P20 Optimised dystrophin mini-constructs for gene delivery

The aim of our project is to obtain an optimal dystrophin construct that will be lentivirally-transduced into skeletal muscle stem cells derived from Duchenne Muscular Dystrophy (DMD) patients. These stem cells would restore functional dystrophin expression following their transplantation into dystrophin-deficient skeletal muscles. To this end, we have cloned a number of different dystrophin truncated forms ranging from 4.2 kb to 8.4 kb cDNA, encoding various components of the dystrophin molecule that may be used in a lentiviral system focusing on elongated transcript length and selected dystrophin domains. The constructs were assessed using in vivo and in vitro tests for functionality, such as membrane stability following osmotic shock as measured by creatine kinase release in myotubes and electroporation into mdx muscle. The construction, cloning strategy and characterisation of the constructs as well as first functional results will be presented. We are aware of challenges to gene transfer approaches and know that further long-term experiments are required to assess the potential of this strategy, but this project may bring us closer to a robust and reproducible system for testing functionality of both engineered and naturally occurring dystrophin mutants.