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Protein tyrosine phosphatase non-receptor type 22 (PTPN22) is a negative regulator of T-cell activation associated with several autoimmune diseases, including systemic lupus erythematosus (SLE). Missense rs2476601 is associated with SLE in individuals with European ancestry. Since the rs2476601 risk allele frequency differs dramatically across ethnicities, we assessed robustness of PTPN22 association with SLE and its clinical sub-phenotypes across four ethnically diverse populations. Ten SNPs were genotyped in 8220 SLE cases and 7369 controls from in European-Americans (EA), African-Americans (AA), Asians (AS), and Hispanics (HS). We performed imputation-based association followed by conditional analysis to identify independent associations. Significantly associated SNPs were tested for association with SLE clinical sub-phenotypes, including autoantibody profiles. Multiple testing was accounted for by using false discovery rate. We successfully imputed and tested allelic association for 107 SNPs within the PTPN22 region and detected evidence of ethnic-specific associations from EA and HS. In EA, the strongest association was at rs2476601 (P = 4.7 × 10(-9), OR = 1.40 (95% CI = 1.25-1.56)). Independent association with rs1217414 was also observed in EA, and both SNPs are correlated with increased European ancestry. For HS imputed intronic SNP, rs3765598, predicted to be a cis-eQTL, was associated (P = 0.007, OR = 0.79 and 95% CI = 0.67-0.94). No significant associations were observed in AA or AS. Case-only analysis using lupus-related clinical criteria revealed differences between EA SLE patients positive for moderate to high titers of IgG anti-cardiolipin (aCL IgG >20) versus negative aCL IgG at rs2476601 (P = 0.012, OR = 1.65). Association was reinforced when these cases were compared to controls (P = 2.7 × 10(-5), OR = 2.11). Our results validate that rs2476601 is the most significantly associated SNP in individuals with European ancestry. Additionally, rs1217414 and rs3765598 may be associated with SLE. Further studies are required to confirm the involvement of rs2476601 with aCL IgG.

Systemic lupus erythematosus (SLE) is an inflammatory autoimmune disease with a strong genetic component. African-Americans (AA) are at increased risk of SLE, but the genetic basis of this risk is largely unknown. To identify causal variants in SLE loci in AA, we performed admixture mapping followed by fine mapping in AA and European-Americans (EA). Through genome-wide admixture mapping in AA, we identified a strong SLE susceptibility locus at 2q22–24 (LOD = 6.28), and the admixture signal is associated with the European ancestry (ancestry risk ratio ∼1.5). Large-scale genotypic analysis on 19,726 individuals of African and European ancestry revealed three independently associated variants in the IFIH1 gene: an intronic variant, rs13023380 [Pmeta = 5.20×10−14; odds ratio, 95% confidence interval = 0.82 (0.78–0.87)], and two missense variants, rs1990760 (Ala946Thr) [Pmeta = 3.08×10−7; 0.88 (0.84–0.93)] and rs10930046 (Arg460His) [Pdom = 1.16×10−8; 0.70 (0.62–0.79)]. Both missense variants produced dramatic phenotypic changes in apoptosis and inflammation-related gene expression. We experimentally validated function of the intronic SNP by DNA electrophoresis, protein identification, and in vitro protein binding assays. DNA carrying the intronic risk allele rs13023380 showed reduced binding efficiency to a cellular protein complex including nucleolin and lupus autoantigen Ku70/80, and showed reduced transcriptional activity in vivo. Thus, in SLE patients, genetic susceptibility could create a biochemical imbalance that dysregulates nucleolin, Ku70/80, or other nucleic acid regulatory proteins. This could promote antibody hypermutation and auto-antibody generation, further destabilizing the cellular network. Together with molecular modeling, our results establish a distinct role for IFIH1 in apoptosis, inflammation, and autoantibody production, and explain the molecular basis of these three risk alleles for SLE pathogenesis., Author Summary African-Americans (AA) are at increased risk of systemic lupus erythematosus (SLE), but the genetic basis of this risk increase is largely unknown. We used admixture mapping to localize disease-causing genetic variants that differ in frequency across populations. This approach is advantageous for localizing susceptibility genes in recently admixed populations like AA. Our genome-wide admixture scan identified seven admixture signals, and we followed the best signal at 2q22–24 with fine-mapping, imputation-based association analysis and experimental validation. We identified two independent coding variants and a non-coding variant within the IFIH1 gene associated with SLE. Together with molecular modeling, our results establish a distinct role for IFIH1 in apoptosis, inflammation, and autoantibody production, and explain the molecular basis of these three risk alleles for SLE pathogenesis.

We previously established an 80 kb haplotype upstream of TNFSF4 as a susceptibility locus in the autoimmune disease SLE. SLE-associated alleles at this locus are associated with inflammatory disorders, including atherosclerosis and ischaemic stroke. In Europeans, the TNFSF4 causal variants have remained elusive due to strong linkage disequilibrium exhibited by alleles spanning the region. Using a trans-ancestral approach to fine-map the locus, utilising 17,900 SLE and control subjects including Amerindian/Hispanics (1348 cases, 717 controls), African-Americans (AA) (1529, 2048) and better powered cohorts of Europeans and East Asians, we find strong association of risk alleles in all ethnicities; the AA association replicates in African-American Gullah (152,122). The best evidence of association comes from two adjacent markers: rs2205960-T (P = 1.71×10-34, OR = 1.43[1.26-1.60]) and rs1234317-T (P = 1.16×10-28, OR = 1.38[1.24-1.54]). Inference of fine-scale recombination rates for all populations tested finds the 80 kb risk and non-risk haplotypes in all except African-Americans. In this population the decay of recombination equates to an 11 kb risk haplotype, anchored in the 5′ region proximal to TNFSF4 and tagged by rs2205960-T after 1000 Genomes phase 1 (v3) imputation. Conditional regression analyses delineate the 5′ risk signal to rs2205960-T and the independent non-risk signal to rs1234314-C. Our case-only and SLE-control cohorts demonstrate robust association of rs2205960-T with autoantibody production. The rs2205960-T is predicted to form part of a decameric motif which binds NF-κBp65 with increased affinity compared to rs2205960-G. ChIP-seq data also indicate NF-κB interaction with the DNA sequence at this position in LCL cells. Our research suggests association of rs2205960-T with SLE across multiple groups and an independent non-risk signal at rs1234314-C. rs2205960-T is associated with autoantibody production and lymphopenia. Our data confirm a global signal at TNFSF4 and a role for the expressed product at multiple stages of lymphocyte dysregulation during SLE pathogenesis. We confirm the validity of trans-ancestral mapping in a complex trait. © 2013 Manku et al.

Objective. To investigate immunoreactivity of systemic lupus erythematosus (SLE) sera with apolipoprotein A1, (Apo A1), the major lipid-binding protein of high-density lipoprotein (HDL). Methods. Since early attempts to identify Apo A1 autoantibodies using standard enzyme-linked immunosorbent assay (ELISA) and immunoblotting techniques had been unsuccessful, a mouse complementary DNA lambda phage expression library was screened. Results. A selected clone (MA1) was found to have 82% DNA sequence homology to a segment of human Apo A1. Since there were nonconservative substitutions in the MA1 protein and lack of a complete sequence, it was possible that the SLE patient'S antibodies were binding MA1 epitopes that were shared by the complete human protein but had not been conformationally accessible using the earlier techniques. Thus, gamma-irradiated ELISA plates were used as an alternative antigen-binding surface for intact human Apo A1, and high-titer anti-human Apo A1 autoantibodies were then identified in the sera of 5 more SLE patients. Conclusion. These findings show that Apo A1 is immunogenic. Apo A1 antibodies may play a role in the decreased HDL levels and Apo A1:Apo B ratios previously reported to occur in subgroups of SLE patients.

