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Objective: Systemic lupus erythematosus (SLE) is associated to boosted atherosclerosis development and a higher cardiovascular disease risk. This study aimed to delineate the role of anti-double stranded DNA (anti-dsDNA) antibodies on the molecular profile and the activity of immune and vascular cells, as well as on their enhanced cardiovascular risk. Approach and Results: Eighty SLE patients were included. Extensive clinical/analytical evaluation was performed, including cardiovascular disease parameters (endothelial function, proatherogenic dyslipidemia, and carotid intima-media thickness). Gene and protein expression profiles were evaluated in monocytes from patients diagnosed positive or negative for anti-dsDNA antibodies by using NanoString and cytokine arrays, respectively. NETosis and circulating inflammatory profile was assessed in both neutrophils and plasma. Positivity and persistence of anti-dsDNA antibodies in SLE patients were associated to endothelial dysfunction, proatherogenic dyslipidemia, and accelerated atherosclerosis. In parallel, anti-dsDNA antibodies were linked to the aberrant activation of innate immune cells, so that anti-dsDNA(+) SLE monocytes showed distinctive gene and protein expression/activity profiles, and neutrophils were more prone to suffer NETosis in comparison with anti-dsDNA(−) patients. Anti-dsDNA(+) patients further displayed altered levels of numerous circulating mediators related to inflammation, NETosis, and cardiovascular risk. In vitro, Ig-dsDNA promoted NETosis on neutrophils, apoptosis on monocytes, modulated the expression of inflammation and thrombosis-related molecules, and induced endothelial activation, at least partially, by FcR (Fc receptor)-binding mechanisms. Conclusions: Anti-dsDNA antibodies increase the cardiovascular risk of SLE patients by altering key molecular processes that drive a distinctive and coordinated immune and vascular activation, representing a potential tool in the management of this comorbidity.

To characterize the splicing machinery (SM) of leukocytes from primary antiphospholipid syndrome (APS), systemic lupus erythematosus (SLE) and antiphospholipid syndrome with lupus (APS + SLE) patients, and to assess its clinical involvement.Monocytes, lymphocytes and neutrophils from 80 patients (22 APS, 35 SLE and 23 APS + SLE) and 50 HD were purified, and 45 selected SM components were evaluated by qPCR-microfluidic array. Relationship with clinical features and underlying regulatory mechanisms were assessed.APS, SLE and APS + SLE leukocytes displayed significant and specific alterations in SM-components (SMC), associated with clinical features [autoimmune profiles, disease activity, lupus nephritis (LN), and CV-risk markers]. A remarkable relationship among dysregulated SMC in monocytes and the presence of LN in SLE was highlighted, revealing a novel pathological mechanism, which was further explored. Immunohistology analysis of renal biopsies highlighted the pathological role of the myeloid compartment in LN. Transcriptomic analysis of monocytes from SLE-LN(+) vs SLE-LN(-) identified 271 genes differentially expressed, mainly involved in inflammation and IFN-signaling. Levels of IFN-related genes correlated with those of SMC in SLE-LN(+). These results were validated in two external SLE-LN(+) datasets of whole-blood and kidney biopsies. In vitro, SLE-LN(+)-serum promoted a concomitant dysregulation of both, the IFN signature and several SMC, further reversed by JAKinibs treatment. Interestingly, IFNs, key inflammatory cytokines in SLE pathology, also altered SMC. Lastly, the over/down-expression of selected SMC in SLE-monocytes reduced the release of inflammatory cytokines and their adhesion capacity.Overall, we have identified, for the first time, a specific alteration of SMC in leukocytes from APS, SLE and APS + SLE patients that would be responsible for the development of distinctive clinical profiles.

Background: This prospective multicenter study developed an integrative clinical and molecular longitudinal study in Rheumatoid Arthritis (RA) patients to explore changes in serologic parameters following anti-TNF therapy (TNF inhibitors, TNFi) and built on machine-learning algorithms aimed at the prediction of TNFi response, based on clinical and molecular profiles of RA patients.Methods: A total of 104 RA patients from two independent cohorts undergoing TNFi and 29 healthy donors (HD) were enrolled for the discovery and validation of prediction biomarkers. Serum samples were obtained at baseline and 6 months after treatment, and therapeutic efficacy was evaluated. Serum inflammatory profile, oxidative stress markers and NETosis-derived bioproducts were quantified and miRNomes were recognized by next-generation sequencing. Then, clinical and molecular changes induced by TNFi were delineated. Clinical and molecular signatures predictors of clinical response were assessed with supervised machine learning methods, using regularized logistic regressions.Results: Altered inflammatory, oxidative and NETosis-derived biomolecules were found in RA patients vs. HD, closely interconnected and associated with specific miRNA profiles. This altered molecular profile allowed the unsupervised division of three clusters of RA patients, showing distinctive clinical phenotypes, further linked to the TNFi effectiveness. Moreover, TNFi treatment reversed the molecular alterations in parallel to the clinical outcome. Machine-learning algorithms in the discovery cohort identified both, clinical and molecular signatures as potential predictors of response to TNFi treatment with high accuracy, which was further increased when both features were integrated in a mixed model (AUC: 0.91). These results were confirmed in the validation cohort.Conclusions: Our overall data suggest that: 1. RA patients undergoing anti-TNF-therapy conform distinctive clusters based on altered molecular profiles, which are directly linked to their clinical status at baseline. 2. Clinical effectiveness of anti-TNF therapy was divergent among these molecular clusters and associated with a specific modulation of the inflammatory response, the reestablishment of the altered oxidative status, the reduction of NETosis, and the reversion of related altered miRNAs. 3. The integrative analysis of the clinical and molecular profiles using machine learning allows the identification of novel signatures as potential predictors of therapeutic response to TNFi therapy.

ObjectivesTo characterise splicing machinery (SM) alterations in leucocytes of patients with rheumatoid arthritis (RA), and to assess its influence on their clinical profile and therapeutic response.MethodsLeucocyte subtypes from 129 patients with RA and 29 healthy donors (HD) were purified, and 45 selected SM elements (SME) were evaluated by quantitative PCR-array based on microfluidic technology (Fluidigm). Modulation by anti-tumour necrosis factor (TNF) therapy and underlying regulatory mechanisms were assessed.ResultsAn altered expression of several SME was found in RA leucocytes. Eight elements (SNRNP70, SNRNP200, U2AF2, RNU4ATAC, RBM3, RBM17, KHDRBS1 and SRSF10) were equally altered in all leucocytes subtypes. Logistic regressions revealed that this signature might: discriminate RA and HD, and anti-citrullinated protein antibodies (ACPAs) positivity; classify high-disease activity (disease activity score-28 (DAS28) >5.1); recognise radiological involvement; and identify patients showing atheroma plaques. Furthermore, this signature was altered in RA synovial fluid and ankle joints of K/BxN-arthritic mice. An available RNA-seq data set enabled to validate data and identified distinctive splicing events and splicing variants among patients with RA expressing high and low SME levels. 3 and 6 months anti-TNF therapy reversed their expression in parallel to the reduction of the inflammatory profile. In vitro, ACPAs modulated SME, at least partially, by Fc Receptor (FcR)-dependent mechanisms. Key inflammatory cytokines further altered SME. Lastly, induced SNRNP70-overexpression and KHDRBS1-overexpression reversed inflammation in lymphocytes, NETosis in neutrophils and adhesion in RA monocytes and influenced activity of RA synovial fibroblasts.ConclusionsOverall, we have characterised for the first time a signature comprising eight dysregulated SME in RA leucocytes from both peripheral blood and synovial fluid, linked to disease pathophysiology, modulated by ACPAs and reversed by anti-TNF therapy.

Objectives: This study employed genetic and functional analyses using OASIS meta-analysis of multiple existing GWAS and gene-expression datasets to identify novel SLE genes. Methods: Four hundred and ten genes were mapped using SNIPPER to 30 SLE GWAS loci and investigated for expression in three SLE GEO-datasets and the Cordoba GSE50395-dataset. Blood eQTL for significant SNPs in SLE loci and STRING for functional pathways of differentially expressed genes were used. Confirmatory qPCR on SLE monocytes was performed. The entire 12p11 locus was investigated for genetic association using two additional GWAS. Expression of 150 genes at this locus was assessed. Based on this significance, qPCRs for DNM1L and KRAS were performed. Results: Fifty genes were differentially expressed in at least two SLE GEO-datasets, with all probes directionally aligned. DDX11, an RNA helicase involved in genome stability, was downregulated in both GEO and Cordoba datasets. The most significant SNP, rs3741869 in OASIS locus 12p11.21, containing DDX11, was a cis-eQTL regulating DDX11 expression. DDX11 was found repressed. The entire 12p11 locus showed three association peaks. Gene expression in GEO datasets identified DNM1L and KRAS, besides DDX11. Confirmatory qPCR validated DNM1L as an SLE susceptibility gene. DDX11, DNM1L and KRAS interact with each other and multiple known SLE genes including STAT1/STAT4 and major components of IFN-dependent gene expression, and are responsible for signal transduction of cytokines, hormones, and growth-factors, deregulation of which is involved in SLE-development. Conclusion: A genomic convergence approach with OASIS analysis of multiple GWAS and expression datasets identified DDX11 and DNM1L as novel SLE-genes, the expression of which is altered in monocytes from SLE patients. This study lays the foundation for understanding the pathogenic involvement of DDX11 and DNM1L in SLE by identifying them using a systems-biology approach, while the 12p11 locus harboring these genes was previously missed by four independent GWAS.

