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Principal Investigator: Gaurav Sahay
Organization: OREGON STATE UNIVERSITY
Fiscal Year: 2021
Award: $183,227
Funding agency: National Eye Institute
ABSTRACT
Gene therapy strategies including augmentation, editing or knockdown, can lead to restoration of vision for
patients suffering from inherited retinal degenerations (IRDs). Viral and non-viral vectors efficiently deliver
genes through the subretinal route of administration, but this requires a surgical procedure that detaches the
fragile degenerating retina. Intravitreal injections offer a safer ocular route for gene delivery and is a widely
used outpatient procedure, but significant biological barriers limit the transport of carriers from the vitreous
chamber to the back of the eye. The vehicle, once delivered in the vitreous chamber, will have to move through
the vitreous humor, which is composed of many proteins that restrict diffusion. If intact, the vehicle must then
penetrate an inner limiting membrane functioning as a sieve and restricting permeability of substances into the
retina. Finally, once the vehicle hits the target cell population, it must get internalized and then escape the
endosome to allow for cytosolic delivery of nucleic acids. After subretinal injection, clinically approved lipid
nanoparticles (LNPs) carrying mRNA can mediate protein expression in the retinal pigment epithelium (RPE)
and photoreceptors, the cell types mainly afflicted in IRDs. However, our preliminary data show that these
LNPs have limited expression post-intravitreal injection. There is a critical need to develop carriers that can
traverse the retinal barriers after intravitreal injection. Our long-term goal is to develop novel, peptide targeted
LNP systems that can deliver genes to the outer retina after an intravitreal injection with limited toxicity. We
hypothesize that nanoparticle physicochemical properties like surface charge, size, stability, and ionizability will
be the critical determinants that enhance permeation towards the retina. We posit that peptides can penetrate
and target specific cell types within the outer retina. Our main objectives are to 1) evaluate the
physicochemical parameters of nanoparticles that are a prerequisite for gene delivery to the outer retina and 2)
identify peptides and their structural features that allow for cell-specific delivery. Individually evaluating each
physicochemical characteristic of LNPs and all peptide moieties available is arduous. Thus, we will generate a
diverse DNA barcoded LNP library and use a phage display peptide library to identify optimal nanocarriers and
peptides, respectively. Most nanocarriers have had limited translation potential due to species-specific
difference in ocular barriers. Therefore, we are evaluating our nanocarriers in an ex-vivo non-human primate
(NHP) model for rapid screening of multiple vectors after intravitreal delivery. Overall, this application will 1)
identify novel peptides and structural features of LNPs that enable gene delivery to the outer retina post-
intravitreal administration, and 2) generate a translational, transfection efficiency platform that can be widely
used for the evaluation of gene delivery systems.
Terms: <Affect><Affinity><Back><Bar Codes><Binding><Bio-Informatics><Bioinformatics><Biological><Blindness><Cell Body><Cells><Characteristics><Charge><Cholesterol><Clinical><DNA><DNA Therapy><Data><Deoxyribonucleic Acid><Development><Diffusion><Dorsum><Encapsulated><Endosomes><Evaluation><Eye><Eyeball><Family><Gene Delivery><Gene Transfer Clinical><Genes><Genetic Intervention><Goals><Immunohistochemistry><Immunohistochemistry Cell/Tissue><Immunohistochemistry Staining Method><Incidence><Individual><Inner Limiting Membrane><Label><Lead><Libraries><Lipids><M mulatta><M. mulatta><Macaca mulatta><Maps><Mediating><Messenger RNA><Mice><Mice Mammals><Modeling><Molecular Interaction><Murine><Mus><NGS Method><NGS system><Neural Retina><Non-Viral Vector><Nucleic Acids><Operative Procedures><Operative Surgical Procedures><Out-patients><Outer pigmented layer of retina><Outpatients><Patients><Pb element><Penetration><Peptide Library><Peptide Phage Display Library><Peptides><Permeability><Phosphatides><Phospholipids><Photoreceptor Cell><Photoreceptors><Photosensitive Cell><Pigment cell layer of retina><Pigmented layer of retina><Population><Procedures><Property><Proteins><Randomized><Rapid screening><Receptosomes><Reporter Genes><Research Priority><Retina><Retina Proper><Retinal Degeneration><Retinal Pigment Epithelium><Retinal gene therapy><Retinal pigment epithelial cells><Rhesus Macaque><Rhesus Monkey><Rods and Cones><Route><Structure><Structure of retinal pigment epithelium><Surface><Surgical><Surgical Interventions><Surgical Procedure><System><Techniques><Toxic effect><Toxicities><Transfection><Translations><Variant><Variation><Vertebrate Photoreceptors><Viral><Visual Receptor><Vitreous Chamber><Vitreous humor><aged><barcode><cell type><cellular targeting><clinical relevance><clinically relevant><degenerative retina diseases><design><designing><develop therapy><developmental><gene delivery system><gene therapy><gene-based therapy><genetic therapy><genome editing><genomic editing><genomic therapy><heavy metal Pb><heavy metal lead><inherited retinal degeneration><intervention development><intravitreal injection><knock-down><knockdown><lipid nanoparticle><mRNA><nano carrier><nano particle><nano-sized particle><nanocarrier><nanoparticle><nanosized particle><next gen sequencing><next generation sequencing><nextgen sequencing><non-human primate><nonhuman primate><nonviral vector><novel><nucleic acid delivery><peptide structure><protein expression><randomisation><randomization><randomly assigned><restore sight><restore vision><retina degeneration><retinal degenerative><retinal degenerative diseases><sight restoration><subretinal injection><surgery><therapy development><treatment development><vector><vision loss><vision restoration><visual loss>