VP6-SUMO Self-Assembly as Nanocarriers for Gastrointestinal Delivery

High proteolytic degradation and poor absorption through epithelial barriers are major challenges to successful oral delivery of therapeutics. Nanoparticle platforms can enhance drug stability and extend the residence time in gastrointestinal (GI) tract. However, drug delivery systems are often inactivated in acidic environment of stomach or suffer poor absorption from intestinal cells due to the mucus layer. To overcome these issues we developed a drug delivery system constituted by a protein construct made by a Rotavirus capsid protein (VP6) and the small ubiquitin-like modifier SUMO. This chimeric construct allows specificity towards intestinal cells, the Rotavirus natural target, combined by an enhanced stability given by the eukaryotic protein transporter SUMO. Furthermore SUMO can act as a molecular switch that facilitates import/export of its ligand to the nucleus, the hypersensitive subcellular site target of many cell killing therapies. In this paper we show that SUMO-VP6 constructs self-assembly into stable nanocarriers. SUMO-VP6 nanocarriers display ideal features for drug delivery: a small size and high monodispersity, a high stability in different pH conditions and a high uptake in the nuclear and cytoplasmic compartment of intestinal cells. These features make SUMO-VP6 nanocarriers a promising novel system for oral delivery of poorly soluble drugs

Potenziale utilizzo di idrolati per il controllo delle infezioni microbiche

Introduzione. Come \ue8 noto, diversi microrganismi costituiscono un diretto pericolo per la nostra salute in quanto possono infettare l'ospite umano: le malattie infettive causate da batteri e funghi sono ogni anno responsabili di morbilit\ue0 e mortalit\ue0 in un numero crescente di individui ospedalizzati e immunocompromessi. Inoltre, negli ultimi anni, la diffusione di ceppi microbici multi-resistenti ai farmaci, (ad es. Staphylococcus aureus, Pseudomonas aeruginosa e Candida spp.), ha provocato infezioni difficili o impossibili da controllare con antimicrobici convenzionali. La necessit\ue0 di nuove terapie per il trattamento delle infezioni microbiche ha portato i ricercatori a concentrarsi su possibili alternative di tipo naturale. L'idrolato (Id), noto anche come idrosol o acqua aromatica, \ue8 la frazione idrofila ottenuta durante il processo di distillazione in corrente di vapore assieme all\u2019olio essenziale (OE), capace di esercitare attivit\ue0 anti-microbiche grazie al contenuto in componenti terpeniche pari a circa l\u20191% v/v. Sebbene entrambe i prodotti della distillazione abbiano una nota azione antimicrobica, non \ue8 mai stata paragonata la loro attivit\ue0 quando estratti dalla stessa pianta. Scopo. Gli scopi di questo lavoro sono stati: (i) quello di paragonare l'attivit\ue0 antimicrobica dell\u2019OE e dell\u2019Id estratti da Monarda citriodora (MC) verso 30 ceppi fungini e batterici coinvolti in infezioni umane, (ii) valutare l\u2019efficacia dell\u2019Id di C. aurantium var. amara (CA) nei confronti di 18 ceppi fungini e batterici potenzialmente responsabili di infezioni cutanee. Materiali e metodi. L\u2019OE e l\u2019Id di MC sono stati ottenuti mediante distillazione in corrente di vapore partendo da piante fiorite coltivate presso l\u2019istituto Scarabelli-Ghini di Imola (fig.1), mentre l\u2019Id-CA era di tipo commerciale. Sono stati eseguiti test di micro-brodo diluizione secondo linee guida internazionali EUCAST per valutare l\u2019efficacia antimicrobica dell\u2019OE-MC (concentrazioni comprese tra 0,0078% e 4%), dell'Id-MC (concentrazioni tra 12,5% e 50%) e dell\u2019Id-CA (concentrazioni tra 3.12% e 50%) vs i ceppi testati. Inoltre, l'OE e gli Id sono stati analizzati in GC-MS per valutarne la composizione terpenica e, nel caso di M. citriodora, paragonare l\u2019efficacia dell\u2019OE a quella dell\u2019Id in funzione del loro contenuto terpenico. Risultati. L'OE-MC e l\u2019Id-CA sono risultati efficaci, a concentrazioni variabili, su tutti i ceppi testati, mentre, l'Id-MC era attivo su tutti i microrganismi tranne che sui ceppi di P. aeruginosa (MIC>4% v/v). Le analisi GC-MS hanno individuato come principali componenti terpeniche il timolo per OE-MC e di Id-MC (19,6% e 66,4% rispettivamente) e il linalolo per Id-CA. Conclusioni. Nonostante sia necessaria una maggiore concentrazione di Id-MC per ottenere lo stesso effetto inibente dell\u2019OE-MC, la quantit\ue0 di terpeni presenti per unit\ue0 di volume nell\u2019Id \ue8 risultata inferiore rispetto all'EO. La maggiore efficacia dei terpeni contenuti nell\u2019Id \ue8 probabilmente dovuta all'ambiente idrofilo che ne promuove una maggiore disponibilit\ue0

