OP0069 FACTORS ASSOCIATED WITH THE RISK OF SEPSIS IN PATIENTS WITH IMMUNE-MEDIATED INFLAMMATORY DISEASES TREATED WITH ANTI-TUMOR NECROSIS FACTOR-ALPHA: A NATIONWIDE, POPULATION-BASED COHORT STUDY

Background:Anti-TNF-α agents have been proven to be effective for patients with immune-medicated inflammatory diseases (IMIDs) including rheumatoid arthritis (RA), ankylosing spondylitis (AS), psoriasis (PsO), psoriatic arthritis (PsA), Crohn’s disease (CD) and ulcerative colitis (UC). Prior studies have shown an increased risk of infection in IMID patients treated with anti-TNF-α but limited studies investigated factors associated with the development of sepsis in patients with IMIDs.Objectives:To investigate factors associated with the development of sepsis in patients with IMIDs using the Taiwanese National Health Insurance Research Database (NHIRD).Methods:We identified all biologic-naïve patients with RA, AS, PsO, PsA, or CD/UC from the claim data via the NHIRD who started their first anti-TNF-α agent (etanercept (ETN), adalimumab (ADA) or golimumab (GOL)) between 2003 and 2017 as study subjects. The index date was the first date of anti-TNF-α prescription. Sepsis was defined based on the sepsis-3 definition. We identified sepsis patients using a validated ICD-9-CM coding system proposed by Angus et al, in which a diagnosis of bacterial/fungal infection with one or more acute organ dysfunction is required to define an episode of sepsis. All study subjects were followed up till the date of first hospitalization due to sepsis, 90 days after the last date of anti-TNF-α prescription, withdrawal from NHIRD or death, whichever came first. We used a Cox regression analysis to assess the associations of covariates with the risk of sepsis shown as hazard ratios (HRs) with 95% confidence interval (CIs). Covariates included anti-TNF-α agent, IMID, age, sex, insured amount, level of urbanization, disease duration, Charlson comorbidity index (CCI), a history of prior hospitalization due to sepsis within 3 months before the index date and medication use within 12 months before the index date and during the follow-up period.Results:We identified 18105 biologic-naïve patients with IMIDs, including 8123 ETN users, 7623 ADA users and 2359 GOL users. The incidence rates (IRs) of sepsis in patients treated with ETN, ADA and GOL were 1080, 1181, and 617 per 105years respectively. Multivariable regression analyses showed that factors associated with an increased risk of sepsis were use of ADA (ETN as reference: HR, 1.21; 95% CI, 1.02–1.42), male (HR, 1.24; 95% CI, 1.04–1.48), age (HR, 1.06; 95% CI, 1.05–1.07), CD/UC (HR, 2.35; 95% CI, 1.57–3.53), CCI (HR, 1.30; 95% CI, 1.23–1.38), prior sepsis (HR, 2.42; 95% CI, 1.78–3.29), prior use of sulfasalazine (HR, 1.25, 95% CI, 1.00-1.55), lower levels of urbanization (level III: HR, 1.37; 95% CI, 1.06–1.77; level IV: HR, 1.68, 95% CI, 1.35–2.10). Factors associated with a decreased risk of sepsis were use of GOL (ETN as reference: HR, 0.59; 95% CI, 0.39–0.84), use of methotrexate (HR, 0.78; 95% CI, 0.65–1.00), and higher insured amount (reference: ≤ 15480 NTD; 15480–28800 NTD: HR, 0.83; 95% CI, 0.68–0.99; 28800–45800 NTD: HR, 0.58; 95% CI, 0.45–0.74; >45800 NTD: HR, 0.33; 95% CI, 0.21–0.54).Conclusion:Our study revealed that among biologic-naïve IMID patients initiating anti-TNF-α treatment, use of ADA, age, sex, CD/UC, CCI, prior sepsis, prior use of sulfasalazine and lower levels of urbanization were associated with an increased risk of sepsis, while use of GOL, use of methotrexate, and higher insured amount were associated with a decreased risk of sepsis.Disclosure of Interests:BO-CHUEN HSU: None declared, Hsin-Hua Chen: None declared, Ching-Heng Lin: None declared, Yi-Ming Chen: None declared, Kuo-Lung Lai: None declared, Der-Yuan Chen: None declared, Wen-Nan Huang: None declared, Yi-Hsing Chen Grant/research support from: Taiwan Ministry of Science and Technology, Taiwan Department of Health, Taichung Veterans General Hospital, National Yang-Ming University, GSK, Pfizer, BMS., Consultant of: Pfizer, Novartis, Abbvie, Johnson & Johnson, BMS, Roche, Lilly, GSK, Astra& Zeneca, Sanofi, MSD, Guigai, Astellas, Inova Diagnostics, UCB, Agnitio Science Technology, United Biopharma, Thermo Fisher, Gilead., Paid instructor for: Pfizer, Novartis, Johnson & Johnson, Roche, Lilly, Astra& Zeneca, Sanofi, Astellas, Agnitio Science Technology, United Biopharma., Speakers bureau: Pfizer, Novartis, Abbvie, Johnson & Johnson, BMS, Roche, Lilly, GSK, Astra& Zeneca, Sanofi, MSD, Guigai, Astellas, Inova Diagnostics, UCB, Agnitio Science Technology, United Biopharma, Thermo Fisher, Gilead.

