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Evaluation of the biological and therapeutic properties of mesoangioblasts-vessel associated progenitor cells- in the cell based therapy of the cystic fibrosis disease
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Vezzali C, Celesti G, Bonfanti C, Antonini S, Barone C, Pesce E, Tomati V, Maiuri L, Egan M, Pedemonte N, Bruscia E, Messina G
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Novel aminoarylthiazole derivatives as correctors of the chloride transport defect in cystic fibrosis: computer assisted drug design, synthesis and biological evaluation
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Liessi N, Cichero E, Pesce E, D’Ursi P, Salis A, Pedemonte N, Damonte G, Galietta LJV, Fossa P, Millo E
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Author(s)
Lentini L, Pibiri I, Melfi R, Tutone M, Pace A, Barone G, Di Leonardo A
Address
Departmentof Biological, Chemical and Pharmaceutical Sciences and Technologies(STEBICEF), University of Palermo Abstract: Background. Cystic Fibrosis patients with nonsense mutation in the CFTR gene generally make virtually no CFTR protein and thus often have a more severe form of CF. Recently, Ataluren (formerly PTC124) was suggested to induce the read-through of premature termination codons (PTCs) mainly the UGA codon. However, despite promising results there is not a general consensus of Ataluren efficacy and mechanism of action.
Hypothesis and objectives. The design of new small molecules (PTC124 related) together with the understanding of their mechanism of action could lead to new pharmacologic approaches for the cure of CF caused by nonsense mutations in the CFTR gene. This project was aimed to evaluate the activity of some PTC124 analogues identified by a virtual screening, by different orthogonal assays with vectors containing PTCs in reporter genes and in CF bronchial epithelial cells. 
Author(s)
de Jonge H and Caldrer S
Address
Department of Gastroenterology & Hepatology, Erasmus University Medical Center, Rotterdam; Cystic Fibrosis Center, Azienda Ospedaliera Universitaria Integrata Verona Abstract: Background. Most physiological assays used for CF diagnosis and as a biomarker in clinical trials measure CFTR-dependent chloride but not bicarbonate transport. Recent studies however reveal that defective bicarbonate rather than chloride transport is the primary cause of impaired mucociliary clearance in the airways and luminal obstruction in the GI tract in CF.
Hypothesis and objectives. Our major aims were (i) development of new protocols to measure bicarbonate transport in rectal biopsies and intestinal organoids from healthy controls and CF patients; (ii) ex vivo testing of CFTR correctors and potentiators for their ability to restore bicarbonate secretion in these models; (iii) validating the concept that CFTR functional variants associated with pancreatitis and sinusitis but not CF have a specific bicarbonate permeation defect. 
Author(s)
Vezzali C, Celesti G, Bonfanti C, Antonini S, Barone C, Pesce E, Tomati V, Maiuri L, Egan M, Pedemonte N, Bruscia E, Messina G
Address
University of Milan, Dept. of Biosciences, Milan, Italy; Yale University, School of Medicine, New Haven (CT), USA; IERFC, Ospedale San Raffaele, Milan, Italy; Gaslini Institute, Genova, Italy Abstract: Background. Since the CFTR gene was cloned in 1989, several strategies for correction of CF lung disease have been explored. Among these, cell-based approaches are under investigation. Endogenous lung stem and progenitor cells have been studied although their contribution in the amelioration of chronic lung diseases is still debated. Stem cell-based approaches to treat CF have not achieved the efficiencies of delivery and engraftment needed for therapy. The reasons rely in the low cell engraftment in the lungs after systemic administration, in the small percentage of differentiated cells in airway epithelia and in the even less percentage of CFTR expression.
Hypothesis and objectives. During our recent works on a class of mouse progenitor cell derived from vessel, named mesoangioblasts4 (mMABs), we observed that mMABs, when systemically transplanted in healthy mice distribute throughout lung, trachea and intestinal epithelium for up to 2 months.The major aim of this study was the evaluation overtime of the engraftment of adult mMABs transplanted in CF mice, in terms of percentage of engraftment and persistence in time. As a parallel purpose, mMABs have been studied in terms of their ability to differentiate in epithelial cells and to express functional CFTR in transplanted CF mice, thus resulting in a general amelioration of the pathology. The whole study aimed to develop a cell therapy approach for patients affected by CF. 
