B. Maurer

OP0034 The Serotonin Receptor 2 Inhibitor Terguride Has Beneficial Effects on Skin Fibrosis: Results from A Phase 2 Proof of Concept Study

Background Circumstantial evidence from preclinical studies indicates a key role of serotonin (5-HT) signaling via the 5-HT-2B receptor in the development of fibrosis. Terguride is an orally available 5-HT-2 receptor inhibitor that has shown beneficial effects in different animal models of systemic sclerosis (SSc). Objectives To evaluate the efficacy of Terguride for the treatment of fibrosis in patients with SSc in an investigator initiated phase 2 proof of concept study. Methods Main inclusion criteria were fulfillment of ACR classification criteria and diffuse cutaneous SSc (dcSSc). Patients with end-stage organ involvement and treatment with potentially disease modifying agents including immunosuppressives were excluded. Patients were treated with Terguride at up to 3 mg/d p.o. or standard of care (post hoc control) for three months. Primary efficacy endpoints were changes of pre-defined skin biopsy biomarkers over the three months treatment period. Secondary efficacy endpoints included change of mRSS and lung function parameters. Serious adverse events (SAEs) and AEs were coded using MedDRA. The study was externally monitored. Results Twelve patients were recruited into the Terguride group and 6 patients into the control group. The primary endpoints, skin biopsy biomarkers, showed a consistent and statistically significant down-regulation compared to the control group (Figure) for dermal thickness, myofibroblast counts and mRNA levels of col1a1, col1a2 as well as for the Lafyatis 4-gene biomarker set (COMP, THSP-1, SIGLEC-1, IFI-44). This was accompanied by a reduction in mRSS of - 32.3% versus baseline in the Terguride group versus stable values in the control group (p<0.05). Lung function parameters did not change significantly. Overall, 33 AEs (n=27 mild and n=6 moderate) and one SAE (pyelonephritis, not related) occurred in the Terguride group, most often consisting of nausea and vomiting (9% and 13% of patients respectively).Twelve patients were recruited into the Terguride group and 6 patients into the control group. The primary endpoints, skin biopsy biomarkers, showed a consistent and statistically significant down-regulation compared to the control group (Figure) for dermal thickness, myofibroblast counts and mRNA levels of col1a1, col1a2 as well as for the Lafyatis 4-gene biomarker set (COMP, THSP-1, SIGLEC-1, IFI-44). This was accompanied by a reduction in mRSS of - 32.3% versus baseline in the Terguride group versus stable values in the control group (p<0.05). Lung function parameters did not change significantly. Overall, 33 AEs (n=27 mild and n=6 moderate) and one SAE (pyelonephritis, not related) occurred in the Terguride group, most often consisting of nausea and vomiting (9% and 13% of patients respectively). Conclusions Terguride was well tolerated in patients with dcSSc. Strong and consistent effects on fibrosis related skin biopsy biomarkers could be observed in this open-label controlled phase 2 proof of concept study, which was further supported by a significant improvement of the mRSS over the control group. These data justify further investigation in an upcoming randomized placebo controlled phase 3 study. Disclosure of Interest O. Distler Grant/research support from: Bayer, Sanofi, Ergonex, Boehringer Ingelheim, Actelion, Pfizer, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, EpiPharm, Ergonex, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, MedImmune, Pharmacyclics, Pfizer, Serodapharm, Sinoxa, B. Maurer: None declared, S. Vettori: None declared, S. Blumhardt: None declared, D. Frey: None declared, A. Distler: None declared, C. Beyer: None declared, J. H. Distler Shareholder of: 4 D Science, Consultant for: Actelion, BMS, Celgene, Bayer Pharma, Boehringer Ingelheim, JB Therapeutics, Sanofi-Aventis, Novartis, UCB, GSK, Array Biopharma and Active Biotech

Manual muscle testing and hand-held dynamometry in people with inflammatory myopathy: An intra- and interrater reliability and validity study

