Recombinant proteins of Zaire ebolavirus induce potent humoral and cellular immune responses and protect against live virus infection in mice.

Infections with filoviruses in humans are highly virulent, causing hemorrhagic fevers which result in up to 90% mortality. In addition to natural infections, the ability to use these viruses as bioterrorist weapons is of significant concern. Currently, there are no licensed vaccines or therapeutics available to combat these infections. The pathogenesis of disease involves the dysregulation of the host's immune system, which results in impairment of the innate and adaptive immune responses, with subsequent development of lymphopenia, thrombocytopenia, hemorrhage, and death. Questions remain with regard to the few survivors of infection, who manage to mount an effective adaptive immune response. These questions concern the humoral and cellular components of this response, and whether such a response can be elicited by an appropriate prophylactic vaccine. The data reported herein describe the production and evaluation of a recombinant subunit Ebola virus vaccine candidate consisting of insect cell expressed Zaire ebolavirus (EBOV) surface glycoprotein (GP) and the matrix proteins VP24 and VP40. The recombinant subunit proteins are shown to be highly immunogenic in mice, yielding both humoral and cellular responses, as well as highly efficacious, providing up to 100% protection against a lethal challenge with live virus. These results demonstrate proof of concept for such a recombinant non-replicating vaccine candidate in the mouse model of EBOV which helps to elucidate immune correlates of protection and warrants further development.

A Novel Synthetic TLR-4 Agonist Adjuvant Increases the Protective Response to a Clinical-Stage West Nile Virus Vaccine Antigen in Multiple Formulations.

West Nile virus (WNV) is a mosquito-transmitted member of the Flaviviridae family that has emerged in recent years to become a serious public health threat. Given the sporadic nature of WNV epidemics both temporally and geographically, there is an urgent need for a vaccine that can rapidly provide effective immunity. Protection from WNV infection is correlated with antibodies to the viral envelope (E) protein, which encodes receptor binding and fusion functions. Despite many promising E-protein vaccine candidates, there are currently none licensed for use in humans. This study investigates the ability to improve the immunogenicity and protective capacity of a promising clinical-stage WNV recombinant E-protein vaccine (WN-80E) by combining it with a novel synthetic TLR-4 agonist adjuvant. Using the murine model of WNV disease, we find that inclusion of a TLR-4 agonist in either a stable oil-in-water emulsion (SE) or aluminum hydroxide (Alum) formulation provides both dose and dosage sparing functions, whereby protection can be induced after a single immunization containing only 100 ng of WN-80E. Additionally, we find that inclusion of adjuvant with a single immunization reduced viral titers in sera to levels undetectable by viral plaque assay. The enhanced protection provided by adjuvanted immunization correlated with induction of a Th1 T-cell response and the resultant shaping of the IgG response. These findings suggest that inclusion of a next generation adjuvant may greatly enhance the protective capacity of WNV recombinant subunit vaccines, and establish a baseline for future development.

The development of recombinant subunit envelope-based vaccines to protect against dengue virus induced disease.

Challenges associated with the interference observed between the dengue virus components within early tetravalent live-attenuated vaccines led many groups to explore the development of recombinant subunit based vaccines. Initial efforts in the field were hampered by low yields and/or improper folding, but the use of the Drosophila S2 cell expression system provided a mechanism to overcome these limitations. The truncated dengue envelope proteins (DEN-80E) for all four dengue virus types are expressed in the S2 system at high levels and have been shown to maintain native-like conformation. The DEN-80E proteins are potent immunogens when formulated with a variety of adjuvants, inducing high titer virus neutralizing antibody responses and demonstrating protection in both mouse and non-human primate models. Tetravalent vaccine formulations have shown no evidence of immune interference between the four DEN-80E antigens in preclinical models. Based on the promising preclinical data, the recombinant DEN-80E proteins have now advanced into clinical studies. An overview of the relevant preclinical data for these recombinant proteins is presented in this review.

Development of a recombinant tetravalent dengue virus vaccine: immunogenicity and efficacy studies in mice and monkeys.

Truncated recombinant dengue virus envelope protein subunits (80E) are efficiently expressed using the Drosophila Schneider-2 (S2) cell expression system. Binding of conformationally sensitive antibodies as well as X-ray crystal structural studies indicate that the recombinant 80E subunits are properly folded native-like proteins. Combining the 80E subunits from each of the four dengue serotypes with ISCOMATRIX adjuvant, an adjuvant selected from a set of adjuvants tested for maximal and long lasting immune responses, results in high titer virus neutralizing antibody responses. Immunization of mice with a mixture of all four 80E subunits and ISCOMATRIX adjuvant resulted in potent virus neutralizing antibody responses to each of the four serotypes. The responses to the components of the tetravalent mixture were equivalent to the responses to each of the subunits administered individually. In an effort to evaluate the potential protective efficacy of the Drosophila expressed 80E, the dengue serotype 2 (DEN2-80E) subunit was tested in both the mouse and monkey challenge models. In both models protection against viral challenge was achieved with low doses of antigen in the vaccine formulation. In non-human primates, low doses of the tetravalent formulation induced good virus neutralizing antibody titers to all four serotypes and protection against challenge with the two dengue virus serotypes tested. In contrast to previous reports, where subunit vaccine candidates have generally failed to induce potent, protective responses, native-like soluble 80E proteins expressed in the Drosophila S2 cells and administered with appropriate adjuvants are highly immunogenic and capable of eliciting protective responses in both mice and monkeys. These results support the development of a dengue virus tetravalent vaccine based on the four 80E subunits produced in the Drosophila S2 cell expression system.