Repeat Aerosol Delivery of Concentrated PEI/pDNA to the Sheep Lung.

McLachlan G, Davies LA, Gordon CM, Vrettou C, Baker E, Tennant P, Baker A, Coles RL, McCormick D, Sumner-Jones SG, Hyde SC, Gill DR, Collie DD

Molecular Therapy, Vol 18, S1, Abstract 880

The American Society of Gene and Cell Therapy Annual Conference, Washington DC, 2010

Considerable advances have been made in lung-directed gene therapy, however it is unlikely that clinical beneft to patients with chronic lung disease can be achieved without repeat administration. For viral vectors, the ability to deliver multiple doses is normally limited by the host immune response, which results in attenuated expression following consecutive exposures. Importantly, aerosol delivery of non-viral gene transfer agents to the lung is not generally associated with the same problem.

We have previously demonstrated that aerosol delivery of plasmid DNA (pDNA) complexed with polyethylenimine (PEI) to sheep, either at the standard (low) concentration or with a novel concentrated formulation (cPEI; Davies et al, 2008, Mol Ther 16:1283) induces a mild transient inflammatory response characterised by increased neutrophils in bronchoalveolar lavage (BAL) fluid. Here we have examined the effect of repeat dosing with cPEI. Two groups of sheep (n=6) were each administered a single dose of cPEI/pCIKLux (32mg DNA at 1.6mg/ml). Expression of luciferase (lux) protein and mRNA were measured either at day 1 (Group 1) or day 15 (Group 2).

Mean lux protein expression at day 1 was 19.4 +/- 6.5 RLU/mg lung protein, which fell to background levels at day 15. A third group was given two doses 14 days apart and gene expression assessed at day 15. Mean Lux expression after a repeat dose was 15.7 +/- 3.7 RLU/mg and was not signifcantly different to day 1 level from a single dose (Group 1). A similar pattern of expression was observed when vector-derived lux mRNA was quantifed in these animals. The mean value from a single dose at day 1 was 62.9 % vector mRNA/endogenous ovCFTR mRNA (%v/e), falling to 0.4 %v/e at day 15.

mRNA expression following two doses was somewhat lower than the level obtained with a single dose at day 1 (11.8 %v/e; p=0.03). Consistent with previous studies, the level of neutrophils in BAL fuid in response to a single dose was 49% on day 1, which decreased to 5% (within normal range) by day 15. Unexpectedly the BAL neutrophil response observed at day 1 following repeat dosing was signifcantly lower than after a single dose (∼23%; p=0.02).

These results indicate that a second aerosol administration of the cPEI/pDNA formulation to the whole lung of a large animal directs repeated transgene expression and induces a reduced BAL neutrophil response compared with the frst dose, suggesting that multiple administrations may be effective and well tolerated.

These data provide the frst evidence that repeated aerosol delivery of a non-viral formulation to the sheep lung is feasible and support the potential use of cPEI formulations in clinical applications where repeated administration to the lung is required.