Manual muscle testing (MMT) and hand-held dynamometry (HHD) are commonly used in people with inflammatory myopathy (IM), but their clinimetric properties have not yet been sufficiently studied. To evaluate the reliability and validity of MMT and HHD, maximum isometric strength was measured in eight muscle groups across three measurement events. To evaluate reliability of HHD, intra-class correlation coefficients (ICC), the standard error of measurements (SEM) and smallest detectable changes (SDC) were calculated. To measure reliability of MMT linear Cohen`s Kappa was computed for single muscle groups and ICC for total score. Additionally, correlations between MMT8 and HHD were evaluated with Spearman Correlation Coefficients. Fifty people with myositis (56±14 years, 76% female) were included in the study. Intra-and interrater reliability of HHD yielded excellent ICCs (0.75–0.97) for all muscle groups, except for interrater reliability of ankle extension (0.61). The corresponding SEMs% ranged from 8 to 28% and the SDCs% from 23 to 65%. MMT8 total score revealed excellent intra-and interrater reliability (ICC>0.9). Intrarater reliability of single muscle groups was substantial for shoulder and hip abduction, elbow and neck flexion, and hip extension (0.64–0.69); moderate for wrist (0.53) and knee extension (0.49) and fair for ankle extension (0.35). Interrater reliability was moderate for neck flexion (0.54) and hip abduction (0.44); fair for shoulder abduction, elbow flexion, wrist and ankle extension (0.20–0.33); and slight for knee extension (0.08). Correlations between the two tests were low for wrist, knee, ankle, and hip extension; moderate for elbow flexion, neck flexion and hip abduction; and good for shoulder abduction. In conclusion, the MMT8 total score is a reliable assessment to consider general muscle weakness in people with myositis but not for single muscle groups. In contrast, our results confirm that HHD can be recommended to evaluate strength of single muscle groups.

OP0088 The role and function of monocyte-derived fibroblast-like cells in multi-organ fibrosis in systemic sclerosis

Background Animal studies indicated bone marrow-derived cells as a source of pathological myofibroblasts in multiple organ fibrosis such as lungs, heart, skin and kidney. Monocytes are implicated in the pathogenesis of systemic sclerosis (SSc), however detailed role of specific monocyte subsets in multi-organ fibrogenesis in SSc remains unclear. Objectives We aimed to determine the role and the contribution of circulating monocytes in the onset and progression of multi-organ fibrosis in SSc. Methods Endomyocardial biopsies (n=10) from SSc patients and healthy controls were screened by immunohistochemistry. CD14+ monocytes isolated from peripheral blood of SSc patients and healthy donors were differentiated towards a myofibroblast phenotype by stimulation with TGF-β1, IL-4, IL-10 and IL-13. In addition, CD14+ monocytes were co-cultured in 2D and 3D models with dermal fibroblasts originating from SSc patients or healthy subjects, or with adult cardiac fibroblasts. TGF-β signalling was blocked by SD208 and A83–01 inhibitors. Profibrotic gene expression and protein secretion were evaluated by qPCR, Western blot, protein array, immunofluorescence and ELISA. Results The myocardia of SSc patients revealed the presence of CD45-expressing infiltrates, an extended collagen I deposition and the presence of CD14-expressing elongated cells in the fibrotic tissue. Stimulated monocytes acquired a myofibroblast-like phenotype with increased expression of collagen I (p<0.0001), fibronectin (p<0.05), and α smooth muscle actin (α-SMA) in comparison to untreated cells. Similarly, CD14+ monocytes exposed to dermal or cardiac fibroblasts acquired spindle shape and expressed higher levels of profibrotic genes. The process of monocyte-to-myofibroblast differentiation employed TGF-β/SMAD signalling. Blocking of the TGF-β receptor I and canonical SMAD-dependent pathway with inhibitors resulted in the abrogation of collagen I secretion by monocytes (p=0.002). CD14+ monocytes from SSc patients were characterised by higher secretion of CXCL10 (p<0.001), which was significantly decreased after cytokine stimulation (p<0.001). Additionally, a tendency towards higher secretion of CCL20, CCL22, Leukemia Inhibitory Factor and neurotrophin-3 was observed for SSc monocytes. Conclusions Here we demonstrated the capability of peripheral blood monocytes to differentiate towards the functional myofibroblast phenotype, designating these cells as one of the potential sources of pathological tissue myofibroblasts in SSc. Additionally, these cells sustained pro-fibrotic cytokines secretion, highlighting their important regulatory functions in the fibrogenesis in SSc. Further studies of monocytes involvement in SSc might lead to novel treatment strategies. Disclosure of Interest M. Rudnik: None declared, M. Stellato: None declared, V. Milleret: None declared, P. Błyszczuk: None declared, B. Maurer Grant/research support from: AbbVie, Protagen, EMDO, Novartis. Congress support from Pfizer, Roche, Actelion. Patent licensed: mir-29 for the treatment of systemic sclerosis, K. Klingel: None declared, J. Henes: None declared, K. Sotlar: None declared, M. Ehrbar: None declared, O. Distler Grant/research support from: Actelion, Bayer, Boehringer Ingelheim, Pfizer, Sanofi. Patent licensed: mir-29 for the treatment of systemic sclerosis, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, ChemomAb, EpiPharm, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, Mepha, MedImmune, Mitsubishi Tanabe Pharma, Pharmacyclics, Pfizer, Sanofi, Serodapharm, Sinoxa, Speakers bureau: AbbVie, iQone Healthcare, Mepha, G. Kania Grant/research support from: Bayer

