C. Fabro

The CC homozygosis of the -174G>C IL-6 polymorphism predicts a lower efficacy of rituximab therapy in rheumatoid arthritis

Identification of genetic biomarkers of response to biologics in rheumatoid arthritis (RA) is a relevant issue. Being IL-6 a key cytokine for B cell survival, the interleukin-6 (IL-6) -174G>C and the IL-6 receptor (IL-6R) D358A gene polymorphisms were investigated in 158 RA patients treated with rituximab (RTX). One hundred and twenty-eight (81.0%) were RF positive and 126 (79.7%) were anti-CCP positive. Response to therapy was evaluated at the end of the sixth month after the first RTX infusion, by using both the EULAR and the ACR criteria. The possible relationship with IL-6 serum levels was also studied. By univariate analysis, lack of response by the EULAR criteria was more prevalent in RA patients with the IL-6 -174 CC genotypes (39.1%), than in the GC/GG patients (18.5%) (OR 2.83; 95%CI=1.10-7.27; p=0.031). A good response was noticed in only one patient (4.3%) with the IL-6 -174 CC genotype, while it was present in 24.4% of GG/GC cases (p=0.06). By stepwise multivariate analysis (including RA duration, baseline DAS28, baseline HAQ, RF status, anti-CCP status and IL-6 genotype as covariates), the IL-6 -174CC genotype was selected as an independent predictor of no response to RTX by both EULAR and ACR≥50 criteria, while the IL-6R polymorphism resulted as not associated. No definite association between gene polymorphisms and IL-6 serum levels was noticed. Present results suggest a possible role for IL-6 genotyping to better plan treatment with RTX in RA, and larger studies are worthwhile.

Transforming growth factor β 869C/T and interleukin 6 -174G/C polymorphisms relate to the severity and progression of bone-erosive damage detected by ultrasound in rheumatoid arthritis

IntroductionSingle nucleotide polymorphisms (SNPs) of transforming growth factor β (TGF-β) and IL-6 genes (respectively, 869C/T and -174G/C) have been associated with radiographic severity of bone-erosive damage in patients with rheumatoid arthritis (RA). Musculoskeletal ultrasound (US) is more sensitive than radiography in detecting bone erosion. We analyzed the association between TGF-β 869C/T and IL-6 -174G/C SNPs and bone-erosive damage, evaluated by US, in a cohort of patients with severely active RA.MethodsSeventy-seven patients were enrolled before beginning anti-TNF treatment. Disease activity was measured using the disease activity score in 28 joints, and the clinical response was evaluated according to the European League Against Rheumatism response criteria. Rheumatoid factor (RF) and anticitrullinated protein/peptide antibodies (ACPAs) were detected. The 869C/T TGF-β and -174G/C IL-6 SNPs were analyzed by PCR amplification. US was performed to assess the bone surfaces of metacarpophalengeal (MCP), proximal interphalangeal (PIP) and metatarsophalangeal (MTP) joints by obtaining multiplanar scans. According to the number of erosions per joint, a semiquantitative score ranging from 0 to 3 was calculated in each anatomical site to obtain a MCP total erosion score (TES), a PIP TES and a MTP TES, all ranging from 0 to 30, and a global patient TES calculated as the sum of these scores (range, 0 to 90).ResultsPatients carrying the TGF-β 869TT genotype showed a statistically significant lower MTP TES than those with the CC or CT genotype (mean MTP TES ± standard deviation for 869TT 6.3 ± 5.7 vs. 869CC/CT 11.7 ± 7.8; P = 0.011). Interestingly, patients with the TT genotype showed dichotomous behavior that was dependent on autoantibody status. In the presence of ACPAs and/or RF, the TT genotype was associated with lower erosion scores at all anatomical sites compared with the CC and CT genotypes. Conversely, the same 869TT patients showed higher erosion scores in the absence of ACPAs or RF.ConclusionsIn RA patients, TGF-β 869C/T SNPs could influence the bone-erosive damage as evaluated by US. The serological autoantibody status (ACPAs and RF) can modulate this interaction.

