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Development of inhalable formulations for delivering peptides to the conductive airways and shielding their interactions with airway barriers, thus enhancing peptide/bacteria interactions, is an important part of peptide-based drug development for lung applications. Here, we report the construction of a biocompatible nanosystem where the antimicrobial peptide SET-M33 is encapsulated within polymeric nanoparticles of poly(lactide-co-glycolide) (PLGA) conjugated with polyethylene glycol (PEG). This system was conceived for better delivery of the peptide to the lungs by aerosol. The encapsulated peptide showed prolonged antibacterial activity, due to its controlled release, and much lower toxicity than the free molecule. The peptide-based nanosystem killed Pseudomonas aeruginosa in planktonic and sessile forms in a dose-dependent manner, remaining active up to 72 h after application. The encapsulated peptide showed no cytotoxicity when incubated with human bronchial epithelial cells from healthy individuals and from cystic fibrosis patients, unlike the free peptide, which showed an EC50 of about 22 µM. In vivo acute toxicity studies in experimental animals showed that the peptide nanosystem did not cause any appreciable side effects, and confirmed its ability to mitigate the toxic and lethal effects of free SET-M33.

Food supplements (FS) containing red yeast rice (RYR) are largely employed to reduce lipid levels in the blood. The main ingredient responsible for biological activity is monacolin K (MoK), a natural compound with the same chemical structure as lovastatin. Concentrated sources of substances with a nutritional or physiological effect are marketed in “dose” form as food supplements (FS). The quality profile of the “dosage form” of FS is not defined in Europe, whereas some quality criteria are provided in the United States. Here, we evaluate the quality profile of FS containing RYR marketed in Italy as tablets or capsules running two tests reported in The European Pharmacopoeia 11 Ed. and very close to those reported in the USP. The results highlighted variations in dosage form uniformity (mass and MoK content) compliant with The European Pharmacopoeia 11 Ed. specifications, whereas the time needed for disintegrating tablets was longer for 44% of the tested samples. The bioaccessibility of MoK was also investigated to obtain valuable data on the biological behaviour of the tested FS. In addition, a method for citrinin (CIT) determination was optimized and applied to real samples. None of the analyzed samples demonstrated CIT contamination (LOQ set at 6.25 ng/mL). Considering the widespread use of FS, our data suggest that greater attention should be paid by fabricants and regulatory authorities to ensure the quality profile and the safe consumption of marketed products.

The potential of intra-venous gallium nitrate (GaN) administration against Pseudomonas aeruginosa pneumonia was recently demonstrated in mice and in cystic fibrosis (CF) patients. Likewise, the added value of direct lung delivery of Ga(III) has been shown in rats. Therefore, the design of a drug delivery system specifically engineered for Ga(III) inhalation is imperative to improve its accumulation in lungs. To this purpose, Ga(III) was efficiently encapsulated into hyaluronic acid/chitosan nanoparticles (Ga_HA/CS NPs), whose features were tuned to facilitate access to the target by overcoming mucus and biofilm surrounding bacteria. Then, to improve in vivo lung deposition, Ga_HA/CS NPs were engineered into mannitol-based NEM (Ga_Man NEM). The powders showed optimal in vitro aerosol performance, and sustained release kinetics in lung lining fluids. Moreover, good tolerability and antimicrobial properties were shown in vitro. Intratracheal insufflation of Ga_Man NEM in rats resulted in a significant improvement of Ga(III) persistence in the lungs coupled to a lower Ga(III) concentration in plasma and urine, compared to GaN solution. Noteworthy, the developed formulation significantly modifies the unfavorable Ga(III) kinetic increasing the Ga(III) to the lung and preventing Ga(III) accumulation in the kidney, key responsible for adverse effects, conclusively demonstrating the benefit of Ga_Man NEM to exploit the therapeutic effect of Ga(III) via inhalation route.

Cystic Fibrosis (CF) is characterized by an airway obstruction caused by a thick mucus due to a malfunctioning Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. The sticky mucus restricts drugs in reaching target cells limiting the efficiency of treatments. The development of new approaches to enhance drug delivery to the lungs represents CF treatment's main challenge. In this work, we report the synthesis and characterization of hybrid core-shell nanoparticles (hNPs) comprising a PLGA core and a dipalmitoylphosphatidylcholine (DPPC) shell engineered for inhalation. We loaded hNPs with a 7-mer peptide nucleic acid (PNA) previously considered for its ability to modulate the post-transcriptional regulation of the CFTR gene. We also investigated the in vitro release kinetics of hNPs and their efficacy in PNA delivery across the human epithelial airway barrier using an ex vivo model based on human primary nasal epithelial cells (HNEC) from CF patients. Confocal analyses and hNPs transport assay demonstrated the ability of hNPs to overcome the mucus barrier and release their PNA cargo within the cytoplasm, where it can perform its biological function.

