A Genetically Modified Adenoviral Vector Exhibits Enhanced Gene Transfer of Human Smooth Muscle Cells

Abstract

Adenoviral vector-based gene therapy is a promising approach for the treatment of restenosis postangioplasty. However, a high concentration of adenoviral vector can cause cellular activation, damage, and an enhanced immune response. One approach to solving this problem is to increase gene transfer efficiency by directing adenoviral vector entry via an alternate receptor system. We have constructed an adenoviral vector, Av9LacZ, that encodes the β-galactosidase gene and contains a chimeric fiber protein that redirects viral vector binding to the Ad3 adenoviral receptor on the host cell. We examined the ability of Av9LacZ to transduce primary human smooth muscle cells (SMC) and found that it showed a 10- to 15-fold higher transduction efficiency when compared to the prototypic adenoviral vector currently used for preclinical and clinical studies. While both vectors were able to transduce rabbit, pig and monkey SMCs, the genetically modified vector transduced human SMC with much higher efficiency. SMC obtained from the aorta, coronary, renal, popliteal and pulmonary arteries were all efficiently transduced by Av9LacZ. Consistent with the data obtained from cultured cells, Av9LacZ also transduced fresh human arterial tissues considerably more efficiently than Av1LacZ. We conclude that the large discrepancy between transduction of animal and human cells by conventional vectors supports a cautious extrapolation of the results of in vivo animal studies to man. Furthermore, the genetically modified AV9 vector may deliver better efficacy and studies in large animal models with this vector could be more predictive of therapeutic efficacy in the treatment of human restenosis.

Gene therapy progress and prospects: adenoviral vectors

“…The most straightforward method of detargeting of CAR binding is to pseudotype human serotypes with capsids from other species [20][21][22] or to simply employ Ad vectors from nonhuman serotypes. 23 This approach has the added benefit of avoiding the neutralization of input vector by preexisting antibodies to human adenoviruses.…”

“…24 A modified approach therefore incorporated the deletion of the entire knob domain of the adenovirus fiber protein and replacing it with two distinct moieties that provide a trimerization function for the knobless fiber and specific binding to the target cell. 25 These approaches have been tested extensively and demonstrated efficient transduction (up to a three log increase) 26 of the desired targets including endothelial 27 and smooth muscle cells, 22 brain microcapillary bed, 28 synovial cells 29 and tumor cells. 30 A Phase I clinical trial is underway in which an Ad vector genetically modified to target integrins via the Arg-Gly-Asp (RGD) peptide motif is being tested in ovarian cancer and recurrent cancer of the oral cavity.…”

Adenoviral Serotype 5 Vectors Pseudotyped with Fibers from Subgroup D Show Modified TropismIn VitroandIn Vivo

“…At present there are 51 known human Ads subdivided into groups A-F. The tropism of Ad serotypes varies widely and indeed some have been exploited to achieve increased localized gene transfer to defined tissues, such as ocular cells (Von Seggern et al, 2003), primary central nervous system cells (Chillon et al, 1999), and vascular smooth muscle and endothelial cells (Havenga et al, 2001(Havenga et al, , 2002Su et al, 2001).…”

A novel combination of promoter and enhancers increases transgene expression in vascular smooth muscle cells in vitro and coronary arteries in vivo after adenovirus-mediated gene transfer

“…Mutations of the fibre protein to include an RGD-motif 17 (allowing binding to a v integrins), a poly-lysine sequence 17 (to enhance binding to heparan-sulphate containing receptors), the Ad3 fibre protein 18 and the Ad16 fibre protein 19 have all increased transgene expression in SMC in vitro. The latter two vectors, however, were associated with reduced transgene expression in pig and/or rat SMC and are clearly of limited use in animal studies where rat carotid and pig coronary and ilio-femoral arteries are commonly used targets for gene transfer.…”