Inhalable, bioresponsive microparticles for targeted drug delivery in the lungs.

OBJECTIVE: There is a growing interest in developing bioresponsive drug delivery systems to achieve greater control over drug release than can be achieved with the conventional diffusion controlled polymeric delivery systems. While a number of such systems have been studied for oral or parenteral delivery, little or no work has been done on bioresponsive delivery systems for inhalation. Using the raised elastase levels present at sites of lung inflammation as a proof-of-concept model, we endeavoured to develop a prototype of inhalable elastase sensitive microparticles (ESMs). METHODS: Microparticles degradable by the enzyme elastase were formed by crosslinking the polymer alginate in the presence of an elastase substrate, elastin, using Ca(+2) ions and subsequent spray drying. KEY FINDINGS: The bioresponsive release of a protein cargo in the presence of elastase demonstrated the enzyme-specific degradability of the particles. The microparticles showed favorable properties such as high drug encapsulation and good powder dispersibility. Potential polymer toxicity in the lungs was assessed by impinging the microparticles on Calu-3 cell monolayers and assessing changes in transepithelial permeability and induction of cytokine release. The microparticles displayed no toxic or immunogenic effects. CONCLUSIONS: With a manufacturing method that is amenable to scale-up, the ability to be aerosolised efficiently from a first-generation inhaler device, enzyme-specific degradability and lack of toxicity, the ESMs show significant promise as pulmonary drug carriers

Bioengineered microparticles for controlled drug delivery to the lungs

Traditional formulations for pulmonary drug delivery mainly focused on two approaches: (i) Dissolving or suspending the drug in a solvent or propellant to produce liquid aerosols or (ii) Blending drug particulates with dry carrier particles typically composed of sugars. Although effective for localised delivery of small drug molecules, these methods did not meet the complex formulation and delivery challenges posed by the newer biotechnology-derived medicines. One of the many avenues being explored to overcome these issues is the use of novel controlled drug release technologies. While such systems are available in the market for the oral and the parenteral routes, lung equivalent models remain elusive. If developed and optimised, these carrier-based formulations have the potential to not only provide controlled drug release, but also offer other advantages such as increased drug penetration to the distal parts of the lung, prolonged residence time of the drug in situ, and improved in vivo drug stability. While a number of biodegradable polymers have been studied for drug delivery to the lungs, little comparative data on the aerodynamic properties or toxicity and immunogenic potential of these polymers is available to allow formulation scientists to assess their usefulness for particular applications, e.g. local versus systemic delivery. In the present study, a range of commonly available polymers was used to prepare inhalable aerosol particles. A comparative evaluation of these particles was performed to judge their suitability as carriers for local or systemic delivery of proteins in the lungs (Chapters 2 and 3). As drug transport across the respiratory epithelium can also be an issue, various permeation enhancers were also screened for their effectiveness in promoting the absorption of a model protein (Chapter 4). Certain polymers showed particular promise for specfi protein delivery needs in the lungs, such as HPC to improve flow properties and sodium hyaluronate, alginate and chitosan for controlled drug release. In general, the polysaccharides showed no in vitro toxicity and immunogenicity as compared to the proteins (gelatin and ovalbumin) or the synthetic polymer, Poly (lactide-co-glycolide). Using a cell-based screening method, the transport of the model protein, parathyroid hormone (PTH) in the presence of permeation enhancers was found to be formulation dependent. While solutions containing sodium taurocholate (STC) and poly-L-arginine showed increased PTH transport, no difference in transport levels were observed when the formulations were administered as a dry powder (Chapter 4). Spray drying, in general, compromised the stability and bioactivity of PTH. Due to their surfactant-like properties, the inclusion of STC and sodium dioctyl sulfosuccinate in the formulations improved the stability of PTH against spray drying induced stresses (Chapter 4). Bioresponsive \u27smart\u27 polymers that respond to environmental stimuli such as the presence of enzymes were explored in the next two chapters. The aim was to develop microparticles degradable by the enzyme elastase as potential drug carriers to treat lung diseases such as emphysema and Chronic Obstructive Pulmonary Disease (COPD) associated with high levels of this enzyme. Two novel technologies were developed: 1) a system based on natural polymers consisting of an interpenetrating network of a protein (elastin) and a polysaccharide (alginate) (Chapter 5) and 2) a synthetic polymer-based system consisting of a crosslinked network of polyethylene glycol (PEG) with elastase sensitive peptide sequences inserted in the backbone (Chapter 6). Although the elastin-alginate system showed favourable properties, including high drug encapsulation, good aerosolisation performance and no cytotoxicity, the use of animal-derived excipients, non-specific drug release and possible degradation by other enzymes are limitations to be overcome (Chapter 5). The PEG-based systems have a peptide sequence in the network that can be cleaved only by the target enzyme (in this case, elastase). These systems are therefore tailored to display high enzyme specificity (Chapter 6). Further work on improving the drug loading and aerosolisabilly of these systems is however, required. The use of relatively safe excipients and mild particle fabrication conditions coupled with high enzyme specificity makes the PEG-based systems better suited as enzyme targeted drug delivery systems

