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Title: The impact of processing on the biophysical properties of synthetic DNA complexes for gene delivery
Author: Mount, Claire Nicole
ISNI:       0000 0001 3427 9781
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 2003
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Research is progressing fast to find safe and effective methods of delivering genes as therapeutic agents. A technical barrier that remains relates to the need for large-scale manufacturing process strategies and process optimisation. The present investigation provides a means of assessing the impact of the formulation process and material factors on the physical properties of plasmid-DNA vector. Response surface methodology (RSM) is combined with experimental techniques to provide new insight into scaleable processing of these systems. To assess the influence of process and material factors on the physical stability of gene complexes two techniques, photon correlation spectrophotometry (PCS) and laser Doppler velocimetry (LDV) were used to characterise DNA complexes. A model formulation was used consisting of ctDNA or pDNA MB113 condensed by poly-L-lysine (30KDa) and a lipopolypeptide formulation, LID, prepared through complexation of DNA with cationic lipid and peptide components. Preliminary experiments showed several individual factors have important effects on the physical stability of synthetic delivery systems; including the mean size and charge of plasmid DNA molecules condensed by cationic agents. These factors include physico-chemical properties of the buffer such as the pH and ionic strength as well as conditions used to prepare the complexes, for example the method and intensity of mixing the components of the complex. Using RSM to analyse experimental data it is shown that the impact of these factors and the effects of their interactions on the physical properties of the complexes are time-dependent. More specifically, for plasmid DNA condensed by poly-L-lysine or cationic lipids, interactions between ionic strength, pH and DNA concentration play a critical role. Whether poly-L-lysine should be used as a condensing agent in the final delivery system or the LID formulation can become clinically and commercially viable remains to be demonstrated. However, the use of RSM combined with the scaleable experimental approach described here represent a rapid and cost effect process tool that may be applied to any delivery system that requires optimisation to retain desirable physical properties.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available