Adenovirus Serotype 5 Fiber Shaft InfluencesIn VivoGene Transfer in Mice

Abstract

Adenoviral vectors used in gene therapy are predominantly derived from adenovirus serotype 5 (Ad5), which infects a broad range of cells. Ad5 cell entry involves interactions with the coxsackie-adenovirus receptor (CAR) and integrins. To assess these receptors in vivo, we mutated amino acid residues in fiber and penton that are involved in receptor interaction and showed that CAR and integrins play a minor role in hepatic transduction but that integrins can influence gene delivery to other tissues. These data suggest that an alternative entry pathway exists for hepatocyte transduction in vivo that is more important than CAR or integrins. In vitro data suggest a role for heparan sulfate glycosaminoglycans (HSG) in adenovirus transduction. The role of the fiber shaft in liver uptake was examined by introducing specific amino acid changes into a putative HSG-binding motif contained within the shaft or by preparing fiber shaft chimeras between Ad5 and Ad35 fibers. Results were obtained that demonstrate that the Ad5 fiber shaft can influence gene transfer both in vitro and to the liver in vivo. These observations indicate that the currently accepted two-step entry pathway, which involves CAR and integrins, described for adenoviral infection in vitro, is not used for hepatic gene transfer in vivo. In contrast, alpha(v) integrins influence gene delivery to the lung, spleen, heart, and kidney. The detargeted vector constructs described here may provide a foundation for the development of targeted adenoviral vectors.

Evaluation of Cell-Penetrating Peptide/Adenovirus Particles for Transduction of CAR-Negative Cells

“…Next, the penton base links with integrin receptors on the cell through the argentine-glycine-aspartic acid (RGD) motif (Wickham, Mathias et al 1993). In addition to CAR and integrin, lysine-lysine-threonine-lysine (KKTK) motifs found in the shaft domain of the fiber protein and heparin sulphate proteoglycans 14 found on the cell plasma membrane aid cell attachment (Smith, Idamakanti et al 2003).…”

Changing the adenovirus fiber for retaining gene delivery efficacy in the presence of neutralizing antibodies

“…27,28 Further studies are needed to dissect the putative role of fiber shaft in the context of infection in the presence of NAb. [29][30][31] Nevertheless, the empiric assays performed here suggest that anti-fiber knob NAbs have an important role in determining the success of gene transfer in vitro and in vivo.…”

Transductional targeting of adenovirus vectors for gene therapy

“…For example, shortening of the native fiber shaft domain of the Ad5 fiber 46 or replacement of the Ad5 shaft with the short Ad3 shaft domain 47 was shown to attenuate liver uptake in vivo following intravenous delivery. In related work, Smith et al 48 These efforts serve to highlight the complexity of vector/host interplay, and have identified important genetic modifications that have important practical implications for designing safer and more effective Adbased vectors for clinical applications. In the absence of a clinically defined upper limit for ectopic liver transduction in humans, it is clear that the concepts of 'de-targeting' and 're-targeting' must be simultaneously employed to allow for maximum vector efficacy at the lowest possible dose.…”