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Licochalcone-A (Lico-A) PLGA NPs functionalized with cell penetrating peptides B6 and Tet-1 are proposed for the treatment of ocular anti-inflammatory diseases. In this work, we report the in vitro biocompatibility of cell penetrating peptides-functionalized Lico-A-loaded PLGA NPs in Caco-2 cell lines revealing a non-cytotoxic profile, and their anti-inflammatory activity against RAW 264.7 cell lines. Given the risk of hydrolysis of the liquid suspensions, freeze-drying was carried out testing different cryoprotectants (e.g., disaccharides, alcohols, and oligosaccharide-derived sugar alcohol) to prevent particle aggregation and mitigate physical stress. As the purpose is the topical eye instillation of the nanoparticles, to reduce precorneal wash-out, increase residence time and thus Lico-A bioavailability, an in-situ forming gel based on poloxamer 407 containing Lico-A loaded PLGA nanoparticles functionalized with B6 and Tet-1 for ocular administration has been developed. Developed formulations remain in a flowing semi-liquid state under non-physiological conditions and transformed into a semi-solid state under ocular temperature conditions (35 °C), which is beneficial for ocular administration. The pH, viscosity, texture parameters and gelation temperature results met the requirements for ophthalmic formulations. The gel has characteristics of viscoelasticity, suitable mechanical and mucoadhesive performance which facilitate its uniform distribution over the conjunctiva surface. In conclusion, we anticipate the potential clinical significance of our developed product provided that a synergistic effect is achieved by combining the high anti-inflammatory activity of Lico-A delivered by PLGA NPs with B6 and Tet-1 for site-specific targeting in the eye, using an in-situ forming gel., R. M. Galindo-Camacho acknowledges the financial support of the Generalitat de Catalunya for the PhD scholarship FI-DGR EMC/2199/2017 (DOGC-7459-2017) and for the scholarship Santander Research/Convocatòria d’estades 2022 per a doctorands de la UB. Author are also thankful to the Spanish Ministry of Economy, Industry and Competitiveness, and the European Regional Development Fund (grants RTI2018-094120-B-I00 and PID2021-122216OB-I00). E. B. Souto wishes to acknowledge the national funds from FCT—Fundação para a Ciência e a Tecnologia, I.P., in the scope of the project UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit on Applied Molecular Biosciences—UCIBIO and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy—i4HB

During the last few decades, several efforts have been made towards the development of biocompatible materials. Among them, peptide amphiphiles (PAs) constitute a novel nanotechnological strategy used in the field of biomedicine since they can provide tissue-specific binding and localization. PAs possess several regions combining hydrophobic and hydrophilic areas that are able to self-assemble in aqueous media forming different tertiary nanostructures able to interact with cellular membranes. Moreover, these molecules can be tuned incorporating collagen, lipids or fluorescent markers. In addition, they can also be used as carriers in order to encapsulate active compounds for drug delivery showing promising features in this area. In this review the self-assembled structures of PAs as well as their pharmacological applications have been summarized. Furthermore, their use as drug delivery systems has been highlighted and the latest advances in this field have been reviewed.

Background: Vaginal drug delivery offers an opportunity for effective microbicide treatments. However, the physiological characteristics of the vagina represent a challenge when preparing microbicide formulations for local delivery that avoid systemic effects. Objective: To review recent advances in vaginal microbicides placing special emphasis on nanocarriers that encapsulate peptide microbicides. The review includes an overview of local treatment and prevention strategies for bactericidal, fungal, protozoal and viral vaginal infections that use peptide microbicides. Methods: Scientific literature and clinical trials published in the PubMed database since 2015 are analyzed. Result: Peptide microbicides are used for vaginal drug delivery as a means of prevention and treatment of several infectious diseases, especially sexually transmitted infections, and it is clear that such peptide-based microbicides constitute a suitable strategy. Their combination with nanotechnological drug delivery systems improves local peptide administration while avoiding their degradation and adverse effects. To date, only a few polymeric and lipid-based nanocarriers have been used for peptide microbicide vaginal delivery. Conclusion: The combination of nanotechnology and peptide microbicides has emerged over recent years, and different platforms are being used for vaginal peptide delivery. Nanostructured systems for peptide delivery via vaginal rings or hydrogels are increasingly used as platforms capable of providing suitable drug delivery. However, there is still a lack of knowledge about nanocarriers aimed at encapsulating peptides for local microbicide delivery., We acknowledge funding received from the Spanish Ministry of the Economy, Industry and Competitiveness (MINECO) and the European Regional Development Fund (grant RTI2018-094120-B-I00).

