OPTIMIZATION OF FORMULATIONS AND CONDITIONS FOR THE AEROSOL DELIVERY OF FUNCTIONAL CATIONIC LIPID-DNA COMPLEXES

We have examined several variables inherent in aerosolizing cationic l ipid:DNA complexes using a jet nebulizer and thereby have optimized th e delivery of functional complexes. Maximal aerosol transfer efficienc y of cationic lipid:pDNA complexes was quantitated and shown to requir e the presence of at least 25 mM NaCl as an excipient. This is possibl y related to effects on the measured zeta potentials of the complex, w hich indicate that the complexes are more highly charged in solutions of physiological ionic strength than in solutions of low ionic strengt h. Inclusion of saline also resulted in retention of the starting lipi d to plasmid DNA (pDNA) ratio following nebulization. These data were used to design in vitro aerosolization experiments with tissue culture cells that resulted in the identification of a cationic lipid:pDNA ra tio of 0.75:1 (mol:mol) as being optimal for aerosolization. This form ulation largely protected pDNA from shear degradation during nebulizat ion and produced a respirable aerosol droplet size (1-3 mu m). It was tested further in a mouse model and shown to result in the dose-depend ent transfection of mouse lungs, generating the equivalent of several picograms of reporter gene activity per mouse lung. The results of the se experiments have provided a set of optimal conditions for nebulizin g cationic lipid:pDNA complexes that can be used as a starting point f or the further evaluation of aerosol delivery of these nonviral gene d elivery vectors in vivo.