Coenzyme Q10 (CoQ10) is a mitochondrial electron carrier and a powerful lipophilic antioxidant located in membranes and plasma lipoproteins. CoQ10 is endogenously synthesized and obtained from the diet, which has raised interest in its therapeutic potential against pathologies related to mitochondrial dysfunction and enhanced oxidative stress. Novel formulations of solubilized CoQ10 and the stabilization of reduced CoQ10 (ubiquinol) have improved its bioavailability and efficacy. Synthetic analogues with increased solubility, such as idebenone, or accumulated selectively in mitochondria, such as MitoQ, have also demonstrated promising properties. CoQ10 has shown beneficial effects in autoimmune diseases. Leukocytes from antiphospholipid syndrome (APS) patients exhibit an oxidative perturbation closely related to the prothrombotic status. In vivo ubiquinol supplementation in APS modulated the overexpression of inflammatory and thrombotic risk-markers. Mitochondrial abnormalities also contribute to immune dysregulation and organ damage in systemic lupus erythematosus (SLE). Idebenone and MitoQ improved clinical and immunological features of lupus-like disease in mice. Clinical trials and experimental models have further demonstrated a therapeutic role for CoQ10 in Rheumatoid Arthritis, multiple sclerosis and type 1 diabetes. This review summarizes the effects of CoQ10 and its analogs in modulating processes involved in autoimmune disorders, highlighting the potential of these therapeutic approaches for patients with immune-mediated diseases.

The aim of this study was to investigate the microRNA (miRNA) expression pattern in neutrophils from rheumatoid arthritis (RA) patients and its contribution to their pathogenic profile and to analyze the effect of specific autoantibodies or inflammatory components in the regulation of miRNA in RA neutrophils and its modulation by biological therapies. Neutrophils were isolated from paired peripheral blood (PB) and synovial fluid samples of 40 patients with RA and from PB of 40 healthy donors. A miRNA array was performed using nCounter technology. Neutrophils from healthy donors were treated in vitrowith antibodies to citrullinated protein antigens isolated from RA patients and tumor necrosis factor-a (TNF-a) or interleukin-6. A number of cytokines and chemokines were analyzed. In vitro treatments of RA-neutrophils with tocilizumab or infliximab were carried out. Transfections with pre-miRNA and DICER downregulation experiments were further performed. RA-neutrophils showed a global downregulation of miRNA and genes involved in their biogenesis, alongside with an upregulation of various potential mRNA targets related to migration and inflammation. Decreased levels of miRNA and DICER correlated with autoimmunity, inflammation and disease activity. Citrullinated protein antigens and TNF-a decreased the expression of numerous miRNA and their biogenesis-related genes, increasing their potential mRNA targets. Infliximab reversed those effects. Transfections with pre-miRNA-223, -126 and -148a specifically modulated genes regulating inflammation, survival and migration whereas DICER depletion influenced the inflammatory profile of neutrophils. Taken together RA-neutrophils exhibited a global low abundance of miRNA induced by autoantibodies and inflammatory markers, which potentially contributed to their pathogenic activation. miRNA biogenesis was significantly impaired in RAneutrophils and further associated with a greater downregulation of miRNA mainly related to migration and inflammation in synovial fluid neutrophils. Finally, anti-TNF-a and anti-interleukin-6 receptor treatments can modulate miRNA levels in the neutrophils, minimizing their inflammatory profile.

Background Juvenile Idiopathic Arthritis (JIA) is one of the most common chronic diseases of childhood that often persists into adulthood and can result in significant long-term morbidity. As a long lasting chronic inflammatory disease, concern has been raised regarding the risk of premature development of cardiovascular disease (CVD) in JIA. This study aims to determine whether adults with JIA in clinical remission display clinical and subclinical signs of CVD risk: inflammatory mediators, adipokines, endothelial dysfunction and oxidative stress markers. Methods This is a cross-sectional study including 25 patients diagnosed with JIA according to the International League of Associations for Rheumatology criteria (ILAR 2001) and 25 age- and sex-matched controls. Remission was determined by JADAS10 Results Mean duration of the disease was 13.47 ± 5.47 years. Mean age was 25.11 ± 7.21. Time in remission was 3.52 ± 3.33 years. Patients were in remission with no treatment (40%) and with treatments (60%). CVD risk factors and CIMT were similar in JIA patients and controls. However, cholesterol levels were significantly elevated in JIA patients. Levels of adipocytokines, oxidative stress and endothelial activation markers were elevated in serum and PBMCs from JIA patients. Serum of those JIA patients induced the activation of adipocytes, endothelial cells and healthy PBMCs. Conclusions JIA adult patients in remission have subclinical signs of inflammation and CVD risk, showed by an increase in the levels of inflammatory cytokines, endothelial activation and oxidative stress markers and adipokines, molecules closely involved in the alteration of the vascular system.

Funder: Europeo de Desarrollo Regional de la Unión Europea, Funder: Rheumatic Diseases Network, OBJECTIVE: To analyze the relationship between complement component 3 (C3) and the prevalence of cardiometabolic risk factors and disease activity in the rheumatic diseases having the highest rates of cardiovascular morbidity and mortality: rheumatoid arthritis (RA), psoriatic arthritis (PsA) and axial spondyloarthritis (axSpA). METHODS: This is a cross-sectional study including 200 RA, 80 PsA, 150 axSpA patients and 100 healthy donors. The prevalence of cardiometabolic risk factors [obesity, insulin resistance, type 2 diabetes mellitus, hyperlipidemia, apolipoprotein B/apolipoprotein A (apoB/apoA) and atherogenic risks and hypertension] was analyzed. Serum complement C3 levels, inflammatory markers and disease activity were evaluated. Cluster analysis was performed to identify different phenotypes. Receiver operating characteristic (ROC) curve analysis to assess the accuracy of complement C3 as biomarker of insulin resistance and disease activity was carried out. RESULTS: Levels of complement C3, significantly elevated in RA, axSpA and PsA patients, were associated with the prevalence of cardiometabolic risk factors. Hard clustering analysis identified two distinctive phenotypes of patients depending on the complement C3 levels and insulin sensitivity state. Patients from cluster 1, characterized by high levels of complement C3 displayed increased prevalence of cardiometabolic risk factors and high disease activity. ROC curve analysis showed that non-obesity related complement C3 levels allowed to identify insulin resistant patients. CONCLUSIONS: Complement C3 is associated with the concomitant increased prevalence of cardiometabolic risk factors in rheumatoid arthritis and spondyloarthritis. Thus, complement C3 should be considered a useful marker of insulin resistance and disease activity in these rheumatic disorders.

Background Systemic lupus erythematosus (SLE) is a complex autoimmune disorder with significant genetic underpinnings. Multiple genome-wide association studies (GWAS) have been conducted with identification of several SLE genes and loci. However, these only explain about 15% of SLE genetic risk indicating that additional loci of modest effect remain to be discovered. Association clustering methods such as OASIS are more apt than single-variant analysis for identifying modest genetic effects. Methods Previously, OASIS analysis of two dbGAP GWAS datasets (6077 subjects; 0.75 million single-nucleotide polymorphisms (SNPs)) identified 30 SLE associated loci. In the present study, 410 genes were mapped to these loci and investigated for gene expression in 3 SLE GEO datasets. A fourth expression dataset, GSE50395, from Cordoba, Spain was used for validation. Expression quantitative trait loci (eQTL) analysis was performed using Blood eQTL browser for significant SNPs in loci harboring differentially expressed genes. Protein network analysis was performed using STRING to determine functional pathways involved. Confirmatory qPCR on monocytes of 12 healthy donors and 12 SLE patients was performed for the significant gene. Results This genetic and functional analysis identified 187 genes in GEO datasets that were differentially expressed in SLE. Of these 55 genes were found to be differentially expressed in at least 2 GEO datasets with all probes directionally aligned. In the Cordoba dataset, 10 genes were found to have altered expression in SLE. DDX11 was significantly down regulated in both the GEO (P=3.60E-02) and the Cordoba (P=8.02E-03) datasets. The most significant SNP, rs3741869 (GWAS P=3.2E-05) in OASIS locus 19 (chromosome 12p11.21), containing the gene DDX11, was found to be a cis-eQTL regulating the expression of DDX11 (P=8.62E-05). DDX11 expression in SLE was confirmed to be down regulated using qPCR (P=0.001), ratifying DDX11 as a novel gene associated with SLE. DDX11 interacted with multiple genes identified using genomic convergence of OASIS loci and expression analysis, including STAT1/STAT4. This study identified several novel SLE candidate genes as well, such as NRF1, TTC9 and BARHL1 which need additional data for verification. Conclusion Genomic convergence approach with a locus-based test, OASIS, and expression studies on multiple datasets can help identify novel SLE genes. This study showed that DDX11, a RNA helicase thought to be involved in genome stability, is repressed in SLE and its expression is modulated by rs3741869.