This study reports novel food-grade granules for co-delivery of L. plantarum 299v and a standardized extract of Olea europaea leaves (Phenolea®) as oral carrier of probiotics and hydroxytyrosol. Different granule formulations containing either L. plantarum 299v (Lac), or the olive leave extract (Phe) or their combination (Lac-Phe) have been successfully produced through wet granulation employing excipients generally regarded as safe as granulating/binding agents. L. plantarum cells withstood the manufacturing process and were stable upon storage at 4 °C for more than 6 months. In vitro dissolution studies in simulated gastro-intestinal fluids showed the capability of the granules to rapidly dissolve and deliver both olive leave phenols and living L. plantarum cells. In simulated digestion conditions, Lac and Lac-Phe granules protected L. plantarum against the harsh environment of the gastro–intestinal tract. Co-administration of Lac and Phe oral granules to healthy mice provided for higher amounts of hydroxytyrosol in urines as compared to Phe granules alone, suggesting that L. plantarum 299v boosted in vivo conversion of oleuropein to hydroxytyrosol. On the other hand, PCR-assisted profiling of the Lactobacillus population in faeces obtained from mice treated with Lac or Lac plus Phe confirmed that the probiotic arrived alive to colon and was there able to exert a sort of perturbing effect on the climax colonic microflora. Overall, these results pave the way towards the development of a nutraceutical useful for combined delivery of bioactive hydroxytyrosol and probiotics to colon site.

Nowadays, the downregulation of genes involved in the pathogenesis of severe lung diseases through local siRNA delivery appears an interesting therapeutic approach. In this study, we propose novel hybrid lipid-polymer nanoparticles (hNPs) consisting of poly(lactic-co-glycolic) acid (PLGA) and dipalmitoyl phosphatidylcholine (DPPC) as siRNA inhalation system. Background: Nowadays, the downregulation of genes involved in the pathogenesis of severe lung diseases through local siRNA delivery appears an interesting therapeutic approach. In this study, we propose novel hybrid lipid-polymer nanoparticles (hNPs) consisting of poly(lactic-co-glycolic) acid (PLGA) and dipalmitoyl phosphatidylcholine (DPPC) as siRNA inhalation system. Methods: A panel of DPPC/PLGA hNPs was prepared by emulsion/solvent diffusion and fully characterized. A combination of model siRNAs against the sodium transepithelial channel (ENaC) was entrapped in optimized hNPs comprising or not poly(ethylenimine) (PEI) as third component. siRNA-loaded hNPs were characterized for encapsulation efficiency, release kinetics, aerodynamic properties, and stability in artificial mucus (AM). The fate and cytotoxicity of hNPs upon aerosolization on a triple cell co-culture model (TCCC) mimicking human epithelial airway barrier were assessed. Finally, the effect of siRNA-loaded hNPs on ENaC protein expression at 72 hours was evaluated in A549 cells. Results: Optimized muco-inert hNPs encapsulating model siRNA with high efficiency were produced. The developed hNPs displayed a hydrodynamic diameter of approximate to 150nm, a low polydispersity index, a negative potential close to -25mV, and a peculiar triphasic siRNA release lasting for 5 days, which slowed down in the presence of PEI. siRNA formulations showed optimal in vitro aerosol performance after delivery with a vibrating mesh nebulizer. Furthermore, small-angle X-ray scattering analyses highlighted an excellent stability upon incubation with AM, confirming the potential of hNPs for direct aerosolization on mucus-lined airways. Studies in TCCC confirmed that fluorescent hNPs are internalized inside airway epithelial cells and do not exert any cytotoxic or acute proinflammatory effect. Finally, a prolonged inhibition of ENaC protein expression was observed in A549 cells upon treatment with siRNA-loaded hNPs. Conclusions: Results demonstrate the great potential of hNPs as carriers for pulmonary delivery of siRNA, prompting toward investigation of their therapeutic effectiveness in severe lung diseases.

In this study, we highlight the potential of the mucoadhesive film made from a poly(ethylene oxide)/hydroxypropyl-β-cyclodextrin (PEO/CD) mixture in the oromucosal delivery of hydrophilic drugs, with a specific focus on dexamethasone phosphate disodium salt (Dexa). CD formed a complex with Dexa in solution and did not interact with mucin as highlighted from the spectrophotometric and spectrofluorimetric analysis. Similarly, CD and PEO did not affect mucin conformation, suggesting no direct interaction between the unstirred water layer and film components. Remarkably, PEO/CD/Dexa films dissolved more slowly than those made of PEO alone also in phosphate-buffered saline (PBS) pH 6.8 and gave a time-control on Dexa delivered dose. These combined effects resulted in a higher amount of Dexa accumulated in the mucosa, which can be highly beneficial in case of local diseases. Furthermore, Dexa amount able to diffuse through porcine buccal mucosa was lower when film contained CD, highlighting how CD can act as a modulator of drug transport also in the case of water-soluble drugs. In summary, our results demonstrate the versatility of PEO/CD films in mucosal delivery of hydrophilic corticosteroids paving the way to a novel approach in the treatment of mouth diseases.