In a previous work, we defined a novel HIV-1 fusion inhibitor peptide (E1P47) with a broad spectrum of activity against viruses from different clades, subtypes, and tropisms. With the aim to enhance its efficacy, in the present work we address the design and synthesis of several peptide amphiphiles (PAs) based on the E1P47 peptide sequence to target the lipid rafts of the cell membrane where the cell–cell fusion process takes place. We report the synthesis of novel PAs having a hydrophobic moiety covalently attached to the peptide sequence through a hydrophilic spacer of polyethylene glycol. Characterization of self-assembly in condensed phase and aqueous solution as well as their interaction with model membranes was analyzed by several biophysical methods. Our results demonstrated that the length of the spacer of polyethylene glycol, the position of the peptide conjugation as well as the type of the hydrophobic residue determine the antiviral activity of the construct. Peptide amphiphiles with one alkyl tail either in C-terminus (C-PAmonoalkyl) or in N-terminus (N-PAmonoalkyl) showed the highest anti-HIV-1 activities in the cellular model of TZM-bl cells or in a preclinical model of the human mucosal tissue explants., Financial support from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO, Spain) and the European Regional Development Fund (Grant RTI2018-094120-B-I00 to I.H.) is gratefully acknowledged. R. P. acknowledges financial support from Ministry of Economy, Industry and Competitiveness, Spain (Grant CTQ2017-88948-P) and the European Regional Development Fund (FEDER). We acknowledge Ignacio Pérez-Pomeda from the Cell Culture Core Facilities of IQAC for his participation in cellular assays and Dr. Yolanda Pérez from the Nuclear Magnetic Resonance Facility of IQAC for performing NMR analyses. Imma Carrera is acknowledged for helping in the surface tension measurements. Jaume Caelles, from the SAXS-WAXS service at IQAC, is acknowledged for technical assistance in the measurements of SAXS. SAXS experiments were performed at NCD beamline at ALBA Synchrotron with the collaboration of ALBA staff (special thanks to Juan Carlos Martínez).

Aim: Globally distributed pelagic fishes are typified by very low to negligible genetic differentiation at oceanic scales arising from high gene flow and (or) large population sizes. However, genomic approaches employing thousands of loci to characterise genetic variation can illuminate subtle patterns of genetic structure and facilitate demographic inference, such that effects arising from gene flow and population size can be partially decoupled. We used a population genomics approach to identify putative stocks in a circumtropical pelagic fish, wahoo, and to assess global connectivity in this species. Location: Indo-Pacific and Atlantic Oceans. Taxon: Wahoo, Acanthocybium solandri (Cuvier, 1832). Methods: Globally distributed wahoo samples from 11 locations (representing a total of 296 individuals) were sequenced using a pool-seq ezRAD approach to obtain 1289–9825 genome-wide SNP loci per population pair for analyses of genetic structure at a minor allele frequency (MAF) >0.05. Demographic inference using a diffusion approximation method (∂a∂i) was performed using 11,495–12,812 SNPs per population pair at a MAF >0.02. Results: Genetic structure, measured as F , was overall low, suggesting very little heterogeneity amongst sample pairs (pairwise F ≤ 0.021). However, there was a clear signal of regional genetic structuring between ocean basins. A principal coordinate analysis separated samples from the Indo-Pacific with those from the Atlantic and an analysis of molecular variance suggested that ~77% of variation in genetic structure was amongst regions. Our demographic analyses found greater support for models including migration over simple models of isolation. Main conclusions: Our study provides the most thorough genetic investigation of wahoo to date. We provide evidence for global connectivity of wahoo populations over their evolutionary history, but we also provide the first indication of subtle regional structure between the Indo-Pacific and Atlantic Oceans, which occurs against a background of high gene flow. The identification of regional stocks will inform new management strategies and guide future investigations in wahoo, an increasingly important species in global fisheries. ST ST

Rheumatoid arthritis (RA) is characterized by the presence of autoantibodies that are of paramount importance for the diagnosis and prognosis of the disease and have been implicated in its pathogenesis. Proteins resulting from post-translational modifications (PTMs) are capable of triggering autoimmune responses important for the development of RA. In this work, we investigate serum antibody reactivity in patients with an established RA against a panel of chimeric peptides derived from fibrin and filaggrin proteins and bearing from one to three PTMs (citrullination, carbamylation and acetylation) by home-designed ELISA tests (anti-AMPA autoantibodies). The role of anti-AMPAs as biomarkers linked to the presence of a more severe RA phenotype (erosive disease with radiological structural damage) and to the presence of interstitial lung disease (ILD), a severe extra-articular manifestation in RA patients entailing a high mortality, was also analyzed. In general, the association with the clinical phenotype of RA was confirmed with the different autoantibodies, and especially for IgA and IgM isotypes. The prevalence of severe joint damage was only statistically significant for the IgG isotype when working with the peptide bearing three PTMs. Furthermore, the median titers were significantly higher in patients with RA-ILD, a finding not observed for the IgG isotype when working with the single- and double-modified peptides., This research was funded by the Spanish Ministry of Economy, Industry and Competitiveness, and the European Regional Development Fund (Grant No. RTI2018-094120-B-I00)

