Disease-modifying antirheumatic drugs (DMARDs) have greatly improved the treatment of rheumatoid arthritis (RA), but strategies to prevent disease onset and recurring flares remain limited. While abatacept (CTLA-4 IgG) can delay RA onset and corticosteroids are used for flare control, the benefit is temporary. We report that combining standard-of-care treatments with a locally administered immunomodulatory agent, termed Agg-CLNP, enhances both disease prevention and flare mitigation. Agg-CLNP consists of polymer nanoparticles conjugated with an immunodominant aggrecan peptide and encapsulate calcitriol. These nanoparticles are optimized for uptake by dendritic cells (DC) in lymph nodes proximal to arthritic joints. In vitro, Agg-CLNP suppressed costimulatory molecules and HLA class II (HLA-2) expression and upregulated CTLA-4 in human monocyte-derived DC from healthy and RA donors. In SKG mice, a T cell-driven RA model, Agg-CLNP combined with CTLA-4 IgG synergistically delayed disease onset and reduced severity. In a dexamethasone (Dex) withdrawal flare model, post-Dex Agg-CLNP treatment reduced flare severity and preserved a regulatory phenotype in DC, while suppressing local pathogenic TH17 cells. Next generation RNA sequencing of lymph node DC revealed Ctla4 upregulation and changes in other immunomodulatory genes linked to flare prevention. These findings highlight Agg-CLNP as a potential therapeutic strategy to address critical unmet needs in RA management.

O056 / #121 Topic:AS23 - SLE-Diagnosis, Manifestations, & Outcomes ABSTRACT CONCURRENT SESSION 09: SLE THERAPY – REVISITING OLD DRUGS AND UNLOCKING HIDDEN POTENTIAL OF NEW MEDICATIONS 24-05-2025 10:40 AM - 11:40 AM Patients with features of systemic lupus erythematosus (SLE) who do not have sufficient criteria to be classified can be designated as having incomplete lupus (ILE). This is a common condition seen in clinical practice and it has further significance as a group that has high risk of progression to SLE. Identification and treatment of those at risk has the potential to reduce the severity and incidence of SLE. Based on previous studies, hydroxychloroquine (HCQ) was chosen as an intervention for a randomized, double-blind, placebo-controlled trial to determine whether the rate of accumulation of clinical and immunologic features of SLE as defined by the 2012 SLICC criteria could be reduced. ILE was defined as ANA positivity with one or 2 additional criteria from the SLICC 2012 list. Males and females 15 to 49 years of age were eligible for enrollment. After baseline evaluation including ophthalmologic exam, participants were randomized 1:1 to HCQ or placebo. Evaluations at 3-month intervals included clinical and laboratory measures as well as patient-reported outcomes (PROs). Treatment was continued for 24 months, but if SLICC criteria were satisfied sooner, patients exited the study. Ophthalmologic exams were carried out at conclusion of treatment. A total of 187 ILE patients were randomized at 7 sites in the USA. After excluding 7 patients found to have SLE criteria at baseline when pending laboratory data were completed, 180 patients were available for analysis; 92 were randomized to HCQ and 88 received placebo. The mean age was 33 years, 91.1% were female and 74.4% were White individuals. At randomization, 65.6% had 2 SLICC criteria; the remainder had 3 SLICC criteria. The most common manifestations involved skin and joints. SLE per criteria developed in 24 participants (13.3%) during the trial who were terminated early and 40 (22%) developed additional SLICC criteria. The primary outcome was the rate of acquisition of SLICC criteria analyzed via a generalized linear mixed-effects model, with an embedded ordinal logistic regression, comparing the changes over time for the 2 arms. This showed similar slopes in the 2 groups (P=0.72). The odds of progressing to a higher SLICC score relative to the previous score was 14% smaller for every 3-month increase in time for the HCQ group and 18% smaller for the placebo group, a difference which was not statistically significant (P=0.69). A key secondary outcome was time to progression to SLE. Using a Cox proportional hazards regression model, the hazard of progressing to SLE was 10% higher for the HCQ group than for placebo, which was not a statistically significant difference (P=0.81). Adverse events were similar in the 2 groups and no serious adverse events related to use of HCQ were recorded. Five individuals were excluded from entry due to abnormal ophthalmologic findings; none developed during the trial. The SMILE results do not endorse the use of HCQ to prevent accumulation of SLICC SLE criteria. However, the definition of ILE used in SMILE does include individuals who are at risk for progressive disease and may be useful in future studies of preventive therapies. Other ongoing analyses will determine whether autoantibodies, inflammatory mediators or PROs were related to progressive illness or use of HCQ.