Author(s)
Venerando A, Villella VR, Cozza G, Esposito S, Maiuri L, Pinna LA
Address
University of Padova and CNR Institute of Neurosciences, European Institute for Research in Cystic Fibrosis. Abstract: Background. CFTR F508 deletion (F508del) is the by far commonest mutation causative of Cystic Fibrosis (CF). F508delCFTR undergoes premature degradation subverting proteostasis regulation and generating fragments which have the potential to up-regulate the protein kinase CK2 that, in turn, can favour CFTR fragmentation and reduce CFTR stability.
Hypothesis & Objectives. The project provide the rationale and proof-of-concept for the design of an original and novel therapeutic strategy that restores CFTR channel function by targeting the specific context in which the mutant CFTR channel fails to traffic to the cell surface. It focuses on the derailed CF proteostatic environment that is driven by the mutant channel, instead of on CFTR protein itself. Our strategy induces a self-sustained positive loop that ameliorates the CF phenotype by inhibiting protein cross linking that blocks autophagy and dampening overactive CK2. The workflow is summarized in the following tasks: 1) Identification and functional characterization of endogenous CK2 targets whose phosphorylation is altered by F508delCFTR; 2) In vivo confirmation of the CK2/CFTR functional link; 3) Analysis of known kinase modulators as a new class of molecules useful to rescue/stabilize F508del-CFTR; 4) In vivo validation of CK2/protein kinases modulators as reagents able to rescue CF phenotypes. 
Author(s)
Liessi N, Cichero E, Pesce E, D’Ursi P, Salis A, Pedemonte N, Damonte G, Galietta LJV, Fossa P, Millo E
Address
University of Genoa, Istituto Giannina Gaslini, Institute for Biomedical Technologies e National Research Council, Italy. Abstract: Background. CF is caused by mutations that abolish the function of CFTR, a protein that is needed to transport chloride and bicarbonate across cell membrane in the epithelial cells. The most frequent mutation, F508del requires a specific pharmacological treatment. Such defects can be addressed with small molecules, known as correctors and potentiators. In previous studies, we identified a class of compounds called aminoarylthiazoles (AATs) that potentially correct the CF basic defect.
Hypothesis and objectives. The main objective of our project is to identify new AATs which could be efficiently able to correct the CFTR protein defect caused by F508del. By using an integrated approach (bioinformatics, chemical synthesis, and functional assays) we will aim at developing drug- like compounds with improved activity on mutant CFTR. 
Author(s)
Sandri A, Bergamini G, Ortombina A, Boschi F, Sorio C, Melotti P, Stellari F, Lleò MM
Address
Università di Verona, Centro Regionale per la Fibrosi Cistica, Chiesi Farmaceutici, Italy Abstract: Background.The possibility of monitoring the inflammatory response in a IL-8 transgenic WT and CFTR knockout (CF), non invasive, animal model has been demonstrated in a previous project conducted by us [1, 2].
Hypothesis and objectives In the past project we shown that the antibiotic azithromycin causes a significant decrease of the lung inflammation in that is capable of inhibiting the synthesis of bacterial virulence factors. Azithromycin is extensively used in cystic fibrosis as an anti-inflammatory molecule but several cases of azithromycin resistance have been reported [1-2], especially in patients using the drug for long times. It seems then important to identify other therapeutic agents to be used in nonresponding patients. 
Author(s)
Mangoni ML, Cappiello F, Casciaro B, Luca V, Chen C, Lin Q, Di PY
Address
Department of Biochemical Sciences, Sapienza University of Rome, Rome-Italy, Department of Environmental and Occupational Health, University of Pittsburgh, USA Abstract: Background. Pseudomonas aeruginosa is the most predominant lung pathogen in patients with cystic fibrosis (CF). It is quite difficult to eradicate because of its acquired resistance to most available antibiotics and ability to form sessile communities or biofilms, in a protective extracellular matrix. Cationic antimicrobial peptides (AMPs) hold promise as future anti-infective agents.
Hypothesis and objectives. Our objective was to develop a short-sized AMP from frog skin, Esc(1-21) and/or its diastereomer containing two D-amino acids, for treatment of P. aeruginosa lung infections. To this aim: (i) an in-vitro analysis of the anti-pseudomonal, anti-inflammatory, wound healing properties of the two peptides, and (ii) preclinical testing to establish both the peptides' safety profiles and therapeutic dosages were performed. 
Author(s)
Berlutti, F.
