16. STAT2 Knockout Syrian Hamsters Support Enhanced Replication and Pathogenicity of Human Adenovirus Type 5, Revealing an Important Role of Type I Interferon Response in Viral Control

Abstract

Oncolytic vectors based on human species C adenovirus type 5 (Ad5) are being developed for cancer gene therapy. Studies on wild-type Ad5 in animal models are likely to provide important insights into the behavior of these vectors in patients. The most frequently used permissive immunocompetent animal model for Ad5 infection is the Syrian hamster. Ad5 (and Ad6) replicates in these animals and causes pathology that is similar to that seen with humans. Here, we report findings with a new Syrian hamster strain in which the STAT2 gene was functionally knocked out (KO) by site-specific CRISPR/Cas9-mediated gene targeting. STAT2 is a critical element of the Type I and Type III interferon signal transduction pathways. STAT2 KO hamsters infected intravenously with Ad5 demonstrated an accentuated pathology compared to the wild-type control animals, and the virus load in the organs (liver, lung, kidney) of STAT2 KO animals was 100- to 1000-fold higher than that in wild-type hamsters. We show that the Type I interferon pathway is disrupted in these hamsters, inasmuch as the interferon response genes PKR, OAS, and Mx2 were induced by Ad5 in the liver of wild-type but not STAT2 KO hamsters, revealing a critical role of interferon-stimulated genes in controlling Ad5 infection. Notably, the adaptive immune response to Ad5 is not adversely affected in STAT2 KO hamsters, and surviving hamsters cleared the infection by 7 to 10 days post challenge. In fact, anti-Ad5 neutralizing antibodies were 10-fold higher at 7 days postinfection in the STAT2 KO hamsters than in wild-type hamsters. T cell infiltration into the liver was similar at 3 and 7 days in the STAT2 KO and wild-type hamsters. Treatment of Ad5-infected STAT2 KO hamsters with high dose cyclophosphamide resulted in markedly increased mortality, pathogenesis, and virus replication in the liver (~1011 TCID50/g liver) at 10 days postinfection, likely because the adaptive immune response as well as other aspects of the innate response were abolished. This is the first study to report findings with a genetically modified Syrian hamster infected with human adenovirus. Further, this is the first study to show that the Type I interferon pathway plays a role in inhibiting Ad5 replication in a permissive animal model. This conclusion is in accord with a previous study in which we showed using a custom microarray platform that there is a robust up-regulation of genes involved in the innate immune response (e.g. OAS, PKR, IFN-inducible protein 10) in the liver at 18 h post-intravenous infection of wild-type Syrian hamsters with Ad5 (Ying, B. et al., Virology 485, 305, 2015). Besides providing an insight into adenovirus infection in humans, our results underscore the usefulness of Syrian hamsters as a permissive model for the study of species C human adenovirus and adenovirus-based vectors. In future studies, STAT2 KO hamsters bearing tumors could be used to explore the role of Type I and Type III interferon signaling on the efficacy of oncolytic adenovirus vectors.