FRI0635-HPR The Myositis Activity Profile – First Results of Content and Construct Validity of The German Version

Background The Myositis Activity Profile (MAP), a questionnaire to assess limitations of activities of daily life in patients with inflammatory myopathy (IM), was developed and tested on psychometric properties in Sweden and later on translated and validated in English1,2. To the best of our knowledge, there is no validated version available for German speaking patients. Objectives To translate and cross-culturally adapt the MAP into German language and to determine content and construct validity. Methods A cross-cultural adaptation of the English version of the MAP into German was performed following international guidelines. To assess content validity of the new German version, patients rated difficulty and importance of items of the MAP using a visual analog scale (VAS range 0–10). For construct validity the correlation between the MAP and five functional tests (Short Physical Performance Battery, one leg stand, Grip Ability Test, upper and lower Progressive Isoinertial Lifting, and the 6 minute walk) were analyzed with the Spearman correlation coefficient. Additionally, for discriminative validity differences between patients and age and gender matched healthy controls were analyzed using the Mann-Whitney- U Test. Item fit within subscales was calculated by Cronbachs alpha. Results Twenty German speaking patients with a diagnosis of acute (20%), subacute (25%) or chronic (55%) myositis were included. Participants were 56.5 ±10.13 years old and 75% of them were female. The median combined difficulty and importance of the 32 items of the MAP was 5.67 (range 5.3–6.5). Correlations between the total score of the MAP and the Short Physical Performance Battery (-0.52), Grip Ability Test (0.52), Progressive Isoinertial Lifting Evaluation (lumbal part: -0.62, cervical part -0.60) and the 6 minute walk test (-0.69) were moderate and the correlation with the one leg stand test was low (-0.26). Healthy participants had significant lower scores (single items, subscales and total score) than patients (p≤0.05). Cronbachs alpha coefficients for the four subscales varied between 0.85 and 0.92. Conclusions This initial validation of the German version of the MAP showed that the 32 items from the English version were also suitable for Swiss conditions. Cronbachs alpha coefficients were similar to those from the English version. As expected the MAP correlated moderately with functional tests, except with the one leg stand. Although balance may be important in patients with IM, the MAP does not cover balance ability. In summary, the MAP seems to be a valid and easy applicable methodology for assessing limitations of activity in daily life. However, further research is required to confirm these preliminary results and to test reliability. References Alexanderson H, Lundberg IE, Stenstrom CH: Development of the myositis activities profile–validity and reliability of a self-administered questionnaire to assess activity limitations in patients with polymyositis/dermatomyositis. J Rheumatol 2002 Alexanderson H, Reed AM, Ytterberg SR: The Myositis Activities Profile-initial validation for assessment of polymyositis/dermatomyositis in the USA. J Rheumatol 2012 Disclosure of Interest None declared

OP0045 Early Detection of Lung Involvement in Systemic Sclerosis Using Molecular Targeted Imaging