Address
Dip. di Salute Pubblica e Malattie Infettive - Università La Sapienza, Roma Abstract: Background. In the airways, the CFTR genetic defect leads to dysregulation of inflammatory and iron homeostasis that precedes bacterial infection that, in turn, worsens host damage. Lactoferrin (Lf), an iron-chelating glycoprotein of the innate immunity present in airway secretions and secreted by neutrophils in infection/inflammation sites, in addition to the well-known antibacterial activity exerts a key role in inflammatory and iron homeostasis.
Hypothesis and objectives. We suppose that aerosolized bLf can exert a key role on inflammatory and iron dysregulation and reduce infection in pre-clinical CF mouse models of lung infection. Moreover, since Lf is a natural molecule, it shows no side effects and toxicity. In this study we employed bovine milk derived Lf (bLf) as it has been yet used in several clinical trials. Since in CF airways human and bacterial proteases may alter Lf integrity reducing its activity, we hypothesize that nano-delivery of bLf can protect the molecule against proteolysis. The objectives are: 1- to evaluate anti-inflammatory and antibacterial activities of bLf administered by aerosol in mouse models of chronic lung infection; 2- to prepare bLf-loaded nanoparticles (bLf-NANOs) to be aerosolized.
Author(s)
Pacello F, D’Orazio M, Battistoni A
Address
Department of Biology, University of Rome Tor Vergata, Roma, Italy Abstract: Background. We have shown that the interaction between Burkholderia cenocepacia and lung epithelial cells is mediated by redox sensitive enzymes of the protein disulphide isomerase (PDI) family, in particular by ERp57. In fact, either thiol reactive agents (DTT, GSH, and DTNB) or ERp57 inhibitors are able to drastically reduce the adhesion and the invasion of B. cenocepacia into epithelial cells and to attenuate the pro-inflammatory response elicited by bacteria. These observations suggest that defects in GSH export in the airway surface liquid, observed in cystic fibrosis (CF) patients, may favour B. cenocepacia infection.
Hypothesis and objectives. ERp57 inhibitors, in particular epigallocatechin-3-gallate (EGCG), the most abundant green tea catechin, could be useful in therapies aimed at controlling Burkholderia lung infection in CF patients. To this aim we have evaluated: 1) the effect of EGCG on Burkholderia infection in different cell types; 2) the effect of EGCG on Burkholderia intracellular replication; 3) the efficacy of EGCG in infected mice. 
Author(s)
Ferrara S, Macchi R, Falcone M, Rossi A, Ranucci S, Bragonzi A, Cigana C, Bertoni G
Address
Department of Biosciences, Università degli Studi di Milano, Milan, Italy, Infections and Cystic Fibrosis Unit, Division of Immunology, Transplantation and Infectious Diseases, San Raffaele Scientific Institute, Milan, Italy Abstract: Background. The comprehension of molecular mechanisms underlying CF airways infection by Pseudomonas aeruginosa is instrumental to the design of clinical protocols to prevent and contrast it. Emerging candidates as molecular regulators of infection and virulence in P. aeruginosa are small RNAs (sRNAs), which in other bacterial pathogens have been shown to play key roles in modulating cellular processes linked to pathogenesis.
Hypothesis and objectives. From this perspective, the main aim of this project was the evaluation of the impact on P. aeruginosa virulence and infection of three new sRNAs recently identified in our lab and named ErsA, ReaL and PesA. 
Author(s)
Bevivino A, Mengoni A, Taccetti G, Fiscarelli EV, De Alessandri A
Address
Department for Sustainability of Production and Territorial Systems, ENEA, Italian National Agency for New Technologies, Energy and Sustainable Economic Development, Rome, Italy Abstract: Background. Cystic fibrosis (CF) is characterized by chronic airway infections involving a complex and dynamic microbial lung community. Though several studies have investigated the taxonomic composition of the airway microbiota, little is still known about the overall functional capabilities of the resident microbial populations and their relation to patient’s lung disease status.
Hypothesis and objectives. Previous studies found changes in CF airway microbiota associated with a severe decline in lung function. The overall goal of this project – which follows the FFC#10/2014 project – is to deeply investigate the airway microbiome of stable (S) and substantial decliners patients (SD) and single patients’ dynamics over a 15-month period, to discover the functional traits harbored by microbial communities and new microbiome-based biomarkers, which may be predictive of decline in lung function. 
Author(s)
Corti A, Melotti P, Sorio C, Bergamini G, Hector A, Griese M, Pompella A
Address
University of Pisa, University of Verona, University of Tübingen Children's Hospital, Ludwig-Maximilians-University Munich Abstract: Background. Inhalation treatments with glutathione (GSH) have gained interest among CF patients. GSH is a primary antioxidant whose levels are significantly decreased in lungs during inflammatory processes. However the results attained so far remain disappointing. One of the possible reasons may lie in the fact that CF lungs often present with increased levels of gammaglutamyltransferase (GGT), an enzyme secreted by inflammatory cells, capable of degrading GSH.
