Effect of adenovirus serotype 5 fiber and penton modifications on in vivo tropism in rats

Abstract

Sequestration of adenovirus serotype 5 (Ad5) in liver restricts its use for gene delivery to other target sites in vivo. To date, no studies have systematically assessed the impact of genetic capsid modifications on in vivo tropism in rats, an important preclinical model for many disease types. We evaluated a panel of Ad5 vectors with capsid mutations or pseudotyped with the short fiber from serotype 41 (Ad41s) for infectivity in Wistar Kyoto rats in vitro and systemically in vivo. In vitro studies demonstrated that both coxsackie and adenovirus receptor (CAR) and heparan sulfate proteoglycan (HSPG) binding were predominant predictors of Ad5 tropism. In vivo, neither CAR nor integrin mutations alone affected liver transduction. The HSPG-binding mutation alone moderately reduced rat liver transgene levels by 2-fold (P < 0.05). This was further substantially decreased by additional mutation of CAR binding (95-fold). Combining CAR and integrin mutations reduced transgene levels by >99% (509-fold, P < 0.01), an effect not observed in parallel experiments in mice and highly variable when studied further in an additional two strains of rat. Ad41s mediated very low liver transduction (58-fold lower than AdCTL). Moreover, CAR-binding mutants (KO1-containing) or pseudotyping 41s eliminated hemagglutination of rat and human red blood cells in vitro. This highlights some important potential species and strain differences dictating Ad5 tropism in vivo and identifies vectors that are substantially detargeted from rat liver in vivo.

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

“…We additionally assessed vessel homogenates for transgene activity using the CPRG Assay Kit that is effective on rat tissue. 25 However, levels of transgene at 14 days were below the limits of detection of the assay for all treatments (data not shown) further enforcing the poor performance of AAV-2, -7 and -8 vectors in vascular tissue. This suggests that additional barriers to efficient transduction by AAV vectors are present in vascular tissue.…”

Adenovirus serotype 5 hexon is critical for virus infection of hepatocytes in vivo

“…delivery, the vast majority of Ad particles are trapped in the liver (26)(27)(28). Unsuccessful attempts to prevent hepatocyte transduction by Ad5-based vectors through ablation of CAR and/or integrin binding implied that another mechanism must exist to allow Ad5 virus particles to enter hepatic cells in a CAR-independent manner (29,30).…”

Polyinosinic acid enhances delivery of adenovirus vectors in vivo by preventing sequestration in liver macrophages

“…Although the early steps of adenovirus infection have been studied extensively in vitro, much is still unknown about the mechanisms governing adenovirus infection in vivo. To date, attempts to modify adenovirus liver tropism by abolishing CAR and/or integrin interactions have not been successful in all animal models analysed (Brunetti-Pierri et al, 2004;Hodges et al, 2005;Lievens et al, 2004;Nicol et al, 2004;Park et al, 2005;Smith et al, 2002). Shayakhmetov et al (2004) demonstrated that coagulation factor IX and the complement component C4-binding protein can bind the adenovirus fiber-knob domain and provide a bridge for virus uptake through cell-surface HSPGs and low-density lipoprotein receptor-related protein.…”

“…Initial attempts to detarget the liver have been made by ablating the CAR-and integrin-based interactions (Akiyama et al, 2004;Koizumi et al, 2003;Martin et al, 2003;Nakamura et al, 2003). Also, fiber-shaft modifications have been introduced to eliminate virus interactions with native receptors (Bayo-Puxan et al, 2006;Breidenbach et al, 2004;Nicol et al, 2004;Shayakhmetov et al, 2004;Smith et al, 2003b;Vigne et al, 2003). However, the results are variable.…”