Cell-based assay directly monitoring viral polymerase activity for drug discovery. - Project Abstract/Summary
Our goal is to develop a new cell-based HTS platform that can be used to improve antiviral drug discovery.
Infection by Dengue virus causes ~100 million illnesses annually, with outbreak severity and frequency
increasing with time. This is true of all RNA viruses, which includes Zika, Ebola, Influenza, Yellow Fever, and
Hepatitis C, to name a few. In addition to the health burden Dengue alone represents, dengue fever constitutes
a large global financial burden (~39 billion USD), and with 20% of the world’s population living in at-risk areas
there is an urgent need for dengue therapeutics. There is currently no approved universal vaccine or dengue
specific antiviral drug. Current antiviral drugs on the market target viral enzymes; viral enzymes often have
structures that are not present in mammalian cells and are absolutely required for their life cycle making them
good targets. High throughput screening assays for dengue drug discovery are either performed in vitro or in
vivo. In vitro assays are target specific, but results do not translate well when performed in cells. In vivo assays
are more clinically relevant, but the target of the drug is unknown, making it hard to predict toxicity and tailor the
drug to improve efficacy. Despite interest in dengue antivirals, no dengue antiviral has successfully passed
clinical testing, suggesting a need for a better antiviral screening platform that can combine the advantages of
in vitro and in vivo assays. We plan to use our propriety RNA aptamer technology to develop a target-specific
cell-based HTS platform targeting the RNA-dependent RNA polymerase (RdRP) of dengue virus. RdRP is an
ideal target in Dengue due to its high identity across serotypes, and an ideal target in RNA viruses because of
its critical role in viral replication, and its lack of a mammalian counterpart. The goal of this application is to
develop a fluorescent sensor capable of monitoring RdRP transcription in a cell line stably expressing viral
proteins to advance dengue antiviral discovery. Phase II of this application is to develop a completely
comprehensive cell line stably expressing both viral proteins and a fluorescent sensor for use in HTS assays.
Phase II will also include the development of similar assays for other viruses, including Zika and Ebola, based
off design knowledge from this Phase I grant.