Background Blocking of the Tumor Necrosis Factor (TNF) activity is a successful therapeutic approach for 50–60% of rheumatoid arthritis (RA) patients. However, there are yet no biomarkers to stratify patients for anti-TNF therapy. Rheumatoid factor (RF) and anti-cyclic-citrullinated antibodies (anti-CCP) have been evaluated as biomarkers of response but the results have shown limited consistency. Anti-carbamylated protein (anti-CarP) and anti-peptidylarginine deiminase type 4 (anti-PAD4) antibodies have been much less studied. Despite being linked to common immune processes, the interaction between these markers has not been evaluated yet. Our aim was to analyze the interaction between these four antibodies in relation to the response to anti-TNF therapy. Methods For this objective, a prospective cohort of n = 80 RA patients starting anti-TNF therapy was recruited. Serum determinations at baseline were performed for RF, anti-CCP, anti-CarP and anti-PAD4 antibodies using enzyme-linked immunosorbent assays (ELISA). The clinical response to anti-TNF therapy was determined at week 12 using the change in DAS28 score. Association was performed using multivariate linear regression adjusting for baseline DAS28, sex and age. Results The interaction between pairs of antibodies was tested by the addition of an interaction term. We found two highly significant antibody interactions associated with treatment response: anti-CarP with anti-PAD4 (p = 0.0062), and anti-CCP with RF (p = 0.00068). The latter antibody interaction was replicated in an independent retrospective cohort of RA patients (n = 199, p = 0.04). Conclusions The results of this study suggest that antibody interaction effects are important factors in the response to anti-TNF therapy in RA., This project was supported by UCB Pharma. The funders had no role in the study design, data analysis, data interpretation or writing the manuscript.

[Introduction]: Palindromic rheumatism (PR) is a form of relapsing/remitting arthritis that may evolve to chronic rheumatic disease, mainly rheumatoid arthritis (RA). The exact nature of PR is unclear, as it may be considered a disease in itself, an abortive form of RA or just a pre-RA stage. [Areas covered]: The authors review the most relevant epidemiological and clinical aspects of PR, especially the pathogenetic role of autoimmunity in PR, with most patients having a characteristic autoantibody profile similar to that observed in RA. The role of autoinflammation is also discussed. A literature review on the rate of RA progression and its prognostic factors was analyzed. Data on the efficacy of drug therapies used to treat PR are presented. PubMed was searched using the terms ‘palindromic rheumatism’ OR ‘palindromic arthritis’. [Expert opinion]: PR is a disease entity with a close but unclear relationship with RA. In PR there is an unmet need, which is to clarify the clinical spectrum and elucidate the risk factors for evolution to RA. The role of autoimmunity and the autoinflammatory component should be investigated. Since most patients evolve to RA, PR may display a unique therapeutic opportunity to avoid this evolution., This paper was not funded. Some of the studies described in this review were supported by two grants from Hospital Clínic de Barcelona, Research, Innovation and Education Department (Emili Letang 052012 and; 37933) and the Spanish Ministry of Economy, Industry and Competitiveness and the European Regional Development Fund (Grant # RTI2018-094120-B-I00 to IH).

[Background]: A restricted response against citrullinated peptides/proteins, with less isotype usage, has been found in palindromic rheumatism (PR) in comparison with rheumatoid arthritis (RA). We hypothesized that this different antibody response may be observed for other post-translational modified proteins. We compared the prevalence and isotype usage of two specificities of anti-carbamylated peptide/protein antibodies (Anti-CarP) in patients with PR and RA., [Methods]: Cross-sectional study including 54 patients with pure PR and 53 patients with RA, matched by sex, age, disease duration and ACPA. Anti-CarP specificities were determined by home-made enzyme-linked immunosorbent assay tests using a synthetic chimeric fibrin/filaggrin homocitrullinated peptide (CFFHP) and fetal calf serum (FCS) homocitrullinated protein as antigens. IgG, IgA and IgM isotypes were measured., [Results]: Anti-CarP were positive (CFFHP or FCS) in 24% and 64% of patients with PR and RA, respectively (p, [Conclusion]: Anti-CarP are found in patients with PR but in a lower proportion and with a different isotype usage from in RA, suggesting a distinct B cell response to homocitrullinated antigens in PR., The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This study was supported by the Hospital Clinic of Barcelona, Research, Innovation and Education Department (Grant # 37933 to RC) and the Spanish Ministry of Economy, Industry and Competitiveness and the European Regional Development Fund (Grant # RTI2018-094120-B-I00 to IH).

© 2020 by the authors., The effective use of fusion inhibitor peptides against cervical and colorectal infections requires the development of sustained release formulations. In this work we comparatively study two different formulations based on polymeric nanoparticles and lipid vesicles to propose a suitable delivery nanosystem for releasing an HIV-1 fusion inhibitor peptide in vaginal mucosa. Polymeric nanoparticles of poly-d,l-lactic-co-glycolic acid (PLGA) and lipid large unilamellar vesicles loaded with the inhibitor peptide were prepared. Both formulations showed average sizes and polydispersity index values corresponding to monodisperse systems appropriate for vaginal permeation. High entrapment efficiency of the inhibitor peptide was achieved in lipid vesicles, which was probably due to the peptide’s hydrophobic nature. In addition, both nanocarriers remained stable after two weeks stored at 4 °C. While PLGA nanoparticles (NPs) did not show any delay in peptide release, lipid vesicles demonstrated favorably prolonged release of the peptide. Lipid vesicles were shown to improve the retention of the peptide on ex vivo vaginal tissue in a concentration sufficient to exert its pharmacological effect. Thus, the small size of lipid vesicles, their lipid-based composition as well as their ability to enhance peptide penetration on vaginal tissue led us to consider this formulation as a better nanosystem than polymeric nanoparticles for the sustained delivery of the HIV-1 fusion inhibitor peptide in vaginal tissues., This work was supported by grant RTI2018-094120-B-I00 from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) and the European Regional Development Fund.

