The COVID-19 pandemic is continuing to cause many serious and fatal cases of pneumonic illness, often ending in multi-organ dysfunction and cardiovascular collapse. Without either vaccine or proven therapy, clinical trials are ongoing to find an effective way to counter this threat. Both existing drugs and new drugs are being tested worldwide to identify potential antivirals with activity against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).
Novel Coronavirus SARS-CoV-2 Transmission electron micrograph of SARS-CoV-2 virus particles, isolated from a patient. Image captured and color-enhanced at the NIAID Integrated Research Facility (IRF) in Fort Detrick, Maryland. Credit: NIAID
The Polyether Ionophores
One compound that has aroused considerable interest is the polyether ionophore (PEI), a compound for which animal data is already available. This family of molecules contains natural products that have many different biological functions. The compounds in this family are best-known for their inhibitory activity against both gram-positive bacteria and coccidian protozoa. As a result, some of them are used as antibiotics in animals.
These compounds also have antiviral activity against both RNA and DNA viruses, including HIV, Zika, and influenza viruses. As far back as the 1970s, research showed that nine polyether ionophore compounds were able to inhibit transmissible gastroenteritis, a coronavirus infection of the porcine small intestine, and some were even found to have a curative effect.
A re-evaluation of this category in 2014 showed two PEIs, namely, salinomycin and monensin, were able to prevent the cytopathogenic effect of MERS-CoV, but could not inhibit SARS-CoV. These were not followed up, and the mechanism of action remains unknown. However, it is thought, on the basis of earlier research, that they block several steps in the replication cycle.
Mechanism of Action of PEIs
The current study published on the preprint server bioRxiv* aims to understand how these compounds affect SARS-CoV-2 in vitro. The researchers from Aarhus University in Denmark examined 11 natural PEIs, with a single synthetic analog, screening them for inhibitory activity against the CPE of SARS-CoV-2 infecting cultured cells with overexpression of TMPRSS2, a serine protease that is instrumental in cleaving the spike protein of the virus.
They found that all the eleven compounds inhibited the viral CPE but with varying selectivity, potency, and cell viability. The synthetic analog HL-201 was a good antibacterial candidate, but an unselective antiviral with low activity. The calcium ionophores ionomycin and calcimycin had modest selectivity, but 50-100-fold selectivity was found for nigericin, indanomycin, and lasalocid, with over hundred-fold selectivity displayed by narasin, salinomycin, monensin, and nanchangamycin,
X-206 – What’s Special?
The compound selected for this study was the ionophore antibiotic X-206, which was both strongly selective and potent, with almost 600 times the selectivity. This has several uncommon substructures, such as three lactol units, which can directly interact with metal ions in the solid-state. The molecule was already shown to inhibit plasmodium parasites.
The current study looked at its inhibitory activity against the replication of the SARS-CoV-2 virus. The endpoints were qRT-PCR and the viral S protein. It was shown to inhibit both viral copy number and S protein formation at the lowest tested concentration of 760 pM. Salinomycin was also tested and proved to be a potent viral inhibitor.
On the other hand, hydroxychloroquine (HCQ) showed little inhibitory activity on the virus in cultured cells expressing TMPRSS2 but effectively inhibited viral replication in wildtype cells. This difference was not seen with the use of X-206.
PEIs are known to build up in lysosomes, inhibiting autophagy