Background Interstitial lung disease (ILD) is one of the leading causes of death in systemic sclerosis (SSc). Since routine diagnostics such as CT and pulmonary function tests only detect impaired organ function and/or damage, there is an unmet need for the non-invasive diagnosis of ILD at earliest, possibly still reversible disease stages. Objectives To assess nuclear imaging for the detection of SSc-ILD by using radiotracers specifically targeting integrin αvβ3 as a pathophysiologic key molecule of early inflammation-dependent fibrosis in the murine model of bleomycin-induced pulmonary fibrosis and in the multisystemic Fra-2 transgenic (tg) mouse model of SSc. Methods Expression of integrin αvβ3 was analysed in lung sections from patients with SSc-ILD, idiopathic pulmonary fibrosis (IPF), healthy controls (n=5) as well as from bleomycin-treated mice, Fra-2 tg mice and respective controls (n=4) using immunohistochemistry. In vivo imaging was performed using 111In-DOTA-RGD radioconjugates specifically targeting integrin αvβ3. SPECT (single photon emission computed tomography) was performed using a small-animal SPECT/CT scanner (NanoSPECT/CT, Mediso). Animals were scanned at early disease time points to visualise inflammation-dependent pulmonary fibrosis. The pulmonary accumulation of the radiotracer was confirmed by ex vivo SPECT/CT, biodistribution, and autoradiography studies. Nonparametric non-related data were expressed as median(Q1,Q3). For statistical analysis, the Mann-Whitney U test was applied. P-values <0.05 were considered statistically significant. Results In lung sections of patients with SSc-ILD and IPF, the expression of integrin αvβ3 was increased by 3.7-fold and 2.9-fold, respectively as compared to healthy controls (p<0.009, p<0.02). Lungs of bleomycin-treated and Fra-2 tg mice, but not of controls showed a significant increase in integrin αvβ3 expression (upregulation by 5.7-fold and 4.8-fold, respectively) as we have similarly observed in SSc-ILD and IPF patients (p<0.03 each, Fig. C). Integrin αvβ3 was primarily expressed on endothelial cells, inflammatory cells and myofibroblasts as indicated by sequential stainings with cell type-specific markers. Notably, at day 7 after intratracheal bleomycin installation, the peak of pulmonary inflammation, nuclear SPECT/CT with 111In-DOTA-RGD targeting integrin αvβ3, successfully visualised pulmonary inflammation and incipient fibrosis in the model of bleomycin-induced pulmonary fibrosis. Similarly, imaging of integrin αvβ3 in Fra-2 tg mice at 13 weeks of age, the starting point of pulmonary fibrosis, showed a higher pulmonary uptake of the radiotracer in Fra-2 tg mice than in controls (Fig. A). Ex vivo SPECT/CT of isolated lungs, biodistribution and autoradiography studies confirmed the in vivo results and validated the specific radiotracer uptake in lungs from bleomycin-challenged mice and Fra-2 tg mice as compared to controls (Fig. B). Conclusions Our data provide the first evidence that targeting pathophysiologic key molecules using nuclear imaging methods for the visualization of inflammation-dependent fibrosis is a promising non-invasive approach for the early detection of lung involvement in SSc. Disclosure of Interest J. Schniering Grant/research support from: Swiss National Science Foundation (S-85605–02–01), S. Haller Grant/research support from: Swiss National Science Foundation (S-85605–02–01), Z. Guo Grant/research support from: respective institution, C. Feghali-Bostwick Grant/research support from: respective institution, R. Schibli Grant/research support from: respective institution, O. Distler Grant/research support from: Bayer, Sanofi, Ergonex, Boehringer Ingelheim, Actelion, Pfizer, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, EpiPharm, Ergonex, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, MedImmune, Pharmacyclics, Pfizer, Serodapharm, Sinoxa, C. Müller Grant/research support from: respective institution, B. Maurer Grant/research support from: respective institution

SAT0244 Active Skin Disease at Baseline Does Not Predict Progression of Skin Fibrosis at One Year Follow Up – A Eustar Analysis

