Abstract
Purpose :
The purpose of this study was to develop a new type of lipid nanoparticle (LNP) for mRNA delivery to the retina, including photoreceptors, in order to treat inherited retinal diseases (IRDs).
Methods :
To achieve this goal, we developed a novel LNPs that are able to deliver mRNA to the retina, including photoreceptors, in rodent and non-human primate models. We generated a small set of LNP variants with various surface modifications. The LNPs were characterized for their hydrodynamic radius, charge, polydispersity index (PDI), and mRNA encapsulation efficiency. We loaded the LNPs with Cre mRNA targeting the floxed stop codon upstream of a tdTomato cassette in Ai9 reporter mice and measured gene recombination using immunosectioning and quantified localization with photoreceptors using FACS and confirmed delivery in NHPs using a reporter mRNA. We delivered Cas9 mRNA and a single guide RNA targeting the stop codon of a loxp site to measure genome editing.
Results :
The most potent LNPs were tested for delivery in rodent and NHPs and were found to be effective in delivering mRNA to the retina, including photoreceptors, in both animal models. The LNPs were able to package more than 90% of nucleic acids and maintained a spherical morphology as demonstrated by cryoEM. The LNPs (80-100 nm in size) with novel surface modifications that led to a 20-fold improvement in gene delivery to photoreceptors. The most potent LNPs enabled Cas9-based gene editing in the retina of rodents amd we currently are testing efficient editing in models of IRDs.
Conclusions :
This study demonstrates the potential of our novel LNPs for mRNA delivery to the retina, including photoreceptors, in rodent and non-human primate models. These LNPs show promising results for gene editing in rodent models and warrant further investigation. In comparison to AAV-based Cas9 delivery, LNP-packaged mRNA has a transitory expression of nucleases that leads to minimal off-targeting and minimal immune response, making it a more tolerable and less immunogenic option for gene editing in the retina. We are currently testing the ability of LNP-based genome editing to provide functional rescue in different animal models of blindness. These novel vectors represent a powerful tool for nucleic acid delivery to the neural retina, independent of species.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.