Yoshitaka Imura

Efficacy of rituximab (anti-CD20) for refractory systemic lupus erythematosus involving the central nervous system

Aim: Neuropsychiatric systemic lupus erythematosus (NPSLE) is a serious treatment-resistant phenotype of systemic lupus erythematosus. A standard treatment for NPSLE is not available. This report describes the clinical and laboratory tests of 10 patients with NPSLE before and after rituximab treatment, including changes in lymphocyte phenotypes. Methods: Rituximab was administered at different doses in 10 patients with refractory NPSLE, despite intensive treatment. Results: Treatment with rituximab resulted in rapid improvement of central nervous system-related manifestations, particularly acute confusional state. Rituximab also improved cognitive dysfunction, psychosis and seizure, and reduced the SLE Disease Activity Index Score at day 28 in all 10 patients. These effects lasted for >1 year in five patients. Flow cytometric analysis showed that rituximab down regulated CD40 and CD80 on B cells and CD40L, CD69 and inducible costimulator on CD4+ T cells. Conclusions: Rituximab rapidly improved refractory NPSLE, as evident by resolution of various clinical signs and symptoms and improvement of radiographic findings. The down regulation of functional molecules on B and T cells suggests that rituximab modulates the interaction of activated B and T cells through costimulatory molecules. These results warrant further analysis of rituximab as treatment for NPSLE.

The RIG-I-like receptor IFIH1/MDA5 is a dermatomyositis-specific autoantigen identified by the anti-CADM-140 antibody

OBJECTIVES Various autoantibodies are detected in the sera of PM/DM patients. Some of them are specific to PM/DM patients and closely associated with clinical manifestations of the diseases. Recently, the anti-CADM-140 antibody was reported to be found specifically in clinically amyopathic DM (C-ADM) patients and to be associated with acute interstitial lung disease (ILD). We assessed the clinical significance of the anti-CADM-140 antibody and then investigated the autoantigen recognized by the anti-CADM-140 antibody. METHODS Autoantibodies were screened in 192 patients with various CTDs and 21 healthy controls using immunoprecipitation with [(35)S]methionine-labelled HeLa cells. Immunoabsorbent column chromatography was used to purify an autoantigen that was subsequently subjected to peptide mass fingerprinting. RESULTS The anti-CADM-140 antibody was revealed to be specific to DM. Most of the anti-CADM-140-positive patients were C-ADM although some of them showed apparent myositis. The anti-CADM-140-positive patients frequently showed hyperferritinaemia and acute progressive ILD with poor prognosis. The anti-CADM-140 antibody was shown to recognize IFN induced with helicase C domain protein 1 (IFIH1), also known as the melanoma differentiation-associated gene 5 (MDA5), which is one of the RIG-I-like receptors and plays a role in innate immune responses. CONCLUSION The anti-CADM-140 antibody was a marker of DM and intractable ILD and recognized IFIH1/MDA5, which is involved in innate immunity. These findings may give a new insight into the pathogenesis of DM.

Anti-aminoacyl-tRNA synthetase antibodies in clinical course prediction of interstitial lung disease complicated with idiopathic inflammatory myopathies

In the treatment of polymyositis and dermatomyositis (PM/DM), the complication of interstitial lung disease (ILD) is an important prognostic factor. It has been reported that autoantibodies against aminoacyl-tRNA synthetases (ARS) are strongly associated with ILD. The aim of this study is to examine the correlation between anti-ARS and the clinical course of ILD. We investigated 41 cases of PM/DM with ILD. The response of ILD to corticosteroids (CS) was determined according to the change in respiratory symptoms, image findings, and pulmonary function between, before and 2 months after the treatment. Anti-ARS (anti-Jo-1, PL-7, PL-12, EJ, OJ and KS) antibodies were screened with the RNA immunoprecipitation assay. In the stratification into ILD-preceding, simultaneous and myopathy-preceding types, anti-ARS antibodies were significantly frequent in the ILD-preceding type (p < 0.05). In the stratification into anti-ARS-positive and negative groups, the response of ILD to CS was significantly better in the positive group (p < 0.05). However, recurrence of ILD was significantly more frequent in the positive group (p < 0.01), and 2 year prognoses of pulmonary function (%VC and %DLCO) were not different between the two groups. In conclusion, screening of anti-ARS may be useful to predict late-onset myopathy in ILD-preceding patients and to predict the clinical course of ILD in PM/DM patients.

