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Deepa Talreja, Siobhan Cashman, Bhanu Dasari, Rajendra Kumar-Singh; Development of a platform protein delivery system for retinal cells in vivo following intravitreal injection. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1193. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
There is currently no efficient method to deliver proteins across the plasmamembrane of photoreceptor or retinal pigment epithelium (RPE) cells in vivo. We have developed a platform protein delivery system that can transport proteins into retinal cells including ganglion cells, photoreceptors and retinal pigment epithelium via intravitreal injection- a current standard of clinical care.
Nucleolin targeting aptamer AS1411 was exploited to introduce proteins intracellularly. Biotinylated AS1411 complexed with Traptavidin was used as a platform to deliver proteins GFP (AS1411/Traptavidin/GFP complex) or X-linked inhibitor of apoptosis (XIAP; AS1411/Traptavidin/XIAP complex) by using intravitreal injection in BALB/C mice. Frozen sections were analyzed for uptake of proteins in the retina. Apoptosis was induced by injecting 20 nmol NMDA into the vitreous. NMDA/XIAP complex injected eyes were compared with NMDA/GFP complex injections to test XIAP function. Retinas were harvested at 24 hours and TUNEL analysis was used to assess apoptosis. Caspase assay was performed to detect the differences in Caspase 3 and 7 activity.
Intravitreal injection of GFP or XIAP complexes enabled delivery of the proteins to ganglion cells, photoreceptors, and retinal pigment epithelium in vivo. XIAP complexes conferred significant protection to ONL and INL in NMDA induced apoptosis when compared to GFP injections. NMDA/XIAP complexes showed fewer apoptotic nuclei by TUNEL analysis relative to NMDA/GFP complex injected eyes. Caspase assay also confirmed a decrease in activity of Caspase 3/7 in XIAP treated eyes compared to GFP.
AS1411/Traptavidin complex can be used as a platform to deliver therapeutic proteins to the retina. Rapid transport of XIAP protein into the cells and inhibition of apoptosis without significant toxicity was observed. This system can be potentially utilized to introduce a large variety of therapeutically relevant proteins that are previously well characterized to maintain the structural integrity and function of retina, thus, preventing vision loss due to ocular complications.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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