Abstract
The establishment of efficient gene delivery to target human tissue is a major obstacle for transition of gene therapy from the pre-clinical phases to the clinic. The poor long-term patency rates for coronary artery bypass grafting (CABG) is a major clinical problem that lacks an effective and proven pharmacological intervention. Late vein graft failure occurs due to neointima formation and accelerated atherosclerosis. Since CABG allows a clinical window of opportunity to genetically modify vein ex vivo prior to grafting it represents an ideal opportunity to develop gene-based therapies. Adenoviral vectors have been frequently used for gene delivery to vein ex vivo and pre-clinical studies have shown effective blockade in neointima development by overexpression of candidate therapeutic genes. However, high titers of adenovirus are required to achieve sufficient gene delivery to provide therapeutic benefit. Improvement in the uptake of adenovirus into the vessel wall would therefore be of benefit. Here we determined the ability of an adenovirus serotype 5 vector genetically-engineered with the RGD-4C integrin targeting peptide inserted into the HI loop (Ad-RGD) to improve the transduction of human saphenous vein smooth muscle cells (HSVSMC), endothelial cells (HSVEC) and intact saphenous vein compared to a non-modified virus (Ad-CTL). We exposed each cell type to virus for 10, 30 or 60 mins and measured transgene at 24 h post infection. For both HSVSMC and HSVEC Ad-RGD mediated increased transduction, with the largest increases observed in HSVSMC. When the experiments were repeated with intact human saphenous vein (the ultimate clinical target for gene therapy), again Ad-RGD mediated higher levels of transduction, at all clinically relevant exposures times (10, 30 and 60 mins tissue:virus exposure). Our study demonstrates the ability of peptide-modified Ad vectors to improve transduction to human vein graft cells and tissue and has important implications for gene therapy for CABG.
Highlights
Long term patency rates for coronary artery bypass grafting (CABG) using autologous saphenous vein are poor, showing 1, 5 and 10 years postCABG rates of 93%, 74% and 41%, respectively [1] and represent a significant clinical problem
In the context of improved gene delivery mediated by the RGD-4C peptide, which was isolated by phage display and targets αv integrins [16], this has been shown for rabbit grafts [17] the vast majority of data is based on gene delivery for cancer [18]
Since smooth muscle cell (SMC) show poor coxsackie and adenovirus receptor (CAR) availability [19], it is relevant that the RGD-4C peptide may circumvent CAR deficiency on target cells to improve levels of transduction
Summary
Long term patency rates for CABG using autologous saphenous vein are poor, showing 1, 5 and 10 years postCABG rates of 93%, 74% and 41%, respectively [1] and represent a significant clinical problem. Pseudotyping the fiber from adenovirus serotype 16, which binds CD46 [12], dramatically improves transduction to vascular cells including intact human saphenous vein allowing lower doses of vector to be used to achieve attractive levels of gene delivery to grafts ex vivo [13]. We assess the ability of RGD-4C-modified adenovirus serotype 5 vectors to enhance gene delivery to human saphenous vein SMC and EC as well as to intact human saphenous vein ex vivo, the ultimate clinical target.
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