Developing regulatable lentiviral vectors for the treatment of nerve injuries and Parkinson’s disease

Abstract

Gene therapy is a promising approach for the treatment of diseases of the central nervous system. However, there is an unmet need for systems to regulate the levels of therapeutic transgenes. In this study, we investigate recently described destabilisation domain technology - novel regulatory systems that may offer advantages over existing methodologies. The destabilisation domain (DD) paradigm is composed of a conditionally stabilised protein domain that can be genetically fused to any protein of interest. Upon translation of this fusion partner in cells, the unstable DD targets the fusion protein to be degraded by the proteasome. The fusion protein is rescued by the presence of a ligand that stabilises the DD, forming a rapid, reversible and tunable regulatory system. We utilised two different forms of DD to create regulatable vectors suitable for use in vitro - to test the ability of the system to regulate fluorescent reporter proteins in cell lines and primary cultured neurons - and in vivo. After choosing the most favourable DD, we developed the technology to regulate levels of therapeutic genes suitable for the treatment of spinal cord injury (the nuclear receptor retinoic acid receptor β2) and Parkinson’s disease (the novel neurotrophic factor CDNF (conserved dopamine neurotrophic factor)). We found we could use the regulatable vector systems to provide dose-dependent gene expression in vitro, and long-term gene expression in vivo by oral administration of the stabilising ligand, with minimal levels of transgenes in the absence of the ligand. We suggest that with further optimisation, DD regulatory technology may provide an efficacious methodology to regulate levels of proteins in the central nervous system.

Date of Award	14 May 2013
Original language	English
Awarding Institution	The University of Bristol
Supervisor	Liang-Fong Wong (Supervisor)