Receptor Interactions Involved in Adenoviral-Mediated Gene Delivery After Systemic Administration in Non-Human Primates

Abstract

Adenovirus serotype 5 (Ad5)-based vectors can bind at least three separate cell surface receptors for efficient cell entry: the coxsackie-adenovirus receptor (CAR), alpha nu integrins, and heparan sulfate glycosaminoglycans (HSG). To address the role of each receptor involved in adenoviral cell entry, we mutated critical amino acids in fiber or penton to inhibit receptor interaction. A series of five adenoviral vectors was prepared and the biodistribution of each was previously characterized in mice. To evaluate possible species differences in Ad vector tropism, we characterized the effects of each detargeting mutation in non-human primates after systemic delivery to confirm our conclusions made in mice. In non-human primates, CAR was found to have minimal effects on vector delivery to all organs examined including liver and spleen. Cell-surface alpha nu integrins played a significant role in delivery of vector to the spleen, lung and kidney. The fiber shaft mutation S*, which presumably inhibits HSG binding, was found to significantly decrease delivery to all organs examined. The ability to detarget the liver corresponded with decreased elevations in liver serum enzymes (aspartate transferase [AST] and alanine transferase [ALT]) 24 hr after vector administration and also in serum interleukin (IL)-6 levels 6 hr after vector administration. The biodistribution data generated in cynomolgus monkeys correspond with those data derived from mice, demonstrating that CAR binding is not the major determinant of viral tropism in vivo. Vectors containing the fiber shaft modification may provide for a detargeted adenoviral vector on which to introduce new tropisms for the development of targeted, systemically deliverable adenoviral vectors for human clinical application.

Expression of the coxsackievirus- and adenovirus receptor in gastrointestinal cancer correlates with tumor differentiation

“…This is in agreement with studies in mice that demonstrated CARindependent uptake of adenovirus by hepatocytes mediated by heparan sulfate glycosaminoglycans. 30 We have described this finding before in the context of the limited hepatotoxicity of systemically administered ONYX-015, which might be influenced by changing accessibility of CAR with repeated virus administrations. 9 In agreement with CAR's function as a tight-junction protein, we found particularly strong expression of the protein around liver canaliculi, bile ducts and pancreatic ducts.…”

Coagulation factor X mediates adenovirus type 5 liver gene transfer in non-human primates (Microcebus murinus)

“…3,17-23 Ablation of CAR-and integrin-binding motifs in the Ad5 capsid did not alter Ad5 biodistribution in cynomolgus monkeys, indicating that an alternate transduction pathway may be responsible for liver transduction in these animals. 19 Alternative serotypes (Ad35) have also shown decreased liver transduction following i.v. delivery in non-human primates, although viral genomes can be detected in hepatic samples.…”

“…24 Other studies have obtained similar results using fiber-pseudotyped Ad5 vectors (Ad5/35 and Ad5/11) where significant gene transduction by these vectors was only observed in the marginal zone of splenic follicles. 18 Importantly, none of the reported modifications incorporated in the Ad5 capsid, via ablation of fiber and/or penton binding to natural receptors, 19 have shown significantly reduced hepatic transduction or viral genome accumulation in non-human primates. In the present study, we document the transduction profiles and viral genome accumulation of Ad5 and FX-binding-ablated Ad5 vectors in Microcebus murinus to identify whether the Ad5-FX pathway modulates Ad5 biodistribution in non-rodent species.…”

“…delivery. 4,5,19 FX-binding-ablated Ad5 vectors have shown substantially reduced hepatocyte transduction in mice and rats following i.v. delivery.…”

Transductional targeting of adenovirus vectors for gene therapy

“…Strategies to inhibit hepatocyte and/or liver Kupffer cell uptake by ablating CAR-or integrin-binding motifs in the Ad capsid have been largely unsuccessful, however, indicating that native Ad tropism determinants contribute little to vector hepatotropism in vivo. [40][41][42][43][44] These data notwithstanding, work by several groups has implicated the fiber protein as a major structural determinant of liver tropism in vivo (reviewed by Nicklin et al 45 ). 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.…”