Objective: To analyse the association between anti-carbamylated protein antibodies (Anti-CarP) and interstitial lung disease (ILD) in rheumatoid arthritis (RA) patients. Methods: Cross-sectional study including RA patients fulfilling the 2010 ACR/EULAR criteria. The main population comprised two groups: (1) RA patients diagnosed with RA-ILD (RA-ILD group); (2) RA patients without ILD (non-ILD RA group). Non-ILD RA patients in whom ILD was suspected underwent a diagnostic work-up and, if ILD was diagnosed, were switched to the RA-ILD group. ILD was diagnosed by high-resolution computed tomography and confirmed by a multidisciplinary committee. An independent replication sample was also obtained. Three Anti-CarP IgG autoantibodies against fetal calf serum (Anti-FCS), fibrinogen (Anti-Fib) and chimeric fibrine/filagrine homocitrullinated peptide (Anti-CFFHP) and one Anti-CarP IgA against FCS (Anti-FCS-IgA) were determined by home-made ELISA. Associations between Anti-CarP and ILD were analysed using multivariable logistic regression adjusted by smoking, sex, age, RA disease duration, rheumatoid factor and anticitrullinated protein antibodies. Results: We enrolled 179 patients: 37 (21%) were finally diagnosed with RA-ILD. Anti-CarP specificities were more frequent in RA-ILD patients (Anti-FCS 70% vs 43%; Anti-Fib 73% vs 51%; Anti-CFFHP 38% vs 19%; Anti-CarP-IgA 51% vs 20%, p, Financial support from the Hospital Clinic of Barcelona, Research, Innovation and Education Department (Grant # 37 933 to RC-M and the Spanish Ministry of Economy, Industry and Competitiveness and the European Regional Development Fund (Grant # RTI2018-094120-B-I00 to IH).

Based on the structure of an HIV-1 entry inhibitor peptide two stapled- and a retro-enantio peptides have been designed to provide novel prevention interventions against HIV transmission. The three peptides show greater inhibitory potencies in cellular and mucosal tissue pre-clinical models than the parent sequence and the retro-enantio shows a strengthened proteolytic stability. Since HIV-1 fusion inhibitor peptides need to be embedded in the membrane to properly interact with their viral target, the structural features were determined by NMR spectroscopy in micelles and solved by using restrained molecular dynamics calculations. Both parent and retro-enantio peptides demonstrate a topology compatible with a shared helix–turn–helix conformation and assemble similarly in the membrane maintaining the active conformation needed for its interaction with the viral target site. This study represents a straightforward approach to design new targeted peptides as HIV-1 fusion inhibitors and lead us to define a retro-enantio peptide as a good candidate for pre-exposure prophylaxis against HIV-1., Financial support from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) and the European Regional Development Fund (Grant RTI2018-094120-B-I00 to I.H.) is gratefully acknowledged. A.M. is supported by a grant SAF2016-75505-R (AEI/MINECO/FEDER, UE) and through the “María de Maeztu” Programme for Units of Excellence in R&D (MDM-2014-0370). The 500-MHz spectrometer was purchased in part through a Research Infraestructure MINECO-FEDER fund (Grant CSIC13-4E-2076). Partial financial support from the Spanish National Research Council (CSIC) is also gratefully acknowledged.

Despite the great effort to decrease the HIV infectivity rate, current antiretroviral therapy has several weaknesses; poor bioavailability, development of drug resistance and poor ability to access tissues. However, molecules such as peptides have emerged as a new expectative to HIV eradication. The vaginal mucosa is the main spreading point of HIV. There are natural barriers such as the vaginal fluid which protects the vaginal epithelium from any foreign agents reaching it. This work has developed and characterized Nanoparticles (NPs) coated with glycol chitosan (GC), loaded with an HIV-1 inhibitor peptide (E2). In vitro release and ex vivo studies were carried out using the vaginal mucosa of swine and the peptide was determined by HPLC MS/MS validated method. Moreover, the peptide was labeled with 5(6)-carboxyfluoresceine and entrapped into the NPs to carried out in vivo studies and to evaluate the NPs penetration and toxicity in the vaginal mucosa of the swine. The mean size of the NPs, ξ and the loading percentage were fundamental features for to reach the vaginal tissue and to release the peptide within intercellular space. The obtained results suggesting that the fusion inhibitor peptides loaded into the NPs coated with GC might be a new way to fight the HIV-1, due to the formulation might reach the human epithelial mucosa and release peptide without any side effects.

