AbstractNanoparticle-mediated drug delivery is a promising approach for the delivery of various drug types to the site of disease in an effort to improve efficacy. Development of polymeric nanoparticles (NP) with slow drug-release kinetics coated with targeting ligands enables cell-specific sustained drug delivery, whilst minimising off-target side effects. In this thesis, a range of novel poly(lactic-co- glycolic acid) (PLGA) NP formulation strategies to deliver various drugs to targeted cells were examined.
A NP formulation for the intracellular delivery of plasmid DNA was developed, whereby the DNA was encapsulated within PLGA NP by combining salting out and emulsion-evaporation processes. This process reduced the requirement for sonication which can induce degradation of the DNA. A monodispersed NP population with a mean diameter of approximately 240 nm was produced, entrapping a model plasmid DNA in both supercoiled and open circular structures. To induce endosomal escape of the NP, a superficial cationic charge was introduced using positively charged surfactants dimethyl-didodecyl-ammonium bromide (DMAS), which resulted in elevated zeta potentials. Fluorescence and transmission electron microscopy demonstrated that the DMAS coated cationic NP were able to evade the endosomal lumen and localise in the cytosol of treated cells. Consequently, DMAS coated PLGA NP loaded with a GFP reporter plasmid exhibited significant improvements in transfection efficiencies with comparison to non-modified particles, highlighting their functional usefulness.
A second study presented in this thesis was focussed on the development of NP covalently conjugated with anti Death Receptor 5 (DR5) monoclonal antibodies (mAbs) to target DR5 expressed on the surface of colorectal tumor cells. Anti DR5 mAbs conjugated NP demonstrated specific targeting and internalisation to colorectal carcinoma HCT116 cells, but more interestingly, also demonstrated activation of the DR5 triggered extrinsic apoptosis pathway. Furthermore Anti DR5 mAbs conjugated NP loaded with camptothecin demonstrated significant cell toxicity; a result of synergistic efficacy between camptothecin delivery and DR5 triggered apoptosis.
In a third study, PLGA NP were conjugated with sialic acid di(a,2^ 8) N- Acetyl-neuraminic acid (o2-8-NANA) to target siglec receptors expressed on immune cells as the basis of a novel anti-inflammatory therapy to treat acute lung injury and the more severe acute respiratory distress syndrome. 02-8-NANA conjugated NP demonstrated RAW 264.7 monocyte cell targeting and internalisation. Co-incubation of these particles in RAW 264.7 cells induced inhibition of the LPS- stimulated release of pro-inflammatory cytokines. Furthermore, 02-8-NANA conjugated NP loaded with camptothecin demonstrated significant targeted THP1 leukemic cells toxicity. Therefore sialic acid conjugated NP may have therapeutic usefulness in the targeting of myeloid cells in a range of disease conditions such as acute lung injury and leukaemia.
In conclusion, the results presented here demonstrate the broad diversity of drug delivery approaches that can be developed with PLGA-based nanoparticles. Furthermore, results obtained with NP conjugated with antibodies and sialic acid demonstrate that NP should not only be considered as a drug carrier anymore, but also as extremely potent drugs that could be included in a global therapeutic strategy.
|Date of Award||Jul 2010|
|Supervisor||Christopher Scott (Supervisor), Brendan Gilmore (Supervisor) & Paul McCarron (Supervisor)|