douglas SH, Burgoyne CH, Greenstein AS, Boylston AW, Emery P, Ponchel F, Isaacs JD. · Academic Unit of Musculoskeletal Disease, IMMECR, Leeds General Infirmary, Great George Street, Leeds LS1 3EX, UK. · Rheumatology (Oxford). · Pubmed #16571607 links to  free full text

Abstract: OBJECTIVE: Our aim was to test the hypothesis that there is a deficit in the CD4+CD25high regulatory T-cell population in early rheumatoid arthritis (RA), either in size or functional activity. METHODS: Peripheral blood mononuclear cells were examined from subjects with early active RA who had received no previous disease-modifying therapy (n = 43), from individuals with self-limiting reactive arthritis (n = 14), from subjects with stable, well-controlled RA (n = 82) and from healthy controls (n = 72). The frequencies of CD4+CD25high T-cells were quantified using flow cytometry, and function was assessed by the ability to suppress proliferation of CD4+CD25- T-cells. Paired blood and synovial fluid was analysed from a small number of RA and reactive arthritis patients. RESULTS: There was a smaller proportion of CD4+CD25high T-cells in the peripheral blood of early active RA patients (mean 4.25%) than in patients with reactive arthritis or in controls (mean 5.90 and 5.30%, respectively, P = 0.001 in each case). Frequencies in stable, well-controlled RA (mean 4.63%) were not significantly different from early active RA or controls. There were no differences in suppressor function between groups. Higher frequencies of CD4+CD25high T-cells were found in synovial fluid than blood in both RA and reactive arthritis. CONCLUSIONS: These data demonstrate a smaller CD4+CD25high regulatory T-cell population in peripheral blood of individuals with early active RA prior to disease-modifying treatment. This may be a contributory factor in the susceptibility to RA and suggests novel approaches to therapy.

von Delwig A, Altmann DM, Isaacs JD, Harding CV, Holmdahl R, McKie N, Robinson JH. · Musculoskeletal Research Group, University of Newcastle upon Tyne, Newcastle upon Tyne, UK. · Arthritis Rheum. · Pubmed #16447222 links to  free full text

Abstract: OBJECTIVE: Type II collagen (CII) is a candidate autoantigen implicated in the pathogenesis of rheumatoid arthritis (RA). Posttranslational glycosylation of CII could alter intracellular antigen processing, leading to the development of autoimmune T cell responses. To address this possibility, we studied the intracellular processing of CII for presentation of the arthritogenic glycosylated epitope CII(259-273) to CD4 T cells in macrophages from HLA-DR1-transgenic mice. METHODS: HLA-DR1-transgenic mice were generated on a class II major histocompatibility complex-deficient background, and T cell hybridomas specific for the glycosylated and nonglycosylated epitope CII(259-273) were developed. Subcellular fractionation of macrophages was used to localize CII degradation to particular compartments and to identify the catalytic subtype of proteinases involved. RESULTS: We showed that the glycosylated CII(259-273) epitope required more extensive processing than did the nonglycosylated form of the same epitope. Dense fractions containing lysosomes were primarily engaged in the processing of CII for antigen presentation, since these compartments contained 1) enzyme activity that generated antigenic CII fragments bearing the arthritogenic glycosylated epitope, 2) the antigenic CII fragments themselves, 3) CII peptide-receptive HLA-DR1 molecules, and 4) peptide/HLA-DR1 complexes that could directly activate T cell hybridomas. Degradation of CII by dense fractions occurred optimally at pH 4.5 and was abrogated by inhibitors of serine and cysteine proteinases. CONCLUSION: Processing of the arthritogenic glycosylated CII(259-273) epitope, which is implicated in the induction of autoimmune arthritis, is more stringently regulated than is processing of the nonglycosylated form of the same epitope. Mechanisms of intracellular processing of the glycosylated epitope may constitute novel therapeutic targets for the treatment of RA.