Lupus nephritis (LN) is a frequent manifestation of systemic lupus erythematosus, and fewer than half of patients achieve complete renal response with standard immunosuppressants. Identifying noninvasive, blood-based immune alterations associated with renal injury could aid therapeutic decisions. Here, we used mass cytometry immunophenotyping of peripheral blood mononuclear cells in 145 patients with biopsy-proven LN and 40 healthy controls to evaluate the heterogeneity of immune activation and identify correlates of renal parameters. Unbiased analysis identified 3 immunologically distinct groups of patients that were associated with different patterns of histopathology, renal cell infiltrates, urine proteomic profiles, and treatment response at 1 year. Patients with enriched circulating granzyme B+ T cells showed more active disease and increased numbers of activated CD8+ T cells in the kidney, yet they had the highest likelihood of treatment response. A second group characterized by a high type I interferon signature had a lower likelihood of response to therapy, while a third group appeared immunologically inactive but with chronic renal injuries. The major immunologic axes of variation could be distilled down to 5 simple cytometric parameters that recapitulate several clinical associations, highlighting the potential for blood immunoprofiling to translate to clinically useful noninvasive metrics to assess immune-mediated disease in LN.

OBJECTIVE:Genome-wide association studies (GWAS) have identified >100 loci for systemic lupus erythematosus (SLE). These loci may also affect age at diagnosis. We aimed to identify genetic variants for age at SLE diagnosis and to complete a GWAS of childhood-onset SLE (cSLE) in children diagnosed <18 years of age. METHODS:Patients met American College of Rheumatology and/or Systemic Lupus Erythematosus International Collaborative Clinics classification criteria, had documented age at diagnosis, and were genotyped on multiethnic arrays. Ungenotyped single nucleotide polymorphisms (SNPs) and HLA alleles were imputed to multiethnic referents. Ancestry was genetically inferred. We tested known SLE loci (142 non-HLA, 166 HLA) with log-transformed age of SLE diagnosis, adjusted for sex and five principal components (significance threshold P < 1.6 × 10-4). We also completed a GWAS of 346 patients with cSLE and 4,080 children and adolescents without SLE of European and East Asian ancestry (genome-wide significance P < 5 × 10-8). RESULTS:We included 1,489 patients with SLE, 51% of patients had cSLE, 88% of patients were female, 39% of patients were of European ancestry, 19% of patients were of East Asian ancestry, and 17% of patients were of Admixed ancestry. The median age at diagnosis was 17.7 years (interquartile range 14.0-30.9). One SLE-risk SNP, intronic to coiled-coil domain containing 113 (CCDC113) (chromosome 16), was associated with younger age at SLE diagnosis (β = -0.12, SE = 0.03, P = 6.3 × 10-6) and with cSLE versus adult-onset SLE (aSLE). GWAS of cSLE compared to controls without SLE identified significant chromosome 6 SNP rs9268469, intronic to TSBP1-AS1 (odds ratio 2.04 [95% confidence interval 1.59-2.63], P = 1.79 × 10-8) and HLA-DQA1. CONCLUSION:We identified a significant locus for younger age at SLE diagnosis, intronic to CCDC113, among a large multiancestral cohort of children and adults with SLE. In the first GWAS of cSLE, we identified a TSBP1-AS1 locus and an HLA-DQA1 previously identified for aSLE.