The work reports the design and synthesis of a chimeric peptide that is composed of the peptide sequences of two entry inhibitors which target different sites of HIV-1 gp41. The chimeric peptide offers the advantage of targeting two gp41 regions simultaneously: the fusion peptide and the loop both of which are membrane active and participate in the membrane fusion process. We therefore use lipid raft-like liposomes as a tool to specifically direct the chimeric inhibitor peptide to the membrane domains where the HIV-1 envelope protein is located. Moreover, the liposomes that mimic the viral membrane composition protect the chimeric peptide against proteolytic digestion thereby increasing the stability of the peptide. The described liposome preparations are suitable nanosystems for managing hydrophobic entry-inhibitor peptides as putative therapeutics.

We recently described that the presence of various anti-CarPs exhibited a consistent effect toward increasing the odds of ILD in RA. The anti-CarP-IgG isotype was most prevalent; however, anti-CarP-IgA had the highest OR (4.3),3 fueling the hypothesis that RA-related antibodies may originate in the respiratory mucosa, as suggested by Solomon et al. The high proportion of positive IgA-ACPA in IPF (21.3%-24.8%) in their study is noteworthy, in our opinion. This finding may be partially explained by the enzyme-linked immunosorbent assay test sensitivity considering that the proportion of positive IgA-ACPA in the healthy control subjects is also clearly higher (5.6%) than that observed in previous reports (< 2%).

Background: Anti-carbamylated protein antibodies (Anti-CarP) represent a novel autoantibody family present in sera of patients with rheumatoid arthritis (RA) with high specificity. Objectives: To analyse the prevalence of Anti-CarP in an established RA cohort and evaluate their association with the presence of other autoantibodies and disease activity and severity. Methods: Cross-sectional study. Presence of Anti-CarP was analyzed in a cohort of patients with established RA (n: 158) by a home-made ELISA test using fetal calf serum. We investigated the demographic, radiological and current and at disease onset clinical features. Rheumatoid factor (RF) and anti-citrullinated peptide antibodies (ACPA) status were also assessed. Results: Anti-CarP were positive in 46.2% of the patients in our cohort and in 15.1%, 19.2% and 9.1% of the patients negative for ACPA, RF and both respectively. Demographic and clinical features are shown in table 1. The mean titer of ACPA (1361±1054 U/mL vs. 918 ± 1092 U/mL) and RF (258 ± 255 AU vs. 174 ± 205 AU) were significantly higher in Anti-CarP positive patients. No difference in symptoms at RA onset was observed. Current (24.7% vs. 11.8%) and previous (54.8% vs. 37.6%) smoking consumption were significantly associated with Anti-CarP antibodies. Anti-CarP positive patients presented a higher CRP values (1. 0±1.6 mg/dL vs. 0.5 ±0.8 mg/dL), swollen joint count (1.3±2.2 vs 0.7 ±1.8) and patient global health assessment VAS (34.8 ±21.5 mm vs. 26.4 ±17.1 mm); although no between-group differences in disease activity (DAS28, CDAI, SDAI and RAPID3) or demographic features were observed. Anti-CarP positive patients presented significant higher scores in Larsen index (23.6 ± 15.7 vs. 15.7 ± 12.6) and disability (HAQ-DI >1) (23.3% vs. 8.3%). Conclusion: Anti-CarP were present in approximately half of the RA cohort and were also detected in seronegative (RF and/or ACPA) patients. In our cohort, patients with Anti-CarP antibodies presented higher tobacco consumption and poorer disease outcomes. Disclosure of Interests: Raul Castellanos-Moreira Speakers bureau: For Lilly and Merk Sharp and Dohme, Sebastian C Rodriguez-Garcia: None declared, Virginia Ruiz: None declared, Oscar Camacho: None declared, Julio Ramirez: None declared, Andrea Cuervo: None declared, Cristina Garcia-Moreno: None declared, Rosa Morla: None declared, Jose Gomez Puerta Speakers bureau: BMS, Pfizer, Amgen, Juan D. Canete: None declared, Isabel Haro: None declared, Raimon Sanmarti Speakers bureau: PFIZER, SANOFI, LILLY, MSD, UCB, NOVARTIS, JANSSEN

Anti-citrullinated peptide/protein antibodies (ACPAs) are the most specific serological biomarkers for rheumatoid arthritis (RA). They have both diagnostic and prognostic value, and are related to more aggressive joint disease in RA. However, a single biomarker cannot differentiate RA subtypes. So, simultaneous analysis of target citrullinated peptides, using a multiplex array based on chimeric peptides composed of several domains of human proteins, could be useful. In this work, eight chimeric peptides and the corresponding native arginine-containing control peptides were obtained by solid-phase peptide synthesis. The study included RA and psoriatic arthritis (PsA) patients attending the Rheumatology Unit of the Hospital Clinic in Barcelona, as well as healthy blood donors (BD) at the same hospital. Our main aim was to explore the diagnostic value of the novel multiplex array compared to a commercial ELISA-based ACPA assay in a serum-saving way. Using the combination of the eight chimeric peptide antigens in the multiplex array, 61.4% of the RA cohort were positive for 3 or more peptides; while, the healthy BD and PsA cohorts did not show any reactivity with the tested peptides. These results indicate that we have developed a highly specific multiplex assay based of chimeric citrullinated peptides derived from filaggrin, fibrin, vimentin and human enolase proteins for the detection of ACPAs in a serum-saving way., IH, MJG and MJB received financial support from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) and the European Regional Development Fund (Grant CTQ2015-63919-R). MJG, MJB, RS and IH are members of the consolidated PEP-REU research group of the Generalitat de Catalunya (expedient: 2017 SGR 254). MJG, RS and IH belong to the Red Temática de Investigación Cooperativa en Salud (RETICS): “Red de Investigación en Inflamación y Enfermedades Reumáticas” (RIER) financed by the Instituto de Salud Carlos III (Grant RD12/0009/0016).

