Development of Inhibitors Against SARS CoV-2 NSP16 Methyltransferase

Abstract: The recent outbreak of SARS-CoV-2 has caused severe economical and humanitarian loss worldwide after it first emergence in Wuhan, China in 2019. The genetic material of this virus is comprised of enveloped, positive- sense, single-stranded RNA. SARS-CoV-2 genome is evolutionary related to the beta coronavirus SARS-CoV causing the previous outbreak in 2003. Currently several vaccines have been launched but targeted therapy to control viral replication and propagation remains limited. In order to rapidly innovate effective medications for clinical curing of this viral infection, we propose to develop new therapeutic agents against COVID-19. We aim to target the nonstructural protein 16 (Nsp16) of SARS-CoV-2, the ribose 2′-O-methyltransferase enzyme (2′-O- MTase) that is responsible for the formation of viral RNA cap-1 structure. The Nsp16 MTase requires the Nsp10 adaptor protein as a co-factor for its enzyme function. The methylation of viral RNA by the Nsp16/Nsp10 complex is critical for both viral replication and viral evasion by host immune recognition. Thus, inhibitors targeting the viral RNA 5'-capping pathways are ideal for controlling and eliminating the virulence of SARS-CoV-2. In this project, we will perform fluorescence polarization (FP)-based high throughput screening (HTS) of small molecule compound libraries using innovative fluorescent analogs of SAM, the methyl donor. We will follow up with several functional MTase assays including HTRF and 32P-labeled substrates to confirm the identified inhibitory activity of selected novel drug-like molecules against the Nsp16/Nsp10 MTase complex of SARS-CoV-2. Validated Nsp16/Nsp10 2'-O-MTase inhibitors will be tested for antiviral activity against SARS-CoV-2. Medicinal optimization of promising candidates will be carried out. Finally, we will use in vivo animal models to investigate the antiviral efficacy of the most potent candidate. The accomplishment of this drug discovery project is to generate a novel avenue of experimental therapy against the existential COVID-19 pandemic via inhibiting the 5'-capping pathways of the coronavirus.