COVID-19 Antivirals: Japanese Study Shows That Oxidized Cholesterol Derivatives Or Oxysterol Shows Antiviral Activity Against SARS-CoV-2

COVID-19 Antivirals: Japanese researchers from National Institute of Infectious Diseases, Tokyo and the Tokyo University of Science have in a new study found that oxidized cholesterol derivatives or oxysterol exhibits potent antiviral activity against the SARS-Cov-2 coronavirus.


 
Developments of effective antiviral drugs targeting SARS-CoV-2 are urgently needed to combat the COVID-19 disease. Oxysterols, defined as oxidized derivatives of cholesterol, include endogenous (naturally occurring) cholesterol metabolites as well as semi-synthetic oxysterol derivatives.
 
The study team had previously studied the use of semi-synthetic oxysterol derivatives as drug candidates for inhibition of cancer, fibrosis, and bone regeneration.
 
In this study, the team had screened a panel of naturally occurring and semi-synthetic oxysterol derivatives for anti-SARS-CoV-2 activity, using a cell culture infection assay.
 
The team showed that the natural oxysterols, 7-ketocholesterol, 22(R)-hydroxycholesterol, 24(S)-hydroxycholesterol, and 27-hydroxycholesterol, substantially inhibited SARS-CoV-2 propagation in cultured cells. Among semi-synthetic oxysterols, Oxy186 displayed antiviral activity comparable to natural oxysterols.
 
Furthermore related oxysterol analogues Oxy210 and Oxy232 displayed more robust anti-SARS-CoV-2 activities, reducing viral replication more than 90% at 10 μM and 99% at 15 μM, respectively. When orally administered in mice, peak plasma concentrations of Oxy210 fall into a therapeutically relevant range (19 μM), based on the dose-dependent curve for antiviral activity in our cell culture infection assay. Mechanistic studies suggest that Oxy210 reduced replication of SARS-CoV-2 with disrupting the formation of double membrane vesicles (DMVs), intracellular membrane compartments associated with viral replication. Oxy210 also inhibited the replication of hepatitis C virus, another RNA virus whose replication is associated with DMVs, but not the replication of the DMV-independent hepatitis D virus.
 
The study warrants further evaluation of Oxy210 and Oxy232 as a safe and reliable oral medication, which could help protect vulnerable populations with increased risk developing COVID-19.
 
The study findings were published on a preprint server and are currently being peer reviewed. https://www.biorxiv.org/content/10.1101/2021.01.31.429001v1
 
With a bleak background of the ongoing COVID-19 pandemic caused by the SARS-CoV-2 coronavirus, effective antivirals are urgently required to rescue those severely ill following infection with the virus.
 
The study describes the antiviral activity of oxysterols, or oxidized cholesterol derivatives, against SARS-CoV-2. These occur in nature, and are not only involved in lipid signaling pathways that modulate a host of processes within the body, including the immune response and lipid homeostasis.
 
Many researchers have recently shown that these compounds have antiviral activity against a wide range of viruses, including (20(S)-hydroxycholesterol [20(S)-OH C] and 22(S)-hydroxycholesterol [22(S)-OHC]) that hepatitis B virus; herpes simplex virus; human papillomavirus; and rotavirus. https://pubmed.ncbi.nlm.nih.gov/24342612/
 
https://www.sciencedirect.com/science/article/pii/S2213231717301659
 
https://www.nature.com/articles/srep07487
 
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6409565/
 
https://pubmed.ncbi.nlm.nih.gov/30212801/
 
https://pubmed.ncbi.nlm.nih.gov/28314593/
 
The study team studied both endogenous and semi-synthetic compounds.
 
An initial research by the same team has shown that semi-synthetic oxysterols such as Oxy186, Oxy210 and Oxy133 can inhibit cancer cells, fibrosis and bone regrowth, respectively. https://pubmed.ncbi.nlm.nih.gov/31137846/
 
https://pubmed.ncbi.nlm.nih.gov/24591126/
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6829886/
 
This current study screens an array of oxysterols for their activity against SARS-CoV-2 in cell culture.
 
The study team found that natural oxysterols were able to produce a marked reduction in the spread of the virus among cells in culture. These compounds include natural oxysterols, namely, 7-ketocholesterol, 22(R)-hydroxycholesterol, 24(S)-hydroxycholesterol, and 27-hydroxycholesterol.
 
