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Title: Cell replacement and ex vivo gene therapy for photoreceptor regeneration
Author: Cramer, Alona
ISNI:       0000 0004 6061 5111
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2015
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Photoreceptor degeneration due to retinitis pigmentosa (RP) is a primary cause of inherited retinal blindness. Photoreceptor cell-replacement therapies may hold the potential for repair in a degenerate retina, by reinstating light sensitive cells to project and form connections with downstream retinal cells and finally the visual cortex. Patient-specific induced pluripotent stem cells (iPSc) could provide an autologous source of cells to replace lost tissue. However, the use of patient-derived iPSc would require that the disease-causing gene mutation be corrected in cells before transplantation. Ex vivo gene therapy of mouse photoreceptor precursor (PhRP) cells and subretinal transplantation of treated cells are here studied in a disease-specific animal model of RP; rhodopsin was ectopically expressed ex vivo in rod precursor cells, sourced from a transgenic model lacking the rhodopsin gene. Treated rod precursors were here transplanted in mice of the same disease model and are shown to gain expression of rhodopsin and mature to regenerate the absent outer nuclear layer (ONL) of degenerate mice. Visual function, assayed in the same animals before and after transplantation, was restored in animals which had no rod function at baseline. Delivery of the rhodopsin gene by both an adeno associated viral (AAV) vector and a non-viral minicircle DNA vector developed here for ex vivo gene delivery to rod photoreceptor precursors showed comparable efficiency and sustained expression. The non-viral minicircle method provides a novel system for efficient photoreceptor therapy and may offer a platform of genetic treatment of photoreceptor degenerations in which the gene in focus exceeds the size limit for packaging in AAV. Human embryonic stem cell (ESC) and human iPSC-derived PhRPs were also transplanted in mice with complete ONL degeneration and were able to reform the lost photoreceptor layer and mature in the host retina. Human cells developed light-sensitive outer segments, and reconnect with host neurons downstream to improve vision in previously blind mice. Efficient transplantation of ex vivo genetically treated rod precursors and human stem cell-derived PhRPs in animal models of progressive RP may provide a clinically-relevant model for the investigation of cell-replacement therapy for photoreceptor regeneration in retinal disease.
Supervisor: MacLaren, Robert Sponsor: Clarendon Fund
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Photoreceptors ; Retinitis pigmentosa ; Stem cells