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Development of new DNA vaccines utilising polymeric gene delivery system

Li, Xiong Wei; (2006) Development of new DNA vaccines utilising polymeric gene delivery system. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Strategies for gene delivery are comprised of a diverse range of live and synthetic approaches; DNA delivery for the purposes of immunisation in turn covers a large part of this research. This work aimed to investigate a number of synthetic systems for application in the delivery of plasmid DNA vaccines, mainly focusing on chitosan followed by PEI complex and PLGA and combination delivery systems with the aim to attempt to address the weaknesses of these systems and develop novel modified carriers/ vectors. The use of cationic polymers and biodegradable polymer particles as vehicles for DNA vaccine delivery has now been established. These systems, however, have limitations in the form of size control, instability or strong stability in the biological medium, low DNA loading efficiency, as in the case for PLGA, or potential toxicity as related to PEI complexes. Investigations of the mechanism of complex formation based on poly(methylacrylamide) (PMAA) derivatives with different cationic side-chains indicated that the interaction between polymers and DNA increases with increasing the cationic strength of the polymers, and polycation/DNA complexes are better formed at acidic pH. In this work, low molecular weight (11kDa) chitosan and PEI-biodegradable particulate combined delivery systems generated with improved in vitro and in vivo stability and transfection properties. To obtain low molecular weight (MR, weight average) chitosan fractions with molecular weights around 10kDa, higher molecular weight chitosan were put through the degradation process using the sodium nitrite method. The DNA/chitosan polyplexes prepared with this fraction resulted in complex sizes smaller than 100nm. In order to determine which of the carrier formulations that were produced was the most suitable for DNA vaccine delivery, the high MR chitosan and low MR chitosan, DNA polyplexes were evaluated further, initially in cell culture for their transfection abilities and then as vaccine carriers in the mouse model. Cell culture was carried out to evaluate the in vitro effect of the investigated systems by growing several cells lines, 293, CHO-K1, B16 F10, A549 and HeLa. Cell culture studies showed that the low MR chitosan/ DNA polyplexes gave the highest and most reproducible transfection efficiencies. However, when tested in vivo, the chitosan/DNA polyplexes were not as effective. Nevertheless, it was the chitosan polyplexes/gWIZ that were successful in eliciting an immune response, suggesting that they hold promise in the field and warrant further investigation. Plasmid gWIZ is a superpure and supercoid plasmid with the cytomegalovirus promoter. in vivo studies in this project included mucosal DNA vaccine delivery (oral, nasal) and comparative non mucosal delivery (i.m. and i.v.) as a promising approach, in preventing mucosally transmitted diseases. Besides eliciting both local and systemic immunity, mucosal vaccination which is non-invasive in nature, increases the patient compliance and reduces the need for vaccine application by trained personnel. Conclusively, chitosan/DNA nano system will be potential for the development of novel DNA vaccines.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Development of new DNA vaccines utilising polymeric gene delivery system
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
Keywords: Health and environmental sciences; DNA; Delivery; Polymeric; Vaccines
URI: https://discovery.ucl.ac.uk/id/eprint/10105148
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