A Tetrapodal Tryptophan Derivative with Multiple Exposed Free Carboxylic Acids Blocks Host Cell Entry of Omicron SARS-Cov-2 and Respiratory Syncytial Virus

Omicron sublineages of SARS-CoV-2 have accelerated the spread of the virus and facilitated immune escape. In this work, we demonstrate that compound <b>2</b>, a potent HIV and enterovirus A71 (EV-A71) entry inhibitor previously discovered in our research group, also displays potent <i>in vitro</i> activity against different SARS-CoV-2 Omicron variants while showing no activity against the ancestral SARS-CoV-2 Wuhan strain. Moreover, its sodium salt (<b>2-Na salt</b>) exhibited <i>in vivo</i> antiviral activity in a murine model of Omicron BA.1. infection. Indeed, biophysical and cryo-EM studies revealed binding of <b>2-Na salt</b> to the Omicron BA.1 spike (S) protein stabilizing a "closed" form in which ∼75% of the S particles have all RBDs down, unlike the usual "open" form with one RBD up. Such "closed" form decreases S avidity for the cellular receptor ACE2, thus inhibiting viral entry. Computer-assisted modeling studies strongly suggest that <b>2</b> can interact with the intersubunit cavity of the S trimer of the Omicron BA.1 subvariant, making use of the multivalency principle. In this context, compound <b>2</b> can be considered a pharmacological tool for studying and validating new antiviral strategies against Omicron variants. We also identified <b>2</b> as a potent inhibitor of RSV and the Ebola virus. The effectiveness of <b>2</b> against a diverse set of viruses of different families supports its use as a promising lead for the development of entry inhibitors against current and future viral infections, representing a meaningful advance in the field.