Hypothesis and objectives. The hypothesis to be verified was whether the different levels of airways GGT in CF patients might modulate the effects of GSH-based therapies. It is of importance to clarify whether GSH inhalation therapies are truly beneficial, useless or even – in selected conditions – potentially dangerous. 
Author(s)
Gemma S, Brogi S, Brindisi M, Vallone A, De Luca F, Docquier JD
Address
Department of Biotechnology, Chemistry, and Pharmacy, Department of Medical Biotechnology, University of Siena, Italy. Abstract: Background. Pseudomonas aeruginosa is the most dominant pathogen associated with adverse clinical outcomes in patients with cystic fibrosis (CF). Anti-pseudomonal β-lactams are the mainstay of therapy for the treatment of chronic P. aeruginosa lung infections CF patients. Unfortunately, this pathogen becomes increasingly resistant to available antibiotics and the global dissemination of multidrug resistant P. aeruginosa strains with acquired carbapenemase genes is a public health concern.
Hypothesis and objectives. One important mechanism of resistance to β-lactams is the production of one or more β-lactamase(s), belonging to either the active serine enzymes or the metallo-β-lactamases (MBLs), which efficiently inactivate these antibiotics. Acquired MBLs are important resistance factors in P. aeruginosa, conferring resistance to most β-lactams, including the last-resort carbapenems. Considering the increasing prevalence of MBL-producing strains in CF patients (as high as >50% in some settings), our goal is to perform the preclinical investigation and optimization of MBLs inhibitors to be used in combination with currently-available β-lactam antibiotics 
Author(s)
Rimessi A, Patergnani S, Giorgi C, Marchi S, Bonora M, Bragonzi A, Cabrini G, Pinton P
Address
University of Ferrara, Italy, San Raffaele Scientific Institute, University-Hospital of Verona, Italy. Abstract: Background. The chronic airway infection by Pseudomonas aeruginosa (P. aeruginosa) in Cystic Fibrosis (CF) is a pathological manifestation associated with an excessive inflammatory response. The NOD-like receptors (NLRs) are intracellular pattern-recognition receptors that recognize pathogen-associated molecular patterns activating pro-inflammatory response through the assembly of large caspase-1-activating complex, called inflammasome. Recent data suggest that the mitochondria integrate distinct pro-inflammatory signals and relay this information to a key molecular complex related to inflammation, the inflammasome. It has been demonstrated that mitochondria are the main source of inflammasome-activating radical species and as such may constitute the signal-integrating organelle for inflammasome recruitment and activation in P. aeruginosa-driven inflammation in CF. Moreover, in the previous project we have demonstrated that mislocalized CFTR is associated with an increase in intracellular Ca2+ content and that it favored the mitochondrial Ca2+ uptake, predisposing the organelle to a major stress responsivity.
Hypothesis and objectives. The overall goal of this project is to broaden our knowledge on role of mitochondria to decode and integrate the pro-inflammatory signals generated during P. aeruginosa infection in CF. We have performed experiments aimed at obtaining a deeper insight into the complex mechanism that controls the mitochondrial homeostasis, in particular the Ca2+ signal, and the inflammasome activation induced by pathogen in CF 
Author(s)
Bosso A, Pirone L, Di Gaetano S, Pedone E, Notomista E, Pizzo E
Address
Department of Biology, University Federico II, Naples, Italy, IBB, CNR, 80134, Naples, Italy. Abstract: Background. Host defence peptides (HDPs) are short cationic molecules produced by the immune systems of most multicellular organisms. They exhibit a wide range of biological activities from direct killing of invading pathogens to modulation of immunity. Chronic infection with P. aeruginosa is the main proven perpetrator of lung function decline and ultimate mortality in CF patients. HDPs, acting as anti-inflammatory molecules, could be able to block LPS pro-inflammatory activity attenuating inflammation and so limiting damage to host tissues. New potential HDPs have been identified by an in silico method and their biochemical and pharmacological features are under investigation.
Hypothesis and objectives. It has been demonstrated that many human proteins, with functions not necessarily related to host defense, are reservoirs of active host defense peptides. The aim of this study was to characterize structural propensity, antimicrobial activity, LPS neutralizing and anti-inflammatory properties of new human HDPs identified by a bioinformatic procedure that we have developed. 