Importantly these compounds reduced viral RNA production by up to 86% relative to controls, but without observable cytotoxicity at doses up to 30 μM, the highest concentration. However, natural oxysterols are unstable during metabolism, and have beneficial as well as potentially harmful actions in the body.
 
For example, 25-OHC suppresses viral replication but also enhances immune cell activation as well as exaggerating the release of immune mediators that cause atherosclerosis. For this reason, semi-synthetic oxysterols were preferred.
 
It was also found that semi-synthetic oxysterols are not only more metabolically stable, but have more potent drug activity, are more selective in their activity, and are safer.
 
These semi-synthetic oxysterols with antiviral activity includes Oxy186 and oxysterol analogues Oxy210 and Oxy232. Oxy186 had very similar antiviral activity relative to that of the natural oxysterols, that is, an 83% reduction in viral RNA relative to controls.
 
Significantly, the analogues had even stronger anti-SARS-CoV-2 activity than the natural oxysterols, in fact, with an over 90% drop in viral replication at a concentration of 10 microM, and almost complete suppression at 15 microM. Both cytopathic effects and virus propagation were suppressed by Oxy210, but not by Oxy133.
 
However Oxy186 and Oxy210 were not associated with a reduction in cell viability, but there was a slightly cytotoxic effect with high concentrations of Oxy232.
 
The study team selected the compound Oxy210 for further studies because it was the most bioavailable, while having powerful inhibitory activity against SARS-CoV-2 replication.
 
It was seen in a mouse study that oral administration of Oxy210 achieved peak plasma concentrations of 19 microM, which is within the therapeutic range, as shown by the dose-response curve for antiviral activity in the cell culture inhibition assay.
 
Though Oxy210 inhibited Hedgehog (Hh) and transforming growth factor β signaling pathways within fibroblasts and cancer cells, the latter was not seen with Oxy232 despite its structural similarity to Oxy210. This indicates that the antiviral activity is not mediated by TGFβ inhibition.
 
It should be noted that this is further supported by the lack of efficacy of the TGFβ inhibitor SB431542, in viral RNA inhibition, or the Hh pathway inactivation by the compounds HPI-1 or GDC0449. This is another advantage of Oxy232, since unwanted off-target effects can be averted.
 
The drug Remdesivir which is a claimed ‘inhibitor of viral RNA replication’ within the cell, does not affect viral entry. This is mirrored by the activity of Oxy210, which reduces viral RNA over the whole lifecycle at any point following entry, but not at the time of viral entry. Thus, the target of Oxy210 is viral replication after viral entry.
 
But where remdesivir has to be given via intravenous infusion in a hospital, the oxysterols may be given orally, at the time when SARS-CoV-2 infection is confirmed. This would prevent disease progression as well as viral spread in infected individuals and their contacts.
 
Importantly oxysterols shift cell metabolism away from viral replication, thus reducing the chances of the emergence of drug-resistant mutants. And finally, oxysterols can be manufactured in a simple process compared to remdesivir.
Antiviral activity mediated by DMV inhibition
 
The actual mechanism of inhibition by Oxy210 is suggested to be via the inhibition of SARS-CoV-2 replication by preventing the normal formation of intracellular membrane compartments within which viral replication proceeds efficiently. These are called double-membrane vesicles (DMVs).
 
It was found that similar inhibitory activity was also seen with the RNA virus hepatitis C, which also replicates within DMVs. Expectedly, the hepatitis D virus, which does not depend on DMVs for replication, was not inhibited by this compound. Further work will be required to determine if Oxy210 has a direct effect on the formation of DMV.
 
The study team suggests that the powerful antiviral activities observed in Oxysterol derivatives merit further research in the area.
 
The study team commented, “Our study warrants further evaluation of Oxy210 and Oxy232 as a safe and reliable oral medication, which could help protect vulnerable populations with increased risk developing COVID-19. Semi-synthetic oxysterol derivatives, such as Oxy210 and Oxy232, could be promising leads in the search for COVID-19 drug candidates, used alone, or in combination with other therapies currently FDA approved or under investigation, such as RDV, convalescent plasma or antibody treatments.”
 
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