Monica Caronni

Analysis of Class II human leucocyte antigens in Italian and Spanish systemic sclerosis

OBJECTIVEnTo determine the role of Class II HLAs in SSc patients from Italy and Spain and in SSc patients of Caucasian ancestry.nnnMETHODSnNine hundred and forty-four SSc patients (Italy 392 patients; Spain 452 patients) and 1320 ethnically matched healthy controls (Italy 398 patients; Spain 922 patients) were genotyped up to the fourth digit by PCR with sequence-specific oligonucleotides for HLA-DRB1, DQA1 and DQB1 loci. Patients included 390 ACA-positive and 254 anti-topo I-positive subjects. Associations between SSc or SSc-specific antibodies and HLA alleles or HLA haplotypes were sought via the chi-square test after 10u2009000-fold permutation testing. A meta-analysis including this study cohort and other Caucasoids samples was also conducted.nnnRESULTSnIn both the cohorts, the strongest association was observed between the HLA-DRB1*1104 allele and SSc or anti-topo I antibodies. The HLA-DRB1*1104 -DQA1*0501 -DQB1*0301 haplotype was overrepresented in Italian [odds ratio (OR)u2009=u20092.069, 95% asymptotic CIs (CI(95)) 1.486, 2.881; Pu2009<u20090.001] and in Spanish patients (ORu2009=u20096.707, CI(95) 3.974, 11.319; Pu2009<u20090.001) as well as in anti-topo-positive patients: Italy (ORu2009=u20092.642, CI(95) 1.78, 3.924; Pu2009<u20090.001) and Spain (ORu2009=u200920.625, CI(95) 11.536, 36.876; Pu2009<u20090.001). In both the populations we also identified an additional risk allele (HLA-DQB1*03) and a protective allele (HLA-DQB1*0501) in anti-topo-positive patients. The meta-analysis showed different statistically significant associations, the most interesting being the differential association between HLA-DRB1*01 alleles and ACAs (ORu2009=u20091.724, CI(95) 1.482, 2.005; Pu2009<u20090.001) or topo I antibodies (ORu2009=u20090.5, CI(95) 0.384, 0.651; Pu2009<u20090.001).nnnCONCLUSIONSnWe describe multiple robust associations between SSc and HLA Class II antigens in Caucasoids that may help to understand the genetic architecture of SSc.

Role of class II human leucocyte antigens in the progression from early to definite systemic sclerosis

OBJECTIVEnHLAs have been extensively associated with SSc susceptibility but their role in the progression of the disease is poorly understood. In 2013 the ACR and European League Against Rheumatism (EULAR) jointly defined criteria for the classification of SSc that allow the early identification of definite SSc patients. In this study we investigated the role of HLA class II antigens in the progression from early to definite SSc.nnnMETHODSnOne hundred and fifty-eight subjects with early SSc according to LeRoy and Medsger criteria and no other manifestation indicative of definite SSc at referral were considered. All the patients underwent high-resolution HLA class II typing and the appraisal of definite SSc was retrospectively conducted in a prospective manner. Lifetime analysis was conducted to gauge the effect of genetic and clinical characteristics on progression of the disease.nnnRESULTSnThe median estimated time to progression was 45 months from referral; the 5 and 10 year estimates of progression were 59.8% and 80%, respectively. ACAs were associated with a reduced risk of progression [median survival 55 vs 23 months for ACA-positive vs ACA-negative patients, P = 0.035; hazard ratio (HR) 0.67 (95% CI 0.458, 0.979)]. HLA alleles within the HLA DQ5-DR1 haplotype [HLA-DRB1*0101-HLA-DQA1*0101(4)-HLA-DQB1*0501] reduced the risk of progression of the disease [median survival 108 vs 44 months for DQ5-DR1 carriers vs DQ5-DR1 non-carriers; HR 0.388 (CI 0.211, 0.712), P = 0.001, corrected P = 0.014]. In multivariate models, the effect of genetics was found to be independent of ACA positivity or other baseline factors; additive risks were observed when the DQ5-DR1 haplotype and ACA were jointly considered.nnnCONCLUSIONnHLA class II alleles within the HLA DQ5-DR1 haplotype are associated with lower rates of progression from early to definite SSc.

Serum levels of vascular dysfunction markers reflect disease severity and stage in systemic sclerosis patients

