Chu, C; (2015) Modulation of the intraocular immune environment by viral gene transfer in mouse models of uveitis. Doctoral thesis , UCL (University College London).

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Abstract

Treatment options for severe uveitis are predominantly limited to systemic immunosuppressants, which are often accompanied by significant side effects. Ocular gene transfer, though, may serve as a sustained local immunomodulating therapy. New adeno-associated virus (AAV) serotypes were used to investigate gene therapy for uveitis in mouse models. Experimental autoimmune uveoretinitis (EAU) was used to model human disease; whilst endotoxin induced uveitis (EIU) was employed to dissect factors affecting immune suppression by AAV. Refinement of AAV production processes improved the quality of vectors including AAV8, AAV9 and ShH10. Technical issues were assessed, confirming apical transgene secretion from RPE and examining the effects of endotoxin contamination. New methods for the evaluation and quantitative scoring of disease severity in both animal models were developed, based on flow cytometry and optical coherence tomography (OCT) for EIU and EAU in C57BL/6J mice, respectively. The complexity of the role of IL-10 in EAU became evident, and pronounced species differences between murine and human IL-10 were identified in EIU. Despite generating intraocular levels comparable to those of recombinant protein known to be effective, AAV-derived human IL-10 did not suppress EIU. Gene delivery of soluble monomeric TNF receptors did not suppress EAU or EIU, and in vitro exhibited less TNF neutralising effects than an equivalent dimeric Fc-fusion protein. Local expression of this dimeric protein from different vectors however was also unable to attenuate EAU. An NFκB-motif based promoter to regulate the expression of AAV-delivered transgenes in uveitis was developed in parallel. It demonstrated activation only in the presence of, and in proportion to the degree of inflammation in EAU. Despite these advances, neither IL-10 nor TNF-inhibitors proved optimal targets for local suppression of intraocular inflammation by AAV. Further work is needed to assess the response of the eye to viral gene transfer in the context of uveitis.

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