Project 4: Nuclease Inhibitors for Viruses of Pandemic Concern

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Project 4 – Nuclease Inhibitors for Viruses of Pandemic Concern Abstract SARS-CoV-2 (SARS2) and highly pathogenic arenaviruses including Lassa virus (LASV), Junin virus, and Machupo virus share a structurally and functionally related 3'-to-5' exoribonuclease (ExoN) domain, which plays essential roles in proofreading during RNA syntheses by the error-prone viral RNA-dependent RNA polymerase (RdRp) and suppressing host antiviral responses. Project 4 is pursuing chemical inhibition of ExoN from these viruses of pandemic concern, with the goal of blocking viral replication by lethal mutagenesis as well as mitigating viral pathogenesis by reactivating host’s interferon responses. Our team has contributed extensively to the structural and functional understanding of these viral ExoN enzymes, including the elucidation of their first atomic structures and characterization of catalytic mechanisms. We have also developed robust fluorescence-based assays to quantitatively analyze the ExoN catalytic activities, including a novel assay featuring fluorogenic RNA aptamer substrates that enables a gain-of-function readout in ultra-high-throughput screening (uHTS). Furthermore, we have used DNA-encoded chemistry technology (DEC-Tec) to obtain selective binders to SARS2 ExoN and LASV nucleoprotein (NP) containing the ExoN domain. Building on these prior and preliminary studies, we will continue to work closely with Core B and use 3 complementary approaches (uHTS, DEC-Tec, and Virtual screening) to identify first-in-class viral ExoN inhibitors. We will then leverage the deep expertise of Core C and Core D in medicinal chemistry and structural biology, respectively, to enhance the potency, selectivity, and pharmacodynamic/kinetic properties of hit compounds for detailed antiviral studies in cell and animal models by Core E. These studies will deliver antiviral drug candidates with a distinct mechanism of action to complement those developed against established antiviral targets including RdRp, helping to address the critical need for novel antiviral drugs against both SARS2 and the highly pathogenic arenaviruses that cause fatal hemorrhagic fever infections.