In vivo animal models for drug delivery across the lung mucosal barrier.

Over recent years the research focus within the field of respiratory drug delivery has broadened to include a wide range of potential applications for inhalation by delivering drugs not just onto the lung mucosa but across it. The range of drugs being assessed is broad and includes both current and novel therapies and there are a growing number of additives that appear capable of enhancing systemic absorption. Comprehensive characterisation of drug delivery to the lungs is a complex task involving the determination of delivered, deposited and (for systemically-targeted drugs) absorbed dose. As it is difficult to simulate in vitro, in vivo whole animal models are still key to inhaled drug development. Because of the anatomical complexities and interspecies differences in the lungs, the appropriate choice of species and drug delivery method is vital during study design. New delivery devices designed specifically for animal studies as well as more sophisticated methods to determine drug deposition and absorption after inhalation are improving the information derived from these studies

A comparative study of a range of polymeric microspheres as potential carriers for the inhalation of proteins

The aim of this study was to compare protein-loaded inhalable microparticles manufactured using a range of biocompatible polymers including hydroxypropyl cellulose (HPC), chitosan, hyaluronic acid, alginate, gelatin, ovalbumin and poly (lactide-co-glycolide)(PLGA). Spray drying was used to prepare microparticles containing bovine serum albumin labeled with flourescein isothiocyanate (BSA-FITC). Particles of respirable size and high protein loading were obtained. No evidence of BSA degradation was seen from PAGE analysis. The microparticles were mixed with mannitol as a carrier and powder aerosolization was assessed with a multi-dose dry powder inhaler (DPI) using a multi-stage cascade impactor. The mass median aerodynamic diameter (MMAD) ranged between 2.9-4.7μm. Potential polymer toxicity in the lungs was compared by impinging the particles on Calu-3 monolayers and assessing the cytotoxicity, induction of cytokine release, changes in transepithelial permeability and electrical resistance. No toxic effects were observed with most of the polymers though some evidence of compromised cell monolayer integrity was seen for PLGA and ovalbumin. PLGA and gelatin microparticles caused a significant increase in IL-8 release. Of the polymers studied, PLGA showed the greatest toxicity. Certain polymers showed particular promise for specific protein delivery needs in the lungs, such as HPC to improve flow properties, sodium hyaluronate for controlled release, and chitosan and ovalbumin for systemic delivery

The Effects of Excipients and Particle Engineering on the Biophysical Stability and Aerosol Performance of Parathyroid Hormone (1-34) Prepared as a Dry Powder for Inhalation

