7 th Australasian gene therapy society meeting

Targeted corrective gene conversion (TCGC) holds much promise as a future therapy for many hereditary diseases in humans, but there still remain significant impediments to effective mutation correction and it is clear that significant work remains to improve TCGC to levels where it can be considered for translation to the clinical setting. Nevertheless, mutation correction frequencies varying between 0.0001% and 40% have been reported using chimeraplasty, oligoplasty, triplex-forming oligonucleotides, and small corrective PCR amplicons. We have investigated potential mechanisms that impede effective gene correction and potential methods by which to improve the efficiency of dystrophin gene mutation correction in the mdx mouse model of Duchenne Muscular Dystrophy (DMD). Arrest of cell cycle using Hydroxyurea and double Thymidine blockade significantly improved TCGC frequency in mdx myoblasts at the gene level, but it was clearly evident that transfection reagents rendered TCGC efficiency variable with some, albeit limited expression of the corrected loci. The latter aspect was investigated by adjustment of transfection conditions, resulting in more robust gene correction with better (but still variable) expression of the corrected locus in a proportion of the treated cells, but still affected by the toxic chemistries used to mediate improved gene correction. These studies report new methods and strategies for improvement of TCGC by cell cycle arrest using molecular methods less toxic than transfection reagents or chemical cell cycle inhibitors: Molecular technologies such as PNA chemistry, advanced electromaterials, and CNA motifs that present lesional signals to the DNA metabolic machinery are potential technologies that may impart levels of efficiency that translate more efficiently from gene to protein levels and more readily facilitate consideration towards clinical application