This is a proposal to develop a highly selective and potent antiviral benzodiazepine compound against yellow fever virus (YFV) as a therapeutic agent for treatment of yellow fever and understand its antiviral mechanism. Yellow fever is an acute viral hemorrhagic disease which threatens approximately one billion people living in tropical areas of Africa, Central and South America. Although a highly effective yellow fever vaccine has been available for more than seven decades, the low vaccination rate fails to prevent outbreaks in at-risk regions. It has been estimated that up to 1.7 million YFV infections occur in Africa each year, resulting in 29,000 to 60,000 deaths, which outnumber the estimated death toll of global dengue virus infection. Thus far, there is no specific antiviral treatment for yellow fever. To cope with this medical challenge, we discovered an acetic acid benzodiazepine (BDAA) compound that potently inhibited YFV replication in both mammalian and insect cells by targeting the viral non-structural 4B (NS4B) protein. We have already demonstrated in YFV-infected hamsters that oral administration of BDAA significantly protected the animals from death, reduced viral load by greater than 2 logs and attenuated viral infection-induced liver injury and body weight loss. Building on these encouraging preclinical studies, we propose in this project to optimize the lead compound to obtain benzodiazepine derivatives with improved solubility and reduced plasma protein binding ability to achieve improved in vivo antiviral efficacy and safety profile. Meanwhile, we will continue our efforts toward understanding the molecular mechanism by which BDAA and its derivatives inhibit YFV replication. Specifically, impacts of the compound on NS4B protein metabolism, integrity and function of viral RNA replication complex as well as YFV interaction with host cellular innate immune response will be investigated. At the completion of this project, we will nominate an optimized benzodiazepine derivative as well as backup compounds for further preclinical/clinical development. In addition to knowing how the compound inhibits YFV replication, our studies proposed in this project should also enrich our knowledge on the function of NS4B protein in YFV replication and evasion of host innate immunity.
With the unmet medical challenge of periodical yellow fever outbreaks in Africa, Central and South America, there is a pressing need for antiviral drugs to treat patients and reduce the mortality of the disease. Recently, we discovered a novel benzodiazepine compound, designated as BDAA, which potently inhibited yellow fever virus (YFV) replication and efficiently prevented the death of YFV-infected hamsters. Mechanistically, BDAA specifically targets viral non-structural protein 4B (NS4B), a key component and organizer of YFV RNA replication complex. In this application, we propose to chemically optimize the lead molecule BDAA to obtain benzodiazepine derivatives that can achieve improved in vivo antiviral efficacy and safety profile as candidates for further preclinical and clinical development. Meanwhile, we will extend our studies on the mode of action of BDAA and further investigate our hypothesis that by specific interaction with NS4B protein, BDAA impairs the integrity of YFV replication complex, which results in inhibition of viral RNA replication and enhanced activation of innate immune response. Hence, the proposed studies will not only lead to development of an orally available first-in-class antiviral agent against YFV, but also generate knowledge on the role and mechanism of NS4B protein in YFV replication and evasion of host innate immune response.