OBJECTIVEnTo improve knowledge of vasculopathy in SSc through the assessment of serum levels of circulating angiogenetic and endothelial dysfunction markers in patients at different stages of the disease.nnnMETHODSnSera from 224 subjects were obtained and concentrations of angiopoietin-2, chemokine (C-X-C motif) ligand (CXCL)-16 (CXCL16), E-selectin, soluble intercellular adhesion molecule-1, IL-8 (CXCL8), soluble vascular adhesion molecule-1 and VEGF were determined by a Luminex assay. Subjects included 43 healthy controls, 47 early SSc patients according to LeRoy and Medsger without other signs and symptoms of evolutive disease, 48 definitive SSc (defSSc) patients according to the 2013 ACR/EULAR criteria without skin or lung fibrosis, 51 lcSSc subjects and 35 dcSSc subjects.nnnRESULTSnThe four groups of patients showed well-distinct clinical and laboratory characteristics, with a linear decreasing trend in forced vital capacity and diffusing capacity for carbon monoxide % predicted values from early SSc to defSSc to lcSSc and to dcSSc, and a linear increasing trend in ESR, and in the prevalence of abnormal CRP, serum gamma globulins and lung fibrosis (all P < 0.0001). Highly significant linear trends pointing to an increase in angiopoietin-2 (P < 0.0001), CXCL16 (P < 0.0001), E-selectin (P = 0.001) and soluble intercellular adhesion molecule-1 (P = 0.002) in relation to the different disease subsets were observed.nnnCONCLUSIONnMarkers characterizing vascular activation are found to be increased in SSc patients from the earliest stages of disease when clinical and laboratory findings of advanced disease cannot yet be detected. These abnormalities progress with the appraisal of the first sclerodermatous manifestation in defSSc and further increase with the onset of fibrotic manifestations.

Preliminary safety and efficacy profile of prucalopride in the treatment of systemic sclerosis (SSc)-related intestinal involvement: results from the open label cross-over PROGASS study

BackgroundProkinetics are used to treat enteric dismotility symptoms in systemic sclerosis (SSc) patients, but they often lack adequate efficacy. The most effective prokinetics belonging to the serotonin (5-HT4) receptor agonists class were withdrawn due to cardiac toxicity in relation to modest 5-HT4 receptor affinity. Prucalopride is a high-affinity 5-HT4 receptor agonist with no major cardiac issues, for which the efficacy in SSc has not yet been assessed.MethodsForty patients with self-reported mild to moderately severe enteric symptoms were enrolled in a cross-over 2u2009×u20092 study. Subjects were randomized 1:1 to prucalopride 2 mg/day or no treatment for one month and vice versa after a 2-week washout period. Before and after each sequence the patients compiled the University of California Los Angeles gastrointestinal tract (UCLA GIT) 2.0 questionnaire and the numbers of complete intestinal movements were recorded. Oro-cecal transit time (OCTT) was evaluated by lactulose breath test in a subgroup of patients. Data were evaluated by mixed linear models corrected for the number of laxatives used during the study periods.ResultsThere were 29 subjects who completed the study; 7 subjects withdrew due to side-effects and 4 subjects were not compliant with the study procedures. As compared to dummy treatment, prucalopride was associated with more intestinal evacuations (pu2009<u20090.001), improvement of UCLA GIT constipation (-0.672u2009±u20090.112 vs 0.086u2009±u20090.115; pu2009<u20090.001), reflux (-0.409u2009±u20090.094 vs 0.01u2009±u20090.096; pu2009<u20090.005) and bloating (-0.418u2009±u20090.088 vs -0.084u2009±u20090.09; pu2009=u20090.01) scores. Treatment was ranked moderately to more than moderately effective by 22 patients (72.4%). OCTT was significantly reduced during prucalopruide consumption (prucalopride: -20.1u2009±u200920.1 vs no treatment: 45.8u2009±u200921.3 minutes; treatment effectu2009=u2009-65.9 minutes; pu2009=u20090.035).ConclusionsThe safety profile of prucalopride in SSc is similar to what is known from the literature. In patients with mild to severe gastrointestinal problems, prucalopride may be effective in treating dismotility symptoms, increasing the number of complete bowel movements and improving bowel transit, reducing reflux disease and bloating.Trial registrationEU Clinical Trial Registry, EudraCT2012-005348-92. Registered on 19 February 2013.

Carbohydrate antigen 15.3 as a serum biomarker of interstitial lung disease in systemic sclerosis patients

BACKGROUNDnTo determine the usefulness of Ca 15.3 as a candidate biomarker in systemic sclerosis (SSc) patients with interstitial lung disease (ILD).nnnMETHODSnTwo-hundred-twenty-one SSc patients with Ca 15.3 determinations were considered; 168 had evidence of interstitial lung involvement on high-resolution computed tomography (HRCT); digitalized scans were available for scoring in 84 subjects. Discrimination between patients with or without ILD, was assessed by receiving operating characteristics (ROC) analysis; correlations between HRCT scores and Ca 15.3 were performed. Survival and serial pulmonary function tasting (PFT) data were used for prognostication.nnnRESULTSnCa 15.3 serum levels strongly correlated with HRCT scores (r=0.734, p<0.0001) which were predictors of survival at the 20% threshold (p=3.1∗10(-4)). Ca 15.3 had an area under ROC to detect the meaningful 20% fibrosis extent equal to 0.927 and abnormal Ca 15.3 values were capable of differentiating between patients at hi- or low-risk for progression in the group with undetermined disease extent (HR=3.209, confidence interval [CI95]=1.56-6.602, p=0.002). Ca 15.3 outperformed other PFT measures in providing a separation of survival estimates where HRCT scans are unavailable. The combined use of HRCT scores and Ca 15.3 in SSc-ILD patients was more discriminatory (HR=4.824, CI95=2.612-8.912, p<0.0001) than the staging system based on HRCT scores plus FVC (HR=2.657, CI95=1.703-4.147, p<0.0001) and characterized by lower prediction errors (0.2134 vs 0.2234).nnnCONCLUSIONnCa 15.3 is a rapid and inexpensive candidate biomarker for SSc-ILD being proportional to the extent of lung injury and specific and sensitive in assessing meaningful extents of the disease with prognostic significance.

