3rd Minicircle & DNA Vector Conference., Bielefeld, 2014
Few promoters can direct high level, sustained expression from non-viral vectors in the murine lung. However, such expression is possible following aerosol delivery of CpG-free plasmids containing hCEFI, a synthetic enhancer/promoter designed as a hybrid of CpG free versions of the human CMV enhancer (hC) and human Elongation Factor 1a (EF) promoter [Hyde et al, 2008, Nat Biotech 26(5):549].
This promoter is currently being utilised in a clinical trial for CFTR gene delivery to the lungs of patients with Cystic Fibrosis. Deletion experiments have identified only the 3’ 93bp of the full-length 302bp promoter to be necessary for high-level and persistent transgene expression. A matrix of plasmids comparing combinations of CpG-free or CpG-rich backbones, with CpG-rich or CpG-free transgenes, were evaluated for reporter gene expression up to 10 weeks post-delivery. In every case, CpG motifs in the transgene significantly reduced luciferase expression, independent of TLR-9 pathway or the gene transfer agent used.
Additionally, when CpG-rich or CpG-free transgenes were evaluated as minicircles, the results mirrored conventional plasmids, confirming that the level and persistence of reporter gene expression in the lung is markedly affected by CpG content.