Morgan AW, Barrett JH, Griffiths B, Subramanian D, Robinson JI, Keyte VH, Ali M, Jones EA, Old RW, Ponchel F, Boylston AW, Situnayake RD, Markham AF, Emery P, Isaacs JD. · Institute of Molecular Medicine, Epidemiology and Cancer Research, University of Leeds, St James's University Hospital, Beckett Street, Leeds, LS9 7TF, UK. · Arthritis Res Ther. · Pubmed #16356189 links to  free full text

Abstract: The Fcgamma receptors play important roles in the initiation and regulation of many immunological and inflammatory processes, and genetic variants (FCGR) have been associated with numerous autoimmune and infectious diseases. The data in rheumatoid arthritis (RA) are conflicting and we previously demonstrated an association between FCGR3A and RA. In view of the close molecular proximity with FCGR2A, FCGR2B and FCGR3B, additional polymorphisms within these genes and FCGR haplotypes were examined to refine the extent of association with RA. Biallelic polymorphisms in FCGR2A, FCGR2B and FCGR3B were examined for association with RA in two well characterized UK Caucasian and North Indian/Pakistani cohorts, in which FCGR3A genotyping had previously been undertaken. Haplotype frequencies and linkage disequilibrium were estimated across the FCGR locus and a model-free analysis was performed to determine association with RA. This was followed by regression analysis, allowing for phase uncertainty, to identify the particular haplotype(s) that influences disease risk. Our results reveal that FCGR2A, FCGR2B and FCGR3B were not associated with RA. The haplotype with the strongest association with RA susceptibility was the FCGR3A-FCGR3B 158V-NA2 haplotype (odds ratio 3.18, 95% confidence interval 1.13-8.92 [P = 0.03] for homozygotes compared with all genotypes). The association was stronger in the presence of nodules (odds ratio 5.03, 95% confidence interval 1.44-17.56; P = 0.01). This haplotype was also more common in North Indian/Pakistani RA patients than in control individuals, but not significantly so. Logistic regression analyses suggested that FCGR3A remained the most significant gene at this locus. The increased association with an FCGR3A-FCGR3B haplotype suggests that other polymorphic variants within FCGR3A or FCGR3B, or in linkage disequilibrium with this haplotype, may additionally contribute to disease pathogenesis.

Ali S, Robertson H, Wain JH, Isaacs JD, Malik G, Kirby JA. · The Applied Immunobiology and Transplantation Research Group, Medical School, University of Newcastle, Newcastle upon Tyne NE2 4HH, United Kingdom. · J Immunol. · Pubmed #16002730 links to  free full text

Abstract: A non-glycosaminoglycan (GAG)-binding variant of the pleiotropic chemokine CCL7 was generated by mutating to alanine the basic (B) amino acids within an identified (44)BXBXXB(49) GAG-binding motif. Unlike wild-type (wt) CCL7, the mutant sequence had no affinity for heparin. However, the mutant retained a normal affinity for CCR1, CCR2b, and CCR3, and produced a normal calcium flux in mononuclear leukocytes. Both the wt and mutant proteins elicited an equal leukocyte chemotactic response within a solute diffusion gradient but, unlike the wt protein, the mutant failed to stimulate cell migration across a model endothelium. The number of leukocytes recruited to murine air pouches by the mutant sequence was lower than that recruited by wt CCL7. Furthermore, the presence of a mixture of a mutant and wt CCL7 within the air pouch elicited no significant cell accumulation. Cell recruitment also failed using a receptor-sharing mixture of mutant CCL7 and wt CCL5 or a nonreceptor sharing mixture of mutant CCL7 and wt CXCL12. The potential of the mutant sequence to modulate inflammation was confirmed by demonstration of its ability to inhibit the chemotactic response generated in vitro by synovial fluid from patients with active rheumatoid arthritis. A further series of experiments suggested that the non-GAG-binding mutant protein could potentially induce receptor desensitization before, and at a site remote from, any physiological recognition of GAG-bound chemokines. These data demonstrate that GAG binding is required for chemokine-driven inflammation in vivo and also suggest that a non-GAG-binding chemokine receptor agonist can inhibit the normal vectorial leukocyte migration mediated by chemokines.