Background In clinical trials of systemic sclerosis (SSc), there is an unmet need for better inclusion criteria to enrich for patients with progressive skin fibrosis [1,2]. Recent trials on skin fibrosis frequently used active skin disease at baseline as an enrichment parameter. However, there is little data available supporting such an approach. Objectives To investigate the relationship between active skin disease at baseline and progression of skin fibrosis after one year in patients with diffuse cutaneous SSc (dcSSc). Methods A longitudinal analysis of the EUSTAR registry was performed. The inclusion criteria were: dcSSc, fulfillment of ACR criteria, baseline mRSS≥7, available data for mRSS at 12±2 months. Skin progression was defined as increase in mRSS of >5 points AND ≥25% within 1 year [1]. Activity of skin fibrosis was assessed by patient-reported parameters (worsening of skin within the past month), by using a modified skin thickness progression rate (defined as MRSS at baseline visit/disease duration (years)) and by following patients with progression of mRSS in the recent 12 months for another 12 months. Additionally, non skin specific activity parameters such as the Valentini index, elevated serum inflammatory markers, and other patient reported activity parameters were also analyzed. The Mann-Whitney and the Chi-square tests were used. Results From the 637 patients included, 9.7% had progressive skin fibrosis after 1 year. The patient-reported worsening of skin fibrosis within the past month prior to baseline was not significantly associated with progressive skin disease after 1 year (p=0.774). Similarly, the modified skin progression rate at baseline did not differ between progressive and non-progressive patients at 1 year follow up (p=0.323). Most interestingly, patients with progression of skin fibrosis in the previous year were not significantly more likely to show progression of skin fibrosis in the following year (p=0.385, none of the initially progressive patients at one year showed further progression). In addition, other more general activity parameters at baseline including the Valentini activity index (p=0.673) and inflammatory markers (Erythrocyte sedimentation rate, p=0.287; C-reactive protein p=0.358) were also not associated with progression of skin fibrosis after one year (Figure 1). Conclusions dcSSc patients with recently active skin disease have a similar or even lower likelihood to show further progression of skin fibrosis in a 12 months trial – most likely because they have reached their peak skin score already. Recently active skin disease should not be used as an enrichment criterion in clinical trials targeting skin fibrosis. References Maurer B, Graf N, Michel BA, et al. Prediction of worsening of skin fibrosis in patients with diffuse cutaneous systemic sclerosis using the EUSTAR database. Ann Rheum Dis. 2015;74(6):1124–31. Allanore Y, Distler O. Systemic sclerosis in 2014: Advances in cohort enrichment shape future of trial design. Nature Rev Rheumatol. 2015;11(2):72–4. Disclosure of Interest R. Dobrota: None declared, B. Maurer: None declared, N. Graf: None declared, O. Kowal-Bielecka Consultant for: Abbvie, Actelion, Bayer, Biogen, Pfizer, Roche, Speakers bureau: Abbvie, Actelion, Bayer, Biogen, Pfizer, Roche, M. Matucci-Cerinic: None declared, P. Airò: None declared, P. Caramaschi: None declared, P. Carreira: None declared, G. Riemekasten: None declared, E. Rosato: None declared, Y. Allanore Grant/research support from: Bristol-Myers Squibb, Roche/Genentech, Inventiva, Pfizer, Sanofi, and Servier, Consultant for: Actelion, Bayer, Roche/Genentech, Inventiva, Medac, Pfizer, Sanofi, Servier, and UCB, O. Distler Grant/research support from: Bayer, Sanofi, Ergonex, Boehringer Ingelheim, Actelion, Pfizer, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, EpiPharm, Ergonex, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, MedImmune, Pharmacyclics, Pfizer, Serodapharm, Sinoxa

SAT0198 The Desscipher Project in Systemic Sclerosis (SSC): Observational Data on Digital Ulcers (DU) Prevention from The Eustar Group

