A Novel Viral Assembly Inhibitor Blocks SARS-CoV-2 Replication in Airway Epithelial Cells

The ongoing evolution of SARS-CoV-2 to evade vaccines and therapeutics underlines the need for novel therapies with high genetic barriers to resistance. The small molecule PAV-104, identified through a cell-free protein synthesis and assembly screen, was recently shown to target host protein assembly machinery in a manner specific to viral assembly. Here, we investigated the capacity of PAV-104 to inhibit SARS-CoV-2 replication in human airway epithelial cells (AECs). Our data demonstrate that PAV-104 inhibited >99% of infection with diverse SARS-CoV-2 variants in primary and immortalized human AECs. PAV-104 suppressed SARS-CoV-2 production without affecting viral entry or protein synthesis. PAV-104 interacted with SARS-CoV-2 nucleocapsid (N) and interfered with its oligomerization, blocking particle assembly. Transcriptomic analysis revealed that PAV-104 reversed SARS-CoV-2 induction of the type-I interferon response and the maturation of nucleoprotein signaling pathway known to support coronavirus replication. Our findings suggest that PAV-104 is a promising therapeutic candidate for COVID-19. Overall design: To understand the transcriptional impact of PAV-104 in the setting of SARS-CoV-2 infection and immunopathology, we performed RNA-seq analysis on ALI-cultured primary AECs from 5 different healthy donors infected for 36 hours with SARS-CoV-2 (MOI=0.1) in the presence or absence of PAV-104. Uninfected, untreated cells (Control) were characterized as a reference for both experimental conditions. The effects of PAV-104 administration alone (in the absence of SARS-CoV-2 infection) were evaluated in three additional donors.