O061 / #515 Topic:AS15 - Lupus Nephritis-Clinical ABSTRACT CONCURRENT SESSION 10: INTEGRATING PROTEOMIC & TRANSCRIPTOMICS IN SLE 24-05-2025 10:40 AM - 11:40 AM Lupus nephritis (LN) can cause severe complications and early mortality in SLE patients. Despite advancements, treatments for LN are not consistently effective and often have adverse effects. This study aims to develop a robust, noninvasive biomarker panel to predict treatment response, using high-throughput proteomics and machine learning to model serum protein expression, as part of the Accelerating Medicines Partnership RA/SLE Network. Over 5,000 proteins were measured in the serum of 158 LN patients at the time of diagnostic kidney biopsy (baseline) and 12 weeks post-biopsy using Olink Explore HT. Clinical response was determined at 52 weeks as complete (CR; n=25), partial (PR; n=22), or no response (NR; n=49). Multivariate logistic regression adjusting for age, gender, and genetic ancestry and machine learning algorithm approaches (Extreme Gradient Boosting) were used to generate a robust prediction model of treatment responsiveness. At baseline, patients with NR exhibited 515 (p-value < 0.05; 160 upregulated) dysregulated proteins within the innate immune system, platelet activation, and pathways seen in neurodegeneration compared to CR. In addition, 1227 (p-value < 0.05; 1180 upregulated) proteins involving the TGFb, IL-10, Th1/Th2 differentiation, platelet activation, and leukocyte transendothelial migration pathways at 12 weeks post-biopsy were differentially expressed in patients with NR compared to CR. Proteins involved in T cell proliferation, Th17 cell differentiation, and TNF, Wnt, EGFR, and IL-10 signaling were persistently elevated at week 12 in NR compared to CR using paired analysis. The proteomic profiles of CR and NR are easily distinguishable at 52 weeks (AUC, 0.85 ± 0.10 with cross-validation accuracy of 76.1% ± 10.2%). While the ML models at baseline and 12 weeks showed less robust prediction performance with 65.9% ± 7.6% and 70.4% ± 7.3% (Figure 1A-C), the model that incorporates the baseline protein levels and the changes from baseline to 12 weeks post treatment showed the most robust prediction with AUC of 0.89 ± 0.06 and accuracy of 81.5% ± 6.2%. In particular, patients with a CR had a significant reduction in CD27, VEGF, HAVCR2, MEGF11, and VSIG4 from baseline to 12 weeks post-biopsy (Figure 1D-E). Furthermore, preliminary trajectory analyses have demonstrated the rapid decline of these proteins before treatment with the levels plateauing near the levels seen in healthy controls throughout the 52-week of study trial. Gene regulatory network analyses of the top predictors demonstrated significantly downregulated lymphocyte activation/differentiation (CD27, IL-7, IL-3, IL-16, CD83, and IL-10) and cellular migration pathways (NRP1, IL-16, PDGFB, CSF1, DDR1, FSTL1) in patients with a CR at week 12. Figure 1. Early downregulation of specific immune pathways upon treatment precedes future clinical response in LN. Changes in serum protein expression, especially the soluble surface receptors shed upon cellular activation, at 12 weeks post-biopsy may serve as noninvasive biomarkers of 52-week treatment response.

OBJECTIVE:Patients with systemic lupus erythematosus (SLE) experience photosensitivity, with exposure to UVB light driving lupus flares and triggering symptoms like joint pain, fatigue, and cutaneous lesions. Although the mechanism(s) linking UVB exposure to systemic effects are unclear, type I interferons (IFNs) are known to play a role. Our previous work has shown that TRIM21, an autoantigen in SLE, functions as a negative regulator on the pathways driving IFN expression. Here we explore how TRIM21 functions to regulate both local and systemic inflammation following UVB exposure and how altered expression may drive cutaneous inflammation and photosensitivity in SLE. METHODS:Wild-type (WT; C57BL/6) and Trim21-/- mice were irradiated with UVB (100 mJ/cm2) on the shaved dorsal region on consecutive days for 1 and 3 weeks, and UVB-induced local cutaneous manifestations and systemic inflammation in blood, spleen, and kidney were examined by messenger RNA expression of inflammatory and type I IFN response genes, histology, and flow cytometry. Mechanistic studies were performed in bone marrow-derived macrophages (BMDMs) and murine dermal fibroblasts (MDFs) from WT and Trim21-/- mice and TRIM21-/- THP-1 cells. RESULTS:Infiltration of inflammatory cells and induction of type I IFN developed in UVB-exposed areas in both sets of mice. Most notably after UVB exposure, we observed splenomegaly and enhanced expression of IFN-stimulated genes in the blood and spleen of Trim21-/- mice. Inflammatory chemokines CXCL10 and CXCL12 were also detected at significantly higher levels in serum of Trim21-/- mice after UVB exposure. Trim21-/- mice exposed to UVB also demonstrated enhanced total IgG levels in serum accompanied by increased skin and kidney deposition of IgG and increased glomerular cellularity and size. To determine the mechanism, we assessed UVB- and cyclic GMP-AMP-dependent Ifnb1 expression in Trim21-/- BMDMs and MDFs, noting increased responses compared with WT cells. This effect was lost in BMDMs from Trim21/Sting1 double knockout mice and skin explants, in keeping with the ability of TRIM21 to regulate cytoplasmic DNA sensing. In keeping with previous reports, we found that degradation of both DDX41 and STING levels were affected in stimulated Trim21-/- BMDMs. CONCLUSION:Taken together, our results indicate that TRIM21 protects against IFN induction at both local and systemic levels by restricting STING signaling.