Author(s)
Luini, A.
Address
Centro Nazionale Ricerche, Dip. di Scienze Biomediche, Istituto di Biochimica delle Proteine, Napoli Abstract: Background. Cystic fibrosis (CF) is a frequent and lethal genetic disease caused by mutations associated with the CF transmembrane regulator (CFTR), a chloride channel located in the apical membrane of epithelial cells lining the ducts. In nearly 70% of the CF patients, the mutation involved is a deletion of phenylalanine at position 508 of the protein (DF-CFTR). The mutant protein cannot fold properly leading to its intracellular retention and degradation. Pharmacological screening approaches have identified small molecule “correctors” (which promote a modest level of DF-CFTR arrival at the plasma membrane), some of which are in clinical trials. Unfortunately, their mode of action is not known.
Hypothesis and objectives. We proposed to develop a rational basis for the pharmacological correction of DF-CFTR defects, by characterizing the mechanism of action of correctors, to identify molecular components and pathways involved in the correction and then targeting them by efficient ways. 
Author(s)
Satriano L, Cossu C, Baroni D, Zegarra-Moran O, Ford F, Pollock N, Moran O
Address
Istituto di Biofisica, CNR, Genova, Italy, Unità Operativa di Genetica Medica, Istituto G. Gaslini, Genova, Italy, Faculty of Life Sciences, University of Manchester, Manchester, UK Abstract: Background and rationale. CFTR is an anionic channel expressed in epithelia whose mutations cause cystic fibrosis, but its structure is still unknown.
Hypothesis and objectives. We explored a set of structural parameters of the wild type and mutated (F508del) CFTR by physical methods.
Essential methods. WT and F508del CFTR were over-expressed in yeast, solubilised in the detergent LPG-14 and purified. The detergent-CFTR complexes were studied by SAXS techniques using a solvent of variable density. 
Author(s)
Pagani, F.
Address
Centro Internazionale di Ingegneria Genetica e Biotecnologie - ICGEB, Trieste, Italy Abstract: Background. Aberrant splicing represents a common disease-causing mechanism in CF and exon skipping is one of the most frequent defect. We recently reported that modified U1 core spliceosomal components (named ExSpe U1) , when specifically engineered to bind in the intron downstream the 5'ss rescues splicing of defective exons. This therapeutic activity can be observed in CFTR exon 13 and 5, where the modified U1s efficiently correct different splicing defects present either at the 5'ss or at the exonic splicing regulatory elements.
Hypothesis. The molecular mechanism of ExSpeu1 -mediated splicing enhancement and effect in vivo are largely unknown. 
Author(s)
Prandini P, De Logu F, Fusi C, Provezza L, Nassini R, Montagner G, Materazzi S, Munari S, Gilioli E, Bezzerri V, Finotti A, Lampronti I, Tamanini A, Dechecchi MC, Lippi G, Ribeiro CM, Rimessi A, Pinton P, Gambari R, Geppetti P, Cabrini G
Address
University Hospital of Verona, Italy, University of Florence, Italy; University of Ferrara, Italy; Marsico Lung Institute and University of North Carolina, Chapel Hill, NC, U.S.A. LTTA of the University of Ferrara, Italy. Abstract: Background. Pseudomonas aeruginosa colonization, prominent inflammation with massive expression of the neutrophil chemokine IL-8 and luminal infiltrates of neutrophils are hallmarks of chronic lung disease in Cystic Fibrosis (CF) patients. The nociceptive Transient Receptor Potential Ankyrin 1 (TRPA1) calcium channels have been recently found involved in non-neurogenic inflammation.
Hypothesis and objectives. TRPA1 channels could be involved in the pro-inflammatory signaling pathways activated in CF airway epithelial cells infected by P. aeruginosa. The principal objective is to verify whether TRPA1 channels could be relevant molecular targets for innovative anti-inflammatory therapies in CF patients. 
Author(s)
Forti F, Briani F, Horner D, Ghisotti D
Address
Department of Biosciences, University of Milan, Italy Abstract: Background and rationale. Pseudomonas aeruginosa is the most common pathogen found in the lung of cystic fibrosis patients (CF). The quality of life of CF patients largely depends on the success or failure of antibiotic treatment. Now, the alarming diffusion of isolates of P. aeruginosa multi-resistant to the antibiotics currently in use makes urgently need to develop new antibacterial therapies.