Background:TNF inhibitors (TNFi) represent an extraordinary advance in the management of Rheumatoid Arthritis (RA). Despite their benefits, there is a percentage of patients (20–40%) that do not achieve clinical improvement. Therefore, it is necessary to search for new and easily accessible biomarkers predictive of therapeutic response that might guide precision medicine.Objectives:1. To explore changes in the molecular profile of RA patients following TNFi therapy in serum samples. 2. To search for new and reliable biomarkers predictive of TNFi response, based on clinical and molecular profiles of RA patients, by using machine learning algorithms.Methods:In a prospective multicenter study, 79 RA patients undergoing TNFi and 29 healthy donors (HD) were enrolled. Twenty-two RA patients were further included as a validation cohort. Serum samples were obtained before and after 6 months of treatment, and therapeutic efficacy was evaluated. Patients’ response was determined following EULAR response criteria. Serum inflammatory profile was analyzed by a multiplex immunoassay, along with oxidative and NETotic profiles, evaluated by commercial kits. A circulating miRNA array was also performed by next-generation sequencing. Clustering analysis was carried out to identify groups of patients with distinctive molecular signatures. Then, clinical and molecular changes induced by TNFi were delineated after 6 months of therapy. Finally, integrative clinical and molecular signatures as predictors of response were assessed at baseline by supervised machine learning methods, using regularized logistic regressions.Results:Inflammatory, oxidative stress and NETosis-derived biomolecules were found altered in RA patients versus HD, closely interconnected and associated with several deregulated miRNAs. This altered molecular profile at baseline allowed the unsupervised division of three clusters of RA patients with distinctive clinical phenotypes, further linked to TNFi effectiveness. Cluster 1 included RA patients with low levels of pro-inflammatory cytokines, associated with a medium-low disease activity score and good clinical response. Clusters 2-3 comprised patients with high levels of pro-inflammatory cytokines, associated with a high disease activity and a non-response rate of 30%.After 6 months of therapy the molecular profile found altered in RA patients was reversed in responder patients, who achieved a molecular phenotype similar to HDs. However, non-responder patients’ molecular profile remained significantly deregulated, including alterations in inflammatory mediators (IL-6, L-8, TNFα, VEGF, IL-1RA, IL-5, IL-15, GMCSF, GCSF, FGFb), oxidative stress markers (LPO) and NETosis-derived products (Elastase), along with specific miRNAs (miR-199a-5p). These molecular changes further correlated with changes in disease activity score. Machine-learning algorithms identified clinical (Creatinine, IgM, Vitamin D, Swollen Joints, C4, Disease Duration and Tryglicerides) and molecular (Nucleosomes, IL-10, miR-106a-5p, IL-13, IL-12p70, IL-15 and LPO) signatures as potential predictors of response to TNFi treatment with high accuracy. Furthermore, the integration of both features in a combined model increased the predictive value of these signatures (AUC: 0.91). These results were further confirmed in an independent validation cohort.Conclusion:1. RA patients display distinctive altered molecular profiles directly linked to their clinical status and associated with TNFi effectiveness. 2. Clinical response was associated with a specific modulation of the inflammatory profile, the reestablishment of the altered oxidative status, the reduction of NETosis and the reversion of related altered miRNAs. 3. The integrative analysis of the clinical and molecular profiles using machine learning allows the identification of novel signatures as potential predictors of therapeutic response to TNFi therapy.Disclosure of Interests:None declared

Background:The presence of cardiovascular disease in psoriatic arthritis (PsA) is of particular concern, as it is considered the leading cause of mortality in PsA. Thus, it is essential to recognize those appropriate therapies that could target this comorbidity, reducing the risk of cardiovascular disease and metabolic alterations.Objectives:To evaluate the efficacy of methotrexate (MTX) and apremilast as monotherapies or in combination, in the clinical manifestations of the disease and the reduction of cardiovascular risk factors in PsA.Methods:Prospective longitudinal study in 30 PsA patients diagnosed according to CASPAR criteria: 10 patients were treated with MTX (12 ± 2,58 mg/week), 10 patients with apremilast (60 mg/day) and 10 were treated with combined therapy for 6 months, recruited in the routine clinical practice at the Reina Sofia Hospital of Cordoba and University Hospital of Jaen, Spain. Clinical and analytical parameters were collected at baseline and after 6 months of treatment: lipid profile (cholesterol, HDL, LDL, TG, ApoA and ApoB), glucose and insulin, body surface area (BSA) affected by psoriasis, number of tender and swollen joints, DAS28, DAPSA, VAS, CRP and ESR.The presence of cardiometabolic risk factors such as metabolic syndrome (MetSyn) was evaluated according to National Cholesterol Education Program (NCEP) adult treatment panel III (ATP III) criteria, meeting 3 of the following characteristics: abdominal obesity (men (>102 cm); women (>88 cm), TG > 150 mg/dL, HDL (men ( 130/85 mmHg, glucose levels > 110 mg/dL). Insulin resistance (HOMA-IR > 2,5), body mass index (BMI), ApoB/ApoA ratio, atherogenic index (AI) and SCORE (age, gender, cholesterol, HDL, smoking habit and diabetes) were also studied.Results:Apremilast or MTX monotherapies caused a moderate reduction of the clinical inflammatory markers (CRP and ESR) and disease activity (VAS, DAPSA and DAS28) after 6 months of treatment. On the other hand, while apremilast significantly reduced the affected BSA, MTX had no significant effect. All those parameters were more significantly reduced after the combined treatment (MTX+ apremilast).Apremilast monotherapy significantly improved alterations in the lipid profile (reducing cholesterol and LDL levels, ApoB/ApoA ratio and AI), insulin resistance and decreased BMI, thus reducing the number of patients with MetSyn. MTX monotherapy treatment had no positive effect on these parameters. None of the treatments had significant effects on SCORE values.The beneficial effects of apremilast on the lipid profile were mitigated after the combination with MTX. Nevertheless, the number of patients with MetSyn decreased even more after the combined therapy of MTX with apremilast compared to apremilast monotherapy.Conclusion:1) In patients with moderate disease activity, treatment with apremilast monotherapy might have some advantages compared to the MTX monotherapy, since it can decrease the percentage of BSA with psoriasis, the lipid profile alteration, IR and weight, thus improving the cardiovascular risk profile. 2) Combined therapy (MTX+ apremilast) can induce a deeper reduction in the disease activity compared to the monotherapies, maintaining, in turn, the positive effects of apremilast on the cardiovascular risk.Funded by ISCIII (PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Iván Arias de la Rosa: None declared, Carmen Torres-Granados: None declared, Maria del Carmen Abalos-Aguilera: None declared, Gómez García Ignacio: None declared, Isabel Añón Oñate: None declared, María José Pérez Galán: None declared, Desiree Ruiz: None declared, Alejandra M. Patiño-Trives: None declared, María Luque-Tévar: None declared, Eduardo Collantes Estevez Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene, Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer., Alejandro Escudero Contreras Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene., María Dolores López Montilla Speakers bureau: Celgene

Rheumatoid arthritis (RA) is an autoimmune and chronic inflammatory disease primarily affecting the joints, and closely related to specific autoantibodies that mostly target modified self-epitopes. Relevant findings in the field of RA pathogenesis have been described. In particular, new insights come from studies on synovial fibroblasts and cells belonging to the innate and adaptive immune system, which documented the aberrant production of inflammatory mediators, oxidative stress and NETosis, along with relevant alterations of the genome and on the regulatory epigenetic mechanisms. In recent years, the advances in the understanding of RA pathogenesis by identifying key cells and cytokines allowed the development of new targeted disease-modifying antirheumatic drugs (DMARDs). These drugs considerably improved treatment outcomes for the majority of patients. Moreover, numerous studies demonstrated that the pharmacological therapy with biologic DMARDs (bDMARDs) promotes, in parallel to their clinical efficacy, significant improvement in all these altered molecular mechanisms. Thus, continuous updating of the knowledge of molecular processes associated with the pathogenesis of RA, and on the specific effects of bDMARDs in the correction of their dysregulation, are essential in the early and correct approach to the treatment of this complex autoimmune disorder. The present review details basic mechanisms related to the physiopathology of RA, along with the core mechanisms of response to bDMARDs.

Background: Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by excess morbidity and mortality from cardiovascular disease. In addition, metabolic alterations have been observed in these patients, which significantly contributes to the cardiovascular risk burden. Thus, the identification of therapies able to mitigate the cardiometabolic alterations in RA is essential. In addition, numerous studies suggest that cardiometabolic risks are mediated through adipocytokines. However, this relationship is not completely defined in RA. Objectives: 1) To evaluate the relationship among cardiometabolic risk factors and the levels of adipocytokines and autoantibodies in RA patients. 2) To analyze the effects anti-TNFα and anti-IL6 therapies on the cardiometabolic alterations. Methods: 1.- A cross-sectional study including 100 RA patients and 50 age-matched healthy donors was carried out. Different parameters related to the cardiometabolic risk were analyzed, including: lipid profile, atherogenic index, ratio ApoB/ApoA, insulin resistance (IR), obesity, hypertension, and the SCORE. Levels of adipocytokines (TNFα, IL6, IL1β, visfatin, adiponectin, leptin and resistin) were evaluated in serum. Carotid intima media thickness (CIMT) was evaluated as atherosclerosis marker. 2.-A prospective study in 15 RA patients before and after 3 months of anti-TNFα therapy and 15 RA patients before and after 3 months of treatment with tocilizumab (TCZ) was performed. All the parameters evaluated in the cross-sectional cohort were tested in this prospective study. Results: RA patients had elevated levels of leptin/adiponectin ratio, visfatin, resistin and inflammatory markers in serum. Our cohort of RA patients displayed increased rates of cardiometabolic risk factors, such as insulin resistance, hypertension, SCORE, pathologic CIMT, atherogenic index and ratio apoB/apoA. The alteration in the levels of adipocytokines were closely related to the autoimmunity, disease activity and clinical inflammatory markers. Of note, visfatin and C3 complement levels were determinant for IR, high levels of SCORE, increased parameters of CVD risk defined by apoB/apoA ratio and pathologic CIMT. Both biological therapies reduced clinical inflammatory markers and disease activity after 3 months of treatment. Anti-TNFα therapy modulated the cytoadipokine profile, reducing serum levels of IL-6, IL-1β, resistin and visfatin, decreasing IR. After treatment with tocilizumab, serum levels of C3 complement, IL-1β and resistin were reduced. Conclusion: 1) Altered adipocytokine profile is closely related to the increased cardiometabolic risk factors associated with RA, where autoimmunity and systemic inflammation play a key role. 2) Anti-TNFα and anti-IL6R therapies, administered for 3 months, could have beneficial effects in the reduction of cardiometabolic risk factors in RA. Acknowledgement: Funded by ISCIII (PI18/00837, CP15/00158, PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER Disclosure of Interests: None declared

