Iemsam-Arng, J;
(2013)
Poly(ethylene) glycol based Delivery Systems for Nucleic Acid Therapies.
Doctoral thesis , UCL (University College London).
Abstract
Our work is aimed at developing a synthetic biocompatible gene and siRNA delivery system for the treatment of primary and metastatic tumours. To facilitate delivery of nucleic acid based drugs into the cell, one strategy is to formulate the naked gene with an amine based non-viral gene delivery system via the counterion interaction. The delivery systems including 4arm-PEG-amine, 4arm-PEG-N-2-ethylamine, 8arm-PEG-amine and 8arm-PEG-N-2-ethylamine were synthesised, characterised and complexed with a reporter gene (β-gal plasmid DNA) in phosphate buffer pH 6.0. The resulting complexes were sized and their zeta potential measured (Malvern Zetasizer 3000HS, Malvern Instruments, UK). The complexes were also imaged using transmission electron microscopy and characterised for DNA binding and DNA protection using gel electrophoresis and the ethidium bromide displacement assay. The in vitro transfection efficiency and cell cytotoxicity of the complexes were determined in the A431 and HeLa cells. Additionally, in vivo therapeutic studies in female nude tumour bearing mice were carried out. A promising DNA-polymer complex of 4arm-PEG-N-2-ethylamine produced a complex of 200-300 nm in diameter (polydispersity < 0.6). Complexes had a zeta potential of +19.8 mV (n=3) and were spherical, fibrillar and toroidal in shape. The new gene delivery complex protected DNA from degradation in serum up to 2 hours and was as efficient as poly(ethylenimine) (PEI) in transfecting the A431 cell line, but it was more than 3 orders of magnitude less cytotoxic than PEI. In vivo a gene medicine, comprising the polymer and the tumour necrosis factor alpha gene, was tumouricidal. When complexed with siRNA, the siRNA polymer complex demonstrated a trend of gene silencing activity. A new synthetic gene delivery polymer of 4arm-PEG-N-2-ethylamine has been synthesised. This polymer is biocompatible to cells and is an efficient in vitro and in vivo gene transfer agent.
| Type: | Thesis (Doctoral) |
|---|---|
| Title: | Poly(ethylene) glycol based Delivery Systems for Nucleic Acid Therapies |
| Language: | English |
| Additional information: | Permission for digitisation WITHHELD. |
| Keywords: | gene therapy, non-viral vector, tumouricidal activity, biocompatibility |
| UCL classification: | UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > UCL School of Pharmacy |
| URI: | https://discovery.ucl.ac.uk/id/eprint/1381001 |
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