Faculty Bibliography

OBJECTIVE: Membrane endothelial protein C receptor (mEPCR) is highly expressed in peritubular capillaries of kidneys from patients with active and poorly responsive lupus nephritis (LN). We investigated the hypothesis that changes in the microvasculature are widespread with extension to the dermal vasculature. METHODS: Skin biopsies from uninvolved skin (buttocks) were performed in 27 patients with LN and 5 healthy controls. Sections were stained with specific antibodies reactive with mEPCR, adiponectin, intercellular adhesion molecule-1 (ICAM-1), and CD31; then assessed by enumeration of stained blood vessels (percentage positive blood vessels) blinded to knowledge of clinical information. RESULTS: There was a significant increase in the prevalence of blood vessels that stained for mEPCR and ICAM-1 in patients compared to controls [94% vs 59% (p = 0.045) and 81% vs 67% (p = 0.037), respectively]. Adiponectin staining and CD31 staining were similar between the groups (45% vs 43% and 98% vs 92%). Dermal staining for mEPCR was greater in patients with proliferative glomerulonephritis than in those with membranous disease (96% vs 60%; p = 0.029). A composite of poor prognostic renal markers and death was significantly associated with greater expression of mEPCR staining. CONCLUSION: These data are consistent with the notion that in patients with LN, activation of the microvasculature extends beyond the clinically targeted organ. The insidious expression of this widespread vasculopathy may be a contributor to longterm comorbidities.

OBJECTIVE: Several confirmed genetic susceptibility loci for lupus have been described. To date, no clear evidence for genetic epistasis is established in lupus. We test for gene-gene interactions in a number of known lupus susceptibility loci. METHODS: Eighteen SNPs tagging independent and confirmed lupus susceptibility loci were genotyped in a set of 4,248 lupus patients and 3,818 normal healthy controls of European descent. Epistasis was tested using a 2-step approach utilizing both parametric and non-parametric 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 lupus patients. 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-score= 3.95, P= 7.8x10(-5) (FDR</=0.05), P(MDR) = 5.9x10(-45) ). Importantly, our data suggest that in lupus patients 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 0.0047). 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

OBJECTIVES: Altered signalling in B cells is a predominant feature of systemic lupus erythematosus (SLE). The genes BANK1 and BLK were recently described as associated with SLE. BANK1 codes for a B-cell-specific cytoplasmic protein involved in B-cell receptor signalling and BLK codes for an Src tyrosine kinase with important roles in B-cell development. To characterise the role of BANK1 and BLK in SLE, a genetic interaction analysis was performed hypothesising that genetic interactions could reveal functional pathways relevant to disease pathogenesis. METHODS: The GPAT16 method was used to analyse the gene-gene interactions of BANK1 and BLK. Confocal microscopy was used to investigate co-localisation, and immunoprecipitation was used to verify the physical interaction of BANK1 and BLK. RESULTS: Epistatic interactions between BANK1 and BLK polymorphisms associated with SLE were observed in a discovery set of 279 patients and 515 controls from northern Europe. A meta-analysis with 4399 European individuals confirmed the genetic interactions between BANK1 and BLK. As BANK1 was identified as a binding partner of the Src tyrosine kinase LYN, the possibility that BANK1 and BLK could also show a protein-protein interaction was tested. The co-immunoprecipitation and co-localisation of BLK and BANK1 were demonstrated. In a Daudi cell line and primary naive B cells endogenous binding was enhanced upon B-cell receptor stimulation using anti-IgM antibodies. CONCLUSION: This study shows a genetic interaction between BANK1 and BLK, and demonstrates that these molecules interact physically. The results have important consequences for the understanding of SLE and other autoimmune diseases and identify a potential new signalling pathway

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/CONCLUSIONS:Disease activity in newly diagnosed patients decreases over their first 5 years, while damage increases. Antibody positivity ran variable courses over this period.

OBJECTIVE: To examine the relationship between circulating B lymphocyte stimulator (BLyS) levels and humoral responses to influenza vaccination in systemic lupus erythematosus (SLE) patients, as well as the effect of vaccination on BLyS levels, and to investigate clinical and serologic features of SLE that are associated with elevated BLyS levels. METHODS: Clinical history, disease activity measurements, and blood specimens were collected from 60 SLE patients at baseline and after influenza vaccination. Sera were tested for BLyS levels, lupus-associated autoantibodies, serum interferon-alpha (IFNalpha) activity, 25-hydroxyvitamin D (25[OH]D), and humoral responses to influenza vaccination. RESULTS: Thirty percent of the SLE patients had elevated BLyS levels, with African American patients having higher BLyS levels than white patients (P = 0.006). Baseline BLyS levels in patients were not correlated with humoral responses to influenza vaccination (P = 0.863), and BLyS levels increased postvaccination only in the subset of patients with BLyS levels in the lowest quartile (P = 0.0003). Elevated BLyS levels were associated with increased disease activity, as measured by the SLE Disease Activity Index, physician's global assessment, and Systemic Lupus Activity Measure in white patients (P = 0.035, P = 0.016, and P = 0.018, respectively), but not in African Americans. Elevated BLyS levels were also associated with anti-nuclear RNP (P = 0.0003) and decreased 25(OH)D (P = 0.018). Serum IFNalpha activity was a significant predictor of elevated BLyS in a multivariate analysis (P = 0.002). CONCLUSION: Our findings indicate that African American patients with SLE have higher BLyS levels regardless of disease activity. Humoral response to influenza vaccination is not correlated with baseline BLyS levels in SLE patients, and only those patients with low baseline BLyS levels demonstrate an increased BLyS response after vaccination