Autoantibodies in idiopathic inflammatory myopathy: an update on clinical and pathophysiological significance.

Purpose of reviewIdiopathic inflammatory myopathy is characterized by the production of autoantibodies to various cellular constituents. These autoantibodies closely correlate with certain clinical conditions and prognosis of disease. This review examines recent progress in myositis-specific autoantibodies, particularly in their clinical significance and pathophysiological roles. Recent findingsDuring the 1-year review period, novel myositis-specific autoantibodies were identified in clinically amyopathic dermatomyositis (anti-CADM-140 antibody) and malignancy-associated myositis (anti-p155 and anti-p155/p140 antibodies). These new autoantibodies are extremely important because it is thought that myositis-specific autoantibodies are negative in these subgroups, and may enable a new classification of idiopathic inflammatory myopathy. New clinical aspects of other myositis-specific autoantibodies (anti-aminoacyl-tRNA synthetases, anti-signal recognition particles and anti-Mi-2) are also described. The possibility was raised that the high expression of myositis-specific autoantigens in regenerating muscle cells and certain cancers may be involved in initiating and perpetuating the autoimmune response in myositis. SummaryMyositis-specific autoantibodies are useful markers for clinical diagnosis, classification and predicting prognosis of idiopathic inflammatory myopathy. To understand the etiopathogenic mechanisms of the disease it is particularly important to elucidate the nature of target autoantigens recognized by these myositis-specific autoantibodies.

Comprehensive microRNA Analysis Identifies miR-24 and miR-125a-5p as Plasma Biomarkers for Rheumatoid Arthritis

MicroRNAs (miRNAs) are present in human plasma and known as a non-invasive biomarker for cancer detection. Our study was designed to identify plasma miRNAs specific for rheumatoid arthritis (RA) by a comprehensive array approach. We performed a systematic, array-based miRNA analysis on plasma samples from three RA patients and three healthy controls (HCs). Plasma miRNAs with more than four times change or with significant (P<0.05) change in expression, or detectable only in RA plasma, were confirmed with plasma from eight RA patients and eight HCs using real-time quantitative PCR. Consistently detectable miRNAs that were significantly different between RA patients and HCs were chosen for further validation with 102 RA patients and 104 HCs. The area under curves (AUC) were calculated after plotting the receiver operating characteristic (ROC) curves. To determine if these miRNAs are specific for RA, the concentrations of these miRNAs were analyzed in 24 patients with osteoarthritis (OA), and 11 patients with systemic lupus erythematosus (SLE). The array analysis and the subsequent confirmation in larger patient cohort identified significant alterations in plasma levels of seven miRNAs. The highest AUC was found for miR-125a-5p, followed in order by miR-24 and miR-26a. Multivariable logistic regression analysis showed that miR-24, miR-30a-5p, and miR-125a-5p were crucial factors for making detection model of RA and provided a formula for Estimated Probability of RA by plasma MiRNA (ePRAM), employing miR-24, miR-30a-5p and miR-125a-5p, which showed increased diagnostic accuracy (AUC: 0.89). The level of miR-24, miR-125a-5p, and ePRAM in OA and SLE patients were lower than that in RA. There was no significant difference in detection for anti-citrullinated protein antibody (ACPA)-positive and ACPA-negative RA patients. These results suggest that the plasma concentrations of miR-24 and miR-125a-5p, and ePRAM are potential diagnostic markers of RA even if patients were ACPA-negative.

