Dendrimer nanocarriers for transport modulation across models of the pulmonary epithelium.

The purpose of this study was to determine the effect of PEGylation on the interaction of poly(amidoamine) (PAMAM) dendrimer nanocarriers (DNCs) with in vitro and in vivo models of the pulmonary epithelium. Generation-3 PAMAM dendrimers with varying surface densities of PEG 1000 Da were synthesized and characterized. The results revealed that the apical to basolateral transport of DNCs across polarized Calu-3 monolayers increases with an increase in PEG surface density. DNC having the greatest number of PEG groups (n = 25) on their surface traversed at a rate 10-fold greater than its non-PEGylated counterpart, in spite of their larger size. This behavior was attributed to a significant reduction in charge density upon PEGylation. We also observed that PEGylation can be used to modulate cellular internalization. The total uptake of PEG-free DNC into polarized Calu-3 monolayers was 12% (w/w) vs 2% (w/w) for that with 25 PEGs. Polarization is also shown to be of great relevance in studying this in vitro model of the lung epithelium. The rate of absorption of DNCs administered to mice lungs increased dramatically when conjugated with 25 PEG groups, thus supporting the in vitro results. The exposure obtained for the DNC with 25PEG was determined to be very high, with peak plasma concentrations reaching 5 µg·mL(-1) within 3 h. The combined in vitro and in vivo results shown here demonstrate that PEGylation can be potentially used to modulate the internalization and transport of DNCs across the pulmonary epithelium. Modified dendrimers thereby may serve as a valuable platform that can be tailored to target the lung tissue for treating local diseases, or the circulation, using the lung as pathway to the bloodstream, for systemic delivery.

Polymeric nanocarriers for transport modulation across the pulmonary epithelium: dendrimers, polymeric nanoparticles, and their nanoblends.

The purpose of this study was to (a) Determine the cellular transport and uptake of amine-terminated generation 3 (G3) poly(amido amine) (PAMAM) dendrimers across an in vitro model of the pulmonary epithelium, and the ability to modulate their transport by forming nanoblends of the dendrimers with biodegradable solid polymeric nanoparticles (NPs) and (b) to formulate dendrimer nanocarriers in portable oral inhalation devices and evaluate their aerosol characteristics. To that end, fluorescein isothiocyanate (FITC)-labeled G3 PAMAM dendrimer nanocarriers (DNCs) were synthesized, and also encapsulated within poly lactide-co-glycolide nanoparticles (NPs). Transport and uptake of both DNCs encapsulated within NPs (nanoblends) and unencapsulated DNCs were tracked across polarized monolayers of airway epithelial cells, Calu-3. DNCs were also formulated as core-shell microparticles in pressurized metered-dose inhalers (pMDIs) and their aerodynamic properties evaluated by Andersen cascade impaction. The apparent permeability of DNCs across the airway epithelial model was similar to that of a paracellular marker of comparable molar mass--order of 10(-7) cm s(-1). The transport and cellular internalization of the DNCs can be modulated by formulating them as nanoblends. The transport of the DNCs across the lung epithelium was completely suppressed within the time of the experiment (5 h) when formulated as blends. The encapsulation also prevents saturation of the cellular internalization profile. Nanoblending may be a potential strategy to modulate the rate of transport and cellular uptake of DNCs, and thus be used as a design strategy to achieve enhanced local or systemic drug delivery.

Propellant-based inhalers for the non-invasive delivery of genes via oral inhalation.

In this work we describe the development of a propellant-based, portable oral inhalation platform for the pulmonary delivery of genes. A core-shell strategy is utilized to efficiently disperse cationic-polymer-DNA nanoparticles in hydrofluoroalkane propellants, and to generate aerosols from the corresponding pressurized metered-dose inhaler formulations (pMDIs) that have excellent aerosol characteristics, suitable for deep lung deposition. The engineered polyplexes and core-shell structures were fully characterized, and their ability to transfect model lung alveolar epithelium cells in vitro was demonstrated. We also show that the propellant does not affect the biological activity of the plasmid DNA, and that the core-shell formulations have no in vitro cytotoxicity. The relevance of this work stems from the fact that pMDIs are the least expensive and most widely used portable oral inhalation devices, and are thus promising platforms for targeting genes to the lungs for the treatment of medically relevant diseases including asthma, cystic fibrosis, chronic obstructive pulmonary disease, and lung cancer.

Reverse aqueous microemulsions in hydrofluoroalkane propellants and their aerosol characteristics.