Systemic lupus erythematosus (SLE) patients are 90% women and over three times more likely to die of cardiovascular disease than women in the general population. Chest pain with no obstructive cardiac disease is associated with coronary microvascular disease (CMD), where narrowing of the small blood vessels can lead to ischemia, and frequently reported by SLE patients. Using whole blood RNA samples, we asked whether gene signatures discriminate SLE patients with coronary microvascular dysfunction (CMD) on cardiac MRI (n=4) from those without (n=7) and whether any signaling pathway is linked to the underlying pathobiology of SLE CMD. RNA-seq analysis revealed 143 differentially expressed (DE) genes between the SLE and healthy control (HC) groups, with virus defense and interferon (IFN) signaling being the key pathways identified as enriched in SLE as expected. We next conducted a comparative analysis of genes differentially expressed in SLE-CMD and SLE-non-CMD relative to HC samples. Our analysis highlighted differences in IFN signaling, RNA sensing and ADP-ribosylation pathways between SLE-CMD and SLE-non-CMD. This is the first study to investigate possible gene signatures associating with CMD in SLE, and our data strongly suggests that distinct molecular mechanisms underly vascular changes in CMD and non-CMD involvement in SLE.

OBJECTIVE:Patients with features of systemic lupus erythematosus (SLE) who do not fulfill classification criteria can be designated as incomplete lupus erythematosus (ILE). This condition includes individuals with a high risk of progression to SLE. Treatment of ILE may reduce symptoms, severity, and incidence of SLE. METHODS:Hydroxychloroquine (HCQ) was chosen as an ILE intervention for a randomized, double-blind trial to determine whether the rate of accumulation of SLE features defined by the 2012 Systemic Lupus Erythematosus International Collaborating Clinics (SLICC) criteria could be reduced. ILE was defined as antinuclear antibody positivity with one to two additional criteria. Patients 15 to 49 years old were eligible. Randomization was 1:1 HCQ to placebo. Evaluations were at 3-month intervals over 24 months. Meeting SLICC classification sooner required exit. RESULTS:Participants (N = 187) were randomized at seven sites. After excluding 7 patients who met SLE classification at baseline when screening laboratory data were completed, 180 patients were analyzed: 92 receiving HCQ and 88 receiving placebo. Considering all these enrollees, 55 developed additional criteria. Of the 118 participants who exited early with SLE or who completed 24 months of evaluation, SLE classification developed in 24 (13.3%); another 24 developed additional criteria but did not meet classification. The rates of acquisition of SLICC criteria and progression to SLE were similar in the two groups (P = 0.72 and P = 0.98, respectively). Development of SLE was associated with new malar rash, oral ulcers, joint tenderness, or pleurisy (P < 0.04). CONCLUSION:Although the Study of Antimalarials in Incomplete Lupus Erythematosus (SMILE) did not show effects of HCQ on ILE progression, the results offer insights into SLE risk in the ILE population.