Lawson CA, Donaldson IJ, Bowman SJ, Shefta J, Morgan AW, Gough A, Isaacs JD, Griffiths B, Emery P, Pease CT, Boylston AW. · Molecular Medicine Unit, University of Leeds, Leeds, UK. · Ann Rheum Dis. · Pubmed #15708895 links to  free full text

Abstract: OBJECTIVE: To analyse T cell receptor beta variable (TCRBV) gene polymorphisms (insertion/deletion related polymorphism (IDRP) and BV6S7) in primary Sjögren's syndrome (PSS). METHODS: Genomic DNA was extracted from blood samples from patients fulfilling the modified European criteria for PSS (n = 61). Healthy control blood samples were obtained from the Blood Transfusion Service (n = 121). As a disease control group, samples from patients with systemic lupus erythematosus (n = 42) were analysed. BV6S7 was genotyped using an established PCR/RFLP method. The IDRP was determined by comparison of the intensity of PCR product bands from within BV9S2 and an internal control region (BV9S1), to ascertain whether 0, 1, or 2 copies of the insertion were present. RESULTS: There was a decrease (p = 0.018) in the proportion of PSS patients with the deleted/deleted genotype. There was no association with specific BV6S7 alleles or genotypes with either the PSS group or the hypergammaglobulinaemic subgroup. There were no significant differences in haplotype frequencies after Bonferroni correction. CONCLUSIONS: A reduced proportion of patients with PSS have the deleted/deleted genotype. Eighty nine per cent of PSS patients have at least one extra germline copy of BV13S2*1. This may relate to previous observations of increased BV13 specific T cells and mRNA in the salivary glands.

Ponchel F, Verburg RJ, Bingham SJ, Brown AK, Moore J, Protheroe A, Short K, Lawson CA, Morgan AW, Quinn M, Buch M, Field SL, Maltby SL, Masurel A, Douglas SH, Straszynski L, Fearon U, Veale DJ, Patel P, McGonagle D, Snowden J, Markham AF, Ma D, van Laar JM, Papadaki HA, Emery P, Isaacs JD. · Molecular Medicine Unit, University of Leeds, Leeds, UK. · Arthritis Res Ther. · Pubmed #15642146 links to  free full text

Abstract: We previously demonstrated prolonged, profound CD4+ T-lymphopenia in rheumatoid arthritis (RA) patients following lymphocyte-depleting therapy. Poor reconstitution could result either from reduced de novo T-cell production through the thymus or from poor peripheral expansion of residual T-cells. Interleukin-7 (IL-7) is known to stimulate the thymus to produce new T-cells and to allow circulating mature T-cells to expand, thereby playing a critical role in T-cell homeostasis. In the present study we demonstrated reduced levels of circulating IL-7 in a cross-section of RA patients. IL-7 production by bone marrow stromal cell cultures was also compromised in RA. To investigate whether such an IL-7 deficiency could account for the prolonged lymphopenia observed in RA following therapeutic lymphodepletion, we compared RA patients and patients with solid cancers treated with high-dose chemotherapy and autologous progenitor cell rescue. Chemotherapy rendered all patients similarly lymphopenic, but this was sustained in RA patients at 12 months, as compared with the reconstitution that occurred in cancer patients by 3-4 months. Both cohorts produced naive T-cells containing T-cell receptor excision circles. The main distinguishing feature between the groups was a failure to expand peripheral T-cells in RA, particularly memory cells during the first 3 months after treatment. Most importantly, there was no increase in serum IL-7 levels in RA, as compared with a fourfold rise in non-RA control individuals at the time of lymphopenia. Our data therefore suggest that RA patients are relatively IL-7 deficient and that this deficiency is likely to be an important contributing factor to poor early T-cell reconstitution in RA following therapeutic lymphodepletion. Furthermore, in RA patients with stable, well controlled disease, IL-7 levels were positively correlated with the T-cell receptor excision circle content of CD4+ T-cells, demonstrating a direct effect of IL-7 on thymic activity in this cohort.