Improvement of Photoreceptor Targeting via Intravitreal Delivery in Mouse and Human Retina Using Combinatory rAAV2 Capsid Mutant Vectors. Invest Ophthalmol Vis Sci 2017 Dec 01;58(14):6429-6439
Date
12/21/2017Pubmed ID
29260200Pubmed Central ID
PMC5736327DOI
10.1167/iovs.17-22281Scopus ID
2-s2.0-85038855464 (requires institutional sign-in at Scopus site) 29 CitationsAbstract
PURPOSE: Effective intravitreal gene delivery to cells of the central retina (i.e., photoreceptors) would be of substantial benefit for treating patients with retinal diseases, such as achromatopsia, where retinal detachment from a subretinal may be harmful. Previous studies demonstrated that mutation of the recombinant adeno-associated virus (rAAV) capsid through introduction of peptide insertions or amino acid substitutions dramatically alters vector tropism. Herein, we evaluate the photoreceptor transduction efficiency of three rAAV2/2-based capsid mutant vectors: rAAV2/2[7m8], rAAV2/2[QuadYF+TV], and a chimeric vector incorporating both mutations (termed rAAV2/2[MAX]) following intravitreal delivery in mice. Furthermore, we evaluate the transduction efficiency of rAAV2/2[MAX] using explanted human central retinal samples to address clinical translatability.
METHODS: Vectors containing a GFP or mCherry reporter gene were intravitreally injected into C57BL/6J or Nrl-EGFP mice, respectively. Transduction was assessed in vivo utilizing a custom multiline confocal scanning laser ophthalmoscope. Injected Nrl-EGFP mouse retinas were used to quantify transduced photoreceptors using flow cytometry. Postmortem human retinal tissue was cultured following administration of rAAV2/2[MAX]. C57BL/6J retinas and human explants were cryosectioned to determine vector tropism.
RESULTS: The chimeric vector rAAV2/2[MAX] transduced significantly higher proportions of the retina than did either single mutant serotypes following intravitreal delivery in murine retina, including inner retinal cells and photoreceptors. Vector rAAV2[MAX] demonstrated transduction of human photoreceptors and ganglion cells.
CONCLUSIONS: Transduction observed via rAAV2/2[MAX] indicates that combining mutations with complementary mechanisms of action in a single vector results in enhanced transduction. rAAV2/2[MAX] also presented the ability to transduce human photoreceptors and ganglion cells, indicating potential for efficient intravitreal vector delivery.
Author List
Reid CA, Ertel KJ, Lipinski DMAuthor
Daniel M. Lipinski PhD Associate Professor in the Ophthalmology and Visual Sciences department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
AnimalsCapsid
Dependovirus
Gene Transfer Techniques
Genetic Therapy
Genetic Vectors
Humans
Intravitreal Injections
Mice
Mice, Inbred C57BL
Photoreceptor Cells, Vertebrate
Promoter Regions, Genetic
Retina
Transduction, Genetic
Viral Tropism
