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Renee Christine Ryals, Siddharth Patel, Kyle Weller, Mark E Pennesi, Gaurav Sahay; Lipid Nanoparticle Based Messenger RNA Delivery to the Retina. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2916.
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© ARVO (1962-2015); The Authors (2016-present)
Gene therapy development for the treatment of retinal degeneration is quickly advancing. With the advent of messenger RNA (mRNA) therapeutics, limitations to the current adeno-associated virus (AAV) gene delivery can be overcome. Advances in mRNA technology have unlocked high efficacy, transient gene delivery in hard-to-transfect cells for intracellular delivery of genes, gene silencing, or genome editing. Lipid nanoparticles (LNPs) are capable of encapsulating and delivering mRNA intracellularly, are easy to scale up, and have shown excellent clinical efficacy and safety. Developing LNPs that can deliver mRNA to the retinal pigment epithelium (RPE) and photoreceptors will have significant impacts for the treatment of retinal degenerations.
Reporter mRNA, luciferase and mCherry, were encapsulated in lipid nanoparticles. LNP size and encapsulation efficiency was measured. Adult BALB/c mice were subretinally injected with 1ul of LNP-luciferase or PBS as control. From 4 to 168 hours post-injection luciferase activity was measured via bioluminescent imaging. Additionally, 1 ul of LNP-mCherry was subretinally injected in right eyes and left eyes were injected with PBS as control. From 24 to 120 hours post-injection, eyes were harvested, sectioned and stained with antibodies specific for mCherry and RPE65. Confocal images were acquired to localize reporter expression in the retina.
Luciferase activity (average radiance) was found to be 4.3X106, 1.8X107, and 8.5X106 p/s/cm2/sr at 4-, 24- and 48-hours post-injection, respectively. From 96 to 168 hours post-injection, luciferase activity (average radiance) ranged from 5 x 105 to 3 x 104 p/s/cm2/sr. Immunohistochemistry showed co-localization of mCherry and RPE65 from 24 to 120 hours post-subretinal injection.
Our currently designed lipid nanoparticles can deliver mRNA to the RPE. Protein expression is maintained up to 120 hours post-injection. Future studies will focus on engineering advanced LNPs that can lead to intracellular delivery of genes in multiple layers of retina.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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