Autosomal dominantly inherited genetic disorders such as corneal dystrophies are amenable to allele-specific gene silencing with small interfering RNA (siRNA). siRNA delivered to the cornea by injection, although effective, is not suitable for a frequent long-term treatment regimen, whereas topical delivery of siRNA to the cornea is hampered by the eye surface's protective mechanisms. Herein we describe an attractive and innovative alternative for topical application using cell-penetrating peptide derivatives capable of complexing siRNA non-covalently and delivering them into the cornea. Through a rational design approach, we modified derivatives of a cell-penetrating peptide, peptide for ocular delivery (POD), already proved to diffuse into the corneal layers. These POD derivatives were able to form siRNA-peptide complexes (polyplexes) of size and ζ-potential similar to those reported able to undergo cellular internalization. Successful cytoplasmic release and gene silencing in vitro was obtained when an endosomal disruptor, chloroquine, was added. A palmitoylated-POD, displaying the best delivery properties, was covalently functionalized with trifluoromethylquinoline, an analog of chloroquine. This modified POD, named trifluoromethylquinoline-palmitoyl-POD (QN-Palm-POD), when complexed with siRNA and topically applied to the eye in vivo, resulted in up to 30% knockdown of luciferase reporter gene expression in the corneal epithelium. The methods developed within represent a valid standardized approach that is ideal for screening of a range of delivery formulations. © 2019 The Authors, This work was supported by the United Kingdom Fight for Sight grant (to C.B.T.M.), The Belfast Association for the Blind (S.D.A., M.A.N., and T.M.), and Northern Ireland Clinical Research Network Vision Research Translation Research Group (M.A.N. and T.M.). Partial financial support from the Spanish Ministry of Economy, Industry and Competitiveness (MINECO) and the European Regional Development Fund (grant CTQ2015-63919-R to I.H.) is also gratefully acknowledged.

Objective To analyse the prevalence of pre-existing palindromic rheumatism (PR) in patients with established rheumatoid arthritis (RA) and evaluate whether these patients have a distinctive clinical and serological phenotype. Methods Cross-sectional study in patients with established RA. Pre-existing PR was determined using a structured protocol and confirmed by retrospective review of medical records. Demographic, clinical, radiological, immunological and therapeutic features were compared in patients with and without PR. Results 158 patients with established RA (78% female) with a mean disease duration since RA onset of 5.1 ± 2.7 years were included. Pre-existing PR was recorded in 29 patients (18%). The median time from the onset of PR to progression to RA was 1.2 years. No between-group differences in demographic features, current disease activity radiographic erosive disease or disability were observed. Patients with PR had a higher prevalence of smoking (72% vs. 40%). Positive rheumatoid factor, anti-citrullinated peptide antibodies and anti-carbamylated protein antibodies were numerically higher in patients with PR. No differences in treatment were observed except for greater hydroxychloroquine use in patients with PR (38% vs. 6%). Palindromic flares persisted in a significant proportion of patients during the RA course, including patients in clinical remission or receiving biological DMARDs. Conclusion Eighteen percent of patients with RA had a history compatible with PR previous to RA onset. No specific clinical or serological phenotype was identified in these patients, although higher hydroxychloroquine use and smoking prevalence were identified. Palindromic flares may persist during the RA disease course despite treatment.

Background: A slower progression of AIDS and increased survival in GBV-C positive individuals, compared with GBV-C negative individuals has been demonstrated; while the loss of GBV-C viremia was closely associated with a rise in mortality and increased progression of AIDS. Following on from the previous reported studies that support the thesis that GBV-C E2 interferes with HIV-1 entry, in this work we try to determine the role of the GBV-C E1 protein in HIV-1 inhibition. Methods: The present work involves the construction of several overlapping peptide libraries scanning the GBV-C E1 protein and the evaluation of their anti-HIV activity. Results: Specifically, an 18-mer synthetic peptide from the GBV-C E1 protein, E1(139-156), showed similar antiviral activity against HIVs from viruses from clades A, B, C, D and AE. Competitive ELISA using specific gp41-targeting mAbs, fluorescence resonance energy transfer as well as haemolysis assays demonstrated that this E1 peptide sequence interacts with the highly conserved N-terminal region of the HIV-1 gp41 (the fusion peptide) which is essential for viral entry. Conclusions: We have defined a novel peptide lead compound and described the inhibitory role of a highly conserved fragment of the E1 protein. General Significance: The results together allow us to consider the non-pathogenic E1 GBV-C protein as an attractive source of peptides for the development of novel anti-HIV therapies.

