1. ¹Nuffield Department of Clinical Laboratory Sciences, University of Oxford, John Radcliffe Hospital, Oxford, UK
2. ²The UK Cystic Fibrosis Gene Therapy Consortium
3. ³Department of Medical Genetics, College of Medicine and Veterinary Medicine, University of Edinburgh, Edinburgh, UK
4. ⁴Department of Gene Therapy, National Heart and Lung Institute, Imperial College, London, UK
5. ⁵Department of Veterinary Clinical Sciences, College of Medicine and Veterinary Medicine, University of Edinburgh, Edinburgh, UK
6. ⁶Medical Genetics Section, School of Molecular and Clinical Medicine, University of Edinburgh, Edinburgh, UK

Correspondence: Deborah, Gill, Nuffield Department of Clinical Laboratory Sciences, University of Oxford, John Radcliffe Hospital, Oxford OX3 9DU, UK. E-mail: Deborah.Gill@ndcls.ox.ac.uk

Received 14 November 2007; Accepted 7 April 2008; Published online 20 May 2008.

Abstract

A major limitation of many self-assembling nonviral gene transfer formulations is that they are commonly prepared at relatively low component concentrations. While this typically has little impact on their use in cell culture, it can severely limit the progress of in vivo studies. In order to overcome this, we have developed a simple, scalable, pharmaceutically acceptable concentration method that has allowed us to increase the concentration of a commonly used pDNA/PEI formulation from 0.2 to >8 mg/ml plasmid DNA (pDNA). Crucially, the concentration method was found to have only minimal impact on the electrostatic properties or size of the pDNA/PEI particles. When delivered as an aerosol to the mouse lung, the concentrated pDNA/PEI formulations resulted in a 15-fold increase in lung reporter gene expression, with minimal impact in terms of inflammation or toxicity. Importantly, this performance advantage was replicated after aerosol administration to sheep lungs, with reporter gene expression being similarly 15-fold higher than with the conventional pDNA/PEI formulation, and lung inflammation falling to background levels. These findings demonstrate that concentrated pDNA/PEI formulations offer increased aerosol gene transfer with decreased inflammatory sequelae, and represent a promising advance in the field of nonviral lung gene transfer. It seems likely that similar benefits might be achievable with alternative delivery routes and with other nonviral formulations.