Background In SSc, the management of DU is a problem in clinical practice, therefore their prevention is highly warranted. Sildenafil, iloprost, calcium channel blockers (CCB), ACE inhibitors (ACEi) and bosentan, alone or in combination, are used for DU prevention without unanimous agreement. Objectives DeSScipher (“to decipher the optimal management of SSc”) is an EC-funded FP7 research project that consists of five observational trials (OT) addressing different aspects of SSc. The aim of OT1 was to evaluate the efficacy of vasoactive and vasodilating drugs or a combination therapy for the management of DU in SSc. Methods Longitudinal data were collected for up to 24 months by 28 DeSScipher centres. In SSc patients, the efficacy of sildenafil, bosentan, sildenafil+bosentan, iloprost or CCB /ACEi for the prevention of new DU was evaluated. Clinical features, ulcer type and other drugs were also recorded. Time without new DU, proportion of patients without new DU, risk of developing new DU and the mean number of new DU per patient were evaluated at 6 months follow-up. Results In OT1, 1394 patients were enrolled. The history of DU was a significant risk factor for developing new DU (OR=3.146; (95%CI:1.19–8.31), p=0.021), therefore analysis was focused on secondary prevention (prevention of new DU in patients with DU history). 473/1394 (33.9%) had a history of DU and 268/473 (56.7%) had follow up data (58.2% limited and 41.8% diffuse SSc subset). 47/268 (17.5%) were on Bosentan, 33/268 (12.3%) on Sildenafil, 40/268 (14.9%) on Iloprost, 31/268 (11.6%) on Sildenafil and Bosentan combination and 117/268 (43.7%) on CCB/ACEi alone. In patients with history of DU in the last 24 weeks, the treatment with CCB/ACEi alone was associated with 7 fold increased risk of developing new DU compared to all other treatment arms (OR =7.313; 95%CI:1.248–42.85, p=0.027). Patients on concomitant immunosuppressive treatment had a trend towards faster DU recurrence. Conclusions History of DU in the past 24 weeks should prompt a more aggressive preventive strategy other than therapy with CCB/ACEi alone due to its lower efficacy to prevent DU recurrence compared to the other treatment options. Acknowledgement The DeSScipher project was funded by the European Communitys Framework Programme 7 (FP7-HEALTH-2012.2.4.4–2 - Observational trials in rare diseases) under grant agreement N° 305495. We acknowledge the contribution of the following EUSTAR centers: Ancona (member N°34), Assiut (168), Bad Bramstedt (187), Belgrade (55), Bucharest (100), Cluj-Napoca (16), Erlangen (106), Frankfurt (124), Hamburg (64), Iasi (162), Istanbul (21), Istanbul (133), Köln (44), Monserrato (142), Moscow (78), Moscow (190), Padua (31), Rome (94), Salford/Manchester (80), Tübingen (56), Wuppertal (192). Disclosure of Interest None declared

OP0289 Micrornas as Potential Regulators of Monocyte Differentiation and Function in Heart Fibrosis in Systemic Sclerosis

