Adenovirus 5 fibers mutated at the putative HSPG-binding site show restricted retargeting with targeting peptides in the HI loop.

Adenoviral vectors are commonly used for liver-directed gene therapy following systemic administration owing to their strong propensity for hepatocyte transduction. However, many disease applications would benefit from the delivery of adenoviruses to alternate tissues via this route. Research has thus focused on stripping the virus of native hepatic tropism in conjunction with modifying virus capsid proteins to incorporate novel tropism. Recently, the KO1S* adenovirus serotype 5 fiber mutant, devoid of both coxsackie and adenovirus receptor binding in the fiber knob domain and mutated at the putative heparan sulphate proteoglycan binding site in the fiber shaft, was shown to possess strikingly poor hepatic tropism in mice, rats, and non-human primates. Thus, it is an ideal candidate for retargeting strategies. We therefore assessed the ability of peptide-modified KO1S* fibers to retarget adenovirus. Peptide insertions were well tolerated and virions produced to high titers. However, expected retargeting at the level of transduction was not observed, despite cell-binding studies showing enhanced vector targeting at the cell surface. Cy3 labeling studies showed retarded trafficking of S*-containing fibers. Taken together, our data demonstrates that KO1S* mutant fibers are ineffective for cell retargeting strategies

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

Transduction of the vascular endothelium by adeno-associated virus (AAV) vectors would have broad appeal for gene therapy. However, levels of transduction by AAV serotype-2 are low, an observation linked to deficiencies in endothelial cell binding, sequestration of virions in the extracellular matrix and/or virion degradation by the proteasome. Strategies to improve transduction of endothelial cells include AAV-2 capsid targeting using small peptides isolated by phage display or the use of alternate serotypes. Previously, we have shown that AAV serotypes-3 through -6 transduce endothelial cells with poor efficiency. Recently, AAV serotypes-7 and -8 have been shown to mediate efficient transduction of the skeletal muscle and liver, respectively, although their infectivity profile for vascular cells has not been addressed. Here, we show that AAV-7 and -8 also transduce endothelial cells with poor efficiency and the levels of transgene expression are markedly enhanced by inhibition of the proteasome. In both cases proteasome blockade enhances the nuclear translocation of virions. We further show that this is vascular cell-type selective since transduction of smooth muscle cells is not sensitive to proteasome inhibition. Analysis in intact blood vessels corroborated these findings and suggests that proteasome degradation is a common limiting factor for endothelial cell transduction by AAV vectors

Gene therapy targeting to tumor endothelium.

Tumor-associated vasculature is a relatively accessible component of solid cancers that is essential for tumor survival and growth, providing a vulnerable target for cancer gene therapy administered by intravenous injection. Several features of tumor-associated vasculature are different from normal vasculature, including overexpression of receptors for angiogenic growth factors, markers of vasculogenesis, upregulation of coagulation cascades, aberrant expression of adhesion molecules and molecular consequences of hypoxia. Many of these differences provide candidate targets for tumor-selective 'transductional targeting' of genetically- or chemically modified vectors and upregulated gene expression can also enable 'transcriptional targeting', regulating tumor endothelia-selective expression of transgenes following nonspecific gene delivery. Tumor vasculature also represents an important site of therapeutic action by the secreted products of antiangiogenic gene therapies that are expressed in non-endothelial cells. In this review we assess the challenges faced and the vectors that may be suitable for gene delivery to exploit these targets. We also overview some of the strategies that have been developed to date and highlight the most promising areas of research