Rhodopsin Genomic Loci DNA Nanoparticles Improve Expression and Rescue of Retinal Degeneration in a Model for Retinitis Pigmentosa.

Abstract

The use of gene therapy may allow replacement of the defective gene. Minigenes, such as cDNAs, are often used. However, these may not express normal physiological genetic profiles due to lack of crucial endogenous regulatory elements. We constructedDNA nanoparticles (NPs) that contain either themouse or human full-length rhodopsin genomic locus, including endogenous promoters, all introns, and flanking regulatory sequences of the 15-16 kb genomic rhodopsin DNA inserts. We transduced the NPs into primary retinal cell cultures from the rhodopsin knockout (RKO) mouse invitro and into the RKO mouse invivo and compared the effects on different functions to plasmid cDNA NP counterparts that were drivenby ubiquitous promoters. Our results demonstrate that genomic DNA vectors resulted in long-term high levels of physiological transgene expression over a period of 5months. In contrast, the cDNA counterparts exhibited low levels of expression with sensitivity to the endoplasmic reticulum (ER) stress mechanism using the same transgene copy number both invitro and invivo. This study demonstrates for the first time the transducing of the rhodopsin genomic locus using compacted DNA NPs.