THU0319 Determination of the Serological Profiles in Anti-Ro-52-Positive Patients with Systemic Autoimmune Rheumatic Diseases.

Background

Ro-52 is an interferon (IFN)-inducible protein of the tripartite motif (TRIM) family. Antibodies against Ro-52 have been described in patients with different autoimmune diseases such as systemic lupus erythematosus (SLE) and Sjogren's syndrome (SS), which are often associated with anti-Ro-60 antibodies. However, in inflammatory myositis, anti-Ro-52 antibodies are frequently present without anti-Ro-60 antibodies. Ro-52 autoantigen is extraordinarily antigenic and its autoantibodies are directed againstboth linear and conformational epitopes.

Objectives

The first aim of the study was to evaluate antibodies against the full length of the Ro-52 antigen (Ro-52) and its five fragments, and their association with serological and clinical data. The second aim was to assess the prevalence of anti-full-length Ro-52 and its five fragments in different systemic autoimmune rheumatic diseases (SARDs).

Methods

Serum samples were obtained from 78 SARDs patients and 68 controls (50 healthy donors and 18 patients with other autoimmune diseases). Ro-52 antibodies were measured using four different Methods: Bioplex2200 Multiplex-Biorad (Laboratories Inc. Hercules, CA, USA), ANA Profile3-Blot, Profile Autoimmune Liver Diseases-Blot, and Myositis Profile3-Blot EUROIMMUN AG. (EUROIMMUN AG, Lubeck, Germany). Serum was considered positive for anti-Ro-52 when at least one of these methods revealed their presence. Full-length Ro-52 antigen was expressed in insect cells using the baculovirus system, and five recombinant Ro-52 antigen fragments [Ro-52-1, Ro-52-2, Ro-52-3, Ro-52-4 (partly overlapping Ro-52-1 and -2) and Ro-52-5 (partly overlapping Ro-52-2 and -3)] expressed in E. coli, were coated onto nitrocellulose membranes in a line immunoassay system. Sera were incubated in accordance with the manufacturers standard protocol (30 min serum, 30 min anti-human IgG/alkaline phosphatase, 10 min NBT/BCIP substrate). Prevalence and reaction intensities were automatically evaluated using the commercial computer programme EUROLineScan from EUROIMMUN AG.

Results

All of the samples obtained from the SARDs patients were positive for antibodies against the full-length Ro52 and fragment 2, except those obtained from the myositis patients. None of the sera samples of the patients and controls were positive for anti-Ro-52-3. Moreover, all of the control samples were negative for the full-length Ro52 and all of the fragments. Myositis group (n=3) gave an isolated but very weak reaction with the full length Ro-52 and displayed no reaction with the fragments. In the SARDs patients, the fragment prevalence rates were Ro-52-1= 4/82 (4.9%), Ro-52-2= 79/82 (96.3%), Ro-52-3= 0/82 (0%), Ro-52-4= 6/82 (7.3%) and Ro-52-5= 22/82 (26.8%).

Conclusions

The main epitope is localised on fragment 2, and all diseases have antibodies that target this fragment. Antibody positivity for full-length Ro52 and negativity for all five fragments suggests a diagnosis of myositis. As all of the samples were positive for fragment 2, and only some for fragments 5 or 4, it seems that the target epitope is localized in the middle of fragment 2 or in the area between fragments 4 and 5. Further studies involving a large number of patients are suggested to confirm our findings.

Disclosure of Interest

None Declared