Novel Chikungunya Vaccine Candidate with an IRES-Based Attenuation and Host Range Alteration Mechanism

Chikungunya virus (CHIKV) is a reemerging mosquito-borne pathogen that has recently caused devastating urban epidemics of severe and sometimes chronic arthralgia. As with most other mosquito-borne viral diseases, control relies on reducing mosquito populations and their contact with people, which has been ineffective in most locations. Therefore, vaccines remain the best strategy to prevent most vector-borne diseases. Ideally, vaccines for diseases of resource-limited countries should combine low cost and single dose efficacy, yet induce rapid and long-lived immunity with negligible risk of serious adverse reactions. To develop such a vaccine to protect against chikungunya fever, we employed a rational attenuation mechanism that also prevents the infection of mosquito vectors. The internal ribosome entry site (IRES) from encephalomyocarditis virus replaced the subgenomic promoter in a cDNA CHIKV clone, thus altering the levels and host-specific mechanism of structural protein gene expression. Testing in both normal outbred and interferon response-defective mice indicated that the new vaccine candidate is highly attenuated, immunogenic and efficacious after a single dose. Furthermore, it is incapable of replicating in mosquito cells or infecting mosquitoes in vivo. This IRES-based attenuation platform technology may be useful for the predictable attenuation of any alphavirus

Heterologous Immunization With Defined RNA and Subunit Vaccines Enhances T Cell Responses That Protect Against Leishmania donovani

The rapid generation of strong T cell responses is highly desirable and viral vectors can have potent CD8+ T cell-inducing activity. Immunity to leishmaniasis requires selective T cell responses, with immunization schemes that raise either CD4 or CD8 T cell responses being protective in small animal models. We have defined the leishmaniasis vaccine candidate recombinant fusion antigens, LEISH-F2 and LEISH-F3+, that when formulated in a stable emulsion with a Toll-like receptor (TLR) 4 agonist, induce protective CD4+ T cell responses in animal models as well as providing therapeutic efficacy in canine leishmaniasis and in clinical trials in leishmaniasis patients. We used the genetic sequences of these validated vaccine antigens to design RNA vaccine constructs. Immunization of mice with the RNA replicons induced potent, local innate responses that were surprisingly independent of TLR7 and activated antigen-presenting cells (APC) to prime for extremely potent antigen-specific T helper 1 type responses upon heterologous boosting with either of the subunit vaccines (recombinant antigen with second generation glucopyranosyl lipid A in stable oil-in-water emulsion; SLA-SE). Inclusion of RNA in the immunization schedule also generated MHCI-restricted T cell responses. Immunization with LEISH-F2-expressing RNA vaccine followed later by subunit vaccine afforded protection against challenge with Leishmania donovani. Together, these data indicate the utility of heterologous prime-boost immunization schemes for the induction of potent antigen-specific CD4 and CD8 T cell responses for protection against intracellular pathogens

Protection of Black-Tailed Prairie Dogs (Cynomys ludovicianus) against Plague after Voluntary Consumption of Baits Containing Recombinant Raccoon Poxvirus Vaccine

Prairie dogs (Cynomys spp.) are highly susceptible to Yersinia pestis and significant reservoirs of plague for humans in the western United States. A recombinant raccoon poxvirus, expressing the F1 antigen of Y. pestis, was incorporated into a palatable bait and offered to 18 black-tailed prairie dogs (Cynomys ludovicianus) for voluntary consumption; 18 negative control animals received placebo baits. Antibody titers against Y. pestis F1 antigen increased significantly (P < 0.01) in vaccinees, and their survival was significantly higher upon challenge with Y. pestis than that of negative controls (P < 0.01)

Early Transcriptional Signatures of the Immune Response to a Live Attenuated Tetravalent Dengue Vaccine Candidate in Non-human Primates

<div><p>Background</p><p>The development of a vaccine against dengue faces unique challenges, including the complexity of the immune responses to the four antigenically distinct serotypes. Genome-wide transcriptional profiling provides insight into the pathways and molecular features that underlie responses to immune system stimulation, and may facilitate predictions of immune protection.</p><p>Methodology/Principal Findings</p><p>In this study, we measured early transcriptional responses in the peripheral blood of cynomolgus macaques following vaccination with a live, attenuated tetravalent dengue vaccine candidate, TDV, which is based on a DENV-2 backbone. Different doses and routes of vaccine administration were used, and viral load and neutralizing antibody titers were measured at different time-points following vaccination. All 30 vaccinated animals developed a neutralizing antibody response to each of the four dengue serotypes, and only 3 of these animals had detectable serum viral RNA after challenge with wild-type dengue virus (DENV), suggesting protection of vaccinated animals to DENV infection. The vaccine induced statistically significant changes in 595 gene transcripts on days 1, 3, 5 and 7 as compared with baseline and placebo-treated animals. Genes involved in the type I interferon (IFN) response, including <i>IFI44</i>, <i>DDX58</i>, <i>MX1</i> and <i>OASL</i>, exhibited the highest fold-change in transcript abundance, and this response was strongest following double dose and subcutaneous (versus intradermal) vaccine administration. In addition, modules of genes involved in antigen presentation, dendritic cell activation, and T cell activation and signaling were enriched following vaccination. Increased abundance of gene transcripts related to T cell activation on day 5, and the type I IFN response on day 7, were significantly correlated with the development of high neutralizing antibody titers on day 30.</p><p>Conclusions/Significance</p><p>These results suggest that early transcriptional responses may be predictive of development of adaptive immunity to TDV vaccination in cynomolgus macaques, and will inform studies of human responses to dengue vaccines.</p></div

Groups of animals and vaccination regimen.

<p>Groups of animals and vaccination regimen.</p

Changes in gene transcript abundance following TDV vaccination.

<p>Results from SAM analysis of transcripts from vaccinated animals compared with baseline, and vaccinated animals compared with placebo animals. (A) Numbers of differentially-expressed (DE) genes at each time-point. FDR<0.05; fold-change≥1.3 solid areas, fold-change≥1.5 dashed areas. Red bars indicate increased abundance; blue bars indicate decreased abundance. (B) Top GO terms associated with DE genes by day, p<0.05 after Benjamani correction for multiple testing. Strength of the association is shown on the abscissa. Red bars represent positively enriched GO terms; blue bars represent negatively enriched GO terms.</p

Comparison of changes in transcript abundance following wt DENV infection and TDV immunization.

<p>A) Hierarchical clustering of 135 genes whose transcripts differ in abundance over time following wt infection of sham-immunized animals (FDR<0.05, fold-change≥1.3). Median fold-change in transcript abundance is shown over time for each group; statistical significance (FDR<5% and fold-change ≥1.3) is indicated in right-hand grey column. Red indicates increased abundance; blue indicates decreased abundance. Black vertical bars delineate gene clusters 1, 2 and 3. B) Median fold-change of transcript abundance for each gene cluster by group. wt DENV, wild-type dengue virus; SCd, subcutaneous double dose; SCs, subcutaneous single dose.</p

Antiviral/type I IFN response following TDV vaccination.

<p>A) Median abundances of 38 transcripts over time (days post-vaccination) in all vaccinated animals, and by group. Red indicates an increase in transcript abundance; blue indicates decreased transcript abundance (FDR<0.05, fold-change≥1.3). Genes listed when significant in comparison between intradermal (ID) and subcutaneous (SC) groups, and SC double dose (SCd) and SC single dose (SCs) groups (p<0.05, ranksum). Placebo group, and placebo animals challenged with wt DENV, shown for comparison. B) Median relative transcript abundance over time by vaccination group. Bars indicate standard error.</p