Background Targeted, biologic inhibitors of B-cell Activating Factor (BAFF) have been evaluated in 4 large Phase 3 clinical trials in nearly 4000 patients with Systemic Lupus Erythematosus (SLE). Data from these studies identify that greatest treatment effect is discernable using more stringent endpoints in patients who enter with higher disease activity scores, taking more corticosteroids, and/or have anti-double-stranded DNA (dsDNA) and low complement C3 or C4 1,2,3 . Objectives The CHABLIS-SC1 study (NCT01395745) has completed enrollment and will be the first trial to prospectively evaluate the impact of treatment on the above “responder populations” using the BAFF inhibitor, blisibimod. Methods CHABLIS-SC1 enrolled subjects positive for anti-nuclear and/or anti-dsDNA antibodies with SELENA-SLEDAI≥10 despite corticosteroid therapy. Subjects were randomized to have blisibimod or placebo added to their background medications. Efficacy will be evaluated using the SLE Responder Index-6 (SRI-6) at Week 52, defined as: ≥6 point reduction in SELENA-SLEDAI; no new BILAG A or 2 new BILAG B organ domain scores; and no worsening in Physician9s Global Assessment. Results Table 1 identifies that the baseline characteristics of the 442 subjects enrolled into the CHABLIS-SC1 study more frequently meet all descriptions of “responder populations” identified previously 2–7 . In previous studies, the SRI-6 was associated with lower placebo responder rates and greater benefit of BAFF inhibitors over placebo. Conclusions BAFF inhibitors are known to be at least modestly effective in SLE and impact B Cell-driven immunopathology such as autoantibodies. The CHABLIS-SC1 study design will determine whether enrichment for the hypothesized “responder population,” patients with more severe, immunologically active SLE, will robustly discriminate treatment impact of blisibimod when background therapy is maintained. References van Vollenhoven RF et al. Ann Rheum Dis. 2012;71:1343. Isenberg DA, et al. Ann Rheum Dis. 2015 Sep. Merrill JT et al. Ann Rheum Dis. 2015 Aug. Dooley MA et al Lupus, 2013;22(1):63, Manzi et al Ann Rheum Dis. 2012;71(11):1833, Merrill et al 2015 (EULAR abstract); Benlysta FDA Drug Approval Package. Acknowledgement We wish to thank all of the patients, Investigators and study personnel participating in this trial. Disclosure of Interest J. Merrill Consultant for: Anthera Pharmaceuticals Inc, V. Strand Consultant for: Anthera, C. Hislop Shareholder of: Anthera, Employee of: Anthera, M. Gangal Shareholder of: Anthera, Employee of: Anthera, R. Martin Shareholder of: Anthera, Employee of: Anthera, K. Kalunian Consultant for: Anthera

Background Most clinical trials of new treatments for systemic lupus erythematosus (SLE) have shown weak discrimination between investigational agents and placebo when added to standard of care (SOC). Given that targeted biologics are unlikely to be effective in all patients, it is challenging for these experimental therapies to exceed the high response rates observed with SOC alone in placebo arms. The design of future SLE trials may be improved by considering strategies for reducing placebo response rates and obtaining a better understanding of the within-patient variability in disease activity during follow up. Objectives The goal of this study was to evaluate longitudinal patterns of response in SLE patients who received placebo plus SOC in two completed 52-week clinical trials. Baseline characteristics discriminating early and persistent responders from non-responders to SOC were also examined to identify patient populations who may benefit most from experimental therapies and could be targeted for enrollment in future trials. Methods Data was obtained from the Collective Data Analysis Initiative (CDAI) of the Lupus Foundation of America and included 147 patients from the placebo plus SOC arms of two randomized Phase II/III trials in moderately-to-severely active lupus patients without acute nephritis. A BILAG-based response was evaluated at weeks 12, 24, 36, 48, and 52. Both cross-sectional and longitudinal analyses of response patterns were performed. Clinical variables, including background medications and baseline factors associated with disease severity, that discriminated persistent responders from non-responders were identified using logistic regression. Results The cross-sectional response rates ranged from 37% - 46% between 12 - 52 weeks for patients treated with placebo plus SOC, similar to the placebo response rates estimated in most non-nephritis lupus trials. The response rate decreased to 14.3% (95% CI: 8.6% - 19.9%) when the criterion was complete response, i.e., response at all visits between 12 - 52 weeks. Agreement between response status at 12 weeks and 36–52 weeks was low (kappa =0.15 - 0.25); furthermore only 31% of initial 12 week responders maintained response at all subsequent visits. Factors contributing to non-response to SOC at all visits included more severe disease, as suggested by more organ systems active at baseline, and low C3 levels at baseline. Less aggressive immunosuppression was also associated with non-response. Conclusions An endpoint based on a sustained rather than cross-sectional response may reduce high placebo response rates in SLE trials that continue aggressive SOC and may improve discrimination between effective experimental treatments and SOC. Although there is increasing interest in conducting efficient 12 or 24 week early phase trials, the observed lack of stability in response to SOC over time highlights a potential weakness with shorter studies that use an endpoint of improvement. Our data also confirm earlier reports that the likelihood of response in the placebo group depends on the severity of disease and the aggressiveness of background treatments. Disclosure of Interest M. Kim: None declared, J. Merrill: None declared, K. Kalunian Consultant for: Bristol-Myers Squibb, MedImmune, AstraZeneca, Merck Serono, UCB, and Genentech, B. Hahn Grant/research support from: Bristol-Myers Squibb, Consultant for: Eli Lilly, A. Roach: None declared, P. Izmirly: None declared

Background Increased activity of the type I interferon (IFN) pathway is central to the pathogenesis of SLE. Blocking the type I receptor may reduce SLE activity more effectively than inhibiting IFN-α alone. Objectives Efficacy and safety of anifrolumab were assessed in a Phase IIb, randomized, double-blind, placebo-controlled study of adults with moderate to severe SLE (the MUSE study). Methods 305 patients were treated for 48 weeks with intravenous anifrolumab (300 mg or 1000 mg) or placebo, in addition to standard-of-care medications. Randomization was stratified by SLE Disease Activity Index 2000 (SLEDAI-2K) score ( Results The primary endpoint was met by more anifrolumab-treated patients [placebo vs. 300 mg vs. 1000 mg: 17.6%, 34.3% ( p =0.014), 28.8% ( p =0.063)] with greater effect sizes observed in the 75% of patients who had a high baseline IFN signature [13.2%, 36.0% ( p =0.004), 28.2% ( p =0.029)]. At Day 365 more anifrolumab-treated patients achieved SRI(4) responses [40.2%, 62.6%, ( p p =0.043)], BILAG-based Composite Lupus Assessment (BICLA) [25.7%, 53.5% ( p p =0.018)], modified SRI(6) [28.4%, 49.5% ( p =0.002), 44.7% ( p =0.015)], and SLEDAI-2K ≤2 [17.6%, 35.4% ( p =0.004), 32.7% ( p =0.012)]. Major clinical response (BILAG “C” or better in all domains at Day 169 maintained to Day 365) was achieved by 6.9%, 19.2% ( p =0.012), and 17.3% ( p =0.025) of patients. BILAG “A” flares were reported in more placebo- vs. anifrolumab-treated patients (16.7%, 9.1%, 10.6%). In patients with baseline Cutaneous Lupus Erythematosus Disease Area and Severity Index activity score ≥10, more anifrolumab-treated patients attained a ≥50% reduction by Day 365 [30.8%, 63.0% ( p =0.013), 58.3% ( p =0.077)]. In patients with ≥8 swollen and ≥8 tender joints at baseline more anifrolumab-treated patients achieved ≥50% decrease in joint count [48.6%, 69.6% ( p =0.038), 64.6% ( p =0.156)]. Although steroid tapering was not mandated, OCS reduction to ≤7.5 mg/d at Day 365 was achieved by 26.6%, 56.4%, and 31.7% of patients. The observed benefits were driven by results in the IFN-high subpopulation (figure). Median suppression of 21 IFN-regulated genes was ∼90% for both doses of anifrolumab. Patients in the placebo group had the lowest incidence of Herpes zoster (2.0%, 5.1%, 9.5%) and cases reported as influenza (2.0%, 6.1%, 7.6%); there were no differences in the incidence of serious adverse events (18.8%, 16.2%, 17.1%). The incidence of infusion-related reactions was similar (5.9%, 2.0%, 3.8%). Conclusions Anifrolumab significantly reduced disease activity across all clinical endpoints. Enhanced effects in IFN-high patients support the pathobiology of this treatment strategy. Acknowledgement Funded by MedImmune. Editorial assistance: K Alexander, QXV Comms, an Ashfield business, UK Disclosure of Interest R. Furie Consultant for: MedImmune, J. Merrill Grant/research support from: MedImmune; Genentech/Roche, Consultant for: Medimmune, Genentech/Roche, Neovacs, V. Werth Consultant for: MedImmune, M. Khamashta: None declared, K. Kalunian Grant/research support from: MedImmune, Consultant for: AstraZeneca, P. Brohawn Shareholder of: AstraZeneca, Employee of: MedImmune, G. Illei Shareholder of: AstraZeneca, Employee of: MedImmune, J. Drappa Shareholder of: AstraZeneca, Employee of: MedImmune, L. Wang Employee of: MedImmune, S. Yoo Shareholder of: AstraZeneca; Regenx Bio, Consultant for: Regenx Bio

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease marked by immune dysregulation. Why some patients have only moderate symptoms and others develop organ-threatening manifestations is unclear. This study evaluates the temporal expression of autoantibodies and cytokines in sera from the Department of Defense Serum Repository during transition from preclinical lupus to SLE in patients (n=83) who do or do not present with nephritis. Patients who met renal criteria (n=30; 36%) did so within 5.2 (±5.5) months of SLE classification (range −5.2 to +12 months). Renal cases experienced earlier onset of autoantibody positivity vs. non-renal cases (mean −3.8 vs −2.6 years relative to SLE classification, p=0.0442). Prior to developing nephritis, renal cases exhibited elevated levels of distinct soluble mediators compared to matched non-renal cases and healthy controls, including adaptive mediators IL-4, IL-5, IL-12, and IFN-γ, chemokines IP-10 and MCP-3, and nephritis-associated mediators SCF and shed TNFRII (all p