Hypothesis and objectives. Phage therapy, the use of the natural enemies of bacteria, is garnering renewed interest as bacterial resistance to antibiotics becomes widespread. This therapy, used for decades in Eastern Europe, can be considered as a therapeutic alternative or a complementary treatment to antibiotics in curing lung infections in CF patients. 
Author(s)
Landini, P.
Address
Dep. of Biosciences, University of Milan, Italy Abstract: Background. In cystic fibrosis (CF) patients, Pseudomonas aeruginosa-induced pneumonia requires frequent antibiotic treatment. Low sensitivity to many antibiotics by P. aeruginosa has prompted the search for novel drugs aimed at the specific inhibition of pathogenesis-related processes. Antimetabolite drugs, such as the pyrimidine analogue flucytosine (FC), are effective inhibitors of biofilm formation and of virulence factors’ production, and show promising activity against P. aeruginosa infections in animal models. However, their utilization in therapy is hindered by possible toxicity, occurrence of resistance, and by lack of precise information of their mechanism of action.
Hypothesis and objectives. The main hypothesis behind this work was that antimetabolite drugs other than FC could share its properties and affect P. aeruginosa virulence and persistence in the host. In addition, we wanted to establish the specific mode of action of FC, as a starting point towards the development of new drugs sharing the same mechanism of action and showing lower risks of long term toxicity. 
Author(s)
Recchiuti A, Mari VC, Codagnone M, Cianci E, Lamolinara A, Iezzi M, Bragonzi A, Moretti P, Nespoli A, Arita M, Romano M
Address
University of Chieti-Pescara, Chieti, Italy; San Raffaele Scientific Institute, Milan, Italy; RIKEN Center for Integrative Medical Sciences, Kanagawa, Japan Abstract: Background. Endogenous resolution mechanisms that promote return to homeostasis are defective in cystic fibrosis (CF) leaving inflammation unrestrained with consequent loss of respiratory function, disability, and premature death of patients. Resolvin (Rv) D1 is and endogenous pro-resolution mediator that has entered clinical trials for the treatment of inflammatory diseases. Therefore, its therapeutic potential in CF is of wide interest.
Hypothesis and Objectives. Overarching hypotheses tested in this project are that RvD1 represents a potent therapeutic candidate for limiting lung inflammation and enhancing resolution of P. aeruginosa infection in CF. Main objectives are to determine, in preclinical models of CF, if RvD1 a) dampens lung inflammation and damage, b) promotes resolution and clearance of bacterial infection 
Author(s)
Marilena Pariano, Matteo Puccetti and Luigina Romani
Address
Department of Experimental Medicine, University ofPerugia, Perugia, Italy Abstract: Background. In patients with Cystic Fibrosis (CF), the progressive decline of pulmonary function is due to a vicious cycle of airways infection and inflammation. Inflammation can be more damaging than the insult itself if uncontrolled, excessive, or prolonged. Although there is debate over whether inflammation in the CF lung is primary (i.e., caused by CFTR mutations) or secondary to chronic infection, inflammation remains the single most significant contributor to disease progression, and its control is crucial for improving patient outcomes. Building upon the results from our past projects–indicating how the application of a systems biology approach and new findings from the laboratory may translate into the development of new therapeutics and rationales for their use–we have proposed a preclinical evaluation study of anakinra, a recombinant, non-glycosylated version of human IL-1 receptor I antagonist (IL-1Ra) in CF. Since 2001, anakinra has proved to be efficacious in a broad spectrum of auto-inflammatory diseases with a remarkable record of safety.
Hypothesis and Objectives. 1. The relative contribution of different inflammasomes to fungal and bacterial infections and inflammation in CF mice. 2.The evaluation of the efficacy of anakinra in experimental and preclinical models of CF. 3. The definition of the molecular mechanisms underlying anakinra activity. 4. The screening of CF patients for IL-1Ra deficiency and the definition of anakinra-responsive signatures through microarray gene expression profiling. 
Author(s)
Lorè NI, Sipione B, Mott R, Iraqi F, Bragonzi A
Address
Infection and Cystic Fibrosis Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS - San Raffaele Scientific Institute, Milan, Italy, Division of Biosciences, UCL Genetics Institute, London, England, Department of Clinical Microbiology and Immunology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel Abstract: Background. The remarkable heterogeneity among CF patients in the time of onset as well as in the severity of P. aeruginosa lung disease raised the question whether other genetic loci in addition to the CFTR can contribute to the clinical variation. Recently, Collaborative Cross (CC) mouse population has been generated as an innovative and powerful source to model the diversity of the human population.