The inflammatory process plays a crucial role in the onset and progression of several lung pathologies, including cystic fibrosis (CF),andthe involvement of NF-κBis widely recognized.Thespecific inhibition of NF-κB by decoy oligonucleotides delivered within the lung may be beneficial, although rationally designed systems are needed to optimize their pharmacological response. Prompted by this need, we have developed and tested in vivo an inhalable dry powder for the prolonged delivery of a decoy oligodeoxynucleotide to NF-κB (dec-ODN), consisting of large porous particles (LPPs) based on poly(lactic-co-glycolic) acid. First, LPPs containing dec- ODN (dec-ODN LPPs) were engineered to meet the aerodynamic criteria crucial for pulmonary delivery, to gain an effective loading of dec-ODN, to sustain its release, and to preserve its structural integrity in lung lining fluids. We then investigated the effects of dec- ODN LPPs in a rat model of lung inflammation induced by the intratracheal aerosolization of LPS from Pseudomonas aeruginosa. Theresults show that a single intratracheal insufflation of dec-ODN LPPs reduced the bronchoalveolar neutrophil infiltration induced by LPS for up to 72 hours, whereas naked dec-ODN was able to inhibit it only at 6 hours. The persistent inhibition of neutrophil infiltrate was associated with reduced NF-κB/DNA binding activity, as well as reduced IL-6, IL-8, and mucin-2 mRNA expression in lung homogenates. We consider it noteworthy that the developed LPPs, preventing the accumulation of neutrophils and NF-κB-related gene expression, may provide a new therapeutic option for the local treatment of inflammation associated with lung disease. Copyright © 2013 by the American Thoracic Society.

We have recently demonstrated that the specific inhibition of nuclear factor-κB by a decoy oligonucleotide (dec-ODN) delivered through inhalable large porous particles (LPP) made of poly(lactic-co-glycolic acid) (PLGA) may be highly beneficial for long-term treatment of lung inflammation. Nevertheless, besides chronic inflammation, multifunctional systems aimed to control also infection are required in chronic lung diseases, such as cystic fibrosis (CF). In this work, we tested the hypothesis that engineering PLGA-based LPP with branched poly(ethylenimine) (PEI) may improve LPP properties for pulmonary delivery of dec-ODN, with particular regard to the treatment of Pseudomonas aeruginosa lung infections. After getting insight into the role of PEI on the technological properties of PLGA-based LPP for delivery of dec-ODN, the putative synergistic effect of PEI free or PEI released from LPP on in vitro antimicrobial activity of tobramycin (Tb) and aztreonam (AZT) against P. aeruginosa was elucidated. Meanwhile, cytotoxicity studies on A549 cells were carried out. Results clearly demonstrate that the dry powders have promising aerosolization properties and afford a prolonged in vitro release of both dec-ODN and PEI. The encapsulation of PEI into LPP results in a 2-fold reduction of the minimum inhibitory concentration of AZT, while reducing the cytotoxic effect of PEI. Of note, the developed ODN/PLGA/PEI LPP persisted at lung at least for 14 days after intratracheal administration in rats where they can provide sustained and combined release of dec-ODN and PEI. dec-ODN will likely act as an anti-inflammatory drug, while PEI may enhance the therapeutic activity of inhaled antibiotics, which are commonly employed for the treatment of concomitant lung infections. We have recently demonstrated that the specific inhibition of nuclear factor-kappa B by a decoy oligonucleotide (dec-ODN) delivered through inhalable large porous particles (LPP) made of poly(lactic-co-glycolic acid) (PLGA) may be highly beneficial for long-term treatment of lung inflammation. Nevertheless, besides chronic inflammation, multifunctional systems aimed to control also infection are required in chronic lung diseases, such as cystic fibrosis (CF). In this work, we tested the hypothesis that engineering PLGA-based LPP with branched poly(ethylenimine) (PEI) may improve LPP properties for pulmonary delivery of dec-ODN, with particular regard to the treatment of Pseudomonas aeruginosa lung infections. After getting insight into the role of PEI on the technological properties of PLGA-based LPP for delivery of dec-ODN, the putative synergistic effect of PEI free or PEI released from LPP on in vitro antimicrobial activity of tobramycin (Tb) and aztreonam (AZT) against P. aeruginosa was elucidated. Meanwhile, cytotoxicity studies on A549 cells were carried out. Results clearly demonstrate that the dry powders have promising aerosolization properties and afford a prolonged in vitro release of both dec-ODN and PEI. The encapsulation of PEI into LPP results in a 2-fold reduction of the minimum inhibitory concentration of AZT, while reducing the cytotoxic effect of PEI. Of note, the developed ODN/PLGA/PEI LPP persisted at lung at least for 14 days after intratracheal administration in rats where they can provide sustained and combined release of dec-ODN and PEI. dec-ODN will likely act as an anti-inflammatory drug, while PEI may enhance the therapeutic activity of inhaled antibiotics, which are commonly employed for the treatment of concomitant lung infections.

