1. Bivalent vaccines that confer protection against rabies and other viruses We have been developing a rabies virus based vaccine that expresses the glycoprotein (GP) from ebolavirus, marburgvirus and sudanvirus, the GPC from lassa virus, and the Spike protein from MERS-CoV. Previously we developed (a) replication-competent, (b) replication-deficient, and (c) chemically inactivated rabies virus (RABV) vaccines expressing Zaire ebolavirus (EBOV) glycoprotein (GP) using a reverse genetics system based on the SAD B19 RABV wildlife vaccine in collaboration with Matthias Schnell of Thomas Jefferson University. Immunization with live or inactivated RABV vaccines expressing EBOV GP induced cellular and humoral immunity against each virus and conferred protection from both lethal RABV and EBOV challenge in mice. We have continued our evaluation of the RABV based EBOV vaccine and have found that an adjuvant improves efficacy of the inactivated vaccine in rhesus and cynomolgus macaque challenge models. A clinical trial lot of the rabies-ebolavirus has been produced and evaluated in NHPs. The Marburg-rabies construct has demonstrated immunogenicity in mice and efficacy testing in mice is underway. If successful, then efficacy testing in nonhuman primate models of Marburg virus will be pursued. The RABV-MERS vaccine has demonstrated immunogenicity and efficacy in mice. A study in camels is currently being planned. The RABV-Lassa virus vaccine candidate has demonstrated immunogenicity in mice. Safety and efficacy testing in mice and guinea pigs is scheduled to start in fall of 2016. If successful, then efficacy will be evaluated in NHP models. 2. Animal model development. EVPS is currently developing PET/CT to investigate pathogenesis of EBOV infection in NHPs. We have evaluated 18-fluorodeoxyglucose as a marker of increased glucose metabolism as a marker of inflammation and 18-fluoro-Albumin as a hemodynamic marker. We observed increased metabolism in the spleen, liver, bone marrow and draining lymph nodes that progressed rapidly and plateaued 5 days post-infection and rapidly decreased as subjects met endpoint criteria. 18-fluoro-Albumin imaging indicated congestion in the bone marrow and spleen. These data indicate that PET/CT imaging can be used to evaluate disease progression in real-time, thus establishing biomarkers that can be used to validate potential countermeasures and increase understanding of pathogenesis. 3.Filovirus Molecular Virology We have resolved the secondary structure of the EBOV 3E-5E minigenome by selective hydroxyl acylation by primer extension. Previously, we identified host proteins that interact with the EBOV trailer. Specifically, HSPA8 binds to a specific motif within the EBOV trailer and mutational analysis, chemical inhibition targeting HSPA8, and reverse genetics have demonstrated a role for HSPA8 in the virus lifecycle. We are currently identifying host proteins that interact with the EBOV NCRs from cell lines of varying permissivity to EBOV infection. Based on these data we may identify therapeutic targets as well as establish mechanisms of filovirus lifecycle regulation.
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