Clinical translation of adeno-associated virus (AAV) and recombinant AAV (rAAV)-mediated gene therapy has been held back by a number of issues including cost and safety. AAVs are non-enveloped viruses with a 4.7Kb wild type genome flanked by inverted terminal repeats (ITRs). AAV production into icosahedral virions relies on two genes, rep and cap, which provide the proteins necessary for replication and encapsidation of the viral genome, in addition to adeno-helper proteins usually provided by an external source.
Present methods of production for vectors designed for clinical use utilise live viruses such as adenovirus or herpes simplex virus to supply the adeno-helper functions, which present significant health and safety concerns for in vivo usage.
The present invention provides a hybrid vector system comprising a phagemid/AAV (PAAV) particle. The novel genome construct is a phagemid, in this case a hybrid of a phage-derived genome and rAAV transgenes. Two origins of replication are present, one phage and one bacterial. This hybrid differentiates from conventional phagemids in that viral transgenes, rather than generic non-viral components, are incorporated into the phagemid under the influence of the bacterial promotor.
This novel hybrid contains no structural bacteriophage genes, which comprise over 50% of a conventional phage genome and are redundant in mammalian therapeutic use. Thus, this phagemid particle is dramatically smaller in size than previously developed hybrid constructs, allowing for increased efficacy in gene transduction and flexibility of the vector system, as large or numerous transgene inserts can be accommodated in the phagemid. This makes it a particularly advantageous system for target-specific gene therapy.