Flucytosine (5-fluorocytosine, 5-FC) is a fluorinated analogue of cytosine currently approved for the systemic treatment of fungal infections, which has recently demonstrated a very promising antivirulence activity against the bacterial pathogen Pseudomonas aeruginosa. In this work, we propose novel inhalable hyaluronic acid (HA)/mannitol composite dry powders for repositioning 5-FC in the local treatment of lung infections, including those affecting cystic fibrosis (CF) patients. Different dry powders were produced in one-step by spray-drying. Powder composition and process conditions were selected after in depth formulation studies aimed at selecting the 5-FC/HA/mannitol formulation with convenient aerosolization properties and drug release profile in simulated lung fluids. The optimized 5-FC/HA/mannitol powder for inhalation (HyaMan_FC#3) was effectively delivered from different breath-activated dry powder inhalers (DPI) already available to CF patients. Nevertheless, the aerodynamic assessment of fine particles suggested that the developed formulation well fit with a low-resistance DPI. HyaMan_FC#3 inhibited the growth of the fungus Candida albicans and the production of the virulence factor pyoverdine by P. aeruginosa at 5-FC concentrations that did not affect the viability of both wild type (16HBE14o-) and CF (CFBE41o-) human bronchial epithelial cells. Finally, pharmacokinetics of HyaMan_FC#3 inhalation powder and 5-FC solution after intratracheal administration in rats were compared. In vivo results clearly demonstrated that, when formulated as dry powder, 5-FC levels in both bronchoalveolar lavage fluid and lung tissue were significantly higher and sustained over time as compared to those obtained with the 5-FC solution. Of note, when the same 5-FC amount was administered intravenously, no significant drug amount was found in the lung at each time point from the injection. To realize a 5-FC lung concentration similar to that obtained by using HyaMan_FC#3, a 6-fold higher dose of 5-FC should be administered intravenously. Taken together, our data demonstrate the feasibility to deliver 5-FC by the pulmonary route likely avoiding/reducing the well-known side effects associated to the high systemic 5-FC doses currently used in humans. Furthermore, our results highlight that an appropriate formulation design can improve the persistence of the drug at lungs, where microorganisms causing severe infections are located.

The aim of this work was to investigate the potential of small nanoparticles (NPs) made of a poly(ethylene glycol)-poly(ε-caprolactone)-amphiphilic diblock copolymer (PEG-. b-PCL, PEG. =. 2. kDa and PCL. =. 4.2. kDa) as drug carrier system through the skin. Zinc(II) phthalocyanine (ZnPc), selected as lipophilic and fluorescent model molecule, was loaded inside NPs by a melting/sonication procedure. Loaded NPs with a hydrodynamic diameter around 60. nm, a slightly negative zeta potential and a ZnPc entrapment dependent on polymer/ZnPc ratio were obtained. Spectroscopic investigations evidenced that ZnPc was entrapped in monomeric form maintaining its emission properties. The transport of ZnPc through porcine ear skin was evaluated on Franz-type diffusion cells after treatment with different vehicles (water or PEG 0.4. kDa) containing free ZnPc or ZnPc-loaded NPs without and with (2-hydroxypropyl)-β-cyclodextrin (HPβCD) as permeation enhancer. Independently of the sample tested, ZnPc was transported in the skin without reaching receptor compartment. On the other hand, ZnPc was found in the skin in large amount and also in the viable epidermis when delivered through NPs associated with HPβCD, especially in conditions limiting water evaporation. Fluorescence images of skin samples after 24. h of permeation were in line with ZnPc dosage in the skin and demonstrated the ability of NPs covalently tagged with rhodamine to penetrate the skin and to locate in the intercellular spaces. Insight into skin chemical properties upon application of NPs by confocal Raman spectroscopy demonstrated that HPβCD caused an alteration of water profile in the skin, highly reducing the degree of hydration at stratum corneum/viable epidermis interface which can promote NP transport. Taken together, these results highlight PEG-. b-PCL NPs coupled with HPβCD as a novel vehicle for the skin delivery of highly lipophilic compounds paving the way to several applications.