PV107 / #499 Poster Topic:AS12 - Genetics, Epigenetics, Transcriptomics Lupus nephritis (LN) is one of the most common and severe manifestations of systemic lupus erythematosus (SLE). We performed genome-wide association studies (GWAS) for lupus nephritis and kidney function measures over time. We hypothesized that analyzing a person’s eGFR variability over time would be a good proxy for LN and improve power for detecting genetic loci for LN. We also used local ancestry estimation to facilitate inclusion of admixed individuals. We included SLE patients from several child and adult dedicated lupus databases and the Systemic Lupus International Collaborating Clinics (SLICC) cohort. All met American College of Rheumatology and/or SLICC SLE criteria and were genotyped on a multi-ethnic Illumina array. Ungenotyped SNPs were imputed to the Trans-Omics for Precision Medicine program (TopMed), and local ancestry of chromosomal information was estimated using RFMix and Tractor software. LN was defined by SLE criteria, with a subset confirmed by kidney biopsy. Kidney function (estimated glomerular filtration rate, eGFR) was calculated using the Schwartz formula for measures <18 years and CKD-EPI for >18 years of age. Wilcoxon rank-sum or Chi-square tests were used to compare characteristics between LN and Non-LN patients. We completed separate GWAS for the outcomes of LN, mean eGFR and eGFR variability over time (log of the mean absolute deviation from mean eGFR per participant), in marginal and multivariable-adjusted regression models with sex, site and local principal components using Regenie. Local ancestry analysis was restricted to individuals of European, African and East Asian ancestry using Tractor. We meta-analyzed ancestry-specific results with METAL software (significancep<5x10^-8). We studied 2981 individuals with SLE, 88% female, 46% of European ancestry, 27% childhood-onset SLE, and 45% with LN (Table). Kidney failure was observed in 25 patients over a median follow-up time of 8.9 years (IQR: 4.1,14.8). Within-person eGFR was similar between people with and without LN, but eGFR variability was significantly greater in people with LN (P-value= 2.2e-16). Variability was calculated using a median of 16 [IQR: 8,35] eGFR measurements per person. GWAS of LN did not identify a significant LN locus, yet GWAS of eGFR variability demonstrated a significant peak on chromosome 15, downstream ofSCH4and intronic toSECISBP2L(Figure). The variant was found in a genomic region of African ancestry. Table. Figure. We completed GWAS of LN, eGFR mean and variability over time, generating ancestry-specific estimates and identified a genome-wide significant locus for variability in measures of renal function over time, in a multiethnic cohort of children and adults with SLE. This locus was only found in an African ancestry portion of the genome. Variability in measures of renal function is correlated with LN, yet GWAS of LN did not identify significant loci. Future work includes repeating analyses to include all global ancestries, and to investigate the biologic link between the loci and LN.

Objectives Factors associated with peripheral arthritis and enthesitis, especially Achilles tendonitis and plantar fasciitis, were examined in a longitudinal cohort of 1075 patients with ankylosing spondylitis (AS) (also known as radiographic axial spondyloarthritis). Methods Patients were derived from the Prospective Study of Outcomes in Ankylosing Spondylitis cohort. Disease activity and functional indices, as well as physical examination and medications used, were measured at every study visit. Univariable and multivariable analyses of the association of peripheral arthritis and enthesitis with clinical, sociodemographic factors were performed. Human leucocyte antigen (HLA)-B alleles were analysed by single-stranded conformational polymorphism analysis. Results Those with peripheral arthritis on examination were more likely to have psoriasis (p=0.001, OR=1.68; CI, 1.11, 2.54), greater functional impairment (p<0.001 OR=1.72; CI, 1.31, 2.27), higher erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) levels (p<0.001), greater tumour necrosis factor (TNF) inhibitor (p<0.001, OR=1.50; CI, 1.14, 1.97) and use methotrexate/sulfasalazine (p<0.001, OR=2.25, CI [1.57, 3.23]). Patients with enthesitis were less likely to be male (p<0.001, OR=0.57; CI, 0.43, 0.75) and have peripheral arthritis (p<0.001, OR=2.35; CI, 1.47, 3.75), greater functional impairment (p<0.001, OR=1.91; CI, 1.43, 2.55) and higher ESR/CRP levels (p<0.001). Patients with plantar fasciitis and/or Achilles’ tendonitis on examination were less likely to male (p<0.001 OR=0.57; CI, 0.43, 0.75), to have significant functional impairment (p<0.001), to be using TNF inhibitors (p<0.001, OR=1.48; CI 1.13, 1.93) and to be using either sulfasalazine or methotrexate (p<0.001, OR=1.86, CI, 1.30, 2.67). HLA-B*15 (p=0.03, OR=1.84; CI, 1.05, 3.21) and HLA-B*37 (p=0.04, OR=3.00; CI, 1.03, 8.74) were marginally increased in frequency in those with peripheral arthritis on examination compared with those without. Conclusion There was a higher prevalence of peripheral musculoskeletal manifestations in women with AS, with significant impact on physical function and greater use of methotrexate or sulfasalazine and TNF inhibitors and enrichment for certain non-HLA-B27 HLA-B alleles.

