Gilboy, P.; (2005) Particle-mediated DNA delivery: evaluation of parameters affecting efficacy in vivo and in vitro. Doctoral thesis , University of London.

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Abstract

Particle-mediated DNA delivery (PMDD) is a technique that uses compressed helium gas to accelerate microscopic gold beads coated with DNA into the skin. As a vaccination tool, it provides an attractive approach for generating antigen specific immunotherapy. As the number of applications for this technology in pre-clinical disease models grows, the need for in vitro surrogate models, which in part, can replace in vivo testing, is becoming increasingly important. Additionally, before it can be utilised successfully in man, a number of critical parameters which contribute to overall efficacy in vivo need to be more fully evaluated. To these ends, an in vitro model which featured PMDD as the means of plasmid delivery was developed. The relationship between the efficiency of PMDD to stimulate antigen specific CD8+ T cell responses in vitro and in vivo under conditions of single and dual plasmid immunisation was explored. Important aspects of the technology such as the association between antigen dose and resulting response were investigated. It was found that the in vitro model correlated well with the early stage primary response in vivo but this correlation deteriorated post boost. In addition, the relative contribution of antigenic competition and co-operation in shaping the resulting response was assessed after co-immunising with two independent antigens. Under the experimental conditions investigated, the dominant type of interaction was found to be competitive. Results suggested that competition occurred at the level of the T cell. Competition was most pronounced when both antigens were delivered in a linked manner and when both antigens were presented on the same APC. These results highlight the requirement for optimising key technical parameters in PMDD. Additionally, it demonstrates that interclonal T cell competition is a functionally relevant phenomenon and that it can occur when standard DNA vaccination strategies are employed.

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