Etanercept-induced anti-Jo-1-antibody-positive polymyositis in a patient with rheumatoid arthritis: a case report and review of the literature

Antitumor necrosis factor (TNF) therapy has been associated with adverse immunologic events including systemic lupus erythematosus. However, the development of polymyositis (PM)/dermatomyositis (DM) associated with anti-TNF therapy is extremely rare. We experienced a case of a 48-year-old female with rheumatoid arthritis (RA) who had anti-Jo-1 antibodies and interstitial lung disease but no previous history of PM/DM and who developed PM soon after the initiation of etanercept (ETN) therapy for RA. The patient recovered upon withdrawal from ETN and corticosteroid (CS) therapies. Only four reports of PM/DM associated with anti-TNF therapy for RA could be found in the literature. The patients described in three of the four reports were positive for anti-Jo-1 antibodies before the initiation of anti-TNF therapy, and in all the cases, recovery occurred after the cessation of anti-TNF-agent administration and CS therapy. These results suggest a relationship between the onset of PM/DM with anti-Jo-1 antibody and anti-TNF therapy for RA.

DIP2 disco-interacting protein 2 homolog A (Drosophila) is a candidate receptor for follistatin-related protein/follistatin-like 1--analysis of their binding with TGF-β superfamily proteins.

Follistatin‐related protein (FRP)/follistatin‐like 1 (FSTL1) is a member of the follistatin protein family, all of which share a characteristic structure unit found in follistatin, called the FS domain. Developmental studies have suggested that FRP regulates organ tissue formation in embryos. Immunological studies showed that FRP modifies joint inflammation in arthritic disease, and modulates allograft tolerance. However, the principle physiological function of FRP is currently unknown. To address this issue, we cloned four FRP‐associated proteins using a two‐hybrid cloning method: disco‐interacting protein 2 homolog A from Drosophila (DIP2A), CD14, glypican 1 and titin. Only DIP2A was expected to be a membrane receptor protein with intracellular regions. Over‐expression of FLAG epitope‐tagged DIP2A augmented the suppressive effect of FRP on FBJ murine osteosarcoma viral oncogene homolog (FOS) expression, and the Fab fragment of IgG to FLAG blocked this effect. Knockdown of Dip2a leaded to Fos gene up‐regulation, and this was not affected by exogenous FRP. These in vitro experiments confirmed that DIP2A could be a cell‐surface receptor protein and mediate a FOS down‐regulation signal of FRP. Moreover, molecular interaction analyses using Biacore demonstrated that FRP bound to DIP2A and CD14, and also with proteins of the TGF‐β superfamily, i.e. activin, TGF‐β, bone morphogenetic protein 2/4 (BMP‐2/4), their receptors and follistatin. FRP binding to DIP2A was blocked by CD14, follistatin, activin and BMP‐2. FRP blocked the ligand–receptor binding of activin and BMP‐2, but integrated itself with that of BMP‐4. This multi‐specific binding may reflect the broad physiological activity of FRP.

A Clinical, Pathological, and Genetic Characterization of Methotrexate-associated Lymphoproliferative Disorders

Objective. Methotrexate-associated lymphoproliferative disorders (MTX-LPD) often regress spontaneously during MTX withdrawal, but the prognostic factors remain unclear. The aim of our study was to clarify the clinical, histological, and genetic factors that predict outcomes in patients with MTX-LPD. Methods. Patients with MTX-LPD diagnosed between 2000 and 2012 were analyzed retrospectively regarding their clinical course, site of biopsy, histological typing, Epstein-Barr virus (EBV) in situ hybridization and immunostaining, and HLA type. Results. Twenty-one patients, including 20 with rheumatoid arthritis (RA) and 1 with polymyositis, were analyzed. The mean dose of MTX was 6.1 mg/week and the mean duration of treatment was 71.1 months. Clinically, 5 patients were diagnosed with EBV-positive mucocutaneous ulcer (EBVMCU) and had polymorphic histological findings. The proportion of those patients successfully treated solely by withdrawal of MTX was significantly greater than that of those without EBVMCU (75% vs 7.7%, p = 0.015). The HLA-B15:11 haplotype was more frequent in patients with EBV+ RA with MTX-LPD than in healthy Japanese controls (p = 0.0079, Bonferroni’s method). EBV latency classification and HLA typing were not associated with the prognosis of MTX-LPD in our cohort. Conclusion. Our data demonstrate that patients in the EBVMCU, a specific clinical subgroup of MTX-LPD, had a better clinical outcome when MTX was withdrawn than did other patients with MTX-LPD.