Objectives 1) To compare the real-world effectiveness among RA patients starting 3 biologic DMARDs (bDMARDs). 2) To evaluate whether patients’ clinical characteristics influence the response to specific bDMARDs. 3) To analyze the impact of bDMARDs on inflammatory profiles. Methods Eighty RA patients initiating TNFi, 22 patients treated with TCZ and 28 treated with RTX, the last ones after switching from TNFi and given as second bDMARDs, were included in the study. Effectiveness was assessed before and after 3 and 6 months of treatment by the European League Against Rheumatism (EULAR) criteria. Adjusted differences were estimated with multivariable linear regression. Differences in treatment outcomes were calculated as a function of patients’ characteristics, through regression modelling. Extensive analysis of the inflammatory profile using a multiplex were also performed. Results A high proportion of patients starting TNFi showed early EULAR Good Response (65,8% responders at 3 months). After 6 months of therapy this percentage even increased (70,6%), so that tender joints’ counts, swollen joints count, SDAI and HAQ improved significantly. After switching from a first TNFi, RTX treatment promoted better early response (81,8% responders after 3 months) than TCZ (67% responders). Differences remained significant after adjusting for potential confounders. Yet, after 6 months, efficacy was similar among both non-TNFi DMARDs, even improving the effectiveness in TCZ-treated RA patients based on EULAR response rate (DAS28 change: 5.09±1.4 to 2.7±1,17) vs RTX-treated patients (DAS28 change: 4.8±1.3 to 3-1±1.3). Patients older, and those suffering arterial hypertension (AH) or hyperlipidemia showed less effectiveness for both, TNFi and non-TNFi DMARDs, most remarkable for RTX and after 6 months of treatment. Analysis of the inflammatory profile, by evaluating 27 key biomarkers of inflammation, showed, in patients’ responders to therapy, specific profiles depending on the biologic DMARD administered. Thus, after 6 months of RTX, a generalized reduction in the expression levels of 22/27 proteins evaluated was observed, while TCZ mainly inhibited proteins involved in cell maturation, proliferation and leukocytes recruitment (IL6, GCSF, IP-10, IFNγ, RANTES). On the other hand, TNFi mainly reduced molecules related to leukocytes’ recruitment (IL-2, IL-8, MIP1β, RANTES). Conclusion 1) Treatment outcomes among RA patients treated with non-TNFi bDMARDs in standard clinical practice, are in line with a superior early effectiveness of RTX compared with TCZ. However, effectiveness in TCZ-treated RA patients increases after 6 months of treatment vs RTX. 2) Patients characteristics influence the response to bDMARDs, suggesting the need of evaluating the benefit risk/ratio based on a personalized clinical evaluation. 3) Anti-inflammatory benefits associated with bDMARDs treatments can differ despite similar drug efficacy against RA, indicating that drug-specific mechanisms are involved in regulating these processes. Acknowledgement Funded by PI-0285-2017, ISCIII, PI15/00837 and RIER RD16/0012/0015, co-funded with FEDER Disclosure of Interests Chary Lopez-Pedrera: None declared, Nuria Barbarroja Puerto: None declared, Carlos Perez-Sanchez: None declared, Alejandro Ibanez-Costa: None declared, Maria Luque-Tevar: None declared, Alejandra Patino-Trives: None declared, Ivan Arias de la Rosa: None declared, Maria del Carmen Abalos-Aguilera: None declared, Rafaela Ortega Castro: None declared, Alejandro Escudero Contreras: None declared, Perez Sanchez Laura: None declared, Carlos Rodriguez-Escalera: None declared, Dolores Ruiz-Montesinos: None declared, Carmen Maria Romero-Barco: None declared, Natalia Mena-Vazquez: None declared, Julia Uceda Speakers bureau: abbie, pfizer, novartis, janmsen, Ma Dolores Toledo-Coello: None declared, M angeles Aguirre-Zamorano: None declared, Eduardo Collantes Estevez: None declared

Objectives: 1. To characterize the mRNAs and microRNAs transcriptomes of monocytes from systemic lupus erythematosus (SLE) patients and their association with the pathophysiology of the disease. 2. To evaluate the role of anti-dsDNA antibodies in the regulation of these processes. 3. To investigate the molecular mechanisms involved in the efficacy of Fluvastatin in preventing the atherothrombotic risk. Methods: Monocytes from peripheral blood of 81 SLE patients and 40 healthy donors (HD) were purified by negative immunomagnetic selection. Then, gene expression microarray (Agilent G4112F platform) and nCounter microRNA expression arrays (Nanostring) were performed. Functional categorization of altered genes and miRNAs was made using IPA software, and interaction networks were identified. Genes and miRNAs integrating the networks were validated in the whole cohorts by RT-PCR. Predicted miRNA-mRNA interactions were tested by microRNA over-expression or inhibition experiments. Serum and cellular inflammatory and oxidative profiles were evaluated by multiplex assay, PCR and specific commercial kits, respectively; phosphorylation status of intracellular proteins was analyzed by PathScan array. To evaluate the clinical significance of the parameters analyzed, correlation and association studies were performed. Mechanistic studies were developed to typify the specific effects of the anti-dsDNA antibodies on monocytes. Besides, the beneficial effects of ex vivo Fluvastatin treatment on the monocyte molecular profiles were assessed. Results: Microarray identified 553 altered genes in SLE monocytes. Relevant biofunctions and disorders on which these genes were involved included inflammatory, immunological, cardiovascular, neurological, renal and reproductive disease. Analysis of miRNA profiles showed altered expression of 35 miRNAs in SLE monocytes. Sixty-one genes were inversely correlated and predicted as CVD-related target genes of 26 differentially expressed miRNAs. Transfection studies confirmed the relationship between specific miRNAs and their identified target genes. Association of these genes and miRNAs with the anti-dsDNA positivity, early atherosclerosis and nephropathy, along with correlations with disease activity (SLEDAI), activation of some intracellular signaling proteins, and levels of serum inflammatory and oxidative markers were demonstrated. In vitro studies demonstrated the specific modulation of several genes/miRNAs by anti-dsDNA, along with the increase of prothrombotic and proinflammatory mediators, the induction of apoptosis and the phosphorylation of intracellular proteins participating in renal and CVD-related signaling pathways. Besides, treatment of HD-monocytes with SLE patients’ serum after Fluvastatin supplementation prevented the proinflammatory altered gene/miRNA profiles induced by serum from those patients before treatment. Conclusion: 1. Gene and microRNA expression profiles allowed the identification of relevant genes and pathways altered in monocytes of SLE patients, associated with the pathogenesis of the disease, and modulated by anti-dsDNA. 2. Specific microRNA-miRNA regulatory networks control the biological processes and factors related to the CV pathology in SLE, which are prevented by Fluvastatin treatment. Acknowledgement: Funded by ISCIII, PI15/01333, PI18/00837 and RIER RD16/0012/0015 co-funded with FEDER. Disclosure of Interests: None declared

Background The Splicing, a process of removing introns of RNA to produce mature messenger RNA, is exerted by an intricate macromolecular complex, the spliceosome. The alternative splicing, key mechanism that provides transcription and protein diversity, is dysregulated in various genetic, metabolic and inflammatory diseases. Splicing variants could contribute to the development and severity of pathologies of crucial importance. Objectives The aim of this study was the identification and characterization of alterations in the splicing machinery of leukocyte subsets of Rheumatoid Arthritis (RA) patients, as well as their influence on the activity of the disease and its atherothrombotic profile. Methods Monocytes, lymphocytes and neutrophils from 74 RA patients and 29 healthy donors (HD) were purified, and 45 elements of the splicing machinery were evaluated using a microfluidic qPCR array (Fluidigm). In parallel, the clinical status of the patients and the carotid intima-media thickness (CIMT) were assessed, along with an extensive serological evaluation, including autoantibodies, oxidative stress markers, inflammatory molecules and prothrombotic mediators. Correlation and association studies and logistic models among those clinical and analytical parameters were performed. Finally, leukocytes from HD were incubated with purified ACPAs-IgG from RA patients and changes promoted in both, splicing machinery and leukocyte inflammatory profile, were assessed. Results A significantly altered expression of spliceosome components was found in all the leukocyte subsets: 21, 17 and 27 components were differentially expressed in monocytes, lymphocytes and neutrophils, respectively, in RA patient’s vs HD. The levels of those altered components were associated to ACPA positivity, high disease activity (DAS28>5.1), early atherosclerosis and radiological involvement. Correlations with inflammatory mediators, oxidative stress markers and NETosis were further identified. Interestingly, eight components, belonging to the major spliceosome (SNRNP70, SNRNP200 and U2AF2), minor spliceosome (RNU4ATAC), and splicing factors (RBM3, RBM17, KHDRS1 and SRSF10) were found simultaneously altered in the three leukocyte subtypes. Logistic regression models, built with levels of simultaneously altered spliceosome components, generated several signatures related to RA disease features that were able to: i) discriminate between RA patients and HD; ii) classify patients with high disease activity (DAS28>5.1); iii) recognize patients with radiological involvement; and iv) identify patients showing atheroma plaques. In vitro studies showed that the purified ACPAs of RA patients modulated the expression of these components. Conclusion 1) The splicing machinery is profoundly altered in leukocytes from RA patients and closely related to the activity of the disease, including inflammation, autoimmunity and articular involvement, as well as with the occurrence of early atherosclerosis. 2) Evaluation of the common altered components of the spliceosome in leukocytes subsets might be used to typify the disease, avoiding complementary ultrasound or radiological tests. Acknowledgement MINECO (FJCI-2016-30825) and ISCIII (PI18/00837 and RIER RD16/0012/0015) co-funded with FEDER. Disclosure of Interests None declared