OBJECTIVE: To assess the efficacy/safety of the B lymphocyte stimulator inhibitor belimumab plus standard therapy compared with placebo plus standard therapy in active systemic lupus erythematosus (SLE). METHODS: In a phase III, multicenter, randomized, placebo-controlled trial, 819 antinuclear antibody-positive or anti-double-stranded DNA-positive SLE patients with scores >/=6 on the Safety of Estrogens in Lupus Erythematosus National Assessment (SELENA) version of the SLE Disease Activity Index (SLEDAI) were randomized in a 1:1:1 ratio to receive 1 mg/kg belimumab, 10 mg/kg belimumab, or placebo intravenously on days 0, 14, and 28 and then every 28 days for 72 weeks. The primary efficacy end point was the SLE Responder Index (SRI) response rate at week 52 (an SRI response was defined as a >/=4-point reduction in SELENA-SLEDAI score, no new British Isles Lupus Assessment Group [BILAG] A organ domain score and no more than 1 new BILAG B score, and no worsening in physician's global assessment score versus baseline). RESULTS: Belimumab at 10 mg/kg plus standard therapy met the primary efficacy end point, generating a significantly greater SRI response at week 52 compared with placebo (43.2% versus 33.5%; P = 0.017). The rate with 1 mg/kg belimumab was 40.6% (P = 0.089). Response rates at week 76 were 32.4%, 39.1%, and 38.5% with placebo, 1 mg/kg belimumab, and 10 mg/kg belimumab, respectively. In post hoc sensitivity analyses evaluating higher SELENA-SLEDAI score thresholds, 10 mg/kg belimumab achieved better discrimination at weeks 52 and 76. Risk of severe flares over 76 weeks (based on the modified SLE Flare Index) was reduced with 1 mg/kg belimumab (34%) (P = 0.023) and 10 mg/kg belimumab (23%) (P = 0.13). Serious and severe adverse events, including infections, laboratory abnormalities, malignancies, and deaths, were comparable across groups. CONCLUSION: Belimumab plus standard therapy significantly improved SRI response rate, reduced SLE disease activity and severe flares, and was generally well tolerated in SLE

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-IFNalpha 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 IFNalpha, and exploratory analyses suggest that IFNalpha inhibition may be associated with clinical benefit in SLE. Trial registration number NCT00299819

OBJECTIVE: Candidate gene and genome-wide association studies have identified several disease susceptibility loci in lupus patients. These studies have largely been performed in lupus patients who are Asian or of European ancestry. This study was undertaken to examine whether some of these same susceptibility loci increase lupus risk in African American individuals. METHODS: Single-nucleotide polymorphisms tagging 15 independent lupus susceptibility loci were genotyped in a set of 1,724 lupus patients and 2,024 healthy controls of African American descent. The loci examined included PTPN22, FCGR2A, TNFSF4, STAT4, CTLA4, PDCD1, PXK, BANK1, MSH5 (HLA region), CFB (HLA region), C8orf13-BLK region, MBL2, KIAA1542, ITGAM, and MECP2/IRAK1. RESULTS: We found the first evidence of genetic association between lupus in African American patients and 5 susceptibility loci (C8orf13-BLK, BANK1, TNFSF4, KIAA1542, and CTLA4; P = 8.0 x 10(-6) , P = 1.9 x 10(-5) , P = 5.7 x 10(-5) , P = 0.00099, and P = 0.0045, respectively). Further, we confirmed the genetic association between lupus and 5 additional lupus susceptibility loci (ITGAM, MSH5, CFB, STAT4, and FCGR2A; P = 7.5 x 10(-11) , P = 5.2 x 10(-8) , P = 8.7 x 10(-7) , P = 0.0058, and P = 0.0070, respectively), and provided evidence, for the first time, of genome-wide significance for the association between lupus in African American patients and ITGAM and MSH5 (HLA region). CONCLUSION: These findings provide evidence of novel genetic susceptibility loci for lupus in African Americans and demonstrate that the majority of lupus susceptibility loci examined confer lupus risk across multiple ethnicities