Objective We previously reported that lupus anticoagulant (LAC) is the main predictor of poor pregnancy outcome in antiphospholipid antibody (aPL)-positive patients. We sought to confirm this finding in an independent group of patients who were subsequently recruited into the PROMISSE study. Methods The PROMISSE study is a multicentre, prospective, observational study of pregnancy outcomes in women with aPL and/or systemic lupus erythematosus (SLE) that enrolled patients from 2003 to 2015. All consecutive, aPL-positive patients from the PROMISSE study who completed their pregnancy between April 2011 and January 2015 (after the previous PROMISSE report) are included in the current report. Patients were followed monthly until delivery, and aPL was tested at first, second and third trimesters of pregnancy and at 12 weeks post partum. Adverse pregnancy outcomes (APOs) were defined as fetal death after 12 weeks of gestation, neonatal death, delivery prior to 36 weeks of gestation due to pre-eclampsia or placental insufficiency or small-for-gestational age (birth weight

Over 20% of pregnancies in patients with systemic lupus erythematosus (SLE) and/or antiphospholipid antibodies (APL) result in an adverse pregnancy outcome (APO) related to abnormal placentation. The ability to identify, early in pregnancy, patients who are destined for poor outcomes would significantly impact care of this high-risk population. In nonautoimmune patients, circulating angiogenic factors are dysregulated in disorders of placentation, such as preeclampsia (PE) and fetal growth restriction.We sought to determine whether early dysregulation of circulating angiogenic factors can predict APO in high-risk SLE and/or APL pregnancies.We used data and samples from the Predictors of Pregnancy Outcome: Biomarkers in APL Syndrome and SLE (PROMISSE), a multicenter prospective study that enrolled 492 pregnant women with SLE and/or APL from September 2003 through August 2013. Patients were followed through pregnancy from12 weeks gestation. Circulating levels of soluble fms-like tyrosine kinase-1 (sFlt1), placental growth factor (PlGF), and soluble endoglin were measured monthly and subjects followed up for APO, classified as severe (PE34 weeks, fetal/neonatal death, indicated preterm delivery30 weeks) or moderate (PE ≥34 weeks, indicated preterm delivery 30-36 weeks, growth restriction without PE).Severe APOs occurred in 12% and moderate APOs in 10% of patients. By 12-15 weeks, sFlt1, PlGF, and soluble endoglin levels were markedly altered in women who developed severe APO. After adjusting for clinical risk factors, sFlt1 was the strongest predictor of severe APO among 12-15 week measures (odds ratio, 17.3 comparing highest and lowest quartiles; 95% confidence interval [CI], 3.5-84.8; positive predictive value [PPV], 61%; negative predictive value [NPV], 93%). At 16-19 weeks, the combination of sFlt1 and PlGF was most predictive of severe APO, with risk greatest for subjects with both PlGF in lowest quartile (70.3 pg/mL) and sFlt1 in highest quartile (1872 pg/mL; odds ratio, 31.1; 95% CI, 8.0-121.9; PPV, 58%; NPV, 95%). Severe APO rate in this high-risk subgroup was 94% (95% CI, 70-99.8%), if lupus anticoagulant or history of high blood pressure was additionally present. In contrast, among patients with both sFlt11872 pg/mL and PlGF70.3 pg/mL, rate of severe APO was only 4.6% (95% CI, 2.1-8.6%).Circulating angiogenic factors measured during early gestation have a high NPV in ruling out the development of severe adverse outcomes among patients with SLE and/or APL syndrome. Timely risk stratification of patients is important for effective clinical care and optimal allocation of health care resources.

Co-stimulatory molecules help to regulate interactions between T cells and antigen-presenting cells and may play an important role in the pathogenesis of lupus. Both work in murine models and some early studies in human lupus support further examination of these molecules as therapeutic targets. Complexities of lupus clinical trial variables may have hampered progress in this area but recent developments in the field may make interventional trials more feasible in the near future. To date biologics which provide direct blockade of interactions between CD40 and CD154, B7RP-1 and ICOS, and CD80 or CD86 with CD28 have been assessed in multicenter clinical trials. These data will be reviewed and critiqued.

We previously reported that the G allele of rs3853839 at 3′untranslated region (UTR) of Toll-like receptor 7 (TLR7) was associated with elevated transcript expression and increased risk for systemic lupus erythematosus (SLE) in 9,274 Eastern Asians [P = 6.5×10−10, odds ratio (OR) (95%CI) = 1.27 (1.17–1.36)]. Here, we conducted trans-ancestral fine-mapping in 13,339 subjects including European Americans, African Americans, and Amerindian/Hispanics and confirmed rs3853839 as the only variant within the TLR7-TLR8 region exhibiting consistent and independent association with SLE (P meta = 7.5×10−11, OR = 1.24 [1.18–1.34]). The risk G allele was associated with significantly increased levels of TLR7 mRNA and protein in peripheral blood mononuclear cells (PBMCs) and elevated luciferase activity of reporter gene in transfected cells. TLR7 3′UTR sequence bearing the non-risk C allele of rs3853839 matches a predicted binding site of microRNA-3148 (miR-3148), suggesting that this microRNA may regulate TLR7 expression. Indeed, miR-3148 levels were inversely correlated with TLR7 transcript levels in PBMCs from SLE patients and controls (R2 = 0.255, P = 0.001). Overexpression of miR-3148 in HEK-293 cells led to significant dose-dependent decrease in luciferase activity for construct driven by TLR7 3′UTR segment bearing the C allele (P = 0.0003). Compared with the G-allele construct, the C-allele construct showed greater than two-fold reduction of luciferase activity in the presence of miR-3148. Reduced modulation by miR-3148 conferred slower degradation of the risk G-allele containing TLR7 transcripts, resulting in elevated levels of gene products. These data establish rs3853839 of TLR7 as a shared risk variant of SLE in 22,613 subjects of Asian, EA, AA, and Amerindian/Hispanic ancestries (Pmeta = 2.0×10−19, OR = 1.25 [1.20–1.32]), which confers allelic effect on transcript turnover via differential binding to the epigenetic factor miR-3148., Author Summary Systemic lupus erythematosus (SLE) is a debilitating autoimmune disease contributed to by excessive innate immune activation involving toll-like receptors (TLRs, particularly TLR7/8/9) and type I interferon (IFN) signaling pathways. TLR7 responds against RNA–containing nuclear antigens and activates IFN-α pathway, playing a pivotal role in the development of SLE. While a genomic duplication of Tlr7 promotes lupus-like disease in the Y-linked autoimmune accelerator (Yaa) murine model, the lack of common copy number variations at TLR7 in humans led us to identify a functional single nucleotide polymorphism (SNP), rs3853839 at 3′ UTR of the TLR7 gene, associated with SLE susceptibility in Eastern Asians. In this study, we fine-mapped the TLR7-TLR8 region and confirmed rs3853839 exhibiting the strongest association with SLE in European Americans, African Americans, and Amerindian/Hispanics. Individuals carrying the risk G allele of rs3853839 exhibited increased TLR7 expression at the both mRNA and protein level and decreased transcript degradation. MicroRNA-3148 (miR-3148) downregulated the expression of non-risk allele (C) containing transcripts preferentially, suggesting a likely mechanism for increased TLR7 levels in risk-allele carriers. This trans-ancestral mapping provides evidence for the global association with SLE risk at rs3853839, which resides in a microRNA–gene regulatory site affecting TLR7 expression.

Objectives The Xq28 region containing IRAK1 and MECP2 has been identified as a risk locus for systemic lupus erythematosus (SLE) in previous genetic association studies. However, due to the strong linkage disequilibrium between IRAK1 and MECP2 , it remains unclear which gene is affected by the underlying causal variant(s) conferring risk of SLE. Methods We fine-mapped ≥136 SNPs in a ∼227 kb region on Xq28, containing IRAK1, MECP2 and seven adjacent genes ( L1CAM, AVPR2, ARHGAP4, NAA10, RENBP, HCFC1 and TMEM187 ), for association with SLE in 15 783 case-control subjects derived from four different ancestral groups. Results Multiple SNPs showed strong association with SLE in European Americans, Asians and Hispanics at p −8 with consistent association in subjects with African ancestry. Of these, six SNPs located in the TMEM187-IRAK1-MECP2 region captured the underlying causal variant(s) residing in a common risk haplotype shared by all four ancestral groups. Among them, rs1059702 best explained the Xq28 association signals in conditional testings and exhibited the strongest p value in transancestral meta-analysis (p meta = 1.3×10 −27 , OR=1.43), and thus was considered to be the most likely causal variant. The risk allele of rs1059702 results in the amino acid substitution S196F in IRAK1 and had previously been shown to increase NF-κB activity in vitro. We also found that the homozygous risk genotype of rs1059702 was associated with lower mRNA levels of MECP2 , but not IRAK1 , in SLE patients (p=0.0012) and healthy controls (p=0.0064). Conclusions These data suggest contributions of both IRAK1 and MECP2 to SLE susceptibility.

Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical manifestations characterized by the development of pathogenic autoantibodies manifesting in inflammation of target organs such as the kidneys, skin and joints. Genome-wide association studies have identified genetic variants in the UBE2L3 region that are associated with SLE in subjects of European and Asian ancestry. UBE2L3 encodes an ubiquitin-conjugating enzyme, UBCH7, involved in cell proliferation and immune function. In this study, we sought to further characterize the genetic association in the region of UBE2L3 and use molecular methods to determine the functional effect of the risk haplotype. We identified significant associations between variants in the region of UBE2L3 and SLE in individuals of European and Asian ancestry that exceeded a Bonferroni-corrected threshold (P

ObjectiveSystemic lupus erythematosus (SLE; OMIM 152700) is a chronic autoimmune disease for which the aetiology includes genetic and environmental factors. ITGAM, integrin αM (complement component 3 receptor 3 subunit) encoding a ligand for intracellular adhesion molecule (ICAM) proteins, is an established SLE susceptibility locus. This study aimed to evaluate the independent and joint effects of genetic variations in the genes that encode ITGAM and ICAM.MethodsThe authors examined several markers in the ICAM1–ICAM4–ICAM5 locus on chromosome 19p13 and the single ITGAM polymorphism (rs1143679) using a large-scale case–control study of 17 481 unrelated participants from four ancestry populations. The single-marker association and gene–gene interaction were analysed for each ancestry, and a meta-analysis across the four ancestries was performed.ResultsThe A-allele of ICAM1–ICAM4–ICAM5 rs3093030, associated with elevated plasma levels of soluble ICAM1, and the A-allele of ITGAM rs1143679 showed the strongest association with increased SLE susceptibility in each of the ancestry populations and the trans-ancestry meta-analysis (ORmeta=1.16, 95% CI 1.11 to 1.22; p=4.88×10−10 and ORmeta=1.67, 95% CI 1.55 to 1.79; p=3.32×10−46, respectively). The effect of the ICAM single-nucleotide polymorphisms (SNPs) was independent of the effect of the ITGAM SNP rs1143679, and carriers of both ICAM rs3093030-AA and ITGAM rs1143679-AA had an OR of 4.08 compared with those with no risk allele in either SNP (95% CI 2.09 to 7.98; p=3.91×10−5).ConclusionThese findings are the first to suggest that an ICAM–integrin-mediated pathway contributes to susceptibility to SLE.

An inherited risk for the occurrence of antiphospholipid antibodies (aPLs), with or without the clinical manifestations associated with antiphospholipid syndrome (APS), is supported by multiple lines of evidence. Family members of patients with APS, whether primary or associated with another autoimmune disease, are more likely to have elevated aPL levels. Multiple reports have described multiplex families with two or more family members with APS (or APS and another autoimmune disorder), and there is a strong association between aPL and APS with specific genes and gene alleles in the major histocompatibility complex. Genetic risk factors also appear to contribute to the prothrombotic risk in individual patients with aPL. The data would suggest that the development of these antibodies, as well as the clinical syndrome, reflects a complex interplay of inherited and acquired risk factors. Ongoing studies designed to identify and characterize genes associated with the development of aPL will improve our understanding of this complex autoimmune disorder and may provide a better understanding of which patients with aPL will develop clinical manifestations of the syndrome.

Objective Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibody production and altered type I interferon expression. Genetic surveys and genome-wide association studies have identified >30 SLE susceptibility genes. One of these genes, TNIP1, encodes the ABIN1 protein. ABIN1 functions in the immune system by restricting NF-?B signaling. The present study was undertaken to investigate the genetic factors that influence association with SLE in genes that regulate the NF-?B pathway. Methods We analyzed a dense set of genetic markers spanning TNIP1 and TAX1BP1, as well as the TNIP1 homolog TNIP2, in case-control populations of diverse ethnic origins. TNIP1, TNIP2, and TAX1BP1 were fine-mapped in a total of 8,372 SLE cases and 7,492 healthy controls from European-ancestry, African American, Hispanic, East Asian, and African American Gullah populations. Levels of TNIP1 messenger RNA (mRNA) and ABIN1 protein in Epstein-Barr virus-transformed human B cell lines were analyzed by quantitative reverse transcription-polymerase chain reaction and Western blotting, respectively. Results We found significant associations between SLE and genetic variants within TNIP1, but not in TNIP2 or TAX1BP1. After resequencing and imputation, we identified 2 independent risk haplotypes within TNIP1 in individuals of European ancestry that were also present in African American and Hispanic populations. Levels of TNIP1 mRNA and ABIN1 protein were reduced among subjects with these haplotypes, suggesting that they harbor hypomorphic functional variants that influence susceptibility to SLE by restricting ABIN1 expression. Conclusion Our results confirm the association signals between SLE and TNIP1 variants in multiple populations and provide new insight into the mechanism by which TNIP1 variants may contribute to SLE pathogenesis. Copyright © 2012 by the American College of Rheumatology.

The role of phosphorylation and dephosphorylation events in homotypic neutrophil aggregation mediated by CD11b/CD18 molecules was investigated using okadaic acid, an inhibitor of serine and threonine phosphatases. In the absence of exogenous stimuli the addition of okadaic acid to neutrophils resulted in a dose-dependent increase in phosphorylation of the CD18 beta chain that was further augmented by PMA but unaffected by FMLP. Phosphorylation induced by okadaic acid was reversed by staurosporine and minimally decreased by the less selective PKA/PKC inhibitors, H-7 and H-8. This suggests the existence of constitutive phosphatase and kinase activity emphasizing the dynamic state of phosphorylation and dephosphorylation of the beta 2 integrins. Unlike PMA, okadaic acid did not promote homotypic neutrophil aggregation. Furthermore, both the PMA-induced pathway of irreversible aggregation, blocked by staurosporine, as well as the FMLP-induced pathway of reversible aggregation, augmented by staurosporine, were inhibited by okadaic acid in a dose- and time-dependent manner. These results provide evidence that a phosphatase-dependent step is involved in each of these two distinct pathways that regulate neutrophil aggregation mediated by beta 2 integrin activation.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by autoantibody production and organ damage. Lupus nephritis (LN) is one of the most severe manifestations of SLE. Multiple studies reported associations between renal diseases and variants in the non-muscle myosin heavy chain 9 (MYH9) and the neighboring apolipoprotein L 1 (APOL1) genes. We evaluated 167 variants spanning MYH9 for association with LN in a multiethnic sample. The two previously identified risk variants in APOL1 were also tested for association with LN in European-Americans (EAs) (N=579) and African-Americans (AAs) (N=407). Multiple peaks of association exceeding a Bonferroni corrected P-value of P

B lymphocyte stimulator (BLyS, also known as TNFSF13B and BAFF) is a type II transmembrane protein and a member of the TNF superfamily (1, 2). BLyS is produced by several different cell types, including monocytes, activated neutrophils, T cells, and dendritic cells (3–5), and is expressed as a cell surface protein which can be furin-cleaved and released into the circulation. Although BLyS has been shown to be constitutively expressed, certain inflammatory cytokines such as IL-2, TNF-α, IFN-γ, and IFN-α can enhance its production and secretion (3, 4, 6, 7). BLyS binds to 3 receptors found primarily on B cells (8). Activation of these receptors leads to B cell proliferation, differentiation, survival, and IgG class switching (1, 4, 9). BLyS has been shown to play an important role in primary immune responses, as anti-BLyS treated mice show profoundly reduced numbers of naive B cells with accompanying attenuated responses to both T-dependent and T-independent antigens (10). Transgenic mice that overexpress BLyS display a myriad of autoimmune features, such as high levels of rheumatoid factor, anti-DNA, circulating immune complexes, and immunoglobulin deposition in the kidneys (9). Mice treated with exogenous BLyS develop an increase in B cell numbers, particularly those directed against chromatin (11), and autoreactive cells encountering transitional B cell checkpoints in the spleen require higher concentrations of BLyS to survive than do non-autoreactive B cells (8, 9, 12). Mouse models have also demonstrated that deletion of either BLyS or its receptor severely impairs B cell development beyond the transitional stage with a resulting decrease in peripheral B cell populations (9, 13–15). Elevated circulating BLyS levels have been found in patients with systemic autoimmune disorders such as SLE, rheumatoid arthritis (RA), and Sjoren’s syndrome (16–18). Early reports found increased serum BLyS levels in SLE patients compared to healthy individuals that correlated with anti-dsDNA titers, but not disease activity (16, 17, 19). BLyS levels did, however, decrease following high-dose corticosteroid treatment (19). Other studies found that peripheral blood leukocyte BLyS mRNA levels correlated with SLE disease activity (20, 21). While most of these initial studies, which were performed in mainly Hispanic SLE patients, failed to demonstrate a correlation between serum BLyS levels and disease activity, a later longitudinal study undertaken at 4 different clinical centers that contained ancestral diversity, found an association between increases in plasma BLyS levels from a previous visit and increases in SELENA-SLEDAI scores at the next visit (22). Additionally, another study involving Norwegian SLE patients, also found a correlation with serum BLyS levels and SLEDAI scores (23). A study examining plasma BLyS protein levels in pediatric SLE also demonstrated an association between elevated BLyS levels and increased disease activity (24). IFN-α has proven to be a key cytokine in SLE pathogenesis, and has been shown to increase BLyS expression in antigen-presenting cells (4). Therefore, it is possible that IFN-α plays a role in driving BLyS production in SLE, although a direct correlation between circulating BLyS levels and serum IFN-α activity has yet to be demonstrated in SLE patients. Additionally, several environmental factors have been implicated in the pathogenesis of SLE (25), including vitamin D deficiency (26). As vitamin D has been shown to suppress the expression of the IFN signature in myeloid-derived dendritic cells, as well as suppress B cell activation and immunoglobulin production (27, 28), it is of interest to assess the relationship between vitamin D and BLyS levels. Belimumab is a fully human monoclonal antibody that binds BLyS and inhibits its activity. Two recent large randomized, double-blind, placebo-controlled, multicenter phase 3 trials, BLISS-52 and BLISS-76, evaluated the efficacy of belimumab plus standard of care compared to placebo plus standard of care; positive results were obtained with both studies reaching their primary efficacy endpoints of reductions in disease activity without ancillary organ flares (29, 30). Taken together, these data suggest that excessive BLyS may be an important mechanism underlying SLE pathogenesis and that BLyS targeted therapies seem promising. However, normal B cell function requires some BLyS activity. To date no studies have evaluated whether specific levels of BLyS are required to mount appropriate vaccination responses or whether BLyS levels change after vaccination. The primary objective of this study is to investigate the relationship between circulating BLyS levels and humoral responses made to influenza vaccination, as well as the effect of influenza vaccination on BLyS production in SLE. The current study also examines select demographic, environmental, and clinical features of SLE for association with elevated BLyS levels in an effort to better understand the mechanisms of elevated BLyS in a heterogeneous SLE population.