Background:To date, novel mechanisms such as the involvement of splicing machinery components in lupus nephropathy and its interplay with the transcriptome in innate immune cells have not been evaluated.Objectives:1- To identify altered transcriptomic signatures associated with the immune response of monocytes from SLE patients and its association with clinical features. 2- To evaluate the role of the spliceosome linked to the transcriptomic profile of SLE monocytes. 3- To analyze mechanistically the impact of anti-dsDNA antibodies (Ab) and the modulation of the spliceosome in the SLE monocytes activity.Methods:Sixty SLE patients and forty healthy donors (HD) were included in the study. Infiltration rate of myeloid cells and its association with clinical features were analyzed in kidney biopsies by Immunohistochemistry. In parallel, circulating monocytes were purified from peripheral blood by immune-magnetic selection. The expression of a set of 770 genes related to autoimmune/inflammatory diseases was evaluated using NanoString Technologies. The levels of the main 45 components of the splicing machinery were further analyzed in these samples using a microfluidic qPCR array (Fluidigm). An extensive clinical/serological evaluation was also performed, comprising disease activity, renal involvement parameters, autoAb profile, and the systemic inflammatory status (27-plex Assay). Finally, in vitro studies involving anti-dsDNA-IgG Ab treatment and over/down-expression of splicing machinery components were carried out to analyze their effects in the monocyte activity.Results:Infiltration of CD68 expressing cells was confirmed in kidney biopsies and associated with parameters of kidney failure (C3/C4, chronic index), highlighting the key role of the myeloid compartment in lupus nephropathy. Gene expression profiling recognized 156 genes differentially expressed in SLE monocytes compared with HDs, including 87 genes up-regulated and 69 down-regulated. Functional analysis showed that most dysregulated genes were associated with the IFN response (i.e. IFIT1, IFI44, IFI44L, RSAD2). In parallel, the altered expression of 27 spliceosome components was demonstrated in SLE monocytes compared with HD, including 3 up-regulated and 24 down-regulated. Correlation studies demonstrated that the aberrant expression of splicing machinery components was linked to the altered interferon signature and the plasma inflammatory profile. This aberrant profile at molecular level was associated with the disease activity status, anti-dsDNA positivity and C3/C4 levels. Interestingly, SLE patients with renal disease displayed a simultaneous alteration of both, the IFN and the spliceosome signatures in monocytes, along with an enlarged pro-inflammatory profile in plasma. Logistic regression models that integrated the concomitant alteration of some splicing machinery components and IFNs genes identified lupus nephritis patients with high accuracy. Mechanistic studies showed that in vitro treatment of monocytes from HDs with anti-dsDNA promoted a concomitant deregulation of the IFN signature and the expression of several spliceosome components (i.e. PTB, RBM17, RNU6ATAC). Finally, the over/down-expression of selected spliceosome components (PTB and RBM17) in monocytes from SLE patients reduced the active release of inflammatory cytokines and their adhesion capacity.Conclusion:1) Monocytes from SLE patients with renal involvement exhibit a remarkable alteration of genes associated with the IFN response, further linked with the aberrant expression of several splicing machinery components. 2) Anti-dsDNA promoted the dysregulation in monocytes of both the IFN and spliceosome signatures, along with an active release of pro-inflammatory mediators. 3) The modulation of key splicing components in monocytes from SLE patients reduce their pro-inflammatory status and migration capacity. Ongoing studies may provide novel biomarkers and therapeutic tools to treat lupus nephropathy.Acknowledgements:Funded by ISCIII, PI18/00837 and RIER RD16/0012/0015 co-funded with FEDERDisclosure of Interests:None declared

Background:To date, although multiple molecular approaches have illustrated the various aspects of Primary Antiphospholipid Syndrome (APS), systemic lupus erythematosus (SLE) and antiphospholipid syndrome plus lupus (APS plus SLE), no study has so far fully characterized the potential role of posttranscriptional regulatory mechanisms such as the alternative splicing.Objectives:To identify shared and differential changes in the splicing machinery of immune cells from APS, SLE and APS plus SLE patients, and their involvement in the activity and clinical profile of these autoimmune disorders.Methods:Monocytes, lymphocytes and neutrophils from 80 patients (22 APS, 35 SLE and 23 APS plus SLE) and 50 healthy donors (HD) were purified by immunomagnetic selection. Then, selected elements of the splicing machinery were evaluated using a microfluidic qPCR array (Fluidigm). In parallel, extensive clinical/serological evaluation was performed, comprising disease activity, thrombosis and renal involvement, along with autoantibodies, acute phase reactants, complement and inflammatory molecules. Molecular clustering analyses and correlation/association studies were developed.Results:Patients with primary APS, SLE and APS plus SLE displayed significant and specific alterations in the splicing machinery components in comparison with HD, that were further specific for each leukocyte subset. Besides, these alterations were associated with distinctive clinical features.Hence, in APS, clustering analysis allowed to identify two sets of patients representing different molecular profile groups with respect to the expression levels of splicing machinery components. Principal component analyses confirmed a clear separation between patients. Clinically, cluster 1 characterized patients with higher thrombotic episodes and recurrences than cluster 2 and displayed a higher adjusted global APS score (aGAPSS). Accordingly, these patients showed higher levels of inflammatory mediators than cluster 2.Similarly, in patients with APS plus SLE, clustering analysis allowed to identify two sets of patients showing differential expression of splicing machinery components. Clinical and laboratory profiles showed that cluster 2 characterized patients that had suffered more thrombotic recurrences, most of them displaying an aGAPSS over 12 points and expressing higher levels of inflammatory mediators than cluster 1. The incidence of lupus nephropathy was similarly represented in both clusters.Lastly, in SLE patients, molecular clustering analysis identified two sets of patients showing distinctive clinical features. One cluster characterized most of the patients positive for anti-dsDNA antibodies, further suffering lupus nephropathy, and a high proportion of them also presenting atheroma plaques and high levels of inflammatory mediators.Correlation studies further demonstrated that several deranged splicing machinery components in immune cells (i.e. SF3B1tv1, PTBP1, PRP8 and RBM17) were linked to the autoimmune profile of the three autoimmune diseases, albeit in a specific way on each disorder. Accordingly, in vitro treatment of HD lymphocytes with aPL-IgG or anti-dsDNA-IgG changed the expression of spliceosome components also found altered in vivo in the three autoimmune diseases. Finally, the induced over/downregulated expression of selected spliceosome components in leukocytes modulated the expression of inflammatory cytokines, changed the procoagulant/adhesion activities of monocytes and regulated NETosis in neutrophils.Conclusion:1) The splicing machinery, profoundly altered in leukocytes from APS, APS plus SLE and SLE patients, is closely related to the activity of these diseases, their autoimmune and inflammatory profiles. 2) The analysis of the splicing machinery allows the segregation of APS, APS plus SLE and SLE, with specific components explaining the CV risk and renal involvement in these highly related autoimmune disorders.Acknowledgements:Funded by ISCIII, PI18/00837 and RIER RD16/0012/0015 co-funded with FEDERDisclosure of Interests:None declared

Background:NAD+ is an important cofactor and second messenger for multiple cellular processes that exhibits antioxidant, anti-apoptotic and anti-inflammatory properties. Pre-clinical studies in animal models of Rheumatoid Arthritis (RA) have demonstrated the therapeutic potential of NAD+ boosters in the control of the disease activity. However, to date no studies have been set up to evaluate the NAD+ metabolism and the therapeutic effects of NAD+ boosters in RA patients.Objectives:1- To study the NAD+ metabolism in RA patients and its association with key clinical features. 2- To evaluate the effect of anti-TNF therapy in the NAD+ metabolism.3- To analyze the beneficial effects of NAD+ boosters in leukocytes from active RA patients.Methods:Plasma and PBMCs were purified from 100 RA patients and 50 healthy donors (HDs). Moreover, an additional cohort of 50 RA patients treated with Anti-TNF therapy was analyzed before and after 6 months of treatment. NAD+ levels were determined by using the NAD/NADH-Glo Assay. NAD+-consuming genes expression were analyzed by RT-PCR. In parallel, PBMCs from eight HDs and eight active RA patients were treated ex vivo with 1 mM of NAD+ boosters including nicotinamide (NAM), nicotinamide riboside (NR), and nicotinamide mononucleotide (NMN). After 24 hours, intracellular reactive oxygen species (ROS) levels (DFCHDA) and the percentage of apoptotic PBMCs (annnexin V/PI) were assessed by flow cytometry. Lastly, a panel of key pro-inflammatory genes were evaluated by RT-PCR.Results:NAD+ and NADH levels were significantly reduced in plasma and PBMCs of RA patients compared with HDs and directly related to disease activity (DAS28, CDAI, SDAI). Accordingly, the expression levels of genes involved in the consumption of NAD+ such as SIRT-1, CD38 and PARP-1 were found up-regulated in PBMCs from RA patients. Anti-TNF therapy for 6 months restored the altered NAD+ levels towards those showed by HDs. Furthermore, the clinical response promoted by Anti-TNF therapy (changes in DAS28) correlated with changes in NAD+ levels. The in vitro treatments of PBMCs isolated from active RA patients with NAD+ boosters significantly increased the NAD+ levels and promoted a deep reduction of intracellular ROS levels, the percentage of apoptotic cells and the expression levels of key inflammatory mediators, such as IL-6, IL-8, IL-1ß, TNF-α, CCL2, IL-23, and STAT-3.Conclusion:1. NAD+ metabolism is altered and associated with the disease activity of RA patients, involving both, reduced NAD+ levels and increased expression of NAD+-consuming genes. 2. Anti-TNF therapy restored NAD+ levels, which were directly linked to the clinical effectiveness. 3. NAD+ boosters reduced the oxidative, apoptotic and inflammatory profiles of RA leukocytes through the parallel increase of intracellular NAD+ levels. Thus, NAD+ boosters might be considered novel therapeutic tools for RA patients.Acknowledgements:Supported by PI18/00837, RIER RD16/0012/0015, RTI2018-100695-B-I00, P18-RT-4264 and CVI276, co-funded with FEDER.Disclosure of Interests:None declared

