Preparation and functionalization of magnetic iron oxide nanoparticles for application in gene delivery

Human diseases due to genetic mutations, such as cancer, have become the leading cause of death worldwide. Gene delivery, the strategy to utilise genetic materials for the treatment and circumvention of genetic diseases, holds a promising future in the biomedical application. In this thesis, the use of magnetic iron oxide nanoparticles (MNP) in combination with other viral or non-viral components to generate a multi-component gene delivery agent (vector) was investigated. Different magnetic vector configurations assembled by systematically varying the mixing order of MNP, polyethylenimine (PEI), and plasmid DNA (pEGFP-C1) to transfect mammalian cells (BHK21) under a magnetic field was studied. It was found that the assembly of vector components influenced the vector extra-cellular properties; specifically vector size and surface properties; leading to a significantly different level of vector cellular uptake and gene expression. The vector intracellular transport; including cellular entry mechanism and final intracellular target sites; were investigated to elucidate the resulting gene expression attributed to different vector configurations. The cell viability results using three different assays also highlight the implication of vector dose and configuration on the cell viability. This work presents a comprehensive understanding into the role of each vector component in affecting the vector extra- and intra-cellular characteristics, facilitating the formulation of efficient and safe magnetic vectors. The use of gold-coated MNP vectors as intracellular DNA trafficking tool is presented. Coating the MNP surface with gold before coupling with PEI/DNA complex exhibited excellent gene expression efficiency and negligible impact to cell viability than the uncoated counterpart, demonstrating the efficacy of gold-coated MNP as dual-purpose magnetic probe for gene delivery and intracellular imaging agents with good biocompatibility. The effect of MNP physiochemical structures (core size and morphology) on the efficiency of viral or non-viral vectors in human cells was studied. Each of the synthesized MNP was coated with PEI to transduce deactivated adenovirus (Adv) or transfect plasmid DNA (pCMV-Luc) carrying luciferase protein via a magnetic field into normal (HEK293A) and cancerous (SKOV-3) human cells. The gene activity enhancement varied with MNP physiochemical structures, demonstrating the importance of MNP physiochemical structures selection in determining the transduction and transfection efficiencies