Firewalls Prevent Systemic Dissemination of Vectors Derived from Human Adenovirus Type 5 and Suppress Production of Transgene-Encoded Antigen in a Murine Model of Oral Vaccination.

To define the bottlenecks that restrict antigen expression after oral administration of viral-vectored vaccines, we tracked vectors derived from the human adenovirus type 5 at whole body, tissue, and cellular scales throughout the digestive tract in a murine model of oral delivery. After intragastric administration of vectors encoding firefly luciferase or a model antigen, detectable levels of transgene-encoded protein or mRNA were confined to the intestine, and restricted to delimited anatomical zones. Expression of luciferase in the form of multiple small bioluminescent foci in the distal ileum, cecum, and proximal colon suggested multiple crossing points. Many foci were unassociated with visible Peyer's patches, implying that transduced cells lay in proximity to villous rather than follicle-associated epithelium, as supported by detection of transgene-encoded antigen in villous epithelial cells. Transgene-encoded mRNA but not protein was readily detected in Peyer's patches, suggesting that post-transcriptional regulation of viral gene expression might limit expression of transgene-encoded antigen in this tissue. To characterize the pathways by which the vector crossed the intestinal epithelium and encountered sentinel cells, a fluorescent-labeled vector was administered to mice by the intragastric route or inoculated into ligated intestinal loops comprising a Peyer's patch. The vector adhered selectively to microfold cells in the follicle-associated epithelium, and, after translocation to the subepithelial dome region, was captured by phagocytes that expressed CD11c and lysozyme. In conclusion, although a large number of crossing events took place throughout the intestine within and without Peyer's patches, multiple firewalls prevented systemic dissemination of vector and suppressed production of transgene-encoded protein in Peyer's patches.

Antigen encoded by vaccine vectors derived from human adenovirus serotype 5 is preferentially presented to CD8+ T lymphocytes by the CD8α+ dendritic cell subset.

Different subsets of dendritic cells (DC) elicit qualitatively different immune responses. In mice, two lymphoid tissue-resident subsets, CD8α(+) and CD8α(-), have been implicated in the induction of T helper 1 (Th1) or Th2 responses, respectively. Moreover, CD8α(+) DC appear to play a major role in priming CD8(+) T lymphocyte responses to viral antigens in the course of diverse viral infections. These considerations have been less extensively explored for vaccine vectors derived from viruses. Despite inefficient ex vivo transduction of DC, vectored vaccines derived from human adenoviruses of serotype 5 (Ad5) elicit robust immune responses, predominantly of the Th1 orientation, in humans and mice. At present it is unknown whether Ad5 interacts with DC subsets in a differential manner, thereby influencing the quality of the elicited IR. To address this issue, successive steps (attachment, transgene expression, MHC class I antigen presentation and activation of antigen-specific T lymphocytes) involved in induction of immune responses by Ad5-based vectors have been examined in CD8α(+) and CD8α(-) murine DC subsets. Although in both ex vivo and in vivo experiments CD8α(+) and CD8α(-) DC subsets captured an Ad5-based vector to a similar extent, transgene expression and subsequent MHC class I display of a transgene-encoded antigen were more efficient in CD8α(+) DC. Moreover, following in vivo and ex vivo transduction with an Ad5-based vaccine, antigen-specific CD8(+) T lymphocytes were more efficiently activated by CD8α(+) DC than by CD8α(-) DC. Thus, superior antigen expression and MHC class I display in CD8α(+) DC may contribute to preferred priming of antigen-specific CD8(+) lymphocytes by Ad5-transduced CD8α(+) DC.

Gag-specific immune enhancement of lentiviral infection after vaccination with an adenoviral vector in an animal model of AIDS.

The evaluation of vaccine strategies in animal models is essential for the development of a vaccine against HIV. In efficacy trials conducted in non-human primate models of AIDS, vaccines based on adenoviruses compared favourably with other vaccine vectors. To determine whether this strategy could be transposed to another animal model, and by extension, to humans, we have evaluated the efficacy of adenoviral vectors in a natural model of AIDS, infection of the cat by the feline immunodeficiency virus (FIV). Recombinant canine adenoviruses expressing the envelope glycoproteins or the Gag protein of a primary strain of FIV were constructed. Three groups of six cats were immunised twice with vectors expressing FIV antigens or with a vector expressing an irrelevant antigen, green fluorescent protein, by intramuscular and subcutaneous routes. Humoral responses were elicited against the transgene product in 6/6, 3/6 and 0/6 cats after immunisation against green fluorescent protein, Gag or the envelope glycoproteins, respectively. Six weeks after the second administration, cats were challenged by the intraperitoneal route with the homologous strain, and viral burden in plasma was followed by quantitative RT-PCR. Immunisation with FIV antigens did not afford protection. Rather, viral RNA was detected at earlier time points in cats immunised against Gag than in cats immunised with a vector expressing an irrelevant antigen. Such immune-mediated enhancement did not appear to have a long-range impact on viral set point or inversion of the CD4(+)/CD8(+) ratio. Thus, in the feline AIDS model pre-existing immunity against a viral antigen exacerbated acute phase infection.