PLGA-based carriers for pulmonary drug delivery: key concepts Poly(lactic-co-glycolic acid) (PLGA) is not a newcomer in the drug delivery field. Indeed, among the biodegradable polymers currently developed to formulate polymeric drug carriers, PLGA is acknowledged as the most attractive one due to its excellent biocompatibility as well as the possibility to adjust drug delivery features by varying composition, molecular weight and chemical structure. Several PLGA types with very different in vivo lifetimes, ranging from 3 weeks to over a year, are currently approved by US FDA and European Medicine Agency for sustained delivery of drugs via the parenteral route. Nowadays, well-established formulation conditions and methods of production afford the delivery of various types of drugs, in other words, hydrophilic or hydrophobic small molecules as well as macromolecules (peptides, proteins, nucleic acid therapeutics) [1,2]. Lupron Depot, Nutropin Depot, Sandostatin LAR Depot, Somatuline LA, TrelstarTM Depot, Arestin, Risperidal ConstaTM are only some of the PLGA-based products on the market for parenteral administration of drugs. Most of them are intended for sustained release of protein therapeutics (e.g., leuprolide, octreotide, lanreotide, triptorelin), since PLGA-based carriers may effectively protect the macromolecule from in vivo degradation occurring at the administration site [1]. On the other hand, it is now possible to modify the properties of PLGA particles to drive their fate in the biological environment and/or to target the drug to specific organs or cells after administration. PLGA nanocarriers with a size less than 1 μm represent a well-established tool to achieve targeted drug or gene delivery, with special emphasis to cancer treatment [2]. In the attempt to find out novel strategies for the prolonged release of drugs in the lung, PLGA nanoparticles and microparticles are gaining momentum also as carriers for inhalation. A fundamental understanding of the behavior of the particle in biologically relevant conditions as well as an appreciation of in vitro/in vivo effects of the encapsulated drug are essential in the design and development of biodegradable PLGA carriers bearing bioactive agents for therapeutic applications. Indeed, if adequately engineered, PLGA carriers are able to deliver in a sustained manner the intact drug in the airways, to shield its interactions with lung environment and to enhance drug interactions with the cell target [3]. Excellent reviews have recently summarized all the basic requirements of carriers for pulmonary drug delivery [4,5]. The first concern arises from the inhalation device, that is, nebulizer, pressurized metered dose inhaler or dry powder inhaler (DPI). Among them, a great deal of interest was recently aroused in delivering PLGA carriers directly to the lung surfaces via DPIs. This may allow to avoid nebulization resulting in possible instability phenomena (e.g., particle aggrePLGA carriers for inhalation: where do we stand, where are we headed?

Inhaled antivirulence drugs are currently considered a promising therapeutic option to treat Pseudomonas aeruginosa lung infections in cystic fibrosis (CF). We have recently shown that the anthelmintic drug niclosamide (NCL) has strong quorum sensing (QS) inhibiting activity against P. aeruginosa and could be repurposed as an antivirulence drug. In this work, we developed dry powders containing NCL nanoparticles that can be reconstituted in saline solution to produce inhalable nanosuspensions. NCL nanoparticles were produced by high-pressure homogenization (HPH) using polysorbate 20 or polysorbate 80 as stabilizers. After 20 cycles of HPH, all formulations showed similar properties in the form of needle-shape nanocrystals with a hydrodynamic diameter of approximately 450 nm and a zeta potential of -20 mV. Nanosuspensions stabilized with polysorbate 80 at 10% w/w to NCL (T80_10) showed an optimal solubility profile in simulated interstitial lung fluid. T80_10 was successfully dried into mannitol-based dry powder by spray drying. Dry powder (T80_10 DP) was reconstituted in saline solution and showed optimal in vitro aerosol performance. Both T80_10 and T80_10 DP were able to inhibit P. aeruginosa QS at NCL concentrations of 2.5-10 mu M. NCL, and these formulations did not significantly affect the viability of CF bronchial epithelial cells in vitro at microbiologically active concentrations (i.e.

Pulmonary drug delivery represents the best way of treating lung diseases, since it allows direct delivery of the drug to the site of action, with few systemic effects. Meanwhile, the lungs may be used as a portal of entry to the body, allowing systemic delivery of drugs via the airway surfaces into the bloodstream. In both cases, the therapeutic effect of the inhaled drug can be optimized by embedding it in appropriately engineered inhalable carriers, which can protect the drug against lung defense mechanisms and promote drug transport across the extracellular and cellular barriers. To this purpose, the attention has been very recently focused on polymeric nanoparticles (NPs). The aim of this review is to offer an overview on the recent advances in NPs for pulmonary drug delivery. After a description of the main challenges encountered in developing novel inhaled products, the design rules to engineer polymeric NPs for inhalation, and in so doing to overcome barriers imposed by the lungs anatomy and physiology, are described. Then, the state-of-art on inhalable biocompatible polymeric NPs based on enzymatically-degradable natural polymers and biodegradable poly(ester)s is presented, with a special focus on NP-based dry powders for inhalation. Finally, the in vitro/in vivo models useful to address the never-ending toxicological debate related to the use of NPs for inhalation are described.