The Multicenter Study of a New Assay for Simultaneous Detection of Multiple Anti-Aminoacyl-tRNA Synthetases in Myositis and Interstitial Pneumonia

Objective Autoantibodies to aminoacyl-tRNA synthetases (ARSs) are useful in the diagnosis of idiopathic inflammatory myopathy (IIM) with interstitial pneumonia (IP). We developed an enzyme-linked immunosorbent assay (ELISA) system using a mixture of recombinant ARS antigens and tested its utility in a multicenter study. Methods: We prepared six recombinant ARSs: GST-Jo-1, His-PL-12, His-EJ and GST-KS expressed in Escherichia coli, and His-PL-7 and His-OJ expressed in Hi-5 cells. After confirming their antigenic activity, with the exception of His-OJ, we developed our ELISA system in which the five recombinant ARSs (without His-OJ) were mixed. Efficiency was confirmed using the sera from 526 Japanese patients with connective tissue disease (CTD) (IIM n = 250, systemic lupus erythematosus n = 91, systemic sclerosis n = 70, rheumatoid arthritis n = 75, Sjögren’s syndrome n = 27 and other diseases n = 13), 168 with idiopathic interstitial pneumonia (IIP) and 30 healthy controls collected from eight institutes. IIPs were classified into two groups; idiopathic pulmonary fibrosis (IPF) (n = 38) and non-IPF (n = 130). Results were compared with those of RNA immunoprecipitation. Results: Sensitivity and specificity of the ELISA were 97.1% and 99.8%, respectively when compared with the RNA immunoprecipitation assay. Anti-ARS antibodies were detected in 30.8% of IIM, 2.5% of non-myositis CTD, and 10.7% of IIP (5.3% of IPF and 12.3% of non-IPF). Anti-ARS-positive non-IPF patients were younger and more frequently treated with glucocorticoids and/or immunosuppressants than anti-ARS-negative patients. Conclusion: A newly established ELISA detected anti-ARS antibodies as efficiently as RNA immunoprecipitation. This system will enable easier and wider use in the detection of anti-ARS antibodies in patients with IIM and IIP.

Anti-U1 RNP antibodies in cerebrospinal fluid are associated with central neuropsychiatric manifestations in systemic lupus erythematosus and mixed connective tissue disease.

OBJECTIVE To determine the significance of anti-U1 RNP antibodies in the cerebrospinal fluid (CSF) of patients with systemic lupus erythematosus (SLE) or mixed connective tissue disease (MCTD) who have central neuropsychiatric SLE (NPSLE). METHODS The frequency of antinuclear antibodies including anti-U1 RNP antibodies in the sera and CSF of 24 patients with SLE and 4 patients with MCTD, all of whom had neuropsychiatric syndromes, was determined using an RNA immunoprecipitation assay and an enzyme-linked immunosorbent assay. The frequency of anti-U1 RNP antibodies in the CSF of patients with central NPSLE was examined, and the anti-U1 RNP index ([CSF anti-U1 RNP antibodies/serum anti-U1 RNP antibodies]/[CSF IgG/serum IgG]) was compared with CSF interleukin-6 (IL-6) levels and the albumin quotient (Qalb, an indicator of blood-brain barrier damage). CSF and serum antibodies against U1-70K, U1-A, and U1-C, including autoantigenic regions, were examined, and the U1-70K, U1-A, and U1-C indices as well as the anti-U1 RNP index were calculated. RESULTS CSF anti-U1 RNP antibodies with an increased anti-U1 RNP index showed 64.3% sensitivity and 92.9% specificity for central NPSLE. The anti-U1 RNP index did not correlate with CSF IL-6 levels or the Qalb. The anti-U1-70K index was higher than the anti-U1-A and anti-U1-C indices in the CSF of anti-U1 RNP antibody-positive patients with central NPSLE. The major autoantigenic region for CSF anti-U1-70K antibodies appeared to be localized in U1-70K amino acid 141-164 residue within the RNA-binding domain. CONCLUSION The frequency of anti-U1 RNP antibodies in the CSF and the anti-U1 RNP index are useful indicators of central NPSLE in anti-U1 RNP antibody-positive patients. The predominance of anti-U1-70K antibodies in CSF suggests intrathecal anti-U1 RNP antibody production.