Faculty Collaboration Database

Improving retinal vascular endothelial cell tropism through rational rAAV capsid design. PLoS One 2023;18(5):e0285370

Date

05/11/2023

Pubmed ID

37167304

Pubmed Central ID

PMC10174500

DOI

10.1371/journal.pone.0285370

Scopus ID

2-s2.0-85159655750 (requires institutional sign-in at Scopus site)

Abstract

Vascular endothelial cells (VEC) are essential for retinal homeostasis and their dysfunction underlies pathogenesis in diabetic retinopathy (DR) and exudative age-related macular degeneration (AMD). Studies have shown that recombinant adeno-associated virus (rAAV) vectors are effective at delivering new genetic material to neural and glial cells within the retina, but targeting VECs remains challenging. To overcome this limitation, herein we developed rAAV capsid mutant vectors with improved tropism towards retinal VEC. rAAV2/2, 2/2[QuadYF-TV], and rAAV2/9 serotype vectors (n = 9, capsid mutants per serotype) expressing GFP were generated by inserting heptameric peptides (7AA) designed to increase endothelial targeting at positions 588 (2/2 and 2/2[QuadYF-TV] or 589 (2/9) of the virus protein (VP 1-3). The packaging and transduction efficiency of the vectors were assessed in HEK293T and bovine VECs using Fluorescence microscopy and flow cytometry, leading to the identification of one mutant, termed EC5, that showed improved endothelial tropism when inserted into all three capsid serotypes. Intra-ocular and intravenous administration of EC5 mutants in C57Bl/6j mice demonstrated moderately improved transduction of the retinal vasculature, particularly surrounding the optic nerve head, and evidence of sinusoidal endothelial cell transduction in the liver. Most notably, intravenous administration of the rAAV2/2[QuadYF-TV] EC5 mutant led to a dramatic and unexpected increase in cardiac muscle transduction.

Author List

Periasamy R, Patel DD, Boye SL, Boye SE, Lipinski DM

Author

Daniel M. Lipinski PhD Associate Professor in the Ophthalmology and Visual Sciences department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Capsid
Capsid Proteins
Cattle
Dependovirus
Endothelial Cells
Genetic Therapy
Genetic Vectors
HEK293 Cells
Humans
Mice
Retina
Transduction, Genetic
Tropism