Selection of multi-valent DNA-encoded chemical library against influenza hemagglutinin and selection of dual-display DNA-encoded chemical library for SARS-CoV-2 main protease inhibitor discovery

Abstract

DNA-encoded chemical library (DEL) has come of age and become a powerful drug discovery technology. DELs enable ultra-high-throughput screening of large chemical libraries against protein targets at a minute scale for rapid ligand discovery. In this thesis, we designed and synthesized two novel DELs targeting influenza hemagglutinin and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease (Mpro) and discovered several binders with inhibitory activities. Influenza viruses use hemagglutinin (HA), a trimeric protein complex on the capsid, to bind to the sialic acid (SA) glycans on the membrane of host cells. Therefore, compounds that can inhibit hemagglutinin-SA interactions are expected to be effective agents to inhibit viral entry into host cells and have a high potential to be developed as anti-influenza drugs. However, since viral hemagglutinin trimers bind to multiple SA glycans simultaneously, mono-valent hemagglutinin inhibitors have been proven to be largely ineffective in blocking SA binding. In order to address this issue, we designed a novel multi-valent DNA-encoded chemical library that can display three ligands simultaneously on a three-way junction DNA scaffold, spatially matching the three SA-binding sites on hemagglutinin trimer and realizing high affinity and selectivity with the multi-valent effect. We have built a library containing 24 million DNA-glycan constructs, and the library was selected against several hemagglutinin trimers. For the H3 selection, the top-performing compound 133-133-133 14-11-17 binds to H3 with a dissociation constant (Kd) of 636 nM and shows hemagglutination inhibition activity of 3 μM (Ki). The multi-valent DNA-encoded library can potentially be an effective tool for exploring the unique ligand spaces of other multi-valent biological systems. Mpro is an essential and highly conserved enzyme in SARS-CoV-2 and other coronaviruses. We developed a dual-display DNA-encoded chemical library with an electrophilic warhead targeting the nucleophilic amino acid residues at Mpro’s catalytic site for the discovery of potential covalent inhibitors. A series of DNA-encoded chemical libraries totaling 54 million compounds were selected against Mpro, and the most potent hit compound 10 with the warhead 111 showed an inhibitory activity of 34.7 nM (IC50), which may be further pursued as potential anti-SARS-CoV-2 drug candidates. Collectively, these studies have shown that novel forms of DELs could be designed to interrogate different types of biologicals, thus further expanding the application scope of DELs in drug discovery.