Objective To investigate whether gene signatures discriminate systemic lupus erythematosus (SLE) patients with coronary microvascular dysfunction (CMD) from those without and whether any signaling pathway is linked to the underlying pathobiology of SLE CMD.Methods This study collected whole blood RNA samples from female subjects aged 37 to 57, comprising 11 SLE patients (4 SLE-CMD, 7 SLE-non-CMD) and 10 HC. Total RNA was then used for library preparation and sequencing. Differential gene expression analysis was performed to identify gene signatures associated with CMD in SLE patients using DEseq2 v1.42.0. Gene Set Enrichment Analysis were performed by ClusterProfiler v4.10.0 and pathfindR v2.3.1.Results RNA-seq analysis revealed 143 differentially expressed (DE) genes between the SLE and HC groups. GO analysis indicated associations with virus defense and interferon signaling in SLE. 14 DE genes were identified from comparison between SLE-CMD and SLE-non-CMD with adjusted parameters (padj < 0.1). Notably, SLE-CMD exhibited elevated levels of genes associated with RNA sensing, while downregulated genes in SLE-non-CMD were associated with blood coagulation and cell-cell junction. Further investigation highlighted differences in IFN signaling and ADP-ribosylation pathways between SLE-CMD and SLE-non-CMD, suggesting distinct molecular mechanisms underlying vascular changes in CMD and reduced left ventricular function in non-CMD.Conclusion Our study identified a unique gene signature in SLE-CMD compared to the HC group, highlighting the significant involvement of type 1 interferon, RIG-I family proteins, and chronic inflammation in the progression of SLE-CMD. The intricate relationship between SLE-CMD and these factors underscores their probable role in initiating and advancing SLE-CMD.### Competing Interest StatementThe authors have declared no competing interest.

Objectives: To examine the association of multimorbidity phenotypes at baseline with disease activity and functional status over time in ankylosing spondylitis (AS).Methods: Patient-reported AS morbidities (comorbidities, N = 28 and extra-musculoskeletal manifestations, EMMs, N = 3) within 3 years of enrollment with a prevalence >= 1 %, were included from the Prospective Study of Outcomes in Ankylosing Spondylitis (PSOAS) cohort. We defined multimorbidity as >= 2 morbidities (MM2+) and substantial multimorbidity as >= 5 morbidities (MM5+). Multimorbidity clusters or phenotypes were identified using K-median clustering. Disease activity (ASDAS-CRP) and functional status (BASFI) measures were collected every 6 months. Generalized estimating equation method was used to examine the associations of multimorbidity counts and multimorbidity clusters with measures of disease activity and functional status over time.Results: Among 1,270 AS patients (9,885 visits) with a median follow-up of 2.9 years (IQ range: 1.0-6.8 years), the prevalence of MM2+ and MM5+ was 49 % and 9 % respectively. We identified five multimorbidity clusters: depression (n = 321, 25 %), hypertension (n = 284, 22 %), uveitis (n = 274, 22 %), no morbidities (n = 238, 19 %), and miscellaneous (n = 153, 12 %). Patients in the depression cluster were more likely to be female and had significantly more morbidities and worse disease activity and functional status compared to those with no morbidities.Conclusion: Approximately 49 % of AS patients in the PSOAS cohort had multimorbidity and five distinct multimorbidity phenotypes were identified. In addition to the number of morbidities, the type of morbidity appears to be important to longitudinal outcomes in AS. The depression cluster was associated with worse disease activity and function.