Development of a Multivalent Macrocyclic Peptide That Shows in Vivo Protection Against SARS-CoV-2 Infection Following Intranasal Administration

The risk of zoonotic spillover of coronaviruses poses a constant threat to humanity, as recently exemplified by SARS‑CoV-2. During the pandemic, the rapid emergence of immune escape mutations led to a loss of efficacy of both therapeutic and humoral anti‑spike antibodies, emphasizing the need for therapeutics that engage binding sites within the spike protein that are both conserved among related viruses and not under selective pressure from immune surveillance. Here, we report the development of a first-in-class fusion inhibitor cyclic peptide that targets such a site. Guided by a deep mutational scan of our earlier mRNA display hit we arrived at an analogue with 5 mutations that retains broad activity within the subgenus sarbecovirus, including against SARS-CoV-1, but shows up to 50-fold improved potency and approximately 25-fold improved solubility. Multimerization further improved IC50 by one order of magnitude to give a tetrameric construct (H4v1Az)4 that inhibits Omicron BA.2.86 replication with an IC50 of approximately 10 nM in cellular assays, including human airway epithelial cell primary cultures. Moreover, (H4v1Az)4 demonstrated significant antiviral efficacy in vivo in a Syrian golden hamster model following combined pre- and post-exposure prophylactic intranasal administration. This multimeric peptide represents a promising broadly active candidate suitable both for treatment of SARS-CoV-2 infection and for preparedness against future emerging sarbecoviruses.