A β-cyclodextrin nanosponge cross-linked with pyromellitic dianhydride (βNS-PYRO) is reported for the first time as multifunctional ingredient in semisolid formulations for drug delivery to the skin. The role of βNS-PYRO on solubilization and stabilization of the photosensitizer benzoporphyrin-derivative monoacid ring A (BPDMA) and all-trans retinoic acid (atRA) as well as its effect on skin permeation of diclofenac (DIC) was investigated. Aqueous solutions, gels, and cream-gels were prepared from mixtures of βNS-PYRO with a conventional gelling agent at specific ratios. The incorporation of BPDMA in βNS-PYRO water solutions prevented its aggregation and gave kinetically stable complexes with high photostability and singlet oxygen generation upon irradiation. atRA incorporated in the βNS-PYRO-containing gel demonstrated a remarkable stability as compared with the formulation without βNS-PYRO, resulting in an eightfold increase of its lifetime. Skin permeation studies highlighted that βNS-PYRO in gels and cream-gels containing DIC significantly decreased the amount of drug permeated through the skin while increasing its amount in stratum corneum and viable epidermis. Overall, swellable βNS-PYRO turns to be a multifunctional coingredient with potential in topical monophasic and biphasic formulations to stabilize light-sensitive drugs and to localize the action of highly penetrating drugs in the external layers of skin.

Cystic fibrosis (CF) is the most common autosomal recessive disease in Caucasians associated with early death. Although the faulty gene is expressed in epithelia throughout the body, lung disease is still responsible for most of the morbidity and mortality of CF patients. As a local delivery route, pulmonary administration represents an ideal way to treat respiratory infections, excessive inflammation and other manifestations typical of CF lung disease. Nonetheless, important determinants of the clinical outcomes of inhaled drugs are the concentration/permanence at the lungs as well as the ability of the drug to overcome local extracellular and cellular barriers. This review focuses on emerging delivery strategies used for local treatment of CF pulmonary disease. After a brief description of the disease and formulation rules dictated by CF lung barriers, it describes current and future trends in inhaled drugs for CF. The most promising advanced formulations are discussed, highlighting the advantages along with the major challenges for researchers working in this field.

Several PEGylated polyester-based nanoncologicals have been proposed in the literature, some of them nowadays being under preclinical/clinical trials or marketed. In this review, we describe the main features of PEGylated polyesters and their correspondent nanocarriers. A first part is devoted to intravenously injectable PEGylated nanocarriers, which represent the systems most investigated so far. After describing fundamental design rules dictated by the administration route, PEGylated nanocarriers currently under preclinical/clinical investigation or in the market will be described from a technological point of view and related therapeutic implications discussed. Finally, new perspective of use of PE-Gylated nanocarriers for oral and pulmonary delivery of anticancer drugs will be considered. © 2014 Bentham Science Publishers.

Inspired by the multiple roles cyclodextrins can play in polymeric systems, here we engineered poly(ethylene oxide) (PEO) films with (2-hydroxypropyl)- β-cyclodextrin (CD) as multipurpose ingredient. To shed light on the potential of CD in formulating PEO buccal films for the delivery of poorly water-soluble drugs, we preliminarily assessed thermal and mechanical properties as well as wettability of films prepared at different PEO/CD ratios. PEO/CD platform containing 54% by weight of CD was chosen as the optimized composition since it matched acceptable mechanical properties, in terms of tensile strength and elasticity, with a good wettability. The platform was tested as buccal delivery system for triamcinolone acetonide (TrA), a lipophilic synthetic corticosteroid sparely water soluble. Confocal Raman imaging clearly showed that CD was homogeneously (i.e. molecularly) dispersed in PEO. Nevertheless, homogenous drug distribution in the film without TrA crystallization occurred only in the presence of CD. Finally, CD-containing PEO film placed in simulated buccal fluids provided a useful speed-up of TrA release rate while showing slower dissolution as compared to PEO film. These results, as well as compliance with quality specifications of pharmaceutical manufacturing products, strongly support the soundness of the strategy and prompt toward further applications of PEO/CD films in buccal drug delivery. Crown Copyright © 2013 Published by Elsevier B.V. All rights reserved.