ObjectiveHigh serum interferon α (IFNα) activity is a heritable risk factor for systemic lupus erythematosus (SLE). Auto-antibodies found in SLE form immune complexes which can stimulate IFNα production by activating endosomal Toll-like receptors and interferon regulatory factors (IRFs), including IRF5. Genetic variation in IRF5 is associated with SLE susceptibility; however, it is unclear how IRF5 functional genetic elements contribute to human disease.Methods1034 patients with SLE and 989 controls of European ancestry, 555 patients with SLE and 679 controls of African–American ancestry, and 73 patients with SLE of South African ancestry were genotyped at IRF5 polymorphisms, which define major haplotypes. Serum IFNα activity was measured using a functional assay.ResultsIn European ancestry subjects, anti-double-stranded DNA (dsDNA) and anti-Ro antibodies were each associated with different haplotypes characterised by a different combination of functional genetic elements (OR>2.56, p−14 for both). These IRF5 haplotype-auto-antibody associations strongly predicted higher serum IFNα in patients with SLE and explained >70% of the genetic risk of SLE due to IRF5. In African–American patients with SLE a similar relationship between serology and IFNα was observed, although the previously described European ancestry-risk haplotype was present at admixture proportions in African–American subjects and absent in African patients with SLE.ConclusionsThe authors define a novel risk haplotype of IRF5 that is associated with anti-dsDNA antibodies and show that risk of SLE due to IRF5 genotype is largely dependent upon particular auto-antibodies. This suggests that auto-antibodies are directly pathogenic in human SLE, resulting in increased IFNα in cooperation with particular combinations of IRF5 functional genetic elements.SLE is a systemic autoimmune disorder affecting multiple organ systems including the skin, musculoskeletal, renal and haematopoietic systems. Humoral autoimmunity is a hallmark of SLE, and patients frequently have circulating auto-antibodies directed against dsDNA, as well as RNA binding proteins (RBP). Anti-RBP autoantibodies include antibodies which recognize Ro, La, Smith (anti-Sm), and ribonucleoprotein (anti-nRNP), collectively referred to as anti-retinol-binding protein). Anti-retinol-binding protein and anti-dsDNA auto-antibodies are rare in the healthy population.1 These auto-antibodies can be present in sera for years preceding the onset of clinical SLE illness2 and are likely pathogenic in SLE.3 4

Objective We describe disease activity, damage, and the accrual of key autoantibodies in an inception systemic lupus erythematosus (SLE) cohort. Methods. The Systemic Lupus International Collaborating Clinics (SLICC) International Research Network, comprising 27 centers from 11 countries, has followed an inception cohort of SLE patients yearly according to a standardized protocol. Of these patients, 298 were followed for a minimum of 5 years and constitute the study population. Disease activity was assessed using the SLE Disease Activity Index 2000 (SLEDAI-2K) and damage was assessed using the SLICC/American College of Rheumatology Damage Index (SDI). Antinuclear antibody (ANA), anti-DNA, and anticardiolipin antibody (aCL) levels and lupus anticoagulant were assessed yearly. Descriptive statistics were generated and repeated-measures general linear models were used to evaluate SLEDAI-2K and SDI over time between whites and nonwhites. Results. Of the 298 patients, 87% were women, 55% were white, 12% were African American, 14% were Asian, 16% were Hispanic, and 2% were categorized as "other." At enrollment, the mean age was 35.3 years, the mean SLEDAI-2K score was 5.9, and the mean disease duration was 5.5 months. Mean SLEDAI-2K scores decreased in the first year and then remained low. SLEDAI-2K scores were significantly lower at each year in whites compared to nonwhites. Mean SDI scores increased progressively over 5 years; there was no significant difference between whites and nonwhites. As expected, ANA positivity was high and anti-DNA positivity was relatively low at enrollment, and both increased over 5 years. Although lupus anticoagulant increased slightly over 5 years, aCL positivity did not. Conclusion. Disease activity in newly diagnosed patients decreases over their first 5 years, while damage increases. Antibody positivity ran variable courses over this period.

Objective Several confirmed genetic susceptibility loci for lupus have been described. To date, no clear evidence for genetic epistasis in lupus has been established. The aim of this study was to test for gene–gene interactions in a number of known lupus susceptibility loci. Methods Eighteen single-nucleotide polymorphisms tagging independent and confirmed lupus susceptibility loci were genotyped in a set of 4,248 patients with lupus and 3,818 normal healthy control subjects of European descent. Epistasis was tested by a 2-step approach using both parametric and nonparametric methods. The false discovery rate (FDR) method was used to correct for multiple testing. Results We detected and confirmed gene–gene interactions between the HLA region and CTLA4, IRF5, and ITGAM and between PDCD1 and IL21 in patients with lupus. The most significant interaction detected by parametric analysis was between rs3131379 in the HLA region and rs231775 in CTLA4 (interaction odds ratio 1.19, Z = 3.95, P = 7.8 × 10−5 [FDR ≤0.05], P for multifactor dimensionality reduction = 5.9 × 10−45). Importantly, our data suggest that in patients with lupus, the presence of the HLA lupus risk alleles in rs1270942 and rs3131379 increases the odds of also carrying the lupus risk allele in IRF5 (rs2070197) by 17% and 16%, respectively (P = 0.0028 and P = 0.0047, respectively). Conclusion We provide evidence for gene–gene epistasis in systemic lupus erythematosus. These findings support a role for genetic interaction contributing to the complexity of lupus heritability.

Purpose Systemic lupus erythematosus (SLE) can significantly affect both health and non-health-related quality of life (HRQOL and non-HRQOL). However, of the existent published patient-reported outcome (PRO) tools, none were developed from US patients, an ethnically diverse population. Furthermore, these tools do not address men with SLE or assess non-HRQOL issues. Herein, we present the development and validation of the Lupus Patient-Reported Outcome tool (LupusPRO) and discuss its clinical utility and research value compared with other PRO tools currently available for SLE. Methods Beginning with a conceptual framework, items for LupusPRO were generated using feedback from women and men with SLE. The tool underwent iterations based on patient feedback and clinimetric and psychometric analyses. Validity (content, construct, and criterion) and reliability (internal consistency and test-retest) for the 44-item LupusPRO tool are presented. Results Consistent with the conceptual framework, items were identified that were related to HRQOL and non-HRQOL constructs. HRQOL domains included (1) lupus symptoms; (2) physical health (physical function, role physical); (3) pain–vitality; (4) emotional health (emotional function and role emotional); (5) body image; (6) cognition; (7) procreation; and (8) lupus medications. Non-HRQOL domains were (1) available social support and coping; (2) desires–goals; and (3) satisfaction with medical care. Internal consistency reliability (0.68-0.94), test-retest reliability (0.55-0.92), content, construct ( r > 0.50 with SF-36), and criterion ( r > −0.35 with disease activity) validity were fair to good. Conclusions LupusPRO is a valid and reliable disease-targeted patient-reported health outcome tool that is generalizable to SLE patients in the United States of varied ethnic backgrounds and either gender.