OP0118 DECIPHERING THE ANTI-PROTEIN-ARGININE DEIMINASE (PAD) RESPONSE IDENTIFIES PAD1 AND PAD6 AS NOVEL AUTOANTIGENS IN RHEUMATOID ARTHRITIS

Background:Protein-arginine deiminase (PAD) 4 enzymes play a central role in the pathogenesis of rheumatoid arthritis (RA) and represents an antigenic target. Among the five known family members (PAD1, PAD2, PAD3, PAD4 and PAD6), only PAD2, PAD3 and PAD4 have been described to have autoantigenic properties. Furtheremore, very little is known on the the isotype usage of these autoantibodies. Understanding the molecular basis of the anti-PAD antibody reponse has the potential to open novel approaches for precision medicine in RA.Objectives:The objectives of this study were to screen for the presence of antibodies to the five PAD family members and to evaluate the isotype usage of the anti-PAD4 response in RA.Methods:First, we developed a panel for the detection of anti-PAD IgG based on a particle-based multi-analyte technology (PMAT), that utilized paramagnetic particles coupled with the different human recombinant PAD proteins (PAD1, PAD2, PAD3, PAD4 and PAD6) and anti-human IgG conjugate. This panel was used to test sera from RA patients (n=33) and non-RA controls (n=36). The controls were comprised of apparently healthy individuals (n=10), and patients with infectious diseases (n=10), systemic lupus erythematosus (n=7), systemic sclerosis (n=9) and Sjogren’s syndrome (n=1). Next, the PAD4-coupled beads were tested with anti-human IgM, IgA and IgG conjugates on an extended cohort of RA patients (n=62) and the same non-RA controls.Results:All five anti-PAD IgG (Figure 1) demonstrated the ability to discriminate between RA patients and controls. At greater than 90% specificity, anti-PAD4 IgG, followed by anti-PAD3 IgG, showed the best diagnostic performance. Significantly higher levels of the five antibodies were observed in RAvs.controls (p-values of 0.0041, <0.0001, 0.0014, 0.0039, and 0.0140 for anti-PAD1, 2, 3, 4 and 6, respectively). Significant correlation was observed between all the antibodies, with the highest between anti-PAD1 and anti-PAD4 (Spearman´srho=0.87,p<0.0001) and the lowest between anti-PAD4 and anti-PAD2 (Spearman’srho=0.38,p=0.0015) and anti-PAD4 and anti-PAD6 (Spearman’srho=0.38,p=0.0011). While principal component analysis (PCA) (Figure 2) showed an association between all anti-PAD antibodies, there was further discrimination that displayed closer association between anti-PAD1, 3 and 4 on one hand, and between anti-PAD2 and 6. For the extended testing of anti-PAD4 with IgG, IgA and IgM, all three isotypes were identified in the sera of RA patients. Higher levels of the three isotypes were observed in RA patients with erosive disease when compared with the patients without erosion, but this association was only significant for anti-PAD4 IgA (p=0.0086).Figure 1.Receiver operating characteristics (ROC) analysis of the discrimination between rheumatoid arthritis (RA) and controls of IgG to protein-arginine deiminase (PAD) 1, PAD2, PAD3, PAD4 and PAD6. The area under the curve (AUC) values are shown in brackets for each biomarker.Abbreviations:TPF: true positive fraction; FPF: false positive fractionFigure 2.Two dimensional principal component analysis (PCA) plot of the anti-PAD levels in RA patients (n=33) and controls (n=36). Anti-PAD1, 3 and 4 have the main contribution to PC1, which explains 51.7% of the variance, and anti-PAD2 and 6 to PC2, that represents 20.8% of it.Abbreviations:PC: principal componentConclusion:Our study is the first to describe PAD1 and PAD6 as novel antigenic targets in RA and to demostrate that the anti-PAD4 B-cell immune response uses all three isotypes (IgG, IgA and IgM). The strong and significant association between anti-PAD4 IgA and joint erosion is of particular clinical relevance.Disclosure of Interests:Laura Martinez-Prat Employee of: I am an employee of Inova Diagnostics, an in vitro diagnostics company., Mary Ann Aure Employee of: I am an employee of Inova Diagnostics, an in vitro diagnostics company., Chelsea Bentow Employee of: I am an employee of Inova Diagnostics, an in vitro diagnostics company., David Lucia Employee of: I am an employee of Inova Diagnostics, an in vitro diagnostics company., Marcos Lopez-Hoyos Consultant of: Inova Diagnostics, an in vitro diagnostics company., Michael Mahler Employee of: I am an employee of Inova Diagnostics, an in vitro diagnostics company.

