Purpose: To demonstrate the importance of dose and drug release rate for pulmonary targeting of inhaled glucocorticoids using an animal model of intrapulmonary drug deposition.
Methods: Liposomes composed of 1,2-distearoyl phosphatidylcholine (DSPC), 1,2-distearoyl phosphatidylglycerol (DSPG) and triamcinolone acetonide phosphate (TAP) or liposomes containing triamcinolone acetonide (TA) were prepared by a mechanical dispersion method followed by extrusion through polycarbonate membranes. Encapsulation efficiency was assessed after size exclusion gel chromatography by reverse phase HPLC. The effect of liposome size (200 nm and 800 nm) on the release kinetics of water-soluble encapsulated material was determined in vitro at 37 degrees C using 6-carboxyfluorescein as a marker and Triton X-100 (0.03%) as a leakage inducer. To investigate the relationship between drug release and pulmonary targeting, 100 micrograms/kg of TAP in 800 nm liposomes was delivered to male rats by intratracheal instillation (IT) and the results compared to data for 100 micrograms/kg TA liposomes (recently shown to exhibit a rapid drug release under sink conditions) and to previous studies reported for an equal dose of TAP in solution and TAP in 200 nm (1). Pulmonary targeting was assessed by simultaneously monitoring glucocorticoid receptor occupancy over time in lung and liver using an ex vivo receptor binding assay as a pharmacodynamic measure of glucocorticoid action. To assess the effect of dose on pulmonary targeting experiments were performed using 2.5, 7.5, 25, 100, and 450 micrograms/kg of TAP in 800 nm liposomes.
Results: The in vitro efflux of 6-carboxyfluorescein from (DSPC:DSPG) liposomes after exposure to Triton-X was biexponential. The terminal half-lives of 3.7 h and 9.0 h for the 200 nm and 800 nm liposomes, respectively, demonstrated that larger liposomes promote slower release of encapsulated water-soluble solute while previous results already indicated that encapsulation of lipophilic TA does not result in sustained release. Pulmonary targeting, defined as the difference between cumulative lin and liver receptor occupancies was most pronounced for the 800 nm liposomes (370%xh), followed by the 200 nm preparation (150%xh). No targeting was observed for TAP in solution (30%xh) or the rapid releasing TA liposome preparation. Correspondingly, the mean pulmonary effect time (MET) increased from 2.4-3.0 hr for TA liposomes or TAO in solution to 5.7 h and > 6.2 h for TAP in 200 nm and in 800 nm liposomes, respectively. Escalating doses of TAP encapsulated in 800 nm liposomes revealed a distinct bell shaped relationship between the TAP dose and pulmonary targeting with a maximum occurring at 100 micrograms/kg (370%xh).
Conclusions: The in vivo data presented here confirm that pulmonary residence time and dose affect the extent of lung targeting of glucocorticoids delivered via the lung.