Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

Favoino, Elvira; Digiglio, Liboria; Cuomo, Giovanna; Favia, Isabella; Racanelli, Vito; Valentini, Gabriele; Perosa, Federico

doi:10.1371/journal.pone.0061453

Cited by 10 publications

(17 citation statements)

References 34 publications

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“…Ten serum samples with highest titer were pooled and used for affinity purification of anti-CarP Ab. The specificity of anti-CarP Ab was assessed by ELISA in binding and inhibition assays as previously described [38].…”

Section: Methodsmentioning

confidence: 99%

Anti-carbamylated protein antibodies and skin involvement in patients with systemic sclerosis: An intriguing association

et al. 2018

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Carbamylation is a post-translational modification that mostly affects proteins with low turnover, such as dermal proteins. Carbamylated proteins accumulate in skin in an age-dependent manner, contributing to tissue alterations. As dermis is affected by systemic sclerosis (SSc) and anti-carbamylated protein antibodies (anti-CarP Ab) are found in SSc patients, we sought to evaluate the specificity of anti-CarP Ab and their relationship with clinical parameters reflecting skin involvement in SSc. This study investigated serum samples and clinical data from 124 patients with SSc. Anti-CarP Ab were affinity purified from pooled SSc sera, and their specificity was assessed by western blotting and ELISA with carbamylated proteins from two species (human and bovine albumin; human fibrinogen). Anti-CarP Ab were measured in SSc serum samples and in 41 healthy aged-matched individuals. Affinity-purified anti-CarP Ab recognized carbamylated epitopes irrespective of the protein type or species origin. Anti-CarP Ab levels inversely correlated with the modified Rodnan skin score (mRss) (Spearman’s R = -0.32, p<0.001), independently of patients’ age. Receiver operating characteristics (ROC) analysis identified anti-CarP Ab cut-offs that best discriminated dichotomized clinical variables related to skin involvement: the only clinical variables that were significantly different between groups were mRss (p = 0.001) and scleredema (p<0.001). Low anti-CarP Ab levels were associated with worse skin involvement. Future prospective studies are needed to assess their usefulness in the clinical setting.

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Section: Methodsmentioning

confidence: 99%

Anti-carbamylated protein antibodies and skin involvement in patients with systemic sclerosis: An intriguing association

et al. 2018

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“…Levels of anti-CarP Ab in serum samples were assessed by indirect ELISA, as described with minor modifications. 19 Briefly, polyvinylchloride 96-well microtiter plates were coated with modified and nonmodified BSA at 5 µg/mL. Wells were blocked with PBS containing 0.5% BSA (PBS-BSA), and then incubated for 4 h with serum samples diluted 100 times in PBS-BSA.…”

Section: Methodsmentioning

confidence: 99%

Low mortality rate in Italian rheumatoid arthritis patients from a tertiary center: putative implication of a low anti-carbamylated protein antibodies prevalence

Iacono¹,

Favoino²,

Borgia³

et al. 2018

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Background and objectiveAnti-carbamylated protein antibodies (anti-CarP Ab) represent a novel kind of autoantibodies specificity detectable in the sera of patients with rheumatoid arthritis (RA). They have been recently reported to be associated with increased mortality in Spanish patients with RA. The aim of our study was to compare the incidence mortality rates (IMRs) detected in RA patients from a tertiary Italian center with those reported in other European tertiary centers and to evaluate the putative role of anti-CarP Ab in modulating the low IMR detected in our patients.MethodsClinical charts of patients consecutively admitted to our center, from January 1, 2008, to December 31, 2014, were retrospectively reviewed. The mortality rate (expressed as the number of deaths in the cohort divided by the number of years of IMR follow-up) and causes of death were assessed at December 31, 2015. Sera of 61 patients, representative of the whole cohort, collected at the time of admission to our center were investigated for the presence and the level of anti-CarP Ab. Demographic and clinical features, mortality rates and prevalence of anti-CarP Ab in our series were compared with those reported in other European cohorts.ResultsWe observed 608 patients for a median of 3.51 years. All-cause and cause-specific IMRs in our cohort were significantly lower than the Better Anti-rheumatic Farmaco-therapy and the Spanish cohort, while only all-cause and cardiovascular IMRs were significantly lower in our series with respect to the Leiden Early Arthritis Clinic cohort. Anti-CarP Ab prevalence was significantly lower in our series than in any other European cohorts.ConclusionWe confirm that the mortality rate is lower in our Italian RA cohort with respect to other European cohorts. Whether the low prevalence of anti-CarP Ab might be responsible for this result awaits to be furtherly investigated.