New vessel formation is the result of the complex orchestration of various elements, such as cells, signalling molecules and extracellular matrix (ECM). In order to establish the suitable conditions for an effective cell response, the influence of vascular endothelial growth factor (VEGF) complexation with heparin sodium salt (Hp) on its pro-angiogenic activity has been evaluated by an in vitro capillary-like tube formation assay. VEGF with or without Hp was embedded into collagen gels, and the activated matrices were characterized in terms of VEGF activity and release kinetics. Taking into account the crucial role of Hp in VEGF stability and activity, VEGF/Hp complex was then encapsulated into microspheres based on poly(lactide-co-glycolide) (PLGA), and microsphere properties, VEGF/Hp release kinetics and VEGF in vitro activity over time were evaluated. Integrated microsphere/collagen matrices were developed in order to provide a continuous release of active VEGF/Hp inside the matrix but also a VEGF gradient at the boundary, which is an essential condition for endothelial cell attraction and scaffold invasion. The results confirmed a strong influence of Hp on VEGF configuration and, consequently, on its activity, while the encapsulation of VEGF/Hp complex in PLGA-microspheres guaranteed a sustained release of active VEGF for more than 30 days. This paper confirms the importance of VEGF stability and signal presentation to cells for an effective proangiogenic activity and highlights how the combination of two stabilizing approaches, namely VEGF/Hp complexation and entrapment within PLGA-based microspheres, may be a very effective strategy to achieve this goal. © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Objectives The aim of this review is to summarize the current state-of-the-art in poly(lactic-co-glycolic acid) (PLGA) carriers for inhalation. It presents the rational of use, the potential and the recent advances in developing PLGA microparticles and nanoparticles for pulmonary delivery. The most promising particle engineering strategies are discussed, highlighting the advantages along with the major challenges for researchers working in this field. Key findings Biodegradable polymer carriers, such as PLGA particles, may permit effective protection and long-term delivery of the inhaled drug and, when adequately engineered, its efficient transport to the target. The carrier can be designed for inhalation on the basis of several strategies through the adequate combination of available particle technologies and excipients. In so doing, the properties of PLGA particles can be finely tuned at micro-size and nano-size level to fulfill specific therapeutic needs. This means not only to realize optimal in vitro/in vivo lung deposition of the formulation, which is still crucial, but also to control the fate of the drug in the lung after particle landing. Summary Although many challenges still exist, PLGA carriers may be highly beneficial and present a new scenario for patients suffering from chronic lung diseases and for pharmaceutical companies working to develop novel inhaled products.

Although few experimental studies have been handled so far to exploit the potential of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) in the production of dry powders for antibiotic inhalation, there has been no comprehensive study on the role played by NP composition. In this work, we try to shed light on this aspect by designing and developing a pulmonary delivery system for antibiotics, such as tobramycin (Tb), based on PLGA NPs embedded in an inert microcarrier made of lactose, referred to as nano-embedded micro-particles (NEM). At nanosize level, helper hydrophilic polymers were used to impart the desired surface, bulk and release properties to PLGA NPs prepared by a modified emulsion-solvent diffusion technique. Results showed that poly(vinyl alcohol) (PVA) and chitosan (CS) are essential to optimise the size and modulate the surface properties of Tb-loaded PLGA NPs, whereas the use of alginate (Alg) allows efficient Tb entrapment within NPs and its release up to one month. Optimized formulations display good in vitro antimicrobial activity against P. aeruginosa planktonic cells. Furthermore, spray-drying of the NPs with lactose yielded NEM with peculiar but promising flow and aerosolization properties, while preserving the peculiar NP features. Nonetheless, in vivo biodistribution studies showed that PVA-modified Alg/PLGA NPs reached the deep lung, while CS-modified NPs were found in great amounts in the upper airways, lining lung epithelial surfaces. In conclusion, PLGA NP composition appears to play a crucial role in determining not only the technological features of NPs but, once processed in the form of NEM, also their in vitro/in vivo deposition pattern. © 2011 Elsevier B.V.

Tissue engineering is one of the most promising research areas in bioregenerative medicine. However, the restoration of biological functionalities by implanting bioartificially engineered tissues is still highly limited because of their lack of vascular networks. The use of proangiogenic molecules delivered from a controlled release device is a promising strategy to induce tissue vascularization. Indeed, the controlled release system can enhance the therapeutic effect in vivo of many short half-life drugs, while circumventing the need for repeated administrations. In this work, PLGA:poloxamer blend based micro- and nanoparticles have been developed for the sustained delivery of a recently developed synthetic proangiogenic compound: SHA-2-22. Drug-loaded PLGA:poloxamer blend microparticles were prepared by an oil-in-oil solvent extraction/evaporation technique. Drug-loaded PLGA:poloxamer nanoparticles were prepared by a modified solvent diffusion technique. These drug carriers were characterized with regard to their physicochemical properties, morphology, drug encapsulation efficiency and release kinetics in vitro. The results show that by adjusting the formulation conditions, it is possible to obtain PLGA:poloxamer micro- and nanoparticles with very high drug loadings, and with the capacity to release the active compound in a controlled way for up to one month. In vitro cell assays performed in an endothelial cell model confirmed the bioactivity of SHA-22-2 encapsulated in PLGA:poloxamer microparticles. © 2010 Elsevier B.V.