Analytical and Clinical Comparison of Two Fully Automated Immunoassay Systems for the Diagnosis of Celiac Disease

Objective.Here we compared analytical and clinical performance characteristics of two novel automated assay systems for the detection of celiac disease (CD) specific antibodies: QUANTA Flash (INOVA Diagnostics, Inc.) and EliA (Thermo Scientific).Methods.A total of 74 biopsy-proven CD patients (2 with IgA deficiency) and 138 controls were tested by both methods.Results.Sensitivities of QUANTA Flash assays ranged from 35.1% to 90.5% and specificities from 96.4% to 99.3%, while sensitivities for EliA assays ranged from 37.8% to 90.5% (equivocal considered positive) and specificities from 97.1% to 100.0%. Good qualitative agreement was found between all assays. Thirty-four (50.0%) of the 68 QUANTA Flash h-tTG IgA positive results were higher than 10 times the upper limit of normal (ULN). In contrast, only 22.8% of the EliA tTG IgA positive samples were >10x ULN. Seventy-three (98.6%) biopsy-proven CD patients were correctly identified with the QUANTA Flash h-tTG IgA+DGP IgG combination, while 64 (86.5%) and 72 (97.3%) (depending on equivocal range) were identified with the same combination of EliA assays.Conclusion.The QUANTA Flash CD assays have outstanding clinical performance. Of particular clinical significance, in light of proposals to decrease the absolute necessity of biopsy, was the demonstration that 50% of the QUANTA Flash h-tTG IgA results were >10x ULN.