Background Heart involvement in patients with systemic sclerosis (SSc) resembles the inflammatory dilated cardiomyopathy (iDCM) phenotype with predominance of inflammation, fibrosis, vasculopathy and heart dysfunction. Animal studies of iDCM indicated bone marrow originated cells as a major source of pathological myofibroblasts. MicroRNAs are key regulators of immune cell function and are involved in many cardiac pathological processes. However, their roles in monocyte differentiation and fibrogenesis are unclear. Objectives To determine the role of circulating monocytes and microRNAs in the onset and progression of myocardial fibrosis in SSc, we examined monocyte differentiation and their microRNAs expression profile in SSc. Methods Endomyocardial biopsies from SSc/iDCM patients and healthy controls were screened by immunohistochemistry. CD14+ monocytes isolated from peripheral blood of SSc patients and healthy donors were differentiated towards the myofibroblast phenotype by stimulation with TGF-β1, IL-4, IL-10 and IL-13. In addition, CD14+ monocytes were co-cultured with dermal fibroblasts originated from SSc patients and healthy subjects. After 7 days, myofibroblast gene expression and cytokines secretion profile were evaluated. MicroRNA candidates were selected using DIANA-TarBase v7.0 and TargetScan Release 7.0 and further analysed by qPCR. Results Myocardium of SSc/iDCM patients (n=10) revealed extensive fibrosis and accumulation of inflammatory cells including CD14+ monocytes. Moreover, fibrotic myocardium from SSc patients exhibited the presence of CD14+/Fra-2+ monocyte-derived fibroblast-like cells. Bioinformatical analysis indicated several potential microRNAs being involved in monocyte differentiation, and qPCR confirmed predicted candidates. We observed decreased basal levels of let-7i (Mann-Whitney U test, p=0.005), miR-10b (p=0.005), miR-21 (p=0.005), miR-29a (p=0.005), miR-155 (p=0.002), miR-199a (p=0.008) and miR-544a (p=0.002), and increased levels of miR-29b (p=0.03) in circulating CD14+ monocytes isolated from SSc patients (n=10) compared to healthy controls (n=4). Stimulated monocytes acquired a myofibroblast-like phenotype with increased expression of collagen I, fibronectin, α smooth muscle actin and Fra-2 in comparison to untreated cells. Similarly, CD14+ monocytes exposed to dermal fibroblasts acquired myofibroblast features. CD14+ monocytes from SSc patients were characterised by higher production of IP-10, MIP-3α, LIF and NT-3. The process of monocyte to myofibroblast differentiation employed Fra-2/TGF-β signalling. Inhibition of the canonical SMAD-dependent pathway with TGFβR1 inhibitors resulted in the abrogation of monocyte-to-myofibroblast differentiation. Conclusions Here we demonstrated the capability of peripheral blood monocytes to differentiate towards the myofibroblast phenotype, indicating these cells as one of the potential sources of pathological tissue myofibroblasts in SSc. Different miRNA expression profiles in SSc monocytes indicate a primary activation state. Further studies of miRNAs regulation pathways might lead to novel treatment strategies, particularly for heart involvement. Disclosure of Interest M. Rudnik: None declared, M. Stellato: None declared, P. Blyszczuk: None declared, E. Pachera: None declared, R. Dobrota: None declared, B. Maurer: None declared, K. Klingel: None declared, J. Henes: None declared, K. Sotlar: None declared, O. Distler Grant/research support from: Bayer, Sanofi, Ergonex, Boehringer Ingelheim, Actelion, Pfizer, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, EpiPharm, Ergonex, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, MedImmune, Pharmacyclics, Pfizer, Serodapharm, Sinoxa, G. Kania: None declared

SAT0227 The Efficacy of Vasoactive and Vasodilating Drugs on Digital Ulcers Healing in Systemic Sclerosis: Data from The Desscipher Observational Study of Eustar Group

Background Digital ulcers (DU) are an important driver of poor quality of life in Systemic Sclerosis (SSc). Real life clinical management of DU includes the use of vasoactive (Bosentan) and vasodilating (Sildenafil, Iloprost, CCB and ACE inhibitors) drugs in different combinations, without strong evidence for efficacy. Objectives One of the aims of the EC-funded FP7 research project DeSScipher (acronym for “to decipher the optimal management of SSc”) was to determine the relative effectiveness of the above treatments for the healing of DU in SSc patients. Methods Twenty eight centres in Europe participated to the study. Longitudinal data were collected over 24 months in a specific web based EUSTAR database. The DU distal to PIP joints at the inclusion visit or developed over the follow-up were included. End-points included: mean number of healed ulcers/patient; proportion of patients with healed DU; and time to healing of at least one DU. Results Out of 1394 enrolled patients, 265 met inclusion criteria. Follow-up data were available for 156 patients. 57% of patients were on vasodilators (Sildenafil, Iloprost, CCB, alone or in combination), 5.8% were on single vasoactive drug (Bosentan) and 37.2% patients were on combination therapy. The mean number of DU/patient at the inclusion visit was comparable among the 3 treatment arms. At 12 and 24 months, the mean number of healed DU/patient was higher in the vasodilator group compared to the vasoactive and combination groups (1.8 vs 1 vs 1.2, p=0.013 and 2.4 vs 1.3 vs 1.3, p=0.018 respectively). No significant difference was observed at 6 months. The proportion of patients with all or at least one DU healed was not significantly different between treatment arms. Initial exploratory analysis also suggested that the estimated mean time to healing of at least one DU was 9.71±2.2 months in vasoactive arm, 8.7±0.5 in vasodilator arm and 7.9±0.6 in combination therapy arm. Further, when considered separately, patients on Sildenafil (±CCB) showed the highest healing rate at 6months follow-up, compared to other drug combinations. However, correction for concurrent treatment and confounding factors is still on going. Conclusions Vasodilating treatment achieved a major number of healed DU at 12 and 24 months. The combination treatment seems to be of help in obtaining a shorter time to healing and limiting observation at 6 months may bias overall interpretation of data. Further analysis is ongoing for different drug combination at different time-points. Acknowledgement The DeSScipher project was funded by the European Communitys Framework Programme 7 (FP7-HEALTH-2012.2.4.4–2 - Observational trials in rare diseases) under grant agreement N° 305495. Disclosure of Interest None declared