Origanum vulgare essential oil vs. A commercial mixture of essential oils: In vitro effectiveness on salmonella spp. from poultry and swine intensive livestock

open11noSalmonella spp. represent a public health concern for humans and animals due to the increase of antibiotic resistances. In this scenario, the use of essential oils (EOs) could be a valid tool against Salmonella contamination of meat. This work compares the in vitro effectiveness of an Italian mixture of feed additives based on EOs (GR-OLI) with EO of Origanum vulgare L., recently admitted by European Food Safety Authority (EFSA) for animal use. Twenty-nine Salmonella serotypes isolated from poultry and pig farms were used to assess GR-OLI and O. vulgare EO antimicrobial propeties. O. vulgare EO was active on the disaggregation of mature biofilm, while GR-OLI was capable of inhibiting biofilm formation and disaggregating preformed biofilm. Furthermore, GR-OLI inhibited bacterial adhesion to Caco-2 cells in a dose-dependent manner. Both products showed inhibition of bacterial growth at all time points tested. Finally, the synergistic action of GR-OLI with commonly used antibiotics against resistant strains was investigated. In conclusion, the mixture could be used both to reduce the meat contamination of Salmonella spp. before slaughter, and in synergy with low doses of ciprofloxacin against resistant strains. Although EOs as feed additives are already used in animal husbandry, no scientific study has ever highlighted their real antimicrobial potential.openDi Vito M.; Cacaci M.; Barbanti L.; Martini C.; Sanguinetti M.; Benvenuti S.; Tosi G.; Fiorentini L.; Scozzoli M.; Bugli F.; Mattarelli P.Di Vito M.; Cacaci M.; Barbanti L.; Martini C.; Sanguinetti M.; Benvenuti S.; Tosi G.; Fiorentini L.; Scozzoli M.; Bugli F.; Mattarelli P

Antibiofilm activity of three different irrigation techniques: An in vitro study

The microbial infection of the endodontic space occurs in a necrotic tooth as a result of dental caries, trauma, periodontal disease, or previous root canal therapy. The disruption of the biofilms and the reduction of the bacterial load inside root canals are crucial for the success of root canal therapy. The aim of this study was to compare, in vitro, the antibiofilm efficacy of a novel passive sonic irrigation (PSI) device with passive ultrasonic irrigation (PUI) and conventional needle irrigation (CNI). Forty-four single-rooted human teeth were inoculated with a culture of E. faecalis for 28 days. The specimens were randomly divided into three groups: PUI, CNI, and PSI (n = 12). The activation protocols were performed using both 17% EDTA and 5.25% NaOCl. Residual bacterial biofilm was taken by means of a canal brush and colony-forming unit (CFU) were counted. The data were analyzed using one-way ANOVA and Games-Howell's post hoc tests. A major reduction in CFU was observed in the PSI and PUI groups, in comparison with the CNI group. No difference was found (p > 0.05) in terms of CFU reduction between PSI and PUI. PSI could be as effective as PUI in the removal of bacterial biofilms from straight root canals

A New Strategy for Glioblastoma Treatment: In Vitro and In Vivo Preclinical Characterization of Si306, a Pyrazolo[3,4-d]Pyrimidine Dual Src/P-Glycoprotein Inhibitor