Hypothesis and objectives. By using the CC lines as a novel and high genetically diverse mouse resource population, this project aims to identify genetic factors that may influence the severity of P. aeruginosa respiratory infections in CF. 
Author(s)
Totani L, Plebani R, Patruno S, Piccoli A, Di Silvestre S, Lanuti P, Recchiuti A, Cianci E, Dell’Elba G, Sacchetti S, Guarnieri S, Mariggiò MA, Mari VC, Anile M, Venuta F, Del Porto P, Moretti P, Prioletta M, Mucilli F, Marchisio M, Pandolfi A, Evangelista V, Romano M
Address
G. D’Annunzio University, Chieti-Pescara, Italy; Italian Institute of Technology, Genoa, Italy; University of Rome “Sapienza” Rome, Italy;"Charles Darwin" Sapienza University, Rome, Italy;S. Liberatore Hospital, Italy. Abstract: Background. Despite the involvement of endothelial cells (EC) in cystic fibrosis (CF) pathogenesis is suggested by (1)the emerging evidence of the risk for cardiovascular events in CF patients as well as by (2) our work showing endothelial dysfunction and its correlation with lung disease in CF patients, EC have not been extensively studied in CF.
Hypothesis and objectives. We hypothesized that the elucidation of CFTR signaling in EC may provide clues for innovative pharmacology for CF. Main objectives of this study were to: 1. Uncover mechanisms of CF endothelial dysfunction as molecular targets for novel therapeutics. 2. Elucidate the clinical relevance of circulating endothelial cells (CEC) and endothelial microvesicles (EMV) in CF. 
Author(s)
Ghigo A, Murabito A, Ren K, Pirozzi F, Ciraolo E, Montresor A, Richter W, Wenzel D, Matthey M, Quinneu NL, Fleischmann B, Gentzsch M, Laudanna C, Hirsch E
Address
University of Torino, Torino, Italy;Federico II University, Naples, Italy; University of South Alabama, Mobile USA; University of Bonn, Germany;University of North Carolina at Chapel Hill, Chapel Hill, USA Abstract: Background and Rationale. The underlying cause of cystic fibrosis (CF) is a mutation in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR), a cyclic AMP (cAMP)-stimulated chloride channel. The consequent CFTR dysfunction primarily affects the respiratory system, where the reduced activity of the channel results in obstruction of small airways and, together with airway inflammation and infections, eventually leads to respiratory failure. A number of CFTR correctors and potentiators have been developed, but their ability to rescue the basic defect of CF is still unsatisfactory.
Hypothesis and Objectives. We previously showed that phosphoinositide 3-kinase γ (PI3K γ) acts as a scaffold protein and negatively regulates cAMP. Here, we hypothesized that targeting PI3K γ scaffold activity enhances cAMP in airway smooth muscle, immune and epithelial cells, leading to concomitant (i) bronchodilation, (ii) anti-inflammatory effects and (iii) CFTR potentiation.
Author(s)
Aureli M, Loberto N, Bassi R, Schiumarini D, Munari S, Valsecchi M, Cantù L, Brocca P, Motta S, Ferrami G, Dechecchi C, Sonnino S
Address
Department of Medical Biochemistry and Translational Medicine, University of Milano, Laboratory of Molecular Pathology-Azienda Ospedaliera Universitaria Integrata di Verona, Italy Abstract: Background. Several studies indicate that sphingolipids (SL) play a regulatory role in airway inflammation, the most critical aspect of CF lung disease. Recently, Ceramide (Cer) derived from glycosphingolipids (GSL) has gained more interest since inhibition of the glucocerebrosidase GBA2 and its down-regulation by siRNA, are associated with a significant reduction of IL-8 after P.aeruginosa (PAO) infection, as well as a reduction of the intrinsic inflammatory state in CF human epithelial bronchial cells (CFhEBC).
Hypothesis and Objectives. In our work hypothesis, the aberrant inflammatory response to PAO in CFhEBC starts from alterations in the lipid composition of specific plasma membrane (PM) macromolecular complex by the action of the GSL-hydrolases associated with the cell surface, including GBA2. To figure out the possible molecular mechanism we investigated the effect of PAO infection on specialized membrane area called lipids rafts. Moreover, with the aim to develop a new anti-inflammatory treatment we developed new lipid based-nanoparticles for the delivery of siRNA targeting GBA2. 