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“…[21,22] For the purposes of this study, we purified these Ig fractions from sera of an additional 4 patients using identical methods. Antibody concentration was determined by UV absorption with 1.35 extinction coefficient at 280 nm for 1 mg/mL.…”

Section: Methodsmentioning

confidence: 99%

“…[21] Specifically, these were 2 phage clones obtained with Ig from patient pt4 (pc4.1 and pc4.2) and 2 phage clones corresponding to patient pt14 (pc14.1 and pc14.2). These clones had been stored at −20 °C in 50% glycerol.…”

Section: Methodsmentioning

confidence: 99%

“…[21,22] Briefly, wells were coated with 5 μg/mL purified Ig in an overnight incubation at 4 °C, and then incubated with phage-enriched supernatant diluted 16 times (100 μL/well) for 4 hours at room temperature. After 4 washings with PBS-T20, bound phage particles were detected with HRP-conjugated anti-M13 mAb and o -phenylenediamine, as above.…”

Section: Methodsmentioning

confidence: 99%

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Subspecificities of anticentromeric protein A antibodies identify systemic sclerosis patients at higher risk of pulmonary vascular disease

et al. 2016

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Patients with systemic sclerosis (SSc) who express autoantibodies to centromeric proteins (CENPs) are at risk of developing pulmonary vascular disease and pulmonary arterial hypertension without fibrosis. Currently no biomarkers are available to predict these complications. We previously characterized the fine specificity of anti-CENP-A antibodies in SSc by screening a phage display library (expressing random 12-mer peptides), and identified phage clones whose peptides were differentially recognized by patients’ autoantibodies. Here, we examined if subgroups of SSc patients with different anti-CENP-A antibody subspecificities also differ clinically, and if serum reactivity to phage-displayed peptides can predict pulmonary vascular disease.Clinical data and serum samples were collected from 84 anti-CENP-A-positive SSc patients. Indirect ELISAs were used to test serum reactivity. Pulmonary vascular disease was defined as high systolic pulmonary arterial pressure (sPAP) and low diffusing lung capacity for carbon monoxide (DLCO; percent of predicted values).Sera were screened for reactivity to peptides expressed by phage clones pc4.2 and pc14.1, confirming our earlier observation of differential specificities. Linear regression showed that the levels of antibodies specific for the 2 phage clones were associated with clinical features of pulmonary vascular disease, but in opposite ways: anti-pc4.2 antibodies were positively associated with sPAP and inversely associated with DLCO, whereas anti-pc14.1 antibodies were inversely associated with sPAP and positively associated with DLCO. Anti-pc4.2 and anti-pc14.1 antibody levels predicted sPAP independently of DLCO. These associations were confirmed by logistic regression using antibodies as predictors and dichotomized sPAP (cutoff, 45 mm Hg) as outcome. The ratio of the 2 antibody levels was a useful marker in predicting high sPAP.This study demonstrates that some SSc clinical features associate with subspecificities of anti-CENP-A antibodies. Moreover, it shows that a simple, inexpensive phage-based assay can predict which SSc patients have high sPAP and low DLCO, hence who are at greater risk of developing pulmonary arterial hypertension. The ability to identify these at-risk patients can contribute to clinical efficiency and effectiveness. Further research into the peptides expressed by the phage clones may reveal the molecular mechanisms that put some anti-CENP-A-positive patients at greater risk than others for pulmonary vascular disease.

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Autoantibodies Recognizing the Amino Terminal 1-17 Segment of CENP-A Display Unique Specificities in Systemic Sclerosis

Cited by 10 publications

References 34 publications

Anti-carbamylated protein antibodies and skin involvement in patients with systemic sclerosis: An intriguing association

Anti-carbamylated protein antibodies and skin involvement in patients with systemic sclerosis: An intriguing association

Low mortality rate in Italian rheumatoid arthritis patients from a tertiary center: putative implication of a low anti-carbamylated protein antibodies prevalence

Subspecificities of anticentromeric protein A antibodies identify systemic sclerosis patients at higher risk of pulmonary vascular disease

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