The TTTT B lymphocyte stimulator promoter haplotype is associated with good response to rituximab therapy in seropositive rheumatoid arthritis resistant to tumor necrosis factor blockers

OBJECTIVE To investigate the polymorphisms in the promoter region of the B lymphocyte stimulator (BLyS) gene as markers of response to rituximab (RTX) in rheumatoid arthritis (RA). METHODS The study was first conducted in 152 Italian RA patients and then replicated in an additional 117 RA patients (73 Italian, 44 British). The European League Against Rheumatism response criteria were used to evaluate the response rate at months 4 and 6 after the first cycle of RTX, by means of the Disease Activity Score in 28 joints using the erythrocyte sedimentation rate; patients were classified according to the best response shown between months 4 and 6. BLyS promoter polymorphisms were analyzed by polymerase chain reaction followed by the analysis of the restriction fragments, BLyS promoter haplotypes were analyzed using the expectation-maximization algorithm, and BLyS serum levels were analyzed using enzyme-linked immunosorbent assay. Odds ratios (ORs) were calculated with 95% confidence intervals (95% CIs). RESULTS The TTTT BLyS promoter haplotype appeared to be significantly associated with response to RTX only in the subset of seropositive patients (those positive for rheumatoid factor and/or anti-cyclic citrullinated peptide). The replication study confirmed that this association was limited to seropositive RA patients in whom treatment with anti-tumor necrosis factor (anti-TNF) agents had previously failed. In the whole series of seropositive patients in whom anti-TNF agents had previously failed, patients carrying the TTTT BLyS promoter haplotype were more prevalent in good responders (18 of 43 [41.9%]) than in moderate responders (20 of 83 [24.1%]) or in nonresponders (1 of 21 [4.8%]) (for good responders versus nonresponders, OR 14.4 [95% CI 1.77-117.39], P=0.0028). Furthermore, multivariate analysis selected the TTTT BLyS promoter haplotype as an independent marker of good response to RTX (for good responders versus nonresponders, OR 16.2 [95% CI 1.7-152.5], P=0.01; for good responders versus moderate responders and nonresponders combined, OR 3.1 [95% CI 1.2-7.8], P=0.02). The relationship between BLyS polymorphisms and BLyS serum levels remained unclear. CONCLUSION BLyS promoter genotyping may be suitable for identifying seropositive RA patients who may have a good response to RTX after anti-TNF agents have failed.

Serum levels of anti-CCP antibodies, anti-MCV antibodies and RF IgA in the follow-up of patients with rheumatoid arthritis treated with rituximab.

Rheumatoid arthritis (RA) is characterized by the presence of circulating rheumatoid factor (RF) and anticitrullinated peptide antibodies (ACPA), which are positive in about 70–80% of patients. APCA have a higher specificity and therefore a higher diagnostic power than RF, but are less informative than RF in monitoring the course of the disease in patients under treatment. Recently, it has been reported that the anticitrullinated vimentin (a-MCV) antibody test can identify a particular subgroup of APCA that may be negative for anticyclic citrullinated peptide (a-CCP) antibodies. Concerning RF, the RF IgA isotype has been described as a more specific marker of erosive joint damage than total RF. The aim of our study was to monitor the levels of a-CCP, a-MCV, total RF and RF IgA in the follow-up of patients with RA treated with B-lymphocytedepletive rituximab (RTX), to detect any differences or peculiarities in patterns of these autoantibodies, especially in relation to their potential use as predictive markers of therapeutic response. We studied 30 patients with RA treated with RTX. All patients were previously unresponsive to at least 6 months of therapy with disease-modifying antirheumatic drugs (DMARDs; methotrexate, leflunomide, cyclosporine, chloroquine) and/or at least 6 months of therapy with anti-TNF biologics. The evaluation of response to RTX was made at month +6 using the EULAR criteria (DAS28). a-CCP, a-MCV, total RF and RF IgA were determined at baseline (before the first infusion of RTX) and after 1, 3 and 6 months. In serum samples obtained before treatment two cytokines essential for Blymphocyte proliferation, interleukin 6 (IL-6) and B-lymphocyte stimulator (BLyS) were also determined. In all patients a significant and consistent reduction in all the tested antibodies was found during follow-up, with no differences in respect of the degree of response to RTX. Of note, at baseline, generally a higher titre of all autoantibodies was seen in patients who then showed a better response to RTX. Finally, there were no differences in serum concentrations of IL-6 and BLyS in patients in relation to the presence or absence of the autoantibodies investigated, nor was there any significant correlation between the serum concentrations of the cytokines and the titres of the autoantibodies. Thus, neither a-MCV compared to a- CCP, nor RF IgA compared to routine total RF, provided any additional predictive information in the follow-up of patients with RA treated with RTX.