New blood vessel formation is a critical requirement for treating many vascular and ischemia related diseases, as well as for many tissue engineering applications. Angiogenesis and vasculogenesis, in fact, represent crucial processes for the functional regeneration of complex tissues through tissue engineering strategies. Several growth factors (GFs) and signaling molecules involved in blood vessels formation have been identified, but their application to the clinical setting is still strongly limited by their extremely short half-life in the body. To overcome these limitations, we have developed a new injectable controlled release device based on polymeric nanoparticles for the delivery of two natural proangiogenic GFs: platelet derived growth factor (PDGF-BB) and fibroblast growth factor (FGF-2). The nanoparticle system was prepared by a modified solvent diffusion technique, encapsulating the GF both in presence and in the absence of two stabilizing agents: bovine serum albumin (BSA) and heparin sodium salt (Hp). The developed nanocarriers were characterized for morphology, size, encapsulation efficiency, release kinetics in vitro and GF activity in cell cultures. The results have indicated that the coencapsulation of stabilizing agents can preserve the GF active structure and, in addition, increase their encapsulation efficiency into nanoparticles. Through this optimization process, we were able to raise the encapsulation efficiency of FGF-2 to 63%, and that of PDGF-BB to 87%. These PLGA:poloxamer blend nanoparticles loaded with GFs were able to release PDGF-BB and FGF-2 in a sustained fashion for more than a month. This work also confirms other positive features of PLGA:poloxamer nanoparticles. Namely, they are able to maintain their stability in simulated biological medium, and they are also nontoxic to cell culture models. Incubation of nanoparticles loaded with FGF-2 or PDGF-BB with endothelial cell culture models has confirmed that GFs are released in a bioactive form. Altogether, these results underline the interest of PLGA:poloxamer nanoparticles for the controlled delivery of GFs and substantiate their potential for the treatment of ischemic diseases and for tissue engineering applications. © 2010 American Chemical Society.

The development of a vascular network in tissue-engineered constructs is a fundamental bottleneck of bioregenerative medicine, particularly when the size of the implant exceeds a certain limit given by diffusion lengths and/or if the host tissue shows a very active metabolism. One of the approaches to achieve the vascularization of tissue constructs is generating a sustained release of proangiogenic factors from the ischemic site. This work describes the formation and characterization of hyaluronic acid-chitosan (HA/CS) nanoparticles for the delivery of two pro-angiogenic growth factors: vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF-BB). These nanoparticles were prepared by an ionic gelification technique, and different formulations were developed by encapsulating the growth factors in association with two stabilizing agents: bovine serum albumin or heparin sodium salt. These carriers were characterized with regard to their physicochemical properties, their stability in biological media, and their cytotoxicity in the C3a hepatoma cell line. The results show that nanoparticles around 200nm can be prepared by this method. HA/CS nanoparticles were stable when incubated in EMEM cell culture medium or in water at 37°C for 24h. Cell culture tests confirmed that HA/CS nanoparticles are not cytotoxic within the concentration range used for growth factor delivery. Moreover, HA/CS nanoparticles were able to entrap efficiently both growth factors, reaching association values of 94% and 54% for VEGF and PDGF, respectively. In vitro release studies confirm that PDGF-BB is released from HA/CS nanoparticles in a sustained manner over ∼ 1 week. On the other hand, VEGF is completely released within the first 24h. © 2010 Informa Healthcare USA, Inc.

The success of any tissue engineering implant relies upon prompt vascularization of the cellular construct and, hence, on the ability of the scaffold to broadcast specific activation of host endothelium and guide vessel ingrowth. Vascular endothelial growth factor (VEGF) is a potent angiogenic stimulator, and if released in a controlled manner it may enhance and guide scaffold vascularization. Therefore, the aim of this work was to realize a scaffold with integrated depots able to release VEGF in a controlled rate and assess the ability of this scaffold to promote angiogenesis. VEGF-loaded poly(lactide-co-glycolide) (PLGA) microspheres were produced and included in a collagen scaffold. The release of VEGF from microspheres was tailored to be sustained over several weeks and occurred at a rate of ∼0.6 ng/day per mg of microspheres. It was found that collagen scaffolds bioactivated with VEGF-loaded microspheres strongly enhanced endothelial cell activation and vascular sprouting both in vitro and in vivo as compared with a collagen scaffold bioactivated with free VEGF. This report demonstrates that by finely tuning VEGF release rate within a polymeric scaffold, sprouting of angiogenic vessels can be guided within the scaffolds interstices as well as broadcasted from the host tissues. © 2009 Wiley Periodicals, Inc.