Engineering and development of single cycle adenovirus vectors as mucosal vaccination platforms.

摘要

Adenovirus (Ad) vectors continue to be among the most potent vectors for vaccination in vivo. A majority of studies developing Ad as a vaccine have utilized first generation replication defective (RD-Ad) vectors. These vectors are robust in small animal models, but their impact wanes when scaled up to larger non-human primates and humans. This is likely due to the their inability to replicate their genome and transgene, thus limiting their vaccine potential. Recently, a number of groups have turned to replication competent (RC-Ad) vectors. These vectors retain their ability to replicate their genome and transgene, which leads to more robust immune responses. A major pitfall of these vectors is that in addition to replicating their genome they also produce infectious virions that can cause adenovirus disease. The overall goal of this thesis was to generate an adenovirus vector that is safe but robust. These "single cycle" adenovirus (SC-Ad) vectors were engineered by deleting the gene for IIIa capsid cement protein of low seroprevalence adenovirus serotype 6. In the first application, we engineered SC-Ad6 to express GFP and luciferase. We found SC-Ad6 replicated its genome and transgene similarly to RC-Ad6 in mouse, human, macaque, and hamster cell lines, it did not package its genome nor did it produce infectious virions. When used for mucosal intranasal immunization of Ad-permissive Syrian hamsters, both SC-Ad and RC-Ad expressed transgenes hundreds of times higher than RD-Ad. Surprisingly, for SC-Ad, but not RC-Ad, this translated into higher transgene-specific serum and vaginal wash antibodies than for RD-Ad. After a single sublingual immunization in rhesus macaques, SC-Ad generated higher IFN-gamma ELISPOT responses and higher serum antibody levels to the transgene than RD-Ad. In the second application, we engineered these vectors to express the hemagglutinin protein from influenza A/PR/8/34, and monitored immune responses in Syrian hamsters and ferrets. Here, we showed SC-Ad6 induces higher serum IgG and hemagglutinin inhibition titers than RD-Ad6. We also showed higher antibodies in mucosal vaginal wash and bronchoalveolar lavage than RD-Ad6. Altogether, these data demonstrate that SC-Ad6 amplifies antigen genes and produces better immune responses than RD-Ad against a variety of antigens. This suggests single cycle vectors may have utility for development as vaccines against infectious pathogens.