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Title: Periocular gene therapy for ocular angiogenesis
Author: Demetriades, Anna-Maria
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2005
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Abstract:
Protein delivery to the eye remains an important area in the clinical arena as new therapeutic proteins are being discovered. The focus of this thesis was to establish periocular gene therapy as a novel way of providing localised and sustained release of endogenous anti-angiogenic proteins to the eye. Three endogenous anti-angiogenic proteins of different sizes with established anti-angiogenic activity were chosen for the purpose of our study. The soluble extracellular domain of the vascular endothelial growth factor (VEGF) receptor-1 (sFlt-l; 78kDa), pigment epithelium-derived factor (PEDF; 50kDa) and transforming growth factor beta-I (TGF-B1; 25kDa). In the first part of the thesis, an adenoviral (Ad) vector expressing the reporter gene LacZ was delivered via periocular injection and resulted in infection of extraocular muscles, orbital fat, connective tissues surrounding the eye, and the episclera. Periocular injections of AdsFlt-l, AdPEDF, and AdTGF-B1 were subsequently administered to murine eyes and increased protein levels of sFt-1, PEDF and TGF-B1 were demonstrated in the retina. Thus providing proof-of-principle that a periocular injection of Ad-vectored anti- angiogenic proteins resulted in increased protein levels in the retina and could be a potential method of delivering therapeutic proteins to the posterior segment of the eye. In the second part of the thesis, single periocular inj ections of AdsFlt-l, AdPEDF and AdTGF-J31 were tested in murine models of choroidal neovascularisation, retinal neovascularisation and macular oedema. These studies demonstrated that both a periocular injection of AdsFlt-1 and AdPEDF resulted in significant inhibition of choroidal neovascularisation in an established laser-induced model. However, a periocular injection of AdTGF-B1 had no effect in this chosen model. In addition, a periocular injection of neither AdsFlt-l nor AdPEDF nor AdTGF-B1 resulted in inhibition of retinal neovascularisation. This may have been due to insufficient, non-therapeutic, levels of expressed protein in the retina. A periocular inj ection of AdsFlt-1 resulted in inhibition ofVEGF-induced breakdown of the blood-retinal barrier providing the first proof-of-principle for gene therapy for macular oedema. Interestingly, a periocular injection of AdTGF-J31 resulted in significant inhibition of eye size and development in the neonatal mouse suggesting the involvement of TGF-B1 in postnatal developmental eye growth. In the third part of the thesis, the abili ty to readminister vectors was determined as this would be critical for the application of gene therapy for chronic diseases if a short-acting vector such as Ad was used as the chosen carrier. Repeated periocular delivery of AdPEDF resulted in increased levels ofPEDF in the sclera but not in the choroid and retina. Additional studies are needed to explore the feasibility of repeated injections of Ad vectors including improvement in our understanding of the T cell mediated and humoral immune response in this setting. These studies provide proof-of-principle that periocular delivery of Ad vectors expressing anti-angiogenic transgenes may be a useful approach in the treatment of posterior segment diseases. Further studies are needed to optimize this approach for long-term delivery for the treatment of chronic diseases such as age-related macular degeneration and diabetic retinopathy. With careful consideration, this therapeutic strategy has the potential to be translated into the clinical arena.
Supervisor: Osborne, Neville Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.559857  DOI: Not available
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