Background:Blocking of the Tumor Necrosis Factor (TNF) activity is a successful therapeutic approach for 2 out of 3 Rheumatoid Arthritis patients. Identifying the patients that will not respond to this therapeutic approach is a major translational goal in RA. Association of seropositivity to rheumatoid factor (RF) or anti-cyclic-citrullinated antibodies (anti-CCP) with anti-TNF response has proven inconclusive, suggesting that other yet unexplored biomarkers could be more informative for this goal.Objectives:We tested the association of two recently introduced biomarkers in RA: anti-carbamylated protein antibodies (anti-CarP) and anti-peptidylarginine deiminase type 4 (anti-PAD4).Methods:A prospective cohort of n=80 RA patients starting anti-TNF therapy was recruited and levels for all four autoantibodies -RF, anti-CCP, anti-CarP and anti-PAD4- were measured at baseline. The change in DAS28 score between baseline and week 12 of therapy was used as the clinical endpoint.Results:Single marker-analysis showed no significant association with drug response. However, when testing for interactions between autoantibodies, we found highly significant associations with drug response. Anti-CCP and RF showed a positive interaction with the response to anti-TNF therapy (P=0.00068), and anti-PAD4 and antiCarP titers showed a negative interaction with the clinical response at week 12 (P=0.0062). Using an independent retrospective sample (n=199 patients), we validated the interaction between anti-CCP and RF with the clinical response to anti-TNF agents. (P=0.044).Conclusion:The results of this study show that interactions between antibodies are important in the response to anti-TNF therapy and suggest potential pathogenic relationships.Acknowledgments :We would like to thank the clinical researchers and patients participating in the IMID Consortium for their collaborationDisclosure of Interests:Antonio Julià: None declared, Maria Lopez Lasanta: None declared, Francisco Blanco: None declared, Antonio Gómez: None declared, Isabel Haro: None declared, Antonio Juan Mas: None declared, Alba Erra: None declared, Mª Luz García Vivar: None declared, Jordi Monfort: None declared, Simon Sánchez Fernandez: None declared, Isidoro González-Álvaro Grant/research support from: Roche Laboratories, Consultant of: Lilly, Sanofi, Paid instructor for: Lilly, Speakers bureau: Abbvie, MSD, Roche, Lilly, Mercedes Alperi-López: None declared, Raúl Castellanos: None declared, Antonio Fernandez-Nebro: None declared, Cesar Diaz Torne: None declared, Núria Palau: None declared, Raquel M Lastra: None declared, Jordi Lladós: None declared, Raimon Sanmarti: None declared, Sara Marsal: None declared

Background:Interstitial lung disease (ILD) is associated with a significant increase in morbidity and mortality in patients with rheumatoid arthritis (RA). Therefore, an early diagnosis is fundamental. Anti-carbamylated proteins (Anti-CarP) have been described in different chronic respiratory diseases without a previous history of RA.Objectives:The aim of this study was to analyse the association between Anti-CarP and ILD in RA patients.Methods:We performed a cross-sectional study, including RA patients fulfilling the 2010 ACR/EULAR criteria. The main population comprised 2 groups: 1) RA patients diagnosed with ILD (RA-ILD group) and 2) RA patients without ILD (non-ILD RA group). ILD was diagnosed by high-resolution tomography and confirmed by a multidisciplinary committee. Three IgG Anti-CarP autoantibodies against fetal calf serum (Anti-FCS), fibrinogen (Anti-Fib), and fibrine/filagrine homocitrullinated peptide (Anti-CFFHP) and one IgA against FCS (Anti-FCS-IgA) were determined by home-made ELISA. Associations between Anti-CarPs and ILD were explored using multivariable logistic regression adjusted by a set of variables known to be related to the development of ILD: smoking, sex, age, RA disease duration, RF and ACPA. An independent replication sample was obtained to validate our findings from another hospital.Results:The main population included 179 patients: 37 were included in the RA-ILD group, and 142 in the non-ILD RA group. Most patients were female (79%), with a mean age of 59.7±13 years with a mean disease duration of 6.6±5 years. Baseline features are shown in table 1. The replication sample was composed of 25 patients in the RA-ILD group and 50 patients in the non-ILD RA group. We found that Anti-CarPs specificities were more frequent in RA-ILD patients (Anti-FCS 70% vs. 43%; Anti-Fib 73% vs. 51%; Anti-CFFHP 38% vs. 19%; Anti-CarP-IgA 51% vs. 20%, pTABLE 1.Main population demographic, clinical, therapeutic, and autoantibody status features.RA-ILDn:37Non-ILD RAn:142p valueFemale (%)25 (68)116 (82)NSAge mean (±SD)67.3 (10.1)57.7 (12.9)Mean disease duration (±SD)11.6 (7.1)5.3 (13.3)Ever smokers (%)21 (57)62 (44)NSSmoking cumulative dose (±SD)30.7 (11.1)21.8 (12)Caucasian (%)31 (84)120 (85)NSTreatmentGlucocorticoids (%)25 (68)81 (57)NScsDMARDs (%)33 (89)132 (86)NSMTX (%)20 (54)95 (67)NSbDMARDs (%)11 (30)36 (25)NSMean DAS28 (±SD)3.71 (1.35)2.74 (1.05)Erosive disease (%)26 (70)63 (44)Mean HAQ-DI (CI-95%)0.69 (0.53-0.85)0.31 (0.24-0.38)ACPA positive (%)29 (78)99 (70)NSMedian titer ACPA (IQR) CU674 (2,215)143 (1,132)NSRF positive (%)28 (76)83 (59)NSMedian titer RF (IQR) IU105 (298)34 (110)NSConclusion:A strong association between RA-ILD and Anti-CarP was found independently of cofounders, including RF and ACPA. Our findings suggest a possible link between Anti-CarP and the development of ILD.Disclosure of Interests:Raul Castellanos-Moreira Speakers bureau: Lilly, MSD, Sanofi, UCB, Sebastian Rodriguez-Garcia: None declared, Katherine Cajiao: None declared, Gabriela Jimenez: None declared, Maria Jose Gomara: None declared, Virginia Ruiz Speakers bureau: Lilly, Pfizer, Ivette Casafont-Solé: None declared, Julio Ramirez: None declared, José Gomez Puerta Speakers bureau: Abbvie, Eli Lilly, BMS, Roche and Pfizer, Susana Holgado Pérez: None declared, Juan de Dios Cañete: None declared, Isabel Haro: None declared, Raimón Sanmartí Speakers bureau: Abbvie, Eli Lilly, BMS, Roche and Pfizer