OP0018 Microrna-145 down-regulation in systemic sclerosis is limited to skin fibroblasts and influenced by epigenetic modifications and negative feedback loop by TGF-β

Background We recently found that miR-145 plays anti-fibrotic effects in SSc fibroblasts via the regulation of the TGF-β pathway in a multi-step fashion. Objectives To elucidate the mechanisms regulating the expression of miR-145 in SSc fibroblasts. Methods Basal miR-145 and pri-miR-145 expression was analysed in 12 SSc and 6 healthy control (HC) dermis and fibroblasts by Real-time PCR. MiR-145 expression was further analysed in SSc and HC fibroblasts exposed to hypoxia, stimulated with pro-fibrotic cytokines (TGF-β, PDGF-B, VEGF-A), and treated with either a Smad3 inhibitor (SIS3) or a TGF- βRII neutralizing antibody. Finally, miR-145 expression was analysed in peripheral blood mononuclear cells (PBMCs) of 5 SSc patients and 5 HC. Results MiR-145 was down-regulated in both SSc dermis and fibroblasts by 2.2 and 2.0 fold respectively (p<0.001) as compared to HC. Similar results were obtained for the non-functional primary transcript, pri-miR-145, suggesting a regulation on the transcriptional level rather than post-transcriptional modifications. No significant difference in the expression of miR-145 was found between SSc fibroblasts exposed to hypoxia or pro-fibrotic cytokines TGF-β, PDGF-B, VEGF-A at conventional doses compared to controls. However, treatment of SSc fibroblasts with both SIS3 and TGF-βRII neutralizing antibodies increased miR-145 levels by 2.4 and 1.8 fold, respectively. This prompted us to investigate the expression of miR-145 in HC fibroblasts treated with TGF-β at 5 ng/ml, finding a 1.3 fold down-regulation, suggesting a possible regulatory loop between miR-145 and the TGF-β pathway. In addition, down-regulation of miR-145 in SSc was mediated by epigenetic modifications, as inhibition of DNA methyltransferases by 5aza-C increased miR-145 levels by 2.5 fold in SSc fibroblasts (p<0.01). Finally, to clarify whether miR-145 down-regulation in SSc fibroblasts was a cell-specific finding, we investigated miR-145 expression in SSc PBMCs and found no difference with respect to HC (p>0.05). Conclusions We previously showed that the down-regulation of miR-145 in SSc has strong pro-fibrotic effects in SSc fibroblasts via direct post-transcriptional induction of TGF-β signalling (TGFBR2 and SMAD3) and CTGF, directly interacting with TGFBR2 3’UTR. Our new data suggest that miR-145 down-regulation in SSc is rather specific for skin fibroblasts. In addition, miR-145 expression in SSc fibroblasts appears modulated by epigenetic mechanisms. Most interestingly, there is a feedback regulation between TGF-β and miR-145 which self-enhances the pathologic pathway. Taken together, these data further support the role for miR-145 in the regulation of the TGF-β pathway in SSc, and make the restoration of miR-145 levels an appropriate option for the treatment of the disease. Disclosure of Interest S. Vettori Grant/Research support from: EULAR ODP, M. Brock: None Declared, N. Iwamoto: None Declared, B. Maurer: None Declared, A. Jungel: None Declared, R. Gay: None Declared, M. Calcagni: None Declared, G. Valentini: None Declared, J. Distler: None Declared, S. Gay: None Declared, O. Distler Grant/Research support from: EULAR ODP, Actelion, Pfizer, Ergonex, Sanofi, Consultant for: Actelion, Pfizer, Ergonex, BMS, Sanofi-Aventis, United BioSource Corporation, medac, 4D Science, Boehringer-Ingelheim, Active Biotech, Roche, Speakers Bureau: Actelion, Pfizer, Encysive, Ergonex