Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.602550
Title: Development of freeze-dried nanoparticles incorporating gene therapy for the management of retinopathy of prematurity
Author: Jain , Arvind Kumar
Awarding Body: Queen's University Belfast
Current Institution: Queen's University Belfast
Date of Award: 2013
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Abstract:
Retinopathy of prematurity (ROP) is a posterior eye disorder found in prematurely born infants, that can result in blindness. In this thesis a delivery system, for the prophylactic management of the disease was designed and tested based on the hypothesis that delivery of a hypoxia inducible plasmid to the posterior eye might prevent the pathogenesis of the ROP. Major challenges identified for the project included: characterisation of the hypoxia inducible plasmid, efficient transfection of the target cells and delivering the DNA to the posterior eye with a minimally invasive route. To meet these requirements it was proposed to encapsulate a hypoxia responsive plasmid condensed with a cationic peptide (RALA) within PLA-PEG nanoparticles that can deliver the DNA to the posterior eye via transscleral route for efficient transfection of the retinal endothelial cells. In order to pursue this hypothesis the DNA condensation with RALA was optimized to give nanoparticies (RNPs) with a z-average diameter of 58.7±10.3 nm and PDI of 0.252±0.050. These nanoparticies efficiently transfected a variety of cell lines. A series of PLA-PEG block copolymers were synthesized with various PEG chain length and LA/EG ratio, and formulated into composite nanoparticles (diameters <200 nm and PDI <0.200) containing the RNPs. TEM was used for the first time to confirm the presence of a nanoparticle-in-nanoparticle system by TEM. Furthermore, an in vitro DNA release study demonstrated that the composite nanoparticles were able to release DNA, with faster DNA release in the first 24 hrs comprising 10% of the DNA content, followed by a slow continuous release for a prolonged duration. Among the studied formulations PLA25-PEG5 exhibited the fastest release where >30% of DNA content was released over 6 weeks. Finally these nanoparticles were evaluated for their ability to cross the sclera and the results demonstrated that > 1 0% of PLA20-PEG2 and 8% of PLA25-PEG5 . nanoparticles crossed the sclera in a 4 hrs, whereas PLGA nanoparticles exhibited permeation of just 3.2% in this time. While the selected hypoxia responsive plasmid, pE9/GFP, failed to show its specificity to express only in hypoxia conditions the work presented herein demonstrates that the proposed delivery system is viable. Thus, this thesis demonstrates the preparation of a composite nanoparticle system where inner RNPs demonstrated their efficiency to condense the DNA and enable them express inside cells and outer PLA-PEG nanoparticles demonstrated their efficiency to cross the sclera. This system is a promising candidate for the prophylactic management of ROP and may be suited to other gene therapies.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.602550  DOI: Not available
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