Pulmonary delivery of therapeutic peptides and proteins has many advantages including high relative bioavailability, rapid systemic absorption and onset of action and a non-invasive mode of administration which improves patient compliance. In this study, we investigated the effect of spray-drying (SD) and spray freeze-drying processes on the stability and aerosol performance of parathyroid hormone (PTH) (1-34) microparticles. In this study, the stabilisation effect of trehalose (a non-reducing sugar) and Brij 97 (a non-ionic surfactant) on spray-dried PTH particles was assessed using analytical techniques including circular dichroism (CD), fluorescence spectroscopy, modulated differential scanning calorimetry and an in vitro bioactivity assay. Physical characterisation also included electron microscopy, tap density measurement and laser light diffraction. The aerosol aerodynamic performance of the formulations was assessed using the Andersen cascade impactor. Based on these studies, a formulation for spray freeze-drying was selected and the effects of the two particle engineering techniques on the biophysical stability and aerosol performance of the resulting powders was determined. CD, fluorescence spectroscopy and bioactivity data suggest that trehalose when used alone as a stabilising excipient produces a superior stabilising effect than when used in combination with a non-ionic surfactant. This highlights the utility of CD and fluorescence spectroscopy studies for the prediction of protein bioactivity post-processing. Therefore, a method and formulation suitable for the preparation of PTH as a dry powder was developed based on spray-drying PTH with trehalose as a stabiliser with the bioactivity of SD PTH containing trehalose being equivalent to that of unprocessed PTH

'Smart' non-viral delivery systems for targeted delivery of RNAi to the lungs.

The emergence of RNAi offers a potentially exciting new therapeutic paradigm for respiratory diseases. However, effective delivery remains a key requirement for their translation into the clinic and has been a major factor in the limited clinical success seen to date. Inhalation offers tissue-specific targeting of the RNAi to treat respiratory diseases and a diminished risk of off-target effects. In order to deliver RNAi directly to the respiratory tract via inhalation, 'smart' non-viral carriers are required to protect the RNAi during delivery/aerosolization and enhance cell-specific uptake to target cells. Here, we review the state-of-the-art in therapeutic aerosol bioengineering, and specifically non-viral siRNA delivery platforms, for delivery via inhalation. This includes developments in inhaler device engineering and particle engineering, including manufacturing methods and excipients used in therapeutic aerosol bioengineering that underpin the development of smart, cell type-specific delivery systems to target siRNA to respiratory epithelial cells and/or alveolar macrophages.</p

A comparative study of a range of polymeric microspheres as potential carriers for the inhalation of proteins.

The aim of this study was to compare protein-loaded inhalable microparticles manufactured using a range of biocompatible polymers including hydroxypropyl cellulose (HPC), chitosan, hyaluronic acid, alginate, gelatin, ovalbumin and poly(lactide-co-glycolide) (PLGA). Spray-drying was used to prepare microparticles containing bovine serum albumin labeled with fluorescein isothiocyanate (BSA-FITC). Particles of respirable size and high protein loading were obtained. No evidence of BSA degradation was seen from PAGE analysis. The microparticles were mixed with mannitol as a carrier and powder aerosolization was assessed with a multi-dose dry powder inhaler (DPI) using a multi-stage cascade impactor. The mass median aerodynamic diameter (MMAD) ranged between 2.9 and 4.7 microm. Potential polymer toxicity in the lungs was compared by impinging the particles on Calu-3 monolayers and assessing the cytotoxicity, induction of cytokine release, changes in transepithelial permeability and electrical resistance. No toxic effects were observed with most of the polymers though some evidence of compromised cell monolayer integrity was seen for PLGA and ovalbumin. PLGA and gelatin microparticles caused a significant increase in IL-8 release. Of the polymers studied, PLGA showed the greatest toxicity. Certain polymers showed particular promise for specific protein delivery needs in the lungs, such as HPC to improve flow properties, sodium hyaluronate for controlled release, and chitosan and ovalbumin for systemic delivery.</p