Background Type I interferons (IFNs) appear to play a central role in disease pathogenesis in systemic lupus erythematosus (SLE), making them potential therapeutic targets. Methods Safety profile, pharmacokinetics, immunogenicity, pharmacodynamics and clinical activity of sifalimumab, an anti-IFNα monoclonal antibody, were assessed in a phase I, multicentre, randomised, double-blind, dose-escalation study with an open-label extension in adults with moderately active SLE. Subjects received one intravenous dose of sifalimumab (n=33 blinded phase, 0.3, 1, 3, 10 or 30 mg/kg; n=17 open-label, 1, 3, 10 or 30 mg/kg) or placebo (n=17). Each phase lasted 84 days. Results Adverse events (AEs) were similar between groups; about 97% of AEs were grade 1 or 2. All grade 3 and 4 AEs and all serious AEs (2 placebo, 1 sifalimumab) were deemed unrelated to the study drug. No increase in viral infections or reactivation was observed. Sifalimumab caused dose-dependent inhibition of type I IFN-induced mRNAs (type I IFN signature) in whole blood and corresponding changes in related proteins in affected skin. Exploratory analyses showed consistent trends toward improvement in disease activity in sifalimumab-treated versus placebo-treated subjects. A lower proportion of sifalimumab-treated subjects required new or increased immunosuppressive treatments (12% vs 41%; p=0.03) and had fewer Systemic Lupus Erythematosus Disease Activity Index flares (3% vs 29%; p=0.014). Conclusions Sifalimumab had a safety profile that supports further clinical development. This trial demonstrated that overexpression of type I IFN signature in SLE is at least partly driven by IFNα, and exploratory analyses suggest that IFNα inhibition may be associated with clinical benefit in SLE. Trial registration number NCT00299819.

ObjectiveSystemic lupus erythematosus is a clinically heterogeneous autoimmune disease. A number of genetic loci that increase lupus susceptibility have been established. This study examines if these genetic loci also contribute to the clinical heterogeneity in lupus.Materials and methods4001 European-derived, 1547 Hispanic, 1590 African-American and 1191 Asian lupus patients were genotyped for 16 confirmed lupus susceptibility loci. Ancestry informative markers were genotyped to calculate and adjust for admixture. The association between the risk allele in each locus was determined and compared in patients with and without the various clinical manifestations included in the ACR criteria.ResultsRenal disorder was significantly correlated with the lupus risk allele in ITGAM (p=5.0×10−6, OR 1.25, 95% CI 1.12 to 1.35) and in TNFSF4 (p=0.0013, OR 1.14, 95% CI 1.07 to 1.25). Other significant findings include the association between risk alleles in FCGR2A and malar rash (p=0.0031, OR 1.11, 95% CI 1.17 to 1.33), ITGAM and discoid rash (p=0.0020, OR 1.20, 95% CI 1.06 to 1.33), STAT4 and protection from oral ulcers (p=0.0027, OR 0.89, 95% CI 0.83 to 0.96) and IL21 and haematological disorder (p=0.0027, OR 1.13, 95% CI 1.04 to 1.22). All these associations are significant with a false discovery rate of ConclusionSignifi cant associations were found between clinical manifestations and the FCGR2A, ITGAM, STAT4, TNSF4 and IL21 genes. The findings suggest that genetic profiling might be a useful tool to predict disease manifestations in lupus patients in the future.

Systemic lupus erythematosus is a chronic-relapsing autoimmune disease of incompletely understood etiology. Recent evidence strongly supports an epigenetic contribution to the pathogenesis of lupus. To understand the extent and nature of dysregulated DNA methylation in lupus T cells, we performed a genome-wide DNA methylation study in CD4 (+) T cells in lupus patients compared to normal healthy controls. Cytosine methylation was quantified in 27,578 CG sites located within the promoter regions of 14,495 genes. We identified 236 hypomethylated and 105 hypermethylated CG sites in lupus CD4 (+) T cells compared to normal controls, consistent with widespread DNA methylation changes in lupus T cells. Of interest, hypomethylated genes in lupus T cells include CD9, which is known to provide potent T-cell co-stimulation signals. Other genes with known involvement in autoimmunity such as MMP9 and PDGFRA were also hypomethylated. The BST2 gene, an interferon-inducible membrane-bound protein that helps restrict the release of retroviral particles was also hypomethylated in lupus patients. Genes involved in folate biosynthesis, which plays a role in DNA methylation, were overrepresented among hypermethylated genes. In addition, the transcription factor RUNX3 was hypermethylated in patients, suggesting an impact on T-cell maturation. Protein-protein interaction maps identified a transcription factor, HNF4a, as a regulatory hub affecting a number of differentially methylated genes. Apoptosis was also an overrepresented ontology in these interaction maps. Further, our data suggest that the methylation status of RAB22A, STX1B2, LGALS3BP, DNASE1L1 and PREX1 correlates with disease activity in lupus patients.

OBJECTIVE: Genetic association of the IL2/IL21 region at chromosome 4q27 has previously been reported in lupus and a number of autoimmune and inflammatory diseases. This study was undertaken to determine whether this genetic effect could be localized, using a very large cohort of lupus patients and controls. METHODS: We genotyped 45 tag single-nucleotide polymorphisms (SNPs) across the IL2/IL21 locus in 2 large independent lupus sample sets. We studied a set of subjects of European descent consisting of 4,248 lupus patients and 3,818 healthy controls, and an African American set of 1,569 patients and 1,893 healthy controls. Imputation in 3,004 additional controls from the Wellcome Trust Case Control Consortium was also performed. Genetic association between the genotyped markers was determined, and pairwise conditional analysis was performed to localize the independent genetic effect in the IL2/IL21 locus in lupus. RESULTS: We established and confirmed the genetic association between IL2/IL21 and lupus. Using conditional analysis and transethnic mapping, we localized the genetic effect in this locus to 2 SNPs in high linkage disequilibrium: rs907715 located within IL21 (odds ratio 1.16 [95% confidence interval 1.10-1.22], P=2.17×10(-8)) and rs6835457 located in the 3'-untranslated flanking region of IL21 (odds ratio 1.11 [95% confidence interval 1.05-1.17], P=9.35×10(-5)). CONCLUSION: Our findings establish the genetic association between lupus and IL2/IL21 with a genome-wide level of significance. Further, our findings indicate that this genetic association within the IL2/IL21 linkage disequilibrium block is localized to IL21. If other autoimmune IL2/IL21 genetic associations are similarly localized, then the IL21 risk alleles would be predicted to operate by a fundamental mechanism that influences the course of a number of autoimmune disease processes., Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disorder with a complex pathogenesis in which genetic, hormonal and environmental factors have a role. Rare mutations in the TREX1 gene, the major mammalian 3′–5′ exonuclease, have been reported in sporadic SLE cases. Some of these mutations have also been identified in a rare pediatric neurological condition featuring an inflammatory encephalopathy known as Aicardi–Goutières syndrome (AGS). We sought to investigate the frequency of these mutations in a large multi-ancestral cohort of SLE cases and controls. A total of 40 single-nucleotide polymorphisms (SNPs), including both common and rare variants, across the TREX1 gene, were evaluated in ~8370 patients with SLE and ~7490 control subjects. Stringent quality control procedures were applied, and principal components and admixture proportions were calculated to identify outliers for removal from analysis. Population-based case–control association analyses were performed. P-values, false-discovery rate q values, and odds ratios (OR) with 95% confidence intervals (CI) were calculated. The estimated frequency of TREX1 mutations in our lupus cohort was 0.5%. Five heterozygous mutations were detected at the Y305C polymorphism in European lupus cases but none were observed in European controls. Five African cases incurred heterozygous mutations at the E266G polymorphism and, again, none were observed in the African controls. A rare homozygous R114H mutation was identified in one Asian SLE patient, whereas all genotypes at this mutation in previous reports for SLE were heterozygous. Analysis of common TREX1 SNPs (minor allele frequency (MAF)>10%) revealed a relatively common risk haplotype in European SLE patients with neurological manifestations, especially seizures, with a frequency of 58% in lupus cases compared with 45% in normal controls (P=0.0008, OR=1.73, 95% CI=1.25–2.39). Finally, the presence or absence of specific autoantibodies in certain populations produced significant genetic associations. For example, a strong association with anti-nRNP was observed in the European cohort at a coding synonymous variant rs56203834 (P=2.99E−13, OR=5.2, 95% CI=3.18–8.56). Our data confirm and expand previous reports and provide additional support for the involvement of TREX1 in lupus pathogenesis.

