Crystallographic Fragment Screening Identifies 21 Hits Targeting SARS-CoV-2 Nsp1ⁿᵗᵈ

What is it about?

This research reports the outcomes of two crystallographic fragment screening (CFS) campaigns targeting the N-terminal domain (NTD) of SARS-CoV-2 nonstructural protein 1 (Nsp1), aimed at identifying novel small-molecule binding sites for antiviral drug development. The study describes the design and construction of a new fragment library, the KIT library, composed of 96 structurally diverse compounds selected from over 1100 candidates based on chemical properties and cluster representation. Both the KIT library and the established F2X-Entry Screen were used in parallel CFS campaigns, yielding 11 and 10 fragment hits respectively—together, the highest number of hits reported for Nsp1ⁿᵗᵈ using nontailored libraries. The majority of fragment hits were found to bind a hydrophobic pocket within Nsp1ⁿᵗᵈ, a region adjacent to functionally important residues involved in mRNA cleavage, ribosome interaction, and recognition of the viral 5′ leader sequence. The article highlights the advantages of the KIT library, including compound stability, availability, clear synthetic provenance, and the presence of halogenated fragments that facilitate binding orientation analysis via anomalous diffraction signals. Comparative analysis indicates that the diverse 3D pharmacophore space of both libraries likely contributes to their high hit rates. These findings establish a foundation for further structure-based optimization of Nsp1 inhibitors as potential antivirals targeting SARS-CoV-2 and related viruses.

Featured Image

Photo by Giovanni Crisalfi on Unsplash

Photo by Giovanni Crisalfi on Unsplash

Why is it important?

This research targets the urgent need for new antiviral strategies against SARS-CoV-2 by exploring novel small-molecule inhibitors of the essential viral protein Nsp1, addressing the limitations of current therapeutics that target only a small subset of viral proteins. By focusing on Nsp1, a multifunctional protein crucial for viral replication and immune evasion, the study contributes to the broader goal of expanding the repertoire of antiviral drug targets and preparing for future coronavirus pandemics. Key Takeaways: 1. This study conducts two crystallographic fragment screening (CFS) campaigns against the SARS-CoV-2 Nsp1 N-terminal domain (Nsp1ⁿᵗᵈ), leading to the identification of 21 new fragment hits that can serve as starting points for antiviral drug development. 2. Findings reveal that the newly developed KIT library and the established F2X-Entry Screen both achieved high hit rates against Nsp1ⁿᵗᵈ, attributed to their broad 3D pharmacophore diversity and structural variation, and that the KIT library offers practical advantages due to its academic origins, compound availability, and chemical tractability. 3. The research demonstrates that most fragment hits bind to a hydrophobic pocket on Nsp1ⁿᵗᵈ located near functionally important regions involved in mRNA cleavage and ribosome binding, providing structural insights that can inform the design of selective inhibitors targeting critical aspects of the viral lifecycle.