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Purpose: AAV vectors have restricted packaging capacity and there are many unanswered questions regarding late immune responses. Nonviral vectors provide an alternative that can address these issues. Previous studies have shown that suprachoroidal injections of biodegradable poly nanoparticles (NPs) provide widespread transgene expression in photoreceptors and RPE cells of rats. However, it is crucial to understand the extent and level of gene expression resulting from suprachoroidal injection of PBAE NPs in eyes that are more similar to human eyes. In this study, the distribution and level of expression of a reporter gene was measured in minipig eyes which resemble human eyes in size and other structural aspects.

Methods: Experiments were done in Göttingen and Yucatan minipigs using PBAE NPs containing GPF expression plasmids. PBAE polymer was synthesized and it was used to condense expression plasmids into NPs. Two plasmids were tested, pCAG-eGFP-Z1 which is a standard plasmid containing bacterial sequences, and pCAG-GFP-nP, which is a miniplasmid from which bacterial sequences were removed. The effect of several variables on the distribution and level of expression after suprachoroidal injection of PBAE NPs was tested, including dose, time after injection, multiple injections, and the impact of minimizing bacterial sequences in the plasmid.

Results: Two weeks after suprachoroidal injection of pCAG-GFP-Z1 in minipig eyes, immunofluorescence and immunohistochemistry of serial sections showed expression of GFP in photoreceptors and RPE throughout the entire eye with no toxicity. Two weeks after a …