20siopenOverexpression of P-glycoprotein (P-gp) and other ATP-binding cassette (ABC) transporters in multidrug resistant (MDR) cancer cells is responsible for the reduction of intracellular drug accumulation, thus decreasing the efficacy of chemotherapeutics. P-gp is also found at endothelial cells' membrane of the blood-brain barrier, where it limits drug delivery to central nervous system (CNS) tumors. We have previously developed a set of pyrazolo[3,4-d]pyrimidines and their prodrugs as novel Src tyrosine kinase inhibitors (TKIs), showing a significant activity against CNS tumors in in vivo. Here we investigated the interaction of the most promising pair of drug/prodrug with P-gp at the cellular level. The tested compounds were found to increase the intracellular accumulation of Rho 123, and to enhance the efficacy of paclitaxel in P-gp overexpressing cells. Encouraging pharmacokinetics properties and tolerability in vivo were also observed. Our findings revealed a novel role of pyrazolo[3,4-d]pyrimidines which may be useful for developing a new effective therapy in MDR cancer treatment, particularly against glioblastoma.openFallacara, Anna Lucia; Zamperini, Claudio; Podolski-Renić, Ana; Dinić, Jelena; Stanković, Tijana; Stepanović, Marija; Mancini, Arianna; Rango, Enrico; Iovenitti, Giulia; Molinari, Alessio; Bugli, Francesca; Sanguinetti, Maurizio; Torelli, Riccardo; Martini, Maurizio; Maccari, Laura; Valoti, Massimo; Dreassi, Elena; Botta, Maurizio; Pešić, Milica; Schenone, SilviaFallacara, Anna Lucia; Zamperini, Claudio; Podolski-Renić, Ana; Dinić, Jelena; Stanković, Tijana; Stepanović, Marija; Mancini, Arianna; Rango, Enrico; Iovenitti, Giulia; Molinari, Alessio; Bugli, Francesca; Sanguinetti, Maurizio; Torelli, Riccardo; Martini, Maurizio; Maccari, Laura; Valoti, Massimo; Dreassi, Elena; Botta, Maurizio; Pešić, Milica; Schenone, Silvi

Human Monoclonal Antibodies to a Novel Cluster of Conformational Epitopes on HCV E2 with Resistance to Neutralization Escape in a Genotype 2a Isolate

The majority of broadly neutralizing antibodies to hepatitis C virus (HCV) are against conformational epitopes on the E2 glycoprotein. Many of them recognize overlapping epitopes in a cluster, designated as antigenic domain B, that contains residues G530 and D535. To gain information on other regions that will be relevant for vaccine design, we employed yeast surface display of antibodies that bound to genotype 1a H77C E2 mutant proteins containing a substitution either at Y632A (to avoid selecting non-neutralizing antibodies) or D535A. A panel of nine human monoclonal antibodies (HMAbs) was isolated and designated as HC-84-related antibodies. Each HMAb neutralized cell culture infectious HCV (HCVcc) with genotypes 1–6 envelope proteins with varying profiles, and each inhibited E2 binding to the viral receptor CD81. Five of these antibodies neutralized representative genotypes 1–6 HCVcc. Epitope mapping identified a cluster of overlapping epitopes that included nine contact residues in two E2 regions encompassing aa418–446 and aa611–616. Effect on virus entry was measured using H77C HCV retroviral pseudoparticles, HCVpp, bearing an alanine substitution at each of the contact residues. Seven of ten mutant HCVpp showed over 90% reduction compared to wild-type HCVpp and two others showed approximately 80% reduction. Interestingly, four of these antibodies bound to a linear E2 synthetic peptide encompassing aa434–446. This region on E2 has been proposed to elicit non-neutralizing antibodies in humans that interfere with neutralizing antibodies directed at an adjacent E2 region from aa410–425. The isolation of four HC-84 HMAbs binding to the peptide, aa434–446, proves that some antibodies to this region are to highly conserved epitopes mediating broad virus neutralization. Indeed, when HCVcc were passaged in the presence of each of these antibodies, virus escape was not observed. Thus, the cluster of HC-84 epitopes, designated as antigenic domain D, is relevant for vaccine design for this highly diverse virus

The Event Horizon General Relativistic Magnetohydrodynamic Code Comparison Project

Recent developments in compact object astrophysics, especially the discovery of merging neutron stars by LIGO, the imaging of the black hole in M87 by the Event Horizon Telescope, and high- precision astrometry of the Galactic Center at close to the event horizon scale by the GRAVITY experiment motivate the development of numerical source models that solve the equations of general relativistic magnetohydrodynamics (GRMHD). Here we compare GRMHD solutions for the evolution of a magnetized accretion flow where turbulence is promoted by the magnetorotational instability from a set of nine GRMHD codes: Athena++, BHAC, Cosmos++, ECHO, H-AMR, iharm3D, HARM-Noble, IllinoisGRMHD, and KORAL. Agreement among the codes improves as resolution increases, as measured by a consistently applied, specially developed set of code performance metrics. We conclude that the community of GRMHD codes is mature, capable, and consistent on these test problems