We welcome the comments by Alunno et al on our article about the association between anticarbamylated protein antibody (anti-CarP) specificities and rheumatoid arthritis-associated interstitial lung disease (RA-ILD).1 The authors proposed a ‘mix and match approach’ consisting of the assessment of various antibody specificities at RA diagnosis with the aim of predicting the development of ILD.2 We believe this hypothesis is reasonable, considering that different antimodified protein antibodies (AMPA) including anti-CarP, anticitrullinated and antiacetylated protein antibodies (ACPA and anti-AceP, respectively) have been associated with RA-ILD.1 3 4 Furthermore, a …

Background Anti-carbamylated protein antibodies (anti-CarP) are a newly-identified antibody family present in the sera of rheumatoid arthritis (RA) patients whose specificity is close to that of ACPA but whose sensitivity is lower. Palindromic rheumatism (PR) is an intermittent form of arthritis that may progress to RA. Patients with PR may present the characteristic autoantibody profile seen in RA, even in the absence of evolution to RA. There are no studies on the presence of anti-CarP antibodies in PR. Objectives To analyse the prevalence of anti-CarP in patients with PR and evaluate their clinical phenotype and relationship with ACPA. Methods Patients diagnosed with PR attended by the Arthritis Unit, Rheumatology Service, Hospital Clinic of Barcelona were included according to the criteria of Guerne et al. Only patients considered as pure PR (patients without evidence of RA or other chronic rheumatic disease at the time of the first determination of anti-CarP) were analysed. An established RA control group was included. Anti-CarP were analysed by a home-made ELISA using a fibrin/chimeric filaggrin homocitrulinated peptide as antigen. IgG, IgA and IgM isotypes were measured. ACPA were analysed with a commercial CCP2 test. The clinical and therapeutic characteristics were compared according to anti-CarP positivity/negativity and to the clinical evolution, including the progression to RA during the follow-up. Results Anti-CarP were analysed in 54 patients with pure PR and 53 RA patients. Anti-CarP were detected in 16.7% and 37.7% (p: NS) of patients with PR and RA, respectively. All PR patients positive for anti-CarP were ACPA positive and had higher ACPA titers. IgG was the predominant isotype (100%), only one patient presented IgA (11.1%) and none IgM. In the RA group, the percentages of IgG, IgA and IgM were 55%, 50% and 40%, respectively, in anti-CarP positive patients. More anti-CarP positive patients developed RA (33.3% vs. 13.6%), although the difference was not significant, during a mean follow-up of 3.8±1.1 years. Remission at the last follow-up visit was more frequent in patients who were anti-CarP negative (22% vs. 60.5% p Conclusions This is the first study to demonstrate the presence of anti-CarP in patients with pure PR. Anti-CarP was associated with ACPA positivity and higher ACPA titers. Anti-CarP isotypes differed between PR and RA, with lower isotype use in PR. Anti-CarP positivity in PR patients could indicate a more refractory disease, with lower rates of remission and greater predisposition to evolve to RA. Acknowledgements This study was supported by two awards granted by Hospital Clinic de Barcelona, Research, Innovation and Education Department (Sonia Cabrera-Villalba, Premi Fi de residencia: Emili Letang 052012; Raul Castellanos-Moreira, Premi Fi de residencia: Emili Letang 379_33_PFR_ 250760_2017). Disclosure of Interest None declared