Background:Psoriatic Arthritis (PsA) displays increased traditional cardiovascular (CV) risk factors, such as insulin resistance (IR), metabolic syndrome or obesity. Thus, it is an urgent need to treat and manage these cardiometabolic comorbidities associated. Some evidence points out that methotrexate could improve CV risk due to its anti-inflammatory properties, however the effect of PDE4 inhibitor treatment on CV risk have not been elucidated yet.Objectives:1) To evaluate the effect of conventional therapy and PDE4 inhibitor in PsA patients with high prevalence of cardiometabolic comorbidities. 2) To identify a molecular patient profile susceptible of being benefit from each therapy regarding disease activity and CV risk.Methods:Thirty biological-naïve patients with PsA were treated with the PDE4 inhibitor (Apremilast), Methotrexate or combined therapy (Apremilast and Methotrexate) following the clinical routine practice for 6 months. A cohort of 30 age and sex-matched healthy donors (HDs) was included. Plasma and peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood of patients and HDs. Different parameters related to the cardiometabolic risk were analyzed, including: atherogenic index, ratio apolipoprotein B (apo B)/apolipoprotein A (apo A), insulin resistance (IR), metabolic syndrome, obesity, arterial hypertension, and the SCORE. Clinical and analytical parameters were collected: lipid profile (total cholesterol, HDL-cholesterol, LDL-cholesterol, triglycerides, apo A and apo B), glucose and insulin, body surface area (BSA) affected by psoriasis, number of tender and swollen joints, DAPSA, C-reactive protein and erythrocyte sedimentation rate. A panel of 92 proteins involved in cardiovascular disease (cardiovascular panel II, Olink) and an adipocytokine profile was measured in plasma and PBMCs. Hard cluster analysis was carried out in order to identify two distinctive molecular phenotypes depending on the treatment response related to the reduction of CV risk.Results:Among the 92 CV-related proteins, the higher levels of two molecules, CD163 and FABP4 observed in PsA patients compared to HDs, were strongly associated with elevated rates of CV risk factors such as apolipoprotein B/A and atherogenic risks, metabolic syndrome, obesity, IR, arterial hypertension and smoking. Thus, we could identify two clinical profiles of patients according to the plasma levels of these molecules: cluster 1 defined by 20 patients with low levels of CD163 and FABP4 and low prevalence of CV comorbidities and cluster 2 defined by 10 patients with high levels of CD163 and FABP4 and high prevalence of CVD comorbidities. Regarding cluster 2, those patients that were treated with Apremilast or combined therapy had a significant reduction of CD163 and FABP4 associated with a drop in total cholesterol, apo B, IR state and body mass index. In addition, both PDE4 inhibitor and combined treatment reduced activity disease. However, Methotrexate in monotherapy did not show a beneficial effect in PsA patients displaying higher levels of CD163, FABP4, total cholesterol and no changes in disease activity after treatment. In regard to cluster 1, the three therapy strategies reduced disease activity after 6 months. Even with low rates of comorbidities, those patients treated with Apremilast had reduced levels of total cholesterol and apolipoprotein B and body mass index after 6 months of therapy. However, no changes were observed in the treatment with Methotrexate or Methotrexate combined with Apremilast.Conclusion:1- CD163 and FABP4 could be considered as potential biomarkers of treatment efficacy regarding cardiometabolic comorbidities. 2- Apremilast might target metabolic alterations in PsA modulating lipid profile, insulin resistance and body mass index, decreasing the levels of surrogate CV-related molecules. 3- Apremilast treatment should be considered in PsA patients with higher rates of cardiometabolic comorbidities.Funded by ISCIII (PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:None declared.

Background:Psoriatic arthritis (PsA) is a chronic inflammatory disease associated with an increased prevalence of cardiovascular (CV) events. Traditional CV risk factors do not account for the increased CV disease mortality in PsA. Inflammation seems to have a key role in the development of this comorbidity, however the specific molecular mechanisms involved are not defined yet.Objectives:To evaluate clinical CV risk factors and surrogate markers and their relationship with inflammation, disease activity and metabolic comorbidities in PsA patientsMethods:This is a cross-sectional study including 100 PsA patients without CV disease recruited in the routine clinical practice at the Rheumatology Department, Reina Sofia Hospital of Cordoba and 100 age-matched healthy donors (HDs). Different parameters related to the cardiometabolic risk were analyzed. Clinical and analytical parameters were collected: lipid profile (cholesterol, HDL, LDL, TG, ApoA and ApoB), glucose and insulin, body surface area (BSA) affected by psoriasis, number of tender and swollen joints, DAPSA, VAS, CRP and ESR. To measure the persistence of inflammation, CRP levels were recorded retrospectively once, twice, or three times during the 5 years prior to study and at the moment of the study. Increased levels of CRP in at least 50% of the determinations was considered as persistent inflammation. Plasma and peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood of patients and HDs. A panel of 92 proteins involved in CV disease and an adipocytokine profile was measured in plasma and PBMCs. In addition, activation of 18 intracellular pathways involved in cell activation was also measured in PBMCs. In vitro experiments in adipocytes treated with serum from PsA patients were also carried out.Results:Traditional CV risk factors including atherogenic risk, insulin resistance (IR), metabolic syndrome, smoking, obesity, arterial hypertension, apolipoprotein B/A risk, type 2 diabetes mellitus and the levels of SCORE were significantly increased in PsA patients. The presence of IR was associated with disease activity markers (DAPSA, ESR and CRP). In fact, the HOMA-IR index was related to the CRP persistence. PsA patients with obesity had significantly increased the number of tender and swollen joints, the levels of DAPSA and CRP. Twenty-eight proteins involved in CV disease and six adipocytokines were significantly elevated in the plasma of PsA patients. Several of these cardiovascular molecules were associated with higher levels of DAPSA (CTSD, GAL3, CD163, FABP4, IL6 and IL1RT2), acute phase reactants (GAL3, TNFα, Adiponectin, TNFR1 and IL6), affected body surface area (IL2RA, GAL3, CCL15, TRAP, CSTB, CD163, OPG and CNTN1) and onychopaty (TRAP, VWF, MCP-1, GAL3, LTBR, TFPI, CHI3L1, CTSZ and JAM-A). In addition, the mRNA expression of most of those 28 CV molecules were significantly increased in PBMCs from PsA patients. At intracellular level, the activation of 11 kinases (ERK1/2, AKT, S6 Ribosomal, mTOR, HSP27, Bad, p70 S6 kinase, PRAS40, p53 and caspase-3) involved in insulin signaling, inflammation, cell survival and apoptosis were altered in PBMCs. Finally, serum from PsA patients was able to modify the expression of these molecules in adipocytes.Conclusion:1) Disease activity and inflammatory burden are closely associated with the presence of metabolic alterations, specifically obesity and IR in patients with PsA. 2) The development of IR is extremely related to the persistence of CRP levels in the previous 5 years. 3) Inflammation is closely associated to the adipose tissue dysfunction in PsA and 4) FABP4, CD163 and GAL3 are surrogate CV markers commonly associated with clinical features of PsA, suggesting the role of these molecules linking CVD and PsA pathogenesis.Funded by ISCIII (PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:None declared.

Background:Objectives:1. This study, developed within the Innovative Medicines Initiative Joint Undertaking project PRECISESADS framework, aimed to identify specific molecular profiles involved in the enhanced CV-risk present in SLE patients and to analyze the relevance of the sustained positivity for anti-dsDNA on the establishment of their atherothrombotic status.Methods:One hundred and twenty-four SLE consecutive patients (not including patients with associated antiphospholipid syndrome), belonging to the PRECISESADS project, were evaluated for the presence of CVD and its association with positivity for anti-dsDNA antibodies. A second cohort of 62 SLE patients was included, of which endothelial dysfunction, lipid profile, the presence of atheroma plaques (identified by a pathologic increase in the carotid intimae media thickness -CIMT-), and the frequencies of anti-dsDNA positivity for 7 years, were evaluated. Serum inflammatory and oxidative stress biomolecules, and NETosis-derived bioproducts were further evaluated by multiplex assay and specific commercial kits, respectively. Besides, miRNnomes were identified using next-generation sequencing. Clinical significance of the biomolecules analyzed was explored by correlation/association studies with immunological and CV-risk features.Results:A significant relationship among the incidence of CVD (i.e. thrombosis or cardiac involvement) and the positivity for anti-dsDNA antibodies was recognized in the first SLE cohort. Accordingly, in the second SLE cohort, significantly impaired micro-vascular endothelial function (identified by reduction of hyperemia post-occlusion area), increased atherogenic index and pathologic increase in the CIMT were assessed in patients positive for anti-dsDNA in relation to anti-dsDNA negative patients. Around a 65% of SLE patients displayed a sustained positivity for anti-dsDNA antibodies for more than 7 years. These patients showed a distinctive and specific molecular profile compared with patients that had remained negative for anti-dsDNA, including increased inflammatory profile (IL1B, IL2, IL6, IL17, EOTAXIN, FGF, GMCSF, IFNγ, IP10, RANTES, TNF), enhanced oxidative status (lipoperoxides), and higher NETosis (nucleosomes, elastase). Levels of those biomolecules were closely interconnected and associated to their regulatory miRNAs, which accordingly exhibited differential expression in SLE anti-dsDNA(+)vsanti-dsDNA(-) patients. Finally, the frequency for positivity of anti-dsDNA significantly correlated both with markers of endothelial dysfunction and with the presence of atheroma plaques in SLE patients, pointing at the direct involvement of anti-dsDNA-Abs in the development of these processes.Conclusion:1. Positivity for anti-dsDNA antibodies confers a specific molecular profile linked to an enhanced CV-risk in SLE patients. 2. Moreover, the sustained positivity for anti-dsDNA antibodies fosters the establishment of an atherothrombotic status in these autoimmune patients.Acknowledgments:Supported by the EU/EFPIA –IMI-JU PRECISESADS (n° 115565) and ISCIII (PI18/0837 and RIER RD16/0012/0015), Co-funded with FEDER.Disclosure of Interests:Alejandra M. Patiño-Trives: None declared, Maria A Aguirre: None declared, Carlos Pérez Sánchez: None declared, Pérez Sánchez Laura: None declared, María Luque-Tévar: None declared, Iván Arias de la Rosa: None declared, Rafaela Ortega Castro: None declared, Maria del Carmen Abalos-Aguilera: None declared, Mario Espinosa: None declared, Pedro Seguí Azpilcueta: None declared, Jacques-Olivier Pers: None declared, Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Marta Alarcon-Riquelme: None declared, Eduardo Collantes Estevez Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene, Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer.

