Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.779984
Title: Intracellular drug delivery : a route to more selective and effective treatments for disease
Author: Mai, Quynh N.
ISNI:       0000 0004 7965 6757
Awarding Body: University of Nottingham
Current Institution: University of Nottingham
Date of Award: 2019
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Abstract:
The Neurokinin 1 receptor (NK1R) is a G protein-coupled receptor (GPCR) and member of the tachykinin family expressed in the central nervous system, immune cells, gastrointestinal tract and vascular endothelia. It is associated with neurogenic inflammation, gastrointestinal function and nociceptive transmission. Stimulation of NK1R by the endogenous agonist, substance P, initiates Gαq-mediated signalling, receptor phosphorylation and β-arrestin recruitment to initiate clathrin-dynamin mediated endocytosis. Internalised NK1R has a spatially and temporally dynamic signalling profile which has been demonstrated to underlie pathophysiological outcomes distinct from the plasma membrane. Therefore, endosomal NK1R populations may be considered a therapeutically distinct target. This thesis explores this concept, through the characterisation of NK1R signalling from endosomes, and the development of two novel drug delivery systems: 1) Lipid-conjugation to anchor soluble molecules into membranes and enhance their targeting; 2) pH-sensitive nanoparticles were used as a non-covalent method of packaging NK1R antagonists for targeted endosomal drug delivery. We investigated the inhibitory potential for lipid-conjugation of Spantide I, a first generation NK1R antagonist (Spantide-Cholestanol). Assessment of the endosomal signalling pathways of NK1R suggested that Spantide-Cholestanol inhibited endosomal signalling with greater potency. We propose a mechanism whereby lipid-conjugation provides sustained association with membranes and can be directed to endosomes to improve inhibition of internalised NK1R signalling. Fluorescence correlation spectroscopy and localisation of a lipid-conjugated fluorescent probe (Cy5-Cholestanol) suggested that there was enrichment in endosomes, and at the plasma membrane. Further investigation of this plasma membrane-associated ligand population revealed that Spantide-Cholestanol has inhibitory effects on receptor trafficking and some plasma membrane-delimited signalling events. Therefore, while Spantide-Cholestanol effectively inhibits endosomally localised NK1R signalling, it also has the potential to bind and modulate plasma membrane localised NK1R. An alternative approach for endosomal drug delivery is the non-covalent loading of NK1R antagonists into pH-sensitive, block copolymer nanoparticles (NPs). The nanoparticle core contained a pH responsive monomer which at acidic pH below the pKa resulted in the protonation of the monomers, triggering repulsion and drug release. This pH is known to be achieved in the acidic endosome environment and therefore may be appropriate for endosome-selective drug release. However, cationic NPs reportedly reduce cell viability. To address this issue, we incorporated diethylene glycol (DEG) into block copolymer to shield the cell from toxicity induced by the charged monomeric subunits at endosomal pH. We observed a DEG-dependent increase in cell health, and internalisation of NPs into Rab5-positive endosomes, to demonstrate that these pH responsive nanoparticles may offer a suitable approach for selective delivery drugs into endosomes. The key findings in this thesis emphasise the importance of considering receptor trafficking/localisation in the entire receptor signalling profile. We have observed that lipidation increases drug localisation in endosomes, significantly improving drug potency to inhibit endosomal signalling. However, some of the drug remains at the plasma membrane and can antagonise the population of membrane-localised NK1R. This suggests that nanodelivery of drugs to endosomes may be a more selective therapeutic strategy. Overall, the efficient and selective delivery of drugs to compartmentalised receptor populations will have important implications for drug discovery programs.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.779984  DOI: Not available
Keywords: RS Pharmacy and materia medica
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