B-Lymphocyte Stimulator and a Proliferation-Inducing Ligand Serum Levels in IgA-Deficient Patients With and Without Celiac Disease

IgA deficiency (IgAD) is the most common form of immunodeficiency and frequently associates with autoimmunity, especially with celiac disease (CD). The mechanisms underlying IgAD and the development of autoimmunity are still relatively unknown. Elevated B‐lymphocyte stimulator (BLyS) and APRIL (a proliferation‐inducing ligand) serum levels characterize several autoimmune diseases. We herein investigated BLyS and APRIL serum levels in IgAD patients with and without CD and compared these patients to CD patients with normal IgA and control patients (HBDs). Compared to HBDs, IgAD patients demonstrated a significant increase of BLyS (P < 0.0001) and APRIL (P= 0.003) levels, and no differences were seen between patients with or without CD. While BLyS appeared similarly overexpressed in IgAD and CD patients, APRIL was significantly increased only in IgAD patients. Because APRIL promotes IgA production, its overexpression may represent a physiological mechanism of compensation. BLyS upregulation may be involved in the increased risk of autoimmune disease development characterizing people carrying IgAD.

The -308 TNFα and the -174 IL-6 promoter polymorphisms associate with effective anti-TNFα treatment in seronegative spondyloarthritis.

The genetic predisposition to a long-term efficacy of anti-tumor necrosis factor (TNF)α treatment in seronegative spondyloarthritis (SpA) was investigated by analysing the possible correlation between several single nucleotide gene polymorphisms and the retention rate of anti-TNFα therapies. We compared patients needing to switch the first anti-TNFα (Sw, No. 64) within at least 12 months of follow-up with patients not needing to switch (NSw, No. 123), observing at least 6 months of treatment to establish anti-TNFα failure, leading to treatment change. Response to treatment was evaluated by standardised criteria (BASDAI for axial involvement, DAS28-EULAR for peripheral involvement). The TNFα -308 A allele and the interleukin (IL)-6 -174GG homozygosis resulted as independent biomarkers predicting survival of the first anti-TNFα therapy in SpA patients (P=0.007, odds ratio (OR): 4.4, 95% confidence interval (CI)=1.5–13.1 and P=0.035, OR: 2.1, 95% CI=1.1–4.4). Also, the male gender (P=0.001, OR: 3.4, 95% CI=1.6–7.1) associated with the NSw phenotype, whereas no association was found either with the specific diagnosis or the predominant joint involvement.

FRI0384 Primary sjÖgren’s syndrome stratified based on the severity of patient-reported fatigue

Background fatigue is one of the most common symptoms reported by patients affected by primary Sjögren’s syndrome (pSS), and a major contributor to impaired quality of life. Objectives to analyse the clinical, serological and histological features of pSS patients stratified according to the severity of their self-reported fatigue. Methods among pSS patients undergoing clinical evaluation in our Sjögren’s Clinic in a six-months period (January-June 2017), 86 consecutive unselected patients, fulfilling the latest ACR/EULAR pSS classification criteria, accepted to report their degree of fatigue on a 10 cm VAS (range 0–100) and to complete the ESSPRI questionnaire. Four subgroups of fatigue severity were defined, as previously published1: no fatigue (VAS=0); low fatigue (VAS=1–24); moderate fatigue (VAS=25–74); high fatigue (VAS=75–100). For each subgroup demographic, serological, histological features and ESSDAI score were collected, as well as the prevalence of pSS-related lymphoma, fibromyalgia (FM), autoimmune thyroiditis, and anaemia. Results fatigue was reported by the 87.2% (n=75) of pSS patients, distributed in subgroups as following: 25.3% (n=19) with low fatigue, 58.7% (n=44) with moderate fatigue and 16% (n=12) with high fatigue. Lymphoma was significantly (p=0.0133) more frequent in the pSS subgroup with high fatigue (33.4%, by considering active lymphoma cases; 50%, by considering also the cases with lymphoma in remission). FM patients were a minority (4.7% ; n=4), and never complained of high fatigue, all of them reporting moderate fatigue. A significant correlation was finally found between fatigue severity and ESSPRI (p<0.0001), but not with ESSDAI (p=0.31). No significant age or sex difference was observed between subgroups. Also autoimmune thyroiditis, anaemia, anti-SSA and/or anti-SSB positivity, rheumatoid factor positivity, and cryoglobulinemia showed no significant difference between subgroups. Conclusions when fatigue is better stratified in pSS, it appears that it is usually moderate or severe, rather than mild. Furthermore, it is unrelated to FM. Overall, fatigue appears as a consequence of pSS itself. Of note, severe fatigue was related in this study with the most important complication influencing patient survival in pSS, i.e., lymphoma. Further studies are needed to disclose the pathogenetic events leading to fatigue in pSS, and investigation of lymphoma in pSS might be also helpful to this end. Reference [1] Ng WF, et al. Transcriptional Signature of Fatigue Derived from Patients with Primary Sjögren’s Syndrome. PLoS One2015Dec 22;10(12):e0143970. Disclosure of Interest None declared

