Mitigation of Hydrochloric Acid (HCl)-Induced Lung Injury in Mice by Aerosol Therapy of Surface-Active Nanovesicles Containing Antioxidant and Anti-inflammatory Drugs.

Acute lung injury leading to alveolar inflammation and surfactant dysfunction remains a medical challenge. Surface-active lipid nanovesicles of 200-250 nm size with antioxidant D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) and anti-inflammatory drug dexamethasone disodium phosphate (DXP) dual combination (Dual-NV) were developed for delivery as aerosols by nebulization in acid lung injury models. Drug deposition studies showed Dual-NV deposited ∼2.5 times more DXP compared to equivalent DXP solution. Nanovesicles are actively internalized by A549 cells through ATP- and clathrin-dependent pathways. The nanovesicles could be phagocytosed by RAW 264.7 macrophages and were nonimmunogenic and did not elicit overproduction of TNF-α, IL-1ß, and IL-6. Dual-NV aerosol therapy at 200 mg/kg body weight, in HCl acid-induced lung injury in mice, markedly reduced pulmonary hemorrhage and protein leakage and improved capillary (airway) patency to ∼96%. Dual-NV aerosol therapy also significantly lowered production of inflammatory cytokine IL-1ß, IL-6, and TNF-α and reduced oxidative stress by ∼95% in the injured group. Surface-active Dual-NV aerosol therapy is promising for replenishing the dysfunctional surfactant pool and mitigating inflammation and oxidative stress in lung injuries.

Exploitation of sub-micron cavitation nuclei to enhance ultrasound-mediated transdermal transport and penetration of vaccines.

Inertial cavitation mediated by ultrasound has been previously shown to enable skin permeabilisation for transdermal drug and vaccine delivery, by sequentially applying the ultrasound then the therapeutic in liquid form on the skin surface. Using a novel hydrogel dosage form, we demonstrate that the use of sub-micron gas-stabilising polymeric nanoparticles (nanocups) to sustain and promote cavitation activity during simultaneous application of both drug and vaccine results in a significant enhancement of both the dose and penetration of a model vaccine, Ovalbumin (OVA), to depths of 500µm into porcine skin. The nanocups themselves exceeded the penetration depth of the vaccine (up to 700µm) due to their small size and capacity to 'self-propel'. In vivo murine studies indicated that nanocup-assisted ultrasound transdermal vaccination achieved significantly (p<0.05) higher delivery doses without visible skin damage compared to the use of a chemical penetration enhancer. Transdermal OVA doses of up to 1µg were achieved in a single 90-second treatment, which was sufficient to trigger an antigen-specific immune response. Furthermore, ultrasound-assisted vaccine delivery in the presence of nanocups demonstrated substantially higher specific anti-OVA IgG antibody levels compared to other transdermal methods. Further optimisation can lead to a viable, safe and non-invasive delivery platform for vaccines with potential use in a primary care setting or personalized self-vaccination at home.

Ultrasound-Propelled Nanocups for Drug Delivery.

Ultrasound-induced bubble activity (cavitation) has been recently shown to actively transport and improve the distribution of therapeutic agents in tumors. However, existing cavitation-promoting agents are micron-sized and cannot sustain cavitation activity over prolonged time periods because they are rapidly destroyed upon ultrasound exposure. A novel ultrasound-responsive single-cavity polymeric nanoparticle (nanocup) capable of trapping and stabilizing gas against dissolution in the bloodstream is reported. Upon ultrasound exposure at frequencies and intensities achievable with existing diagnostic and therapeutic systems, nanocups initiate and sustain readily detectable cavitation activity for at least four times longer than existing microbubble constructs in an in vivo tumor model. As a proof-of-concept of their ability to enhance the delivery of unmodified therapeutics, intravenously injected nanocups are also found to improve the distribution of a freely circulating IgG mouse antibody when the tumor is exposed to ultrasound. Quantification of the delivery distance and concentration of both the nanocups and coadministered model therapeutic in an in vitro flow phantom shows that the ultrasound-propelled nanocups travel further than the model therapeutic, which is itself delivered to hundreds of microns from the vessel wall. Thus nanocups offer considerable potential for enhanced drug delivery and treatment monitoring in oncological and other biomedical applications.

Inertial cavitation to non-invasively trigger and monitor intratumoral release of drug from intravenously delivered liposomes.

The encapsulation of cytotoxic drugs within liposomes enhances pharmacokinetics and allows passive accumulation within tumors. However, liposomes designed to achieve good stability during the delivery phase often have compromised activity at the target site. This problem of inefficient and unpredictable drug release is compounded by the present lack of low-cost, non-invasive methods to measure such release. Here we show that focused ultrasound, used at pressures similar to those applied during diagnostic ultrasound scanning, can be utilised to both trigger and monitor release of payload from liposomes. Notably, drug release was influenced by liposome composition and the presence of SonoVue® microbubbles, which provided the nuclei for the initiation of an event known as inertial cavitation. In vitro studies demonstrated that liposomes formulated with a high proportion of 1,2 distearoyl-sn-glycero-3-phosphoethanolamine (DSPE) released up to 30% of payload following ultrasound exposure in the presence of SonoVue®, provided that the exposure created sufficient inertial cavitation events, as characterised by violent bubble collapse and the generation of broadband acoustic emissions. In contrast a 'Doxil'-like liposome formulation gave no such triggered release. In pre-clinical studies, ultrasound was used as a non-invasive, targeted stimulus to trigger a 16-fold increase in the level of payload release within tumors following intravenous delivery. The inertial cavitation events driving this release could be measured remotely in real-time and were a reliable predictor of drug release.

Effect of D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) on surfactant monolayers.

In the present study, the effects of an amphiphilic polymer, d-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) on model surfactant monolayers dipalmitoylphosphatidylcholine (DPPC), a binary mixture of DPPC with palmitoyloleoyl phosphatidylglycerol (DPPC-POPG) 9:1 (w/w) and binary mixture of DPPC and oleic acid (DPPC-OA) were evaluated. The ability of TPGS to act as an antioxidant adjuvant for pulmonary surfactants was also evaluated. Compression isotherms of surfactant monolayers at 37 °C in a Langmuir-Blodgett trough showed that DPPC and DPPC:TPGS mixed monolayers (1:0.25-1:1, w/w) exhibited low minimum surface tensions (MST) of 1-2 mN/m. Similarly [DPPC:POPG (9:1, w/w)]:TPGS mixed films of 1:0.25-1:1 weight ratios reached 1-2 mN/m MST. DPPC:POPG:TPGS liposomes adsorbed to surface tensions of 29-31 mN/m within 1s. While monolayers of DPPC:OA (1:1, w/w) reached high MST of ∼11 mN/m, DPPC:OA:TPGS (1:1:0.25, w/w) film reached near zero MST suggesting that low concentrations of TPGS reverses the effect of OA on DPPC monolayer. Capillary surfactometer studies showed DPPC:TPGS and [DPPC:POPG (9:1, w/w)]:TPGS liposomes maintained 84-95% airway patency. Fluorescence spectroscopy of Laurdan loaded DPPC:TPGS and DPPC:POPG:TPGS liposomes revealed no segregation of lipid domains in the lipid bilayer. Addition of TPGS to soybean liposome significantly reduced thiobarbituric acid reactive substance (TBARS) by 29-39% confirming its antioxidant nature. The results suggest a potential use of TPGS as an adjuvant to improve the surfactant activity as well as act as an antioxidant by scavenging free radicals.