Background/Purpose: We previously validated T Cell-bound C4d (TC4d), T Cell-bound IgG (TIgG), and T Cell-bound IgM (TIgM) biomarkers, demonstrating high sensitivity and specificity for differentiating systemic lupus erythematosus (SLE) from apparently healthy volunteers (AHVs) and other autoimmune rheumatic diseases (ARDs). The current study aims to further evaluate the specificity of these biomarkers in distinguishing SLE from other suspected anti-nuclear antibody (ANA)-associated ARDs within a large cohort.

Methods: Three cohorts were analyzed: 71 clinically diagnosed SLE patients meeting the 2019 ACR/EULAR classification criteria; 162 AHVs; and 31,221 subjects from a de-identified population undergoing autoimmune testing (Jan–Apr 2025). Suspected ARDs were identified based on autoantibody levels exceeding ten times the diagnostic assay cutoffs (anti-CCP for RA; anti-Ro52, anti-Ro60, and anti-SSB/La for Sjögren’s syndrome; anti-centromere protein B for limited scleroderma; anti-Scl-70 for diffuse scleroderma; and anti-Jo-1 for myositis) and negative for anti-dsDNA and anti-Smith antibodies. Semiquantitative flow cytometry was used to measure cell-bound TC4d, TIgG, and TIgM, reported as median fluorescence intensity (MFI). Autoantibody measurements utilized ELISA or enzyme linked fluorescent assay (ELFA) methods per kit recommendations (ThermoFisher, Phadia). Biomarker distributions, sensitivity, specificity, likelihood ratios (LRs), and diagnostic odds ratios (DORs) were assessed at thresholds achieving 95% specificity vs. AHVs, 95% specificity vs. ARDs, and 99% specificity vs. ARDs. Mann-Whitney U tests evaluated significant differences.

Results: SLE patients (N = 71) had a mean (SD) age of 42.6 (9.8) years, were predominantly female (83.1%), ANA IFA-positive (83.1%), and ANA ELISA-positive (97.2%). Suspected ARD groups included RA (N = 648), Sjögren’s (N = 140), limited scleroderma (N = 366), diffuse scleroderma (N = 26), and myositis (N = 22) [Table 1]. At thresholds providing 95% specificity vs. AHVs, the T Cell biomarkers showed specificities of 87.5% (TC4d), 87.3% (TIgG), and 83.5% (TIgM) for differentiating SLE from ARDs (Table 2). Sensitivity for SLE at 95% specificity for SLE vs. ARD was 64.8% (TC4d), 54.9% (TIgG), and 36.6% (TIgM). At thresholds providing 99% specificity vs. ARDs, sensitivities ranged from 11% to 28%. The DORs at 95% specificity vs. AHVs were 24.1 (TC4d), 16.3 (TIgG), and 7.3 (TIgM). Median TC4d, TIgG, and TIgM levels were significantly higher in SLE patients compared to all other ANA-associated ARDs (Figure 1A-F, p < 0.01). At a moderate pre-test probability for SLE (30%), the post-test probabilities following a positive TC4d result were 72.7% (95% specificity vs. AHVs), 84.5% (95% specificity vs. ARDs), and 91.8% (99% specificity vs. ARDs) [Figure 1G-I]. Conversely, negative TC4d results reduced the post-test probability to 9.9%, 13.7%, and 23.7%, respectively.

Conclusion: Cell-bound TC4d, TIgG, and TIgM exhibit robust specificity for differentiating SLE from other ANA-associated ARDs. The availability of multiple threshold levels allows flexible clinical application, enhancing diagnostic decision-making through optimized rule-in or rule-out approaches.

Table 1. Demographic characteristics

Table 2. Performance characteristics

Figure 1. Biomarker distributions and diagnostic performances at varying levels of specificity for SLE

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/specificity-of-cell-bound-t-cell-biomarkers-tc4d-tigg-tigm-in-differentiating-sle-from-ana-associated-rheumatic-diseases/