Lilley, Caroline Elizabeth; (2000) Herpes simplex virus vectors for gene delivery to the CNS: Applications in the study of Alzheimer's disease. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Understanding the interactions between the presenilins and amyloid precursor protein (APP) remains one of the most challenging questions in Alzheimer's research. In vivo studies are currently limited to the introduction of exogenous genes into the germline to produce transgenic mice. The study of gene combinations then requires the generation and subsequent crossing of multiple transgenic lines. This thesis reports the development of a system to offer improvements to both cell culture and animal studies of APP/presenilin interactions. This system is based on disabled herpes simplex virus vectors which can directly deliver genes to neurons either in vitro or in vivo, allowing multiple mutations to be rapidly introduced and evaluated. The viral vectors constructed have been disabled by the deletion or inactivation of genes encoding ICP34.5/ORF P, VP 16, ICP27, ICP4 and vhs and consequently do not express significant amounts of any of the IE gene products. The vectors are non-toxic to cultured neurons at high MOI and are capable of expressing multiple exogenous genes in cell culture or in vivo. Gene expression is maintained in the long term using promoter systems containing elements from the HSV1 latency associated transcript promoter. Following injection into the CNS in vivo, widespread exogenous gene expression occurs as a result of retrograde transport of the vector from the site of inoculation to connected sites within the nervous system. Wild type and mutant APP and presenilin 1 were introduced into these viruses both individually and in combination. The viruses were then used to deliver these genes to primary neuronal cultures from rodent and non-rodent origins. The significance of using non-rodent neurons and observations from these experiments are described. The HSV-1 system described in this thesis should provide powerful means by which gene combinations relevant to Alzheimer's disease can be rapidly introduced and evaluated.

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