In this work we describe the structure and environment of reverse aqueous microemulsions formed in 1,1,1,2-tetrafluoroethane (HFA134a) propellant in the presence of a non-ionic ethoxylated copolymer, and the aerosol characteristics of the corresponding pressurized metered dose inhaler (pMDI) formulations. The activity of selected polypropylene oxide-polyethylene oxide-polypropylene oxide (PO(m)EO(n)PO(m)) amphiphiles at the HFA134a-water interface was studied using in situ high-pressure tensiometry, and those results were used as a guide in the selection of the most appropriate candidate surfactant for the formation of microemulsions in the compressed HFA134a. The environment and structure of the aggregates formed with the selected surfactant candidate, PO(22)EO(14)PO(22), was probed via UV-vis spectroscopy (molecular probe), and small angle neutron scattering (SANS), respectively. High water loading capacity in the core of the nanoaggregates was achieved in the presence of ethanol. At a water-to-surfactant molar ratio of 21 and 10% ethanol, cylindrical aggregates with a radius of 18Å, and length of 254Å were confirmed with SANS. Anderson Cascade Impactor (ACI) results reveal that the concentration of the excipients (C(exp), including surfactant, water and ethanol) has a strong effect on the aerosol characteristics of the formulations, including the respirable fraction, and the mass mean aerodynamic diameter (MMAD), and that the trend in MMAD can be predicted as a function of the C(exp) following similar correlations to those proposed to common non-volatile excipients, indicating that the nanodroplets of water dispersed in the propellant behave similarly to molecularly solubilized compounds. Cytotoxicity studies of PO(22)EO(14)PO(22) were performed in A549 cells, an alveolar type II epithelial cell line, and indicate that, within the concentration range of interest, the surfactant in question decreases cell viability only lightly. The relevance of this work stems from the fact that aqueous-based HFA-pMDIs are expected to be versatile formulations, with the ability to carry a range of medically relevant hydrophilic compounds within the nanocontainers, including high potency drugs, drug combinations and biomacromolecules.

The potential for the noninvasive delivery of polymeric nanocarriers using propellant-based inhalers in the treatment of Chlamydial respiratory infections.

A novel strategy for pulmonary delivery of polymeric nanocarriers (NCs) pressurized-metered dose inhalers (pMDIs) is reported in this work. Core-shell particles consisting of a water soluble, hydrofluoroalkane(HFA)-philic biodegradable copolymer of chitosan and poly(lactic acid), and a core of poly(d,l-lactide-co-glycolide) (PLGA) NCs were prepared by a modified emulsification-diffusion methodology. Dispersions of the core-shell particles in HFA propellant revealed enhanced physical stability compared to polymeric NCs alone, and more importantly, excellent aerosol characteristics as determined by inertial impaction studies. Confocal microscopy revealed that the polymeric NCs from such core-shell particles are capable not only to be taken up by Calu-3 (airway epithelial) cells that have been infected with Chlamydia pneumoniae, an intracellular pathogen, but are also internalized within chlamydial inclusions. Our results suggest that the proposed methodology can be used as a general platform for the delivery of polymeric NCs to the respiratory tract using the inexpensive pMDIs, and that such an approach may be used to target and deliver drugs to treat chlamydial-related infections.

Core-shell particles for the dispersion of small polar drugs and biomolecules in hydrofluoroalkane propellants.

PURPOSE: Demonstrate the applicability of a novel particle-based technology for the development of suspensions of small polar drugs and biomolecules in hydrofluoroalkane (HFA) propellants for pressurized metered-dose inhalers (pMDIs). MATERIALS AND METHODS: Emulsification diffusion was used to prepare core-shell particles. The shell consisted of oligo(lactide) grafts attached onto a short chitosan backbone. The active drug was arrested within the particle core. Colloidal Probe Microscopy (CPM) was used to determine the cohesive forces between particles in a model HFA propellant. The aerosol characteristics of the formulations were determined using an Anderson Cascade Impactor (ACI). Cytotoxicity studies were performed on lung epithelial and alveolar type II cells. RESULTS: CPM results indicate that particle cohesive forces in liquid HFA are significantly screened in the presence of the polymeric shell and correlate well with the physical stability of suspensions in propellant HFA. The proposed formulation showed little or no cytotoxic effects on both Calu-3 and A549 cells. CONCLUSIONS: Core-shell particles with a shell containing the lactide moiety as the HFA-phile showed excellent dispersion stability and aerosol characteristics in HFA-based pMDIs. This is a general strategy that can be used for developing novel suspension pMDIs of both small polar drugs and large therapeutic molecules.

Biocompatible, lactide-based surfactants for the CO2-water interface: high-pressure contact angle goniometry, tensiometry, and emulsion formation.

The unique properties of compressed CO2, including its low cost, nontoxicity, easily tunable solvent strength, and favorable transport properties, make it an environmentally attractive alternative to volatile organic solvents. Suitable surface-active species can be utilized to realize the full potential of clean, CO2-based technologies, by helping to overcome the low solubility typically associated with many solutes of interest in CO2. In this work we synthesize and investigate the interfacial activity of a series of nonionic amphiphiles with a biocompatible and biodegradable CO2-phile at both the CO2-water (C|W) and CO2-water-solid (C|W|S) interfaces. We developed a high-pressure pendant drop tensiometer and contact angle goniometer that allows us to measure both tension and contact angle in tandem. The tension of the C|W interface was measured in the presence of the lactide (LA)-based surface active agents with varying molecular weight and hydrophilic-to-CO2-philic ratios. Emulsion studies with an optimum balanced surfactant were performed. The contact angle of water droplets against a silane-modified (hydrophobic) substrate under CO2 atmosphere was also measured in presence of a selected LA-based amphiphile. The results demonstrate that the nonionic copolymers with the biodegradable and biocompatible LA-based group can significantly reduce the tension of the C|W interface. The LA-based surface active species are also capable of forming stable emulsions of water and CO2 and reducing the angle of the three-phase C|W|S contact line.