Structure-based identification of naphthoquinones and derivatives as novel inhibitors of main protease Mpro and papain-like protease PLpro of SARS-CoV-2 (preprint)

The worldwide COVID-19 pandemic caused by the coronavirus SARS-CoV-2 urgently demands novel direct antiviral treatments. The main protease (Mpro) and papain-like protease (PLpro) are attractive drug targets among coronaviruses due to their essential role in processing the polyproteins translated from the viral RNA. In the present work, we virtually screened 688 naphthoquinoidal compounds and derivatives against Mpro of SARS-CoV-2. Twenty-four derivatives were selected and evaluated in biochemical assays against Mpro using a novel fluorogenic substrate. In parallel, these compounds were also assayed with SARS-CoV-2 PLpro. Four compounds inhibited Mpro with half-maximal inhibitory concentration (IC50) values between 0.41 M and 66 M. In addition, eight compounds inhibited PLpro with IC50 ranging from 1.7 M to 46 M. Molecular dynamics simulations suggest stable binding modes for Mpro inhibitors with frequent interactions with residues in the S1 and S2 pockets of the active site. For two PLpro inhibitors, interactions occur in the S3 and S4 pockets. In summary, our structure-based computational and biochemical approach identified novel naphthoquinonal scaffolds that can be further explored as SARS-CoV-2 antivirals.

Colloidal aggregators in biochemical SARS-CoV-2 repurposing screens (preprint)

To fight the SARS-CoV-2 pandemic, much effort has been directed toward drug repurposing, testing investigational and approved drugs against several viral or human proteins in vitro. Here we investigate the impact of colloidal aggregation, a common artifact in early drug discovery, in these repurposing screens. We selected 56 drugs reported to be active in biochemical assays and tested them for aggregation by both dynamic light scattering and by enzyme counter screening with and without detergent; seventeen of these drugs formed colloids at concentrations similar to their literature reported IC50s. To investigate the occurrence of colloidal aggregators more generally in repurposing libraries, we further selected 15 drugs that had physical properties resembling known aggregators from a common repurposing library, and found that 6 of these aggregated at micromolar concentrations. An attraction of repurposing is that drugs active on one target are considered de-risked on another. This study suggests not only that many of the drugs repurposed for SARS-CoV-2 in biochemical assays are artifacts, but that, more generally, when screened at relevant concentrations, drugs can act artifactually via colloidal aggregation. Understanding the role of aggregation, and detecting its effects rapidly, will allow the community to focus on those drugs and leads that genuinely have potential for treating COVID-19. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/458413v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@bfe5a2org.highwire.dtl.DTLVardef@7d05a1org.highwire.dtl.DTLVardef@1fcc3a5org.highwire.dtl.DTLVardef@190a153_HPS_FORMAT_FIGEXP M_FIG Table of Contents Graphic C_FIG

Bivalent binding of a fully human IgG to the SARS-CoV-2 spike proteins reveals mechanisms of potent neutralization (preprint)

In vitro antibody selection against pathogens from naive combinatorial libraries can yield various classes of antigen-specific binders that are distinct from those evolved from natural infection1-4. Also, rapid neutralizing antibody discovery can be made possible by a strategy that selects for those interfering with pathogen and host interaction5. Here we report the discovery of antibodies that neutralize SARS-CoV-2, the virus responsible for the COVID-19 pandemic, from a highly diverse naive human Fab library. Lead antibody 5A6 blocks the receptor binding domain (RBD) of the viral spike from binding to the host receptor angiotensin converting enzyme 2 (ACE2), neutralizes SARS-CoV-2 infection of Vero E6 cells, and reduces viral replication in reconstituted human nasal and bronchial epithelium models. 5A6 has a high occupancy on the viral surface and exerts its neutralization activity via a bivalent binding mode to the tip of two neighbouring RBDs at the ACE2 interaction interface, one in the "up" and the other in the "down" position, explaining its superior neutralization capacity. Furthermore, 5A6 is insensitive to several spike mutations identified in clinical isolates, including the D614G mutant that has become dominant worldwide. Our results suggest that 5A6 could be an effective prophylactic and therapeutic treatment of COVID-19.

A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug-Repurposing (preprint)

An outbreak of the novel coronavirus SARS-CoV-2, the causative agent of COVID-19 respiratory disease, has infected over 290,000 people since the end of 2019, killed over 12,000, and caused worldwide social and economic disruption1,2. There are currently no antiviral drugs with proven efficacy nor are there vaccines for its prevention. Unfortunately, the scientific community has little knowledge of the molecular details of SARS-CoV-2 infection. To illuminate this, we cloned, tagged and expressed 26 of the 29 viral proteins in human cells and identified the human proteins physically associated with each using affinity-purification mass spectrometry (AP-MS), which identified 332 high confidence SARS-CoV-2-human protein-protein interactions (PPIs). Among these, we identify 66 druggable human proteins or host factors targeted by 69 existing FDA-approved drugs, drugs in clinical trials and/or preclinical compounds, that we are currently evaluating for efficacy in live SARS-CoV-2 infection assays. The identification of host dependency factors mediating virus infection may provide key insights into effective molecular targets for developing broadly acting antiviral therapeutics against SARS-CoV-2 and other deadly coronavirus strains.