Systemic lupus erythematosus (SLE), a complex polygenic autoimmune disease, is associated with increased complement activation. Variants of genes encoding complement regulator factor H (CFH) and five CFH-related proteins (CFHR1-CFHR5) within the chromosome 1q32 locus linked to SLE, have been associated with multiple human diseases and may contribute to dysregulated complement activation predisposing to SLE. We assessed 60 SNPs covering the CFH-CFHRs region for association with SLE in 15,864 case-control subjects derived from four ethnic groups. Significant allelic associations with SLE were detected in European Americans (EA) and African Americans (AA), which could be attributed to an intronic CFH SNP (rs6677604, in intron 11, P meta = 6.6×10−8, OR = 1.18) and an intergenic SNP between CFHR1 and CFHR4 (rs16840639, P meta = 2.9×10−7, OR = 1.17) rather than to previously identified disease-associated CFH exonic SNPs, including I62V, Y402H, A474A, and D936E. In addition, allelic association of rs6677604 with SLE was subsequently confirmed in Asians (AS). Haplotype analysis revealed that the underlying causal variant, tagged by rs6677604 and rs16840639, was localized to a ∼146 kb block extending from intron 9 of CFH to downstream of CFHR1. Within this block, the deletion of CFHR3 and CFHR1 (CFHR3-1Δ), a likely causal variant measured using multiplex ligation-dependent probe amplification, was tagged by rs6677604 in EA and AS and rs16840639 in AA, respectively. Deduced from genotypic associations of tag SNPs in EA, AA, and AS, homozygous deletion of CFHR3-1Δ (P meta = 3.2×10−7, OR = 1.47) conferred a higher risk of SLE than heterozygous deletion (P meta = 3.5×10−4, OR = 1.14). These results suggested that the CFHR3-1Δ deletion within the SLE-associated block, but not the previously described exonic SNPs of CFH, might contribute to the development of SLE in EA, AA, and AS, providing new insights into the role of complement regulators in the pathogenesis of SLE., Author Summary Systemic lupus erythematosus (SLE) is a complex autoimmune disease, associated with increased complement activation. Previous studies have provided evidence for the presence of SLE susceptibility gene(s) in the chromosome 1q31-32 locus. Within 1q32, genes encoding complement regulator factor H (CFH) and five CFH-related proteins (CFHR1-CFHR5) may contribute to the development of SLE, because genetic variants of these genes impair complement regulation and predispose to various human diseases. In this study, we tested association of genetic variants in the region containing CFH and CFHRs with SLE. We identified genetic variants predisposing to SLE in European American, African American, and Asian populations, which might be attributed to the deletion of CFHR3 and CFHR1 genes but not previously identified disease-associated exonic variants of CFH. This study provides the first evidence for consistent association between CFH/CFHRs and SLE across multi-ancestral SLE datasets, providing new insights into the role of complement regulators in the pathogenesis of SLE.

Systemic lupus erythematosus (SLE, MIM152700) is an autoimmune disease characterized by self-reactive antibodies resulting in systemic inflammation and organ failure. TNFAIP3, encoding the ubiquitin-modifying enzyme A20, is an established susceptibility locus for SLE. By fine mapping and genomic re-sequencing in ethnically diverse populations, we fully characterized the TNFAIP3 risk haplotype and identified a TT>A polymorphic dinucleotide (deletion T followed by a T to A transversion) associated with SLE in subjects of European (P = 1.58 × 10-8, odds ratio = 1.70) and Korean (P = 8.33 × 10-10, odds ratio = 2.54) ancestry. This variant, located in a region of high conservation and regulatory potential, bound a nuclear protein complex composed of NF-κB subunits with reduced avidity. Further, compared with the non-risk haplotype, the haplotype carrying this variant resulted in reduced TNFAIP3 mRNA and A20 protein expression. These results establish this TT>A variant as the most likely functional polymorphism responsible for the association between TNFAIP3 and SLE. © 2011 Nature America, Inc. All rights reserved.

Objective T cells from patients with systemic lupus erythematosus (SLE) express increased amounts of PP2Ac, which contributes to decreased production of interleukin-2 (IL-2). Because IL-2 is important in the regulation of several aspects of the immune response, it has been proposed that PP2Ac contributes to the expression of SLE. This study was designed to determine whether genetic variants of PPP2AC are linked to the expression of SLE and specific clinical manifestations and account for the increased expression of PP2Ac. Methods We conducted a trans-ethnic study of 8,695 SLE cases and 7,308 controls of 4 different ancestries. Eighteen single-nucleotide polymorphisms (SNPs) across PPP2CA were genotyped using an Illumina custom array. PPP2CA expression in SLE and control T cells was analyzed by real-time polymerase chain reaction. Results A 32-kb haplotype comprising multiple SNPs of PPP2CA showed significant association with SLE in Hispanic Americans, European Americans, and Asians, but not in African Americans. Conditional analyses revealed that SNP rs7704116 in intron 1 showed consistently strong association with SLE across Asian, European American, and Hispanic American populations (odds ratio 1.3 [95% confidence interval 1.14-1.31], meta-analysis P = 3.8 × 10 -7). In European Americans, the largest ethnic data set studied, the risk A allele of rs7704116 was associated with the presence of renal disease, anti-double-stranded DNA, and anti-RNP antibodies. PPP2CA expression was ?2-fold higher in SLE patients carrying the rs7704116 AG genotype than those carrying the GG genotype (P = 0.007). Conclusion Our data provide the first evidence of an association between PPP2CA polymorphisms and elevated PP2Ac transcript levels in T cells, which implicates a new molecular pathway for SLE susceptibility in European Americans, Hispanic Americans, and Asians. Copyright © 2011 by the American College of Rheumatology.

Publisher Summary Multiple organizations have devised instruments for the purpose of monitoring SLE disease activity, primarily with the goal of following the course of clinical disease expression in epidemiological studies. Several of these indices have been adapted for use in clinical trial outcomes. Validated instruments have been successful at measuring SLE activity. This chapter focuses on several validated instruments of disease activity that have been widely utilized in clinical practice, are accepted outcome indices for clinical trials, and have been used as endpoints for the evaluation of many experimental drugs. These include several iterations of the BILAG (British Isles Lupus Assessment Group) Index, the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) (and modified versions), and ways in which these instruments have been combined in composite outcome measures or modified to specifically address the capturing of flares. The BILAG Index links the grading of disease activity to the presence or absence of an intent to change therapy. It ranks symptoms by organ and by severity. The SLEDAI charts the presence or absence of symptoms but does not have the same level of complexity as the BILAG. Although it is not designed to assess different organs separately, it can be used that way, and it may have an advantage over the BILAG in being able to separate the assessment of multiple features within one organ, especially in the mucocutaneous system. Both instruments can be useful in clinical trials, but an understanding of their strengths and weaknesses is very important in designing outcomes for trials.

Publisher Summary This chapter addresses the problems in trial designs for lupus that may potentially have contributed to the failure of some treatments to demonstrate efficacy and for belimumab to have shown a more pronounced treatment effect. It reviews the evidence for the hypothesis that clinical trials to test even the most promising investigational drugs for lupus are impeded by what at first glance might seem to be reasonable regulatory requirements, and additionally by the heterogeneity of the disease itself, the natural waxing and waning of mild to moderate lupus symptoms, and an unacceptable signal-to-noise ratio caused by disparate background treatments. There has been a sophisticated precision in the preclinical study of biologics, each of which has been finely targeted to correct specific immune imbalances that are known to be present in many (but not all) lupus patients. Still, studies continue to enter patients without regard to their variable, underlying immune states and still omit the use of logical biomarkers to guide optimal treatment regimens. Subsequently, this chapter proposes solutions to each problem, emphasizing that the effects of background treatments on the immune disorder(s) that are specifically targeted by novel interventions should be better understood before decisions are made about which background treatments are suitable in a given clinical trial design and at what point in the protocol they should be given.

Publisher Summary This chapter reviews the impact on endothelial cells from prototypic pathologic features of lupus and addresses the question of whether the spectrum of autoantibodies associated with APS may be directly or indirectly involved in this process. Endothelial damage is implicated in the development of atherosclerosis regardless of whether the disease is incited by hypertension, diabetes, smoking, or poorly regulated lipid metabolism and regardless of the specific alignment of these various risk factors in individual patients. Infections and autoimmune diseases, including systemic lupus erythematosus (SLE) and the primary antiphospholipid syndrome (APS), have become strongly identified with accelerated atherosclerosis, and an extensive literature of in vitro and in vivo studies supports a model in which chronic immune imbalance provides a critical and ubiquitous underpinning to the development of arterial plaque. Furthermore, it is increasingly evident that the progressive events that lead to atherosclerosis are promulgated to a great extent by disordered endothelial cells interacting with inflammatory mediators. The specific risk for premature atherosclerosis in autoimmune patients such as those with lupus is known to include traditional risk factors, chronic generalized inflammation, and target-specific autoimmune mechanisms. The effects of all of these elements might be exacerbated by the chronic use of medications such as corticosteroids that add an additional disease burden by contributing to obesity, hyperglycemia, and hyperlipedemia.

Systemic lupus erythematosus (SLE) is a chronic, relapsing autoimmune connective tissue disease, primarily affecting the skin, joints, kidneys, heart, lungs, nervous system, blood elements, and serosal membranes. SLE is characterized by cytokine dysregulation, polyclonal B-cell activation, autoantibody production, and increased immune complex formation due to aberrations involving hyperactive B cells, T cells, and cells of the monocytic lineage. The symptoms of SLE are often diverse and nonspecific, and timely identification of SLE and associated comorbidities in patients is critical as aggressive monitoring and therapy may be warranted, especially in patients with poor prognoses. Based on the up-to-date understanding of the pathophysiology of SLE, the first targeted biological agent belimumab has been approved by the US Food and Drug Administration (FDA) in more than 50 years, and many targeted agents are being evaluated in late-stage clinical trials. There is a clear need to discuss how and when to incorporate new and emerging biological agents in managing patients with SLE. Additionally, the potential for increased risk of infections is a factor that heavily influences the rheumatologist׳s decision to use biological agents in managing patients with SLE. Hence, in this roundtable educational activity, expert faculty will review and discuss the strategies for timely diagnosis of SLE and associated comorbidities. They will also discuss the current understanding of the pathophysiology of SLE and how new and emerging biological agents help address the underlying pathophysiological aberrations in patients with SLE. The faculty will also review strategies to minimize the risk of infections and other toxicities in patients with SLE.

