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Authors: Cheng, Y-W. et al.

Link to paper: https://papers.ssrn.com/sol3/papers.cfm?abstract_id=3613035

Journal/ Pre-Print: Cell Reports (under review)

Tags: Cell Biology, Drug discovery/Drug repurpose, Therapeutics, Virology

Research Highlights

1. Furin inhibitors CMK and naphthofluorescein prevent cleavage of the SARS-CoV-2 spike (S) protein and syncytia formation in VeroE6 cells expressing WT S protein, in a similar way to mutation of the furin cleavage site

2. CMK and naphthofluorescein suppresses S protein cleavage and syncytia formation upon infection of VeroE6 cells with authentic SARS-CoV-2 virus

3. CMK and TMPRSS2 inhibitor camostat prevented initial viral entry, with CMK also reducing syncytium formation later in the infection, whereas naphthofluorescein affected viral replication after initial entry into host cells

Summary 

This study investigated the antiviral potential of targeting furin-mediated SARS-CoV-2 spike (S) protein cleavage. Mutation of the furin cleavage site or use of furin inhibitors CMK or naphthofluorescein reduced S protein cleavage and syncytia formation in VeroE6 cells expressing SARS-CoV-2-S. Similar results were observed upon CMK or naphthofluorescein treatment of Vero cells infected with authentic SARS-CoV-2. The stage of viral replication also appeared to be important for each inhibitor. CMK and camostat were most effective pre-infection and naphthofluorescein was effective post-infection. Although viral replication was not abolished by CMK treatment post-infection, the authors note that CMK is still able to reduce syncytium formation and may therefore still impede virulence.

Impact for SARS-CoV2/COVID19 research efforts

Understand the virology and/or cell biology of SARS-CoV2/COVID19

Inhibit of SARS-CoV2/COVID19 transmission

Treat of SARS-CoV2/COVID19 positive individuals (proof of concept)

Study Type

· In vitro study - VeroE6 cell lines were infected with authentic SARS-CoV-2 virus

Strengths and limitations of the paper

Novelty: In-depth characterisation of furin inhibitor action in cells infected with authentic virus. Gives greater focus to syncytium formation than new virus production, consistent with previous (non-COVID-19) papers showing that this is a more effective than cell-free spread of viruses

Standing in the field: Nothing controversial. Supportive of role for furin and TMPRSS2 in SARS-CoV-2-S cleavage

Appropriate statistics: No statistical test used

Viral model used: Most experiments conducted using authentic virus in VeroE6 cells. Specific isolate from patients used - hCoV-19/Taiwan/NTU03/2020

Translatability: Yes. The authors suggest that CMK could be used as a prophylactic drug to prevent virus spread to uninfected individuals, due to its role in preventing initial viral entry. However, furin is expressed in multiple tissues and has other functions in tissue homeostasis, so CMK use could be associated with multiple side effects. It is also not an approved drug or currently in clinical use. This is also true to naphthofluorescein, which they suggest could be used to treat COVID-19 patients due to its effect on viral RNA transcription.

Main limitations: 1) Do not state number of experimental repeats

2) Also, no statistical tests were used

3) The authors draw conclusions about mechanism of action of the inhibitors from associations (see pre- and post-infection experiment (Figure 5)) rather than directly testing the molecular mechanism

4) In dose-dependence experiments, effective doses of inhibitors are still quite high (e.g. 50μM for CMK, 20μM for naphthofluorescein)

5) The effect of CMK on virus titres does not seem very big, relative to naphthofluorescein; statistical tests would help to determine whether there was a difference