Background:Objectives:1. This study, developed within the Innovative Medicines Initiative Joint Undertaking project PRECISESADS framework, aimed at identify specific inflammatory and oxidative stress determinants involved in the enhanced CV-risk present in SLE patients and to analyze the relevance of the sustained positivity for anti-dsDNA on the establishment of their atherothrombotic status.Methods:One hundred and twenty-four SLE consecutive patients (not including patients with associated antiphospholipid syndrome), belonging to the PRECISESADS project, were evaluated for the presence of CVD and its association with positivity for anti-dsDNA antibodies. A second cohort of 62 SLE patients was included, of which endothelial dysfunction, lipid profile, the presence of atheroma plaques (identified by a pathologic increase in the carotid intimae media thickness -CIMT-), and the frequencies of anti-dsDNA positivity for 7 years, were evaluated. Serum inflammatory and oxidative stress biomolecules, and NETosis-derived bioproducts were further evaluated by multiplex assay and specific commercial kits, respectively. Besides, miRNnomes were identified using next-generation sequencing. Clinical significance of the biomolecules analyzed was explored by correlation/association studies with immunological and CV-risk features.Results:A significant relationship among the incidence of CVD (i.e. thrombosis or cardiac involvement) and the positivity for anti-dsDNA antibodies was recognized in the first SLE cohort. Accordingly, in the second SLE cohort, significantly impaired micro-vascular endothelial function (identified by reduction of hyperemia post-occlusion area), increased atherogenic index and pathologic increase in the CIMT were assessed in patients positive for anti-dsDNA in relation to anti-dsDNA negative patients. Around a 65% of SLE patients displayed a sustained positivity for anti-dsDNA antibodies for more than 7 years. These patients showed a distinctive and specific molecular profile compared with patients that had remained negative for anti-dsDNA, including increased inflammatory profile (IL1B, IL2, IL6, IL17, EOTAXIN, FGF, GMCSF, IFNγ, IP10, RANTES, TNF), enhanced oxidative status (lipoperoxides), and higher NETosis (nucleosomes, elastase). Levels of those biomolecules were closely interconnected and associated to their regulatory miRNAs, which accordingly exhibited differential expression in SLE anti-dsDNA(+)vsanti-dsDNA(-) patients. Finally, the frequency for positivity of anti-dsDNA significantly correlated both with markers of endothelial dysfunction and with the presence of atheroma plaques in SLE patients, pointing at the direct involvement of anti-dsDNA-Abs in the development of these processes.Conclusion:1. Positivity for anti-dsDNA antibodies confers a specific inflammatory/oxidative profile linked to an enhanced CV-risk in SLE patients. 2. Moreover, the sustained positivity for anti-dsDNA antibodies fosters the establishment of an atherothrombotic status in these autoimmune patients.Acknowledgments:Supported by the EU/EFPIA –IMI-JU PRECISESADS (n° 115565) and ISCIII (PI18/0837 and RIER RD16/0012/0015), Co-funded with FEDER.Disclosure of Interests:Inmaculada Concepcion Aranda-Valera: None declared, Alejandra M. Patiño-Trives: None declared, Roldán Molina Rosa: None declared, Maria A Aguirre: None declared, Pérez Sánchez Laura: None declared, Carlos Pérez Sánchez: None declared, María Luque-Tévar: None declared, Iván Arias de la Rosa: None declared, Maria del Carmen Abalos-Aguilera: None declared, Desiree Ruíz-Vilchez: None declared, Mario Espinosa: None declared, Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Eduardo Collantes-Estévez Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer.

Background:Cardiovascular risk factors are increased in Psoriatic Arthritis (PsA). In fact, around 60% out of PsA patients display insulin resistance (IR), a hallmark of metabolic syndrome, which might significantly contribute to the cardiovascular disease. Latest studies suggested that inflammatory and metabolic disorders may be under epigenetic control, including DNA methylation. DNA methylation is an unexplored area in the field of PsA.Objectives:To study the alterations in the genome-wide DNA methylation profile of CD4+T cells from PsA patients and its relationship with its pathology and the risk of cardiovascular comorbidity.Methods:Twenty healthy controls (HC) and 20 PsA patients were included in the study. PsA patients were classified into insulin resistant and non-insulin resistant according to HOMA-IR index. CD4+T lymphocytes were isolated from peripheral blood by positive immunomagnetic selection. The Illumina Infinium MethylationEPIC Beadchip was used to obtain DNA methylation profiles across approximately 850,000 CpGs (TSS1500, TSS200, 5UTR, 3UTR, first exon, gene body). Beta values (β) estimating methylation levels were obtained at each CpG site, and differentially methylated genes (DMG) between PsA and HC were identified. Functional classification of these genes was carried out through gene ontology analysis (PANTHER database). Gene expression analysis of the selected genes was also evaluated by RT-PCR. Vascular parameters including carotid intima-media thickness (cIMT) and endothelial function was analyzed by ecodoppler and periflux respectively.Results:The genome-wide methylation analysis identified 112 DMGs including 41 hypomethylated and 71 hypermethylated. These differentially methylated genes were enriched with several signaling pathways and disease categories including immune response, metabolic processes, oxidative stress, vascular and inflammatory pathways. The altered gene expression of selected genes with altered methylation levels in PsA was also validated. Correlation and association analysis of these DMGs with clinical and analytical variables, cardiovascular risk factors and endothelial microvascular function revealed that the degree of methylation of these genes was significantly associated with cIMT (IGF1R, NDRG3, SMYD3, HLA-DRB1, WDR70), arterial pressure (METT5D1, NRDG3, ADAM17, SMYD3, WNK1, CBX1), insulin resistance (AKAP13, SEMA6D, PLCB1), altered lipid profile and atherogenic index (MYBL1, METT5D1, MAN2A1, SLC1A7, SEMA6D, PLCB1, TLK1, SDK1, CBX1), inflammation (MYBL1, NDUFA5, METT5D1, SEMA6D, PLCB1, TLK1), and endothelial dysfunction (ADAMST10, GPCPD1, CCDC88A). In addition, this analysis also identified 435 DMGs including 280 hypomethylated and 155 hypermethylated in CD4+T cells from IR-PsA vs non IR-PsA patients. Between these two groups of PsA patients, CHUK, SERINC1, RUNX1, TTYH2, TXNDC11, FAF1, BICD1, SCD5, PDE5A, FAS, NFIA and GRP75 displayed the most significantly altered methylation, suggesting the role of these genes in the metabolic complications associated with PsA.Conclusion:These findings help our understanding of the pathogenesis of PsA and advance epigenetic studies in regards to this disease and the cardiometabolic comorbidities associated. Funded by ISCIII (PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:Iván Arias de la Rosa: None declared, María Dolores López Montilla Speakers bureau: Celgene, Javier Rodríguez: None declared, Esteban Ballester: None declared, Carmen Torres-Granados: None declared, Carlos Perez-Sanchez: None declared, Maria del Carmen Abalos-Aguilera: None declared, Gómez García Ignacio: None declared, Desiree Ruiz: None declared, Alejandra M. Patiño-Trives: None declared, María Luque-Tévar: None declared, Eduardo Collantes-Estévez Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer., Alejandro Escudero Contreras Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene.

Background:Recent studies underlined the crucial role of DNA methylation in several autoimmune diseases by altering gene expression profiles, thus influencing disease severity. Yet, aberrant methylation patterns in monocytes, key players in the pathogenesis of APS patients, has not been evaluated.Objectives:To analyze the genome-wide DNA methylation profile of monocytes from APS patients and its relationship with the cardiovascular (CV) pathology. 2. To evaluate the role of antiphospholipid antibodies (aPL) in the regulation of this process.Methods:Thirty-three APS patients and 15 healthy donors (HD) were included in the study. Monocytes were isolated from peripheral blood by positive immunomagnetic selection. The Illumina Infinium Methylation EPIC Beadchip was used to obtain DNA methylation profiles across approximately 850,000 CpGs (TSS1500, TSS200, 5UTR, 3UTR, first exon, intergenic, gene body). Beta values (β) estimating methylation levels were obtained at each CpG site, and differentially methylated genes (DMG) between APS and HD were identified. Functional classification of that genes was carried out by gene ontology analysis (PANTHER database). Gene expression of selected DMG genes was evaluated by RT-PCR. CV-risk parameters, including carotid intima-media thickness (CIMT) and microvascular endothelial function were further assessed, and correlation/association studies were developed with clinical and analytical variables. The effects of aPLs were also evaluated byin vitrostudies.Results:Genome-wide DNA methylation analysis identified 813 DMG, including 279 hypomethylated and 534 hypermethylated. Functional classification of these genes revealed signatures associated with biological processes and pathways related to their clinical profile, including immune response, adhesion, oxidative stress and vascular signaling. Correlation and association studies showed that the methylation levels of genes related to immune response were associated with the CV-risk score, aGAPSS (CCR2, TXLNB, GLIPR), type of thrombosis (SIGLEC11, COLEC11, LRRC16A, AHSA1, TRIL) and aPL titers (CLEC4G, RGS4, HLA-DPA1, GBP6, RAET1E, HLA-G, HLA-DPA1, HLA-H, TXLNB). Besides, methylation levels of DMG related to vascular signaling and adhesion processes were associated with the presence of thrombotic recurrences (VEGFA, MAPK14, ITGA8, EPCAM, PCDHA6, DLG1) as well as with traditional CV-risk factor such as hypertension and dyslipidemia (ITGA11, DSCAM, CLEC4F, CDH4, LTBP2, PCDHB14). In addition, methylation levels of DMG genes related to oxidative stress (GP2, PGD, ADH1) were associated with microvascular endothelial dysfunction. An altered mRNA expression of some of those genes with aberrant methylation and related to increased CV-risk and thrombotic recurrences in APS was also identified. Both, abnormal methylation and transcription levels of several genes were further associated with a pathological increase of the CIMT. Finally, in vitro studies supported the role of aPLs as key players in the altered methylation and transcriptomic profiles of APS patients.Conclusion:APS patients showed an impaired methylation profile in monocytes of genes associated with clinical features of the disease, including aPL titers, CV risk, thrombotic recurrences, endothelial dysfunction and early atherosclerosis. These results offered a map to the monocytes methylome and shed light on the pathophysiology of APS, paving the way for the development of new, more effective biomarkers and therapeutics.Acknowledgments:Funded by ISCIII (PI18/0837 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:Carlos Perez-Sanchez: None declared, Alejandra M. Patiño-Trives: None declared, Maria A Aguirre: None declared, Pérez Sánchez Laura: None declared, María Luque-Tévar: None declared, Iván Arias de la Rosa: None declared, Carmen Torres-Granados: None declared, Maria del Carmen Abalos-Aguilera: None declared, Pedro Seguí Azpilcueta: None declared, Javier Rodríguez: None declared, Esteban Ballester: None declared, Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Eduardo Collantes Estevez Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene, Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer.

