Small molecule identified as a SARS-CoV-2 inhibitor

By Dr. Liji Thomas, MDJan 6 2022Reviewed by Danielle Ellis, B.Sc.

The coronavirus disease 2019 (COVID-19) pandemic is caused by the beta-coronavirus termed the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Its rapid and extensive spread has strained healthcare systems and the global economy to near breaking point. Without few specific antivirals available at present, the need of the hour is to develop effective drugs that can specifically inhibit viral entry and infection of host cells both before and after exposure.

Study: Compound screen identifies the small molecule Q34 as an inhibitor of SARS-CoV-2 infection. Image Credit: Piloci Studio/Shutterstock

Investigators are exploring the feasibility of suppressing SARS-CoV-2 entry into the target human host cells to prevent infection at an early stage in the viral life cycle. This could help prevent infection as well as propagation of the virus. A new study published in iScience reports the results of a screening strategy to identify small molecules that target viral entry to block infection with this virus.

What did the study show?

This study aimed to pick out compounds from a library of over 260,000 compounds, which are expected to suppress the protein-protein interaction between the human cell receptor for the virus, the angiotensin-converting enzyme 2 (ACE2) and the viral spike protein. The screening strategy led to the identification of compound Q34.

This compound was a potent inhibitor of the cell entry of SARS-CoV-2 but does not cause cytotoxicity. The researchers also tested the effect of the compound on cell entry by pseudovirus bearing either the SARS-CoV-2 spike or the vesicular stomatitis virus glycoprotein (VSV-G).

This showed that Q34 inhibits SARS-CoV-2 spike-mediated cell entry but does not block VSV-G-mediated entry. There was no observable effect on cell viability, indicating a lack of toxicity and suggesting that the inhibition of cell entry was not due to this effect.

When the cells were pretreated with Q34, SARS-CoV-2 replication was dramatically reduced compared to the control-treated cells, but this effect was abolished when the compound was added to already infected cells. This indicates that this compound suppresses the entry of the virus into the cells.

Since the compound is expected to block the ACE2-spike interaction, experimental verification of this activity was performed, which showed no inhibition of this interaction. This unexpected finding was followed by the equally surprising finding that the endosomal proteases cathepsin B and L were also not inhibited by Q34. These were required for priming the virus for entry into cells that lack the more often used protease TMPRSS2.

When TMPRSS2 was overexpressed or combined with the expression of another putative entry receptor, NRP1, the cells showed a higher rate of viral entry irrespective of treatment with either Q34 or control. This seems to rule out the involvement of either of these molecules in the mechanism of entry of the virus into the host cell.

It is evident from these results that the compound successfully suppresses the entry of the pseudovirus bearing the SARS-CoV-2 spike into cells. Moreover, this effect is seen over a wide range of dosages, indicating a good safety profile.

When tested with authentic SARS-CoV-2, the compound could prevent viral entry into hACE2-HEK cells substantially. When used to pretreat cells, Q34 also reduced viral ribonucleic acid (RNA) loads in infected cells compared to those pretreated with the control. The level of inhibition is comparable to that achieved by chloroquine.

The almost complete suppression of SARS-CoV-2 infection with Q34 at levels of 100 μM was not accompanied by cytotoxicity compared to the controls. The supernatant from cell cultures treated with Q34 showed a drastic reduction in viral spike protein levels, supporting the strong inhibition exerted by this compound on viral entry and infection of the target cells.

This effect was similar to that observed in cells treated with chloroquine. Thus, Q34 efficiently disrupts SARS-CoV-2 entry into human cells without causing cytotoxicity in terms of reduced cell number or altered morphology.

The compound also showed inhibition of authentic virus entry into human iPSC-derived neurons and astrocytes, with a steep drop in the infection rate compared to controls. The viral RNA load within these cells was dramatically reduced with Q34 treatment, again without obvious toxic effects on the cells. The pathological changes induced by SARS-CoV-2 infection, like degeneration of neurites, and apoptosis of infected astrocytes, were also largely mitigated by treatment with this compound.

Next, they showed that treatment with this compound prevented infection of Calu-3 cells, as shown by a steep reduction in the percentage of cells containing the spike protein. The levels of viral RNA and spike protein in the supernatant were also reduced in treated cells.

Taken together, these data demonstrate that Q34 efficiently inhibits SARS-CoV-2 infection of COVID disease-relevant human cells.”

What are the implications?

As COVID-19 continues to threaten the world into the near foreseeable future, the search for effective and safe antivirals continues as part of the effort to reduce morbidity and mortality during the waves of infection.

The current study demonstrates the promising results of Q34 on SARS-CoV-2 entry into relevant human cell lines, with marked inhibition of cell entry and the prevention of pathological changes in infected cells, and without obvious cytotoxic effects in treated cells.

The use of the pseudovirus system for screening made it possible to screen safely and rapidly, compared to the use of the authentic virus, besides providing a quantitative readout due to the incorporated luciferase reporter. This robust and sensitive platform bypasses the need to use biosafety level 3 facilities and widens the scope of research into antiviral inhibitors.

In view of the observed potent inhibition of SARS-CoV-2 entry into human cells by Q34, it is possible that “this small molecule compound and its analogs have great potential to be developed into effective prophylactics and therapeutics for COVID-19 and related pandemics in the future.”