18F-FDG uptake in main arterial branches of patients with large vessel vasculitis: visual and semiquantitative analysis

ObjectiveOver the last decade, the contribution of 18F-FDG (FDG) PET/CT imaging to the diagnosis of large vessel vasculitis has been widely investigated. The aim of this study was to evaluate a more extensive role for PET/CT in grading vascular inflammation in patients with different clinical stages of disease.MethodsThe images of 66 PET/CT studies of 34 patients, performed at diagnosis and/or during follow-up were reviewed. FDG uptake in different regions of aorta and in its major branches was visually (regional Score: rS) and semiquantitatively (regional SUVmean: rSUV) assessed. The global vascular uptake was also evaluated for each study by summing all rSs (summed Score; sS) and averaging rSUVs (averaged SUV; aSUV). FDG uptake in 15 PET/CT studies of control age-matched subjects without signs or symptoms of vasculitis was also analyzed.ResultsHigher levels of regional and global FDG uptake were found at diagnosis in comparison with follow-up studies of 12 patients with complete longitudinal observation (p value range 0.0552–0.0026). In the latter group high values were generally observed when disease relapse or incomplete response to therapy (active disease) occurred, whereas lower uptake was found in studies of remitted patients (pxa0=xa0<0.01), whose FDG levels were similar to those of control subjects. At ROC analysis performed on all image dataset, optimal cut-off levels of regional and global FDG vascular uptake provided a good discrimination between 25 patients at diagnosis and 15 control subjects (aSUV greater than 0.697; PPVxa0=xa092.3; NPVxa0=xa092.9). Major overlap was observed among FDG levels of 21 patients with active disease and in remission (aSUV greater than 0.653; PPVxa0=xa058.3; NPVxa0=xa094.1). Similar performances of visual and semiquantitative analyses were found when areas under curves (AUCs) were compared.Conclusions18F-FDG PET/CT has a promising role in grading inflammation in patients with large arteries vasculitis. Nevertheless, a cut-off based analysis of FDG vascular uptake is not sufficient to separate patients with active and inactive disease during follow-up.

AB0204 Serum Levels of Vascular Markers Reflect Disease Severity and Stage in Systemic Sclerosis

Background Vasculopathy is one of the cardinal feature of systemic sclerosis (SSc) that can be observed in every patients from the earliest stages of the disease. Abnormalities in circulating angiogenetic factors and endothelial dysfunction markers in patients at different stages of the disease that include early or non-fibrotic (pre-fibrotic) subjects have not thoroughly described so far. Objectives To characterize vasculopathy in SSc patients through the determination of 7 selected vascular biomarkers in sera from a large cohort of patients with well defined and distinct clinical characteristics. Methods Sera from 224 subjects were obtained and concentrations of Angiopoietin 2, CXCL-16, E-Selectin, ICAM-1, IL-8, VCAM-1, VEGF were determined by a Luminex commercial assay. Subjects included 43 healthy controls (HC), 47 early systemic sclerosis patients (EaSSc) according to LeRoy and Medsger criteria (Raynauds phenomenon + abnormal capillaroscopy patterns and/or SSc-specific antibodies without other signs and symptoms of disease), 48 definitive SSc patients (defSSc) according to the 2013 ACR/EULAR criteria without skin fibrosis, 51 limited cutaneous (lcSSc) and 35 diffuse cutaneous (dcSSc) subjects. Results The four groups of patients showed well-distinct clinical and laboratory characteristics with a linear trend of decrease in FVC and DLco % predicted levels from EaSSc to defSSc to lcSSc and to dcSSc patients, and a linear trend of increase in CRP, ESR, gamamglobulin serum levels as well as the prevalence of lung fibrosis on the other way around. Serum concentrations of the studied analytes are reported in the table; for most vascular factors a characteristics linear trend (ANOVA polynomial test for trend <0.01) could be observed with the lowest level in HC and a gradual increase that reflects the severity if the disease. Conclusions Our study demonstrate, for the first time, that circulating levels of vascular involvement start to increase in SSc patients from the earlies stages of the disease when clinical and laboratory findings of advanced disease cannot yet be detected. These abnormalities progress in parallel with the appraisal of the first sclerodermatous manifestation in defSSc and steadly increase with the onset of fibrotic manifestations. Acknowledgements The present work was financed by a grant from GILS (Gruppo Italiano per la Lotta alla Sclerodermia), theme “La diagnosi precoce della sclerodermia”. Disclosure of Interest None declared