A broad-spectrum macrocyclic peptide inhibitor of the SARS-CoV-2 spike protein.

The ongoing COVID-19 pandemic has had great societal and health consequences. Despite the availability of vaccines, infection rates remain high due to immune evasive Omicron sublineages. Broad-spectrum antivirals are needed to safeguard against emerging variants and future pandemics. We used messenger RNA (mRNA) display under a reprogrammed genetic code to find a spike-targeting macrocyclic peptide that inhibits SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) Wuhan strain infection and pseudoviruses containing spike proteins of SARS-CoV-2 variants or related sarbecoviruses. Structural and bioinformatic analyses reveal a conserved binding pocket between the receptor-binding domain, N-terminal domain, and S2 region, distal to the angiotensin-converting enzyme 2 receptor-interaction site. Our data reveal a hitherto unexplored site of vulnerability in sarbecoviruses that peptides and potentially other drug-like molecules can target.

Inhibition of H1 and H5 Influenza A Virus Entry by Diverse Macrocyclic Peptides Targeting the Hemagglutinin Stem Region.

Influenza A viruses pose a serious pandemic risk, while generation of efficient vaccines against seasonal variants remains challenging. There is thus a pressing need for new treatment options. We report here a set of macrocyclic peptides that inhibit influenza A virus infection at low nanomolar concentrations by binding to hemagglutinin, selected using ultrahigh-throughput screening of a diverse peptide library. The peptides are active against both H1 and H5 variants, with no detectable cytotoxicity. Despite the high sequence diversity across hits, all tested peptides were found to bind to the same region in the hemagglutinin stem by HDX-MS epitope mapping. A mutation in this region identified in an escape variant confirmed the binding site. This stands in contrast to the immunodominance of the head region for antibody binding and suggests that macrocyclic peptides from in vitro display may be well suited for finding new druggable sites not revealed by antibodies. Functional analysis indicates that these peptides stabilize the prefusion conformation of the protein and thereby prevent virus-cell fusion. High-throughput screening of macrocyclic peptides is thus shown here to be a powerful method for the discovery of novel broadly acting viral fusion inhibitors with therapeutic potential.