Highlights on Novel Technologies for the Detection of Antibodies to Ro60, Ro52, and SS-B

Objective. We aimed to compare a chemiluminescent immunoassay (CIA, QUANTA Flash) on BIO-FLASH with a multiplex flow immunoassay (MFI) on BioPlex 2200 for the detection of antibodies to Ro60, Ro52, and SS-B.Methods. The study included 241 samples, from patients suffering from systemic autoimmune diseases (n=108) as well as disease controls (n=133). All samples were tested for anti-Ro52, anti-Ro60, and anti-SS-B (La) antibodies on QUANTA Flash (INOVA Diagnostics, San Diego, USA) and BioPlex 2200 (Bio-Rad Laboratories Inc., Hercules, USA). Discrepant samples were tested by two independent methods: BlueDot/ANA and QUANTRIX Microarray (both D-tek, Belgium).Results. The overall qualitative agreements were 95.4% (95% confidence interval, CI 92.0–97.7%) for anti-Ro52, 98.8% (95% CI 96.4–99.7%) for anti-Ro60, and 91.7% (95% CI 87.5–94.9%) for anti-SS-B antibodies. There were 34 discrepant samples among all assays (20 anti-SS-B, 11 anti-Ro52, 3 anti-Ro60). 30/33 of retested samples (by D-tek dot blot) agreed with the QUANTA Flash results. Similar findings were obtained with QUANTRIX Microarray kit.Conclusion. QUANTA Flash and BioPlex 2200 show good qualitative agreement. The clinical performances were similar for anti-Ro52 and anti-Ro60 autoantibodies while differences were observed for anti-SS-B (La) antibodies.

New Serology Assays Can Detect Gluten Sensitivity among Enteropathy Patients Seronegative for Anti–Tissue Transglutaminase

Background: Some patients with celiac disease (CD) may be seronegative with the commonly used test for IgA anti–tissue transglutaminase (anti-tTG) antibodies. Our aim was to explore whether newer assays incorporating synthetic deamidated gliadin-related peptides (DGPs) or other TG isoenzymes as antigen are useful for detecting gluten sensitivity in IgA anti-tTG–seronegative patients. Methods: We assayed serum samples obtained at diagnosis from (a) anti-tTG–seronegative patients with a CD-like enteropathy (n = 12), (b) skin biopsy–proven dermatitis herpetiformis (DH) patients (n = 26), and (c) IgA anti-tTG–positive CD patients (n = 26). All patients had typical total IgA concentrations. All patients underwent intestinal biopsy and serum testing for (a) detection of IgA and IgG isotypes of both anti-DGP and anti-tTG in a single assay (tTG/DGP Screen; INOVA Diagnostics), (b) simultaneous detection of both IgA and IgG anti-DGP antibody isotypes (DGP Dual; INOVA Diagnostics), and (c) detection of antibodies to transglutaminase 3 (TG3) or transglutaminase 6 (TG6). Results: All anti-tTG–seropositive patients also tested positive in anti-DGP assays. Overall, tTG/DGP Screen detected 6 (31.6%) of the 19 anti-tTG seronegatives, and anti-DGP Dual produced positive results in 5 (26.3%) of these cases. Whereas both assays detected 2 anti-tTG–negative DH patients with partial villous atrophy, they were positive in only 2 of the 5 cases with no histologically discernible mucosal damage. Testing for antibodies to TG3 and TG6 identified 7 (36.8%) of the 19 anti-tTG–negative patients, 5 of which were also positive for anti-DGP. Conclusions: Detection of anti-DGP with tTG/DGP Screen or anti-DGP Dual, or detection of antibodies to other TG isoenzymes, enhances the sensitivity for detecting gluten sensitivity among non–IgA- deficient, anti-tTG–seronegative patients with CD-like enteropathy.

Evaluation of the INOVA Diagnostics Enzyme-Linked Immunosorbent Assay Kits for Measuring Serum Immunoglobulin G (IgG) and IgA to Deamidated Gliadin Peptides

ABSTRACT New assays for antibodies to deamidated gliadin peptides (DGP) expressing celiac disease-specific epitopes were evaluated using 154 sera previously tested for endomysial immunoglobulin A (IgA) (EMA), transglutaminase IgA (TGA), and conventional gliadin antibodies. DGP antibody results showed 97% concordance with EMA and TGA results. Of 56 sera negative for EMA and TGA but positive for conventional gliadin antibodies, 54 (96%) were negative for DGP antibodies.