Screening of Natural Products and Small Molecules Uncovers Novel Coronavirus 1a/1b Frameshifting Inhibitors with Antiviral Properties

The novel severe acute respiratory syndrome-related coronavirus (SARS-CoV-2) and its emerging variants continue to pose a global health emergency. Like most coronaviruses, SARS-CoV-2 replication depends on programmed ribosomal frameshifting (PRF), which, due to its conserved nature, represents a promising antiviral target. Here, we employ a microscale thermophoresis (MST) assay to screen two diverse in-house libraries of natural products and synthetic small molecules for their frameshift RNA-binding properties. We identify trichangion and the ureidothiophene-coupled antimicrobial peptide 25 as potent binders. Using MST and surface plasmon resonance (SPR), we determine the binding affinity and interaction kinetics of the hit compounds. Moreover, we model the binding of the new PRF inhibitors into the structure of the SARS-CoV-2 RNA frameshift element, thus suggesting potential binding sites for RNA-targeting molecules. Furthermore, we demonstrate that these compounds substantially reduce frameshift levels in vitro and suppress viral propagation in SARS-CoV-2-infected cells. This work highlights the potential of natural products, especially the phenothiazine class, and synthetic amphiphilic cationic peptides to target frameshifting RNA elements, which sets the stage for the development of novel antivirals.