Longitudinal ANA and ENA Dynamics in Preclinical Systemic Autoimmune Rheumatic Diseases: A Retrospective Cohort Study

Objectives To characterize longitudinal changes in antinuclear antibody (ANA) titers and patterns, together with the autoantibody profile, in asymptomatic ANA-positive individuals and undifferentiated connective tissue disease (UCTD) patients, and to identify serological predictors of systemic autoimmune rheumatic diseases (SARD) progression. Methods We analyzed autoimmune serology from 225 asymptomatic ANA-positive or UCTD subjects with longitudinal follow-up (1-7 years) to assess changes over time, seroconversion, and progression to (SARD). ANAs were quantified by indirect IF using the Kallestad® HEp-2 kit and specific autoantibodies measured using the Bioplex® 2200 ANA Screening System which assesses the levels of anti-dsDNA, -chromatin, -Ro, -La, -Sm, -SmRNP, -RNP, -Jo-1, -Scl-70, -centromere and -ribosomal P antibodies. Results At baseline, a high ANA titer (≥1:640) predominated in 60% of subjects, with speckled and homogeneous patterns representing the majority of immunofluorescent patterns. The most prevalent baseline autoantibodies were Ro (23.3%) and RNP (15.7%). Among subjects with serial ANA testing (n=77), 60% had initial high titers (≥1:640). Upon follow-up, 53% had declining titers with 6.5% seroconverting to ANA-negative. Of those who seroconverted, 60% had high titers (≥1:640) initially and 40% had a dense fine speckled (DFS) pattern. The median time to ANA loss was 3.3 years. Among the 6 subjects who ever demonstrated a DFS pattern, none of whom progressed to SARD, 33.3% converted to negative, and 33.3% transitioned to other ANA patterns. This variability underscores the dynamic but low-risk nature of DFS pattern. Among the 135 subjects with serial autoantibody profiling, Ro remained the most prevalent antibody (40%) and was usually stably elevated (89% remained persistently positive), showing minimal fluctuation over time. In contrast, RNP (17%) and dsDNA (6.7%) showed the greatest instability, with 44% of dsDNA-positive and 22% of RNP-positive subjects becoming negative. Interestingly, Scl-70, which was positive in 9 patients, also showed instability, with 4 of these patients losing positivity over time. Nine patients progressed to a defined SARD after a median of 31 months. Overall, progressors had high ANA titers (≥1:320), and none became ANA-negative. Among this group, dsDNA (89%) and Ro (SSA) (56%) were the most prevalent autoantibodies. Conclusion Many subjects demonstrated immunological improvements. Subjects who achieved ANA negativity typically began with high ANA titers that declined over time. In contrast, progressors showed persistently high ANA titers, frequently accompanied by rising dsDNA and stable Ro reactivity. These findings support the clinical importance of serial ANA and ENA monitoring to differentiate transient autoimmunity from evolving systemic disease.