Author(s)
Braggion C, and Terlizzi V
Address
Dipartimento di Medicina Pediatrica, Centro Regionale Fibrosi Cistica - Ospedale dei Bambini A. Meyer, Firenze Abstract: Background. A clinical guideline (LG) is a document with the aim of guiding decisions and criteria regarding diagnosis, management, and treatment in specific areas of healthcare; also in the Cystic Fibrosis (CF) world, scientific societies and organizations publish LG to give recommendations; since guidelines may have both methodological problems and conflict of interest our strategy is needed to choose which guidelines should be implemented.
Aims. Evaluate the reliability of every single guideline produced within the CF; identify the accurate documents and to implement its knowledge among the healthcare profession; produce a reasoned synopsis of the existing recommendations.
Author(s)
Pilette C, and De Rose V
Address
Université Catholique de Louvain, Brussels, Dip. di Scienze Biologiche e Cliniche, Università di Torino, Italy. Abstract: Background. Cystic fibrosis (CF) represents the most common lethal autosomal recessive disorder in the white population, mainly affecting the lungs. It affects the Cystic Fibrosis Transmembrane Regulator (CFTR) gene, which encodes a protein expressed on the apical membrane of airway epithelial cells, where it acts as a cAMP-dependent chloride channel and regulator of other channels, including the epithelial Na+ channel (ENaC). Airway colonization of the airways and lung infections are a hallmark of this disease, in particular with Pseudomonas aeruginosa (PA) that affects '70% of CF patients and is associated with poor clinical outcomes. However, the mechanisms underlying the persistence of pathogens in CF airways, remain largely unclear. Secretory-immunoglobulin A (S-IgA) is a major line of mucosal defense, through so-called immune exclusion of inhaled / ingested antigens and pathogens. Following synthesis of polymeric (mainly dimeric) IgA by subepithelial mucosal plasma cells, IgA is transported across the epithelium by a transcellular routing mediated by the polymeric immunoglobulin receptor (pIgR).
Hypothesis and objectives. This project aims to investigate whether the production of secretory IgA (S-IgA) is impaired in the CF lung, through which mechanisms, and whether this defect contributes to the pathogenesis of CF by impairing immunoprotection against respiratory pathogens such as PA. 
Author(s)
Tortoli E, Cariani L, Di Serio C, Niemann S
Address
Istituto Scientifico San Raffaele, Milano; Università Vita-Salute San Raffaele, Milano, Italy. Abstract: Background. M. abscessus (MA) is a species frequently isolated from cystic fibrosis (CF) patients. Three Subspecies (subsp.) of MA exist and their identification is crucial for epidemiology and optimal patient’s management. Although the environment is considered the source of MA infections, its transmission between CF patients has been hypothesized.
Objectives. We aimed to investigate a large collection of MA isolates obtained from CF patients to increase present knowledge about the prevalence of the 3 subsp., their antimicrobial susceptibility, the presence of MA genotypes characterized by increased virulence and responsible for severe clinical manifestations, the transmissibility between patients of the infection. 
Author(s)
Cirilli N, Raia V, De Gregorio F, Di Pietro M, Tosco A, Salvadori L, Sepe A, Buzzetti R, Minicuci N, Rocco I
Address
1Cystic Fibrosis Referral Care, Ancona, Italy; University “Federico II”, Naples, Italy; CNR, Padova, Italy Abstract: Background. The sweat test (ST) remains the main test for Cystic fibrosis (CF) diagnosis and it is now also used to evaluate the function of the CFTR protein in basal conditions and under the influence of CFTR modulators. However it is not yet well-defined the biological variability of the test, both among different individuals (inter-individual) and within the same individual at different times and under different conditions (intra-individual).
Hypothesis and objectives. The project aims to study intra-individual sweat chloride (Cl) biological variability both in subjects with CF and in healthy people and to assess its possible correlation to different variables (diet, seasons, menstrual cycle). 
Author(s)
Tomei P, Masola V, Granata S, Chilosi M, Lupo A, Zaza G
Address
Renal Unit, Department of Medicine, University of Verona, Department of Pathology and Diagnostics, Laboratory of Molecular Pathology, University-Hospital of Verona, Italy. Abstract: Background. MTOR-inhibitors (mTOR-I), Everolimus (EVE) and Sirolimus, immunosuppressants broadly used in transplantation, may determine severe adverse events including pulmonary fibrosis.
Hypothesis and objectives. The pathogenic mechanism of mTOR-I-associated pulmonary toxicity is still unclear, but epithelial to mesenchymal transition (EMT) of bronchial/pulmonary cells may have a role.
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