Receptor-mediated targeting of spray-dried lipid particles coformulated with immunoglobulin and loaded with a prototype vaccine.

PURPOSE: Spray-dried lipid-based microparticles (SDLM) serve as a platform for delivery of a wide variety of compounds including peptides, proteins, and vaccines to the respiratory mucosa. In the present study, we assessed the impact of IgG-mediated targeting to phagocytic cells of inactivated influenza virus formulated in SDLM, on subsequent immune responses. METHODS: SDLM were produced containing inactivated influenza virus strain A/WSN/32/H1N1 (WSN), with or without IgG. Using phagocytic antigen presenting cells (APC) and a T cell hybridoma (TcH) line specific for a dominant influenza virus epitope, we compared the in vitro responses elicited by ligand-formulated (SDLM-IgG-WSN) and non-ligand particles (SDLM-WSN). The effect of including the IgG ligand in the formulation was further characterized by measuring the immune responses of rodents vaccinated with SDLM. RESULTS: SDLM-IgG-WSN were internalized in an Fc receptor (FcR)-dependent manner by phagocytic APC that were then able to effectively present a dominant, class II-restricted epitope to specific T cells. While SDLM-WSN elicited a lower response than administration of plain inactivated virus in saline, the level of the T cell response was restored both in vitro and in vivo by incorporating the APC FcR ligand, IgG, in the SDLM. CONCLUSIONS: Incorporation of FcR ligand (IgG) in SDLM restored the limited ability of formulated virus to elicit T-cell immunity, by receptor-mediated targeting to phagocytes.

Novel lipid-based hollow-porous microparticles as a platform for immunoglobulin delivery to the respiratory tract.

PURPOSE: Delivery of specific antibodies or immunoglobulin constructs to the respiratory tract may be useful for prophylaxis or active treatment of local or systemic disorders. Therefore, we evaluated the utility of lipid-based hollow-porous microparticles (PulmoSpheres) as a potential delivery vehicle for immunoglobulins. METHODS: Lipid-based microparticles loaded with human immunoglobulin (hIgG) or control peptide were synthesized by spray drying and tested for: i) the kinetics of peptide/protein release, using ELISA and bioassays; ii) bioavailability subsequent to nonaqueous liquid instillation into the respiratory tract of BALB/c mice, using ELISA and Western blotting; iii) bioactivity in terms of murine immune response to xenotypic epitopes on human IgG, using ELISA and T cell assays; and iv) mechanisms responsible for the observed enhancement of immune responses, using measurement of antibodies as well as tagged probes. RESULTS: Human IgG and the control peptide were both readily released from the hollow-porous microspheres once added to an aqueous environment, although the kinetics depended on the compound. Nonaqueous liquid instillation of hIgG formulated in PulmoSpheres into the upper and lower respiratory tract of BALB/c mice resulted in systemic biodistribution. The formulated human IgG triggered enhanced local and systemic immune responses against xenotypic epitopes and was associated with receptor-mediated loading of alveolar macrophages. CONCLUSIONS: Formulation of immunoglobulins in hollow-porous microparticles is compatible with local and systemic delivery via the respiratory mucosa and may be used as means to trigger or modulate immune responses.

Hollow porous particles in metered dose inhalers.

PURPOSE: To assess the physical stability and aerosol characteristics of suspensions of hollow porous microspheres (PulmoSpheres) in HFA-134a. METHODS: Cromolyn sodium, albuterol sulfate, and formoterol fumarate microspheres were prepared by a spray-drying method. Particle size and morphology were determined via electron microscopy. Particle aggregation and suspension creaming times were assessed visually, and aerosol performance was determined via Andersen cascade impaction and dose uniformity studies. RESULTS: The hollow porous particle morphology allows the propellant to permeate freely within the particles creating a novel form of suspension termed a homodispersion, wherein the dispersed and continuous phases are identical, separated by an insoluble interfacial layer of drug and excipient. Homodispersion formation improves suspension stability by minimizing the difference in density between the particles and the medium, and by reducing attractive forces between particles. The improved physical stability leads to excellent dose uniformity. Excellent aerosolization efficiencies are also observed with PulmoSpheres formulations, with fine particle fractions of about 70%. CONCLUSIONS: The formation of hollow porous particles provides a new formulation technology for stabilizing suspensions of drugs in hydrofluoroalkane propellants with improved physical stability, content uniformity, and aerosolization efficiency.

Oxidative assessment of phospholipid-stabilized perfluorocarbon-based blood substitutes.

Methods based on the HPLC separation with subsequent UV detection and spectrofluorimetry have been developed to monitor the formation of oxidative decomposition products of phospholipids in perfluorocarbon emulsions. Catalytic, as well as emulsion oxidative stability studies have been conducted utilizing egg yolk phospholipid (EYP) and perfluorocarbons of varied compositions/purity in order to assess their effect on susceptibility to oxidation. Our studies indicate that phospholipid composition, degree of unsaturation, perfluorocarbon purity and the presence of oxygen and trace metals have a significant effect on the formation of oxidative decomposition products. A combination of methods has proven useful in monitoring the levels of oxidative decomposition products of phospholipids in phospholipid-stabilized perfluorocarbon-based blood substitutes. Such an approach has proven beneficial in the development of pharmaceutical agents of potentially higher quality and storage stability.

Stabilization of perflubron emulsions with egg yolk phospholipid.

Egg Yolk Phospholipid(EYP) has been used extensively as the primary surfactant in parenteral fat emulsions for many years. The simplicity, functionality and physiologic tolerance of EYP has contributed greatly to its success in the intravenous emulsion arena. The mechanism of stabilization in triglyceride emulsions is well understood; however, this is not the case with perfluorocarbon emulsions. Interfacial models, as well as emulsion stability studies, have been conducted utilizing EYP of varied composition in order to derive a structure/function relationship. Our studies indicate that minor components, total unsaturation, acyl chain length and presence of charged species have significant impact on the functional properties of EYP and the subsequent stability of the emulsion product. These findings contribute to our ability to design and manipulate natural surfactants with superior properties for use in medical applications of perfluorocarbon emulsions.