Background:Objectives:The aim of this study was to evaluate whether alterations in the splicing machinery of immune cells from Systemic Lupus Erythematosus (SLE) patients could influence the development and activity of the disease and the kidney involvement.Methods:The study was conducted in 41 SLE patients and 34 healthy donors (HD). Monocytes, lymphocytes and neutrophils were purified by immunomagnetic selection. Then, selected elements of the splicing machinery and a set of genes related to inflammation, renal and cardiovascular disease (including interleukins, adipocytokines, chemokines, and oxidative stress markers, among others) were evaluated using a microfluidic qPCR array (Fluidigm). Besides, the inflammatory profile in plasma, including the analysis of 27 proteins, was evaluated by using the Bioplex assay. In parallel, an extensive clinical/serological evaluation was performed; comprising disease activity and cardiovascular and renal involvement along with autoantibodies, complement factors, and acute phase reactants. Correlation and association studies and logistic models among those clinical and analytical parameters were developed. Mechanistic in vitro studies were carried out by incubation of HD-leukocytes with anti-dsDNA-IgG purified from SLE patients and changes promoted in both, splicing machinery and leukocyte inflammatory profile, were assessed.Results:A significantly altered expression of spliceosome components was found in all the leukocyte subsets: 27, 12 and 11 components were differentially expressed in monocytes, lymphocytes and neutrophils, respectively, in SLE patient’s vs HD. In parallel, a number of genes codifying for proteins involved in inflammation, fatty acid metabolism, oxidative stress and migration were found altered and correlations with spliceosome components were further identified. Besides, those statistical analyses demonstrated multiple links among altered spliceosome components and the clinical profile of these patients, such as the activity of the disease (SLEDAI), the occurrence of obstetric complications and the presence of arterial hypertension. Logistic regression and ROC curve analyses identified a signature composed in each leukocyte subset by 3 altered spliceosome components that could differentiate between SLE and HDs. Remarkably, the levels of a high number of those altered components were associated to the presence of lupus nephritis (LN). Moreover, ROC curve analyses allowed to identify several cell-specific spliceosome components as potential biomarkers of renal disease. In patients with LN we could also identify a distinctive inflammatory profile in plasma in relation to patients without renal involvement, which further correlated with the altered expression of a number of spliceosome components in each leukocyte subset. Lastly, the in vitro treatment of HD leukocytes with anti-dsDNA promoted the alteration of several spliceosome components found also altered in vivo in SLE lymphocytes, monocytes and neutrophils.Conclusion:1)The splicing machinery is greatly altered in leukocytes from SLE patients, regulated, at least partially, by anti-dsDNA antibodies and closely related to the activity of the disease, including obstetrical complications, hypertension and lupus nephritis. 2)Specific components of the spliceosome in SLE leukocytes subsets might be used as potential biomarkers to typify the disease, particularly kidney involvement.Acknowledgments :Funded by ISCIII (PI18/00837 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests: :Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer., María Luque-Tévar: None declared, Alejandro Ibañez-Costa: None declared, Alejandra M. Patiño-Trives: None declared, Maria A Aguirre: None declared, Pérez Sánchez Laura: None declared, Maria del Carmen Abalos-Aguilera: None declared, Iván Arias de la Rosa: None declared, Pedro Seguí Azpilcueta: None declared, Mario Espinosa: None declared, Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Eduardo Collantes-Estévez Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Justo Pastor Castaño Fuentes: None declared, Raúl Miguel Luque Huertas: None declared, Carlos Perez-Sanchez: None declared

Objectives:To evaluate the changes promoted in levels of circulating inflammatory mediators in RA patients in response to TNF-α inhibitors (TNFi) and anti-CD20 therapies, in order to identify biomarkers of clinical efficacy and potential predictors of therapeutic response to these drugs.Methods:In a prospective RA cohort multicenter study, we collected serum from RA patients with moderate or high disease activity prior and after 6 months of treatment with TNFi or rituximab (RTX), and analyzed levels of 27 proteins that constitute a multi-biomarker test of the inflammatory profile of these samples, using a multiplex immunoassay. Patients’ response was determined according to the EULAR response criteria (good/moderate/no). We compared basal levels of inflammatory molecules between the differential response patient groups and analyzed their discriminative ability. Logistic prediction models were created to assess the added value of potential inflammatory predictors.Results:Among 111 total RA patients, 50 of 85 (59%) patients in the TNFi group and 18 of 26 patients in the RTX group (69%) responded to the biologic treatment. High DAS28 or SDAI scores, or titers of auto-antibodies (RF or ACPA) at baseline were not predictive of response to any treatment. Instead, smoking habit and hyperlipidemia at baseline were predictors of a worse response to any of these bDMARDs.Of the molecules analyzed by the multiplex assay, 14 inflammatory mediators showed a significant downregulation on patients’ responders to TNFi therapy. Moreover, the decline on 7 biomolecules was related to reduced DAS28. After RTX treatment, 15 inflammatory mediators were reduced in patients with good clinical response; downregulation in 4 of those biomolecules correlated with reduced DAS28.In the search for predictors of response to each bDMARD, by using the MetaboAnalyst software, we could classify patients with distinctive therapeutic response based on the baseline levels of the inflammatory molecules analyzed. Receiver operating characteristic (ROC) analyses for those multiple biomarkers allowed us to further identify specific signatures of inflammatory biomolecules that may serve as predictors of response to each bDMARD therapy with high sensitivity and specificity. Thus, a signature of five molecules was identified as potential predictor of TNFi response [Vascular endothelial growth factor (VEGF), Eotaxin, RANTES, IL7 and IL-17]. Indeed, a signature including three highly expressed cytokines/chemokines in RA serum were identified as predictors of RTX response [interferon-inducible protein 10 (IP10), Eotaxin and monocyte chemotactic protein 1 (MCP-1)].Conclusion:The extensive analysis of serum inflammatory profile allowed to identify specific and distinctive signatures of biomolecules that, in coordination with known clinical and serological profiles, might predict the response of RA patients to TNFi or RTX treatments.Acknowledgments :Funded by Junta de Andalucía (PI-0285-2017), ISCIII, (PI18/00837 and RIER RD16/0012/0015) co-funded with FEDERDisclosure of Interests:María Luque-Tévar: None declared, Carlos Perez-Sanchez: None declared, Font Ugalde Pilar: None declared, Montserrat Romero-Gómez: None declared, Alejandra M. Patiño-Trives: None declared, Desiree Ruiz: None declared, Iván Arias de la Rosa: None declared, Maria del Carmen Abalos-Aguilera: None declared, Rafaela Ortega Castro: None declared, Alejandro Escudero Contreras: None declared, Carlos Rodríguez-Escalera Speakers bureau: Lilly, GSK, Novartis and Sanofi, José Javier Pérez Venegas: None declared, María Dolores Ruiz Montesinos: None declared, Carmen Dominguez: None declared, Carmen Romero Barco: None declared, Antonio Fernandez-Nebro: None declared, Natalia Mena-Vázquez: None declared, Jose Luis Marenco Speakers bureau: ABbvie, Pfzer, lilly, Julia Uceda: None declared, Mª Dolores Toledo-Coello: None declared, Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene., Maria A Aguirre: None declared, Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer., Eduardo Collantes-Estévez Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE, Lilly, Bristol and Celgene.

Rheumatoid Arthritis (RA), Systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS) are the systemic autoimmune diseases (SADs) most associated with an increased risk of developing cardiovascular (CV) events. Cardiovascular disease (CVD) in SADs results from a complex interaction between traditional CV-risk factors, immune deregulation and disease activity. Oxidative stress, dyslipidemia, endothelial dysfunction, inflammatory/prothrombotic mediators (cytokines/chemokines, adipokines, proteases, adhesion-receptors, NETosis-derived-products, and intracellular-signaling molecules) have been implicated in these vascular pathologies. Genetic and genomic analyses further allowed the identification of signatures explaining the pro-atherothrombotic profiles in RA, SLE and APS. However, gene modulation has left significant gaps in our understanding of CV co-morbidities in SADs. MicroRNAs (miRNAs) are emerging as key post-transcriptional regulators of a suite of signaling pathways and pathophysiological effects. Abnormalities in high number of miRNA and their associated functions have been described in several SADs, suggesting their involvement in the development of atherosclerosis and thrombosis in the setting of RA, SLE and APS. This review focusses on recent insights into the potential role of miRNAs both, as clinical biomarkers of atherosclerosis and thrombosis in SADs, and as therapeutic targets in the regulation of the most influential processes that govern those disorders, highlighting the potential diagnostic and therapeutic properties of miRNAs in the management of CVD.