Targeted Gene Delivery to Vascular Tissue In Vivo by Tropism-Modified Adeno-Associated Virus Vectors

Abstract

Background-Gene therapy offers an unprecedented opportunity to treat diverse pathologies. Adeno-associated virus (AAV) is a promising gene delivery vector for cardiovascular disease. However, AAV transduces the liver after systemic administration, reducing its usefulness for therapies targeted at other sites. Because vascular endothelial cells (ECs) are in contact with the bloodstream and are heterogeneous between organs, they represent an ideal target for site-specific delivery of biological agents. Methods and Results-We isolated human venous EC-targeting peptides by phage display and genetically incorporated them into AAV capsids after amino acid 587. Peptide-modified AAVs transduced venous (but not arterial) ECs in vitro, whereas hepatocyte transduction was significantly lower than with native AAV. Intravenous infusion of engineered AAVs into mice produced reduced vector accumulation in liver measured 1 hour and 28 days after injection and delayed blood clearance rates compared with native AAV. Peptide-modified AAVs produced enhanced uptake of virions in the vena cava with selective transgene expression. Retargeting was dose dependent, and coinfusion of either heparin or free competing peptides indicated that uptake was principally independent of native AAV tropism and mediated via the peptide. Conclusions-AAV tropism can be genetically engineered by use of phage display-derived peptides to generate vectors that are selective for the vasculature.

Heart-targeted adeno-associated viral vectors selected by in vivo biopanning of a random viral display peptide library

“…1 Among the tolerated insertion sites for small peptides, insertions in the AAV2 heparin binding domain 26,27 adjacent to amino acids 587 or 588 was most successful. 17,23,[28][29][30][31][32] An inherent problem of genetic modification of the AAV capsid is the design of the targeting peptide. Our knowledge about cell surface receptors and their ligands is incomplete, and the toleration and performance of ligands identified by phage display are so far not predictable within the constraints of the AAV capsid structure, although it was successful in several instances.…”

Adeno-associated virus (AAV)-7 and -8 poorly transduce vascular endothelial cells and are sensitive to proteasomal degradation

“…[15][16][17] Alternate strategies to improve endothelial cell transduction by AAV have been actively sought. 18 These include AAV-2 retargeting using vascular cell-selective peptides 11,19,20 or the use of alternate serotypes. 21 We previously reported that for vascular cells (including endothelial and smooth muscle cells) AAV-2 produced the highest levels of transduction compared to AAV-3, -4, -5 or -6, although these levels were lower than those achieved in the more permissive cell type HeLa.…”

Gene Delivery Systems: Tailoring Vectors to Reach Specific Tissues

“…For AAV2-derived vectors, peptides inserted in positions 1, 34, 138, 161, 459, 584, 587 and 588 (relative to VP1 start codon) are displayed on the vector surface, and allow production of vectors with similar viral titers to those observed for unmodified AAV2 vectors (Shi et al, 2001;Wu et al, 2000b). Most of the studies have used positions 138 (VP2 N-terminal), 587 and 588 (HSPG binding domain) to insert peptides ranging from 5 to 272 amino acids (Loiler et al, 2003;Nicklin et al, 2001a;Perabo et al, 2006;Shi et al, 2001;White et al, 2007;White et al, 2004;Wu et al, 2000b;Yu et al, 2009). Capsid protein modifications have improved gene delivery to lung (Kwon & Schaffer, 2008), endothelial cells (Nicklin et al, 2001a), pancreatic islets (Loiler et al, 2003), vascular tissue (White et al, 2004), atherosclerotic lesions (White et al, 2007), muscle (Yu et al, 2009), myocardium (Yang et al, 2009), and cancer cells .…”