Background/Purpose: Anti-nuclear antibodies (ANA) are present in approximately 90% of sera from systemic sclerosis (SSc) patients and play an important diagnostic and prognostic role in SSc. Besides anti-Scl-70 (topo I), anti-centromere and anti-RNA Polymerase III which are included in the SSc classification criteria, several other antibodies can be found, some of which clearly associate with defined disease phenotypes. Anti-U11/12 Ribonucleoprotein (RNP) antibodies have been reported in a small portion of SSc patients especially with pulmonary fibrosis. RNPC-3, also known as U11/U12 small nuclear ribonucleoprotein, is a 65 KDa protein that has been reported as an autoantibody target and to be associated with a nuclear speckled indirect immunofluorescence (IIF) pattern on HEp-2 cells. Recently we described a novel linear B-Cell epitope that was used for the development of an autoantibody assay to detect a subpopulation of anti-RNPC-3 antibodies. Our aim in this study was to determine the prevalence of anti-RNPC-3 antibodies in a cohort of well characterized SSc patients and to assess clinical associations.

Methods: A total of 299 SSc patient samples from a large research cohort enriched for samples with a nuclear speckled IIF pattern were tested using a novel particle-based multi-analyte technology (PMAT) for antibodies to recombinant RNPC-3 (rRNPC-3) and the newly described RNPC-3 derived peptide (RNPC-3p). Immunoadsorption experiments by liquid phase inhibition was carried out to estimate the contribution of antibodies to the novel B-cell epitope compared with the whole anti-RNPC-3 antibody response. Clinical and serological associations were ascertained.

Results: The levels of antibodies to rRNPC-3 and RNPC-3p correlated moderately (See Figure, Spearman=0.32, p=0.0001; chi-squared p=0.0002). As expected, levels of anti-RNPC-3p antibodies were higher in patients positive for the recombinant antigen (p=0.0013). Both rRNPC-3 and RNPC-3p were associated with a speckled pattern. Immunoadsorption showed significant inhibition using the RNPC-3 derived peptide, but not with a control peptide. Prevalence and levels of anti-RNPC-3 antibodies were higher in anti-Scl-70, anti-Cenp, and/or anti-RNA Pol III triple negative patients [10/106 (9.4%) vs. 6/193 (3.1%), p=0.03; and median titers 122.0 vs. 107.5, p=0.08]. Patients positive for anti-RNPC-3 antibodies tended to have more interstitial lung disease, especially when considering follow-up. None of the clinical associations reached statistical significance most likely due to limited sample size and prevalence of anti-RNPC-3 antibodies.

Conclusion: Our study confirms the association of anti-RNPC-3 antibodies with the nuclear speckled IIF pattern and their increased prevalence in SSc triple negative patients (anti-Scl-70, anti-centromere, and anti RNA-Pol III antibodies). The previously identified RNPC-3 peptide represents a significant target of anti-RNPC-3 antibodies, but does not capture the entire reactivity indicating the presence of additional epitopes. Further studies are needed to verify the clinical utility of the assays.

RNPC3 Figure 1

RNPC3 Figure 2

Table Clinical characteristics

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/anti-rnpc-3-antibodies-are-associated-with-nuclear-speckled-immunofluorescence-pattern-and-enriched-in-triple-negative-systemic-sclerosis-patients/