To describe vascular events during an 8-year followup in a multicenter systemic lupus erythematosus (SLE) inception cohort and their attribution to atherosclerosis.Clinical data, including comorbidities, were recorded yearly. Vascular events were recorded and attributed to atherosclerosis or not. All of the events met standard clinical criteria. Factors associated with atherosclerotic vascular events were analyzed using descriptive statistics, t-tests, and chi-square tests. Stepwise multivariate logistic regression was used to assess the association of factors with vascular events attributed to atherosclerosis.Since 2000, 1,249 patients have been entered into the cohort. There have been 97 vascular events in 72 patients, including: myocardial infarction (n = 13), angina (n = 15), congestive heart failure (n = 24), peripheral vascular disease (n = 8), transient ischemic attack (n = 13), stroke (n = 23), and pacemaker insertion (n = 1). Fifty of the events were attributed to active lupus, 31 events in 22 patients were attributed to atherosclerosis, and 16 events were attributed to other causes. The mean +/- SD time from diagnosis to the first atherosclerotic event was 2.0 +/- 1.5 years. Compared with patients followed for 2 years without atherosclerotic events (n = 615), at enrollment, patients with atherosclerotic vascular events were more frequently white, men, older at diagnosis of SLE, obese, smokers, hypertensive, and had a family history of coronary artery disease. On multivariate analysis, only male sex and older age at diagnosis were associated factors.In an inception cohort with SLE followed for up to 8 years, there were 97 vascular events, but only 31 were attributable to atherosclerosis. Patients with atherosclerotic events were more likely to be men and to be older at diagnosis of SLE.

Purpose It was hypothesised that the coding variant (R77H), rs1143679, within ITGAM could predict specific clinical manifestations associated with systemic lupus erythematosus (SLE). Method To assess genetic association, 2366 patients with SLE and 2931 unaffected controls with European ancestry were analysed. The patients with SLE were coded by the presence or absence of individual American College of Rheumatology criteria. Logistic regression and Pearson χ 2 tests were used to assess statistical significance. Results For overall case-control analysis, a highly significant association was detected (p=2.22×10−21, OR 1.73). Using case-only analysis, a significant association was detected with renal criteria (p=0.0003), discoid rash (p=0.02) and immunological criteria (p=0.04). When patients with SLE were compared with healthy controls, the association became stronger for renal (p=4.69×10−22, OR 2.15), discoid (p=1.77×10−14, OR 2.03) and immunological (p=3.49×10−22, OR 1.86) criteria. Risk allele frequency increased from 10.6% (controls) to 17.0% (SLE), 20.4% (renal), 18.1% (immunological) and 19.5% (discoid). Conclusion These results show a strong association between the risk allele (A) at rs1143679 and renal disease, discoid rash and immunological manifestations of SLE.

Objective. To evaluate the sensitivity and specificity of magnetic resonance imaging (MRI) in detecting erosions, bone edema, and synovitis in the metacarpophalangeal and wrist joints for rheumatoid arthritis (RA).Methods. MRI scans of bilateral hands and wrists of 40 healthy subjects and 40 RA patients were performed using 0.2 T extremity-MRI and read blindly using a modified RA MRI (RAMRIS) system (no contrast injection, imaging in 1 plane only). To determine interreader reliability, images of 10 randomly selected subjects were read independently by a musculoskeletal radiologist.Results. A total of 3360 bones were evaluated. Patients with RA had significantly more erosions as well as higher scores for bone edema and synovitis than healthy subjects. Age had a significant effect on the number of erosions in both groups. However, when disease duration was factored in, age became insignificant in RA patients. Erosion number correlated with positive rheumatoid factor and higher C-reactive protein values. The intraclass correlation coefficient between the 2 readers was 0.76 for individual joints and 0.88 for total scores. When having a single erosion was used as a positive test for RA, the sensitivity of this test was 90%, but the specificity was only 35%. Presence of bone edema provided 65% sensitivity and 82.5% specificity. Eliminating the lunate from scoring for bone edema increased the specificity to 87.5% while decreasing the sensitivity to 62.5%.Conclusion. While MRI is a highly sensitive tool for identifying and tracking the progression of erosions, erosions detected by MRI with measures commonly used in a rheumatologist’s office (no contrast, imaging in 1 plane) provide low specificity for RA. Bone marrow edema is the most specific MRI lesion for RA in this setting.

Objective. To assess the safety, tolerability, biologic activity, and efficacy of belimumab in combination with standard of care therapy (SOC) in patients with active systemic lupus erythematosus (SLE). Methods. Patients with a Safety of Estrogens in Lupus Erythematosus: National Assessment (SELENA) version of the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) score > 4( n 449) were randomly assigned to belimumab (1, 4, or 10 mg/kg) or placebo in a 52-week study. Coprimary end points were the percent change in the SELENA–SLEDAI score at week 24 and the time to first SLE flare. Results. Significant differences between the treatment and placebo groups were not attained for either primary end point, and no dose response was observed. Reductions in SELENA–SLEDAI scores from baseline were 19.5% in the combined belimumab group versus 17.2% in the placebo group. The median time to first SLE flare was 67 days in the combined belimumab group versus 83 days in the placebo group. However, the median time to first SLE flare during weeks 24 –52 was significantly longer with belimumab treatment (154 versus 108 days; P 0.0361). In the subgroup (71.5%) of serologically active patients (antinuclear antibody titer >1:80 and/or anti– double-stranded DNA [anti-dsDNA] >30 IU/ml), belimumab treatment resulted in significantly better responses at week 52 than placebo for SELENA–SLEDAI score (28.8% versus 14.2%; P 0.0435), physician’s global assessment (32.7% versus 10.7%; P 0.0011), and Short Form 36 physical component score (3.0 versus 1.2 points; P 0.0410). Treatment with belimumab resulted in a 63–71% reduction of naive, activated, and plasmacytoid CD20 B cells, and a 29.4% reduction in anti-dsDNA titers (P 0.0017) by week 52. The rates of adverse events and serious adverse events were similar in the belimumab and placebo groups. Conclusion. Belimumab was biologically active and well tolerated. The effect of belimumab on the reduction of SLE disease activity or flares was not significant. However, serologically active SLE patients responded significantly better to belimumab therapy plus SOC than to SOC alone.

In our earlier study, we utilized a Bayesian design to probe the association of approximately 1000 genes (approximately 10,000 single-nucleotide polymorphisms (SNPs)) with systemic lupus erythematosus (SLE) on a moderate number of trios of parents and children with SLE. Two genes associated with SLE, with a multitest-corrected false discovery rate (FDR) of0.05, were identified, and a number of noteworthy genes with FDR of0.8 were also found, pointing out a future direction for the study. In this report, using a large population of controls and adult- or childhood-onset SLE cases, we have extended the earlier investigation to explore the SLE association of 10 of these noteworthy genes (109 SNPs). We have found that seven of these genes exhibit a significant (FDR0.05) association with SLE, both confirming some genes that have earlier been found to be associated with SLE (PTPN22 and IRF5) and presenting novel findings of genes (KLRG1, interleukin-16, protein tyrosine phosphatase receptor type T, toll-like receptor (TLR)8 and CASP10), which have not been reported earlier. The results signify that the two-step candidate pathway design is an efficient way to study the genetic foundations of complex diseases. Furthermore, the novel genes identified in this study point to new directions in both the diagnosis and the eventual treatment of this debilitating disease.

Systemic lupus erythematosus (SLE) is an autoimmune disease with highly variable clinical presentation. Patients suffer from immunological abnormalities that target T-cell, B-cell and accessory cell functions. B cells are hyperactive in SLE patients. An adapter protein expressed in B cells called BANK1 (B-cell scaffold protein with ankyrin repeats) was reported in a previous study to be associated with SLE in a European population. The objective of this study was to assess the BANK1 genotype-phenotype association in an independent replication sample. We genotyped 38 single nucleotide polymorphisms (SNPs) in BANK1 on 1892 European-derived SLE patients and 2652 European-derived controls. The strongest associations with SLE and BANK1 were at rs17266594 (corrected P-value=1.97 x 10(-5), odds ratio (OR)=1.22, 95% CI 1.12-1.34) and rs10516487 (corrected P-value=2.59 x 10(-5), OR=1.22, 95% CI 1.11-1.34). Our findings suggest that the association is explained by these two SNPs, confirming previous reports that these polymorphisms contribute to the risk of developing lupus. Analysis of patient subsets enriched for hematological, immunological and renal ACR criteria or the levels of autoantibodies, such as anti-RNP A and anti-SmRNP, uncovers additional BANK1 associations. Our results suggest that BANK1 polymorphisms alter immune system development and function to increase the risk for developing lupus.