Journal of Cystic Fibrosis, 3 S1 S29
The European Cystic Fibrosis Conference, Birmingham, 2004
As part of the UK Cystic Fibrosis Gene Therapy Consortium, we are aiming to identify gene transfer agents (GTA) that are candidates for delivery of CFTR to human airways. Typically, products identified from the literature or developed by members of the Consortium, including targeting peptides (e.g. HIV-TAT), lipids/liposomes (e.g. GL67) and polymers (e.g. JetPEITM), are tested in a series of assays, through which products can be evaluated systematically. The GTA are screened first in the mouse nose using a luciferase reporter gene to identify promising formulations; these are then taken through to our nPD (nasal potential difference) core facility where changes in conductance of nasal airway are measured in CF mice after treatment with a CFTR-encoding plasmid in the chosen formulation. The next stage is aerosolisation of the GTA in sheep lungs, which models the physical delivery of our chosen GTA, in anticipation of human clinical trials. The initial testing in the mouse nose involves the perfusion of 100?l of GTA into the left nostril of C57BL6 mice over 15'. Our studies are powered at 0.8-0.95 with n=6-8 per group. Luciferase and protein assays (with intra- and inter-assay variability <=10%) are combined to give activity in relative light units/mg protein. The GTA are tested in standardised conditions, including a dose response starting from the Maximum Feasible Dose (MFD) to one tenth of MFD. Additionally, various ratios of components can be tested. In the last 6 months, we have tested over 20 GTA. We anticipate being able to test 1-2 novel products per week. We will show examples of the iterative process required to optimise airway specific gene expression of a particular GTA prior to its progression to the next stage of evaluation.