Branston, Ruth Helen; (2001) Development of HSV-1 vectors for applications in the peripheral nervous system. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Herpes Simplex Virus (HSV) is potentially an ideal vector for gene therapy in the peripheral nervous system for a number of reasons. HSV is neurotrophic with a life cycle that involves retrograde transport of the virion from peripheral nerve terminals to sensory neuron cell bodies. Thus, for gene delivery to cell bodies in dorsal root ganglia there would be no need for vector administration by an invasive surgical route. In addition HSV has evolved the ability to enter life long latency in the nuclei of infected sensory neurons, providing the potential for very long-term transgene expression. This study aimed to identify and optimise the use of attenuated HSV-1 vectors that are non-pathogenic, non- cytotoxic and able to express transgenes during both lytic and latent infection. Several vectors deleted for various combinations of non-essential and/or essential virus genes were designed and evaluated and optimal replication, competent and incompetent vectors for peripheral gene delivery were identified. In addition, transgene expression cassettes consisting of different promoter/reporter gene arrangements were tested for their ability to express single or multiple genes in the long-term. Using this information, optimised vectors containing either the neuropeptide galanin or Cre recombinase were constructed for use in nerve regeneration studies or conditional gene expression respectively. In further studies aimed at developing regulatable gene expression systems for HSV, vectors were generated containing a number of ligand inducible transgene expression cassettes. Of these, the ecdysone regulatable system was found to be most promising and replication competent and incompetent ecdysone inducible vectors were constructed and found to give regulated gene expression in various circumstances. Overall the work described in this thesis resulted in the successful development of HSV vectors for the PNS and the development of regulatable HSV gene expression systems which are highly promising for future work.

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