A HTS Assay for Identifying Anti-Viral Drugs Against Arbovirus Infections

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DESCRIPTION (provided by applicant): In this proposal, we propose to transfer our HTS-ready, florescence based caspases-3/7 assay to identify novel anti-viral drugs against arboviruses infection based on the different host apoptotic responses in BTV infection model system. Arthropod borne viruses (arboviruses) are important human and/or animal pathogens that cause acute virus infections with severe diseases and/or death. Several recent human and/or animal epidemics are caused by arboviruses, including Dengue virus (DNV) in Asia, West Nile virus (WNV) in North America and Bluetongue virus (BTV) in Europe. Arboviruses are unique because they are transmitted to their vertebrate hosts by arthropod vectors, therefore, they must be capable of replicating in two very divergent host taxa--vertebrates and insects. Although arboviruses replicate efficiently in both vertebrate and insect cells, the respective host cellular responses are quite different. For example, BTV infection induces a rapid apoptotic response in vertebrate cells, whereas such apoptotic response is unapparent in insect cells, despite the productive virus replication in both host cells. While apoptosis has been positively linked to arbovirus diseases in infected animals/human, insects show no detectable signs of any diseases. Little attention has been paid to the significance of such different host cellular responses and the possibility of adapting such strategy for drug discovery to the protection of vertebrate hosts. We hypothesize that compounds preventing BTV-induced apoptosis in vertebrate cells could act either via inhibiting apoptosis or via interfering viral life-cycle. A secondary assay will allow us to separate anti-viral hits from the apoptosis inhibitors. These anti-viral compounds could presumably also protect host cells from host apoptotic response induced by other arboviruses, including DNV and WNV. Our long term goal is to develop new prevention and control measures for arbovirus diseases in human. The objectives of this application will be achieved by carrying out the following Specific Aim: To screening designated compound library using the developed efficient HTS assay in the BTV infection model in vertebrate cells. Based on our preliminary data showing that BTV-induced apoptosis in vertebrate cells via intrinsic apoptotic pathway, an efficient HTS assay have been designed, developed, optimized and validated using this model system. We propose to transfer this assay to the designated screening center to reproduce, miniaturize and automate the assay. A secondary assay using apoptosis inducers including Staurosporine will also be implemented to confirm hits and exclude false positives including apoptosis inhibitors. The possible mechanism of these hits will also be examined to prioritize these hits for further investigations against DNV and WNV infection. Arthropod borne viruses (arboviruses) are important human and/or animal pathogens that cause acute virus infections with severe diseases and/or death, several arboviruses cause recent human or animal epidemics, including Dengue virus in Asia, West Nile virus in North America and Bluetongue virus in Europe. In this proposal, we proposed to transfer our HTS-ready, florescence based caspases- 3/-7 assay to identify novel targets against arboviruses infection based on the different host apoptotic responses in arbovirus infection to protect hosts from arboviruses infections, including Dengue virus and Bluetongue virus. Our long term goal is to develop new prevention and control measures for arbovirus diseases in human and animals.