THU0236 The TNF Alpha -308 and the IL-6 -174 Promoter Polymorphisms Associate with Effective Anti-TNF Alpha Treatment in Seronegative Spondyloarthritis

Background One of the most important outcomes of treatment in SpA is the retention rate of therapy in the long term. Many genetic studies in chronic polyarthritis reported a correlation between response to anti-TNF therapy and several single nucleotide polymorphisms (SNPs), but the usefulness of this finding when related to the clinical outcome is ill defined. Objectives The primary endpoint of this study was to establish a possible correlation between several genetic polymorphisms, including ones never investigated before in SpA, and the retention rate of the first TNFa blocking agent, by comparing patients needing to switch the first anti-TNFa agent (Sw) to patients not needing to switch (NSw). To distinguish these two groups, a follow-up of at least one year was required. Methods 187 consecutive SpA patients (124 males and 63 females; mean age 52±30 years) were studied. All the patients had been followed up for at least 12 months after the introduction of the first anti-TNFα agent. The mean disease duration was 30±28 years (range 2-58 years). Seventy-four (39.6%) patients were diagnosed as affected by psoriatic arthritis (PsA), 66 (35.3%) by ankylosing spondylitis (AS) and 47 (25.1%) by undifferentiated spondyloarthritis (uSpA). Six SNPs were analysed: TNFα -308A>G (rs1800629), TNFR2 196M>R, IL-6 -174G>C (rs1800795), IL-6Ra, FCGR3A 158V>F (rs396991) and TGF-beta 869T>C (rs 19822073). The chi-squared or Fishers exact test and Mann–Whitneys U-test were used. Results The TNFα and the IL-6 promoter polymorphisms were significantly associated with the NSw phenotype in SpA. In particular, the TNFα -308 AA/AG genotypes were found in 24.4% NSw vs 10.9% Sw patients (p=0.03) and the GG homozygosis of the IL-6 promoter polymorphism in 49.6% NSw vs 34.4% Sw patients (p=0.047). By multivariate analysis, the TNFα -308A allele as well as the presence of the IL-6 -174GG homozygosis resulted as independent biomarkers predicting survival of the first anti-TNFα therapy in SpA (p=0.007, OR 4.4, 95%CI 1.5-13.1, and p=0.035, OR 2.1, 95%CI 1.1-4.4, respectively). Also the male gender (p=0.001, OR 3.4, 95%CI 1.6-7.1) resulted an independent variable associated with the NSw phenotype. The other SNPs, located in the genes encoding TNFR2, IL-6Ra, TGFb and FCGR3A, did not show any significant association. No association was found between the BASDAI or DAS28 response at month +6 (for axial or peripheral involvement, respectively) and all the investigated polymorphisms. The exposure time to the anti-TNFα therapies was significantly higher in NSw in comparison with Sw (NSw: 58±25.9 months vs Sw: 30.8±20.2 months, p<0.0001); thus, the NSw group was not biased by a shorter follow-up under anti-TNFα therapy. Conclusions The TNFα -308 and the IL-6 -174 promoter polymorphisms appear significantly associated with a higher efficacy of anti-TNFa therapy in SpA, expressed by the retention rate of the first anti-TNFα agent. This confirms the importance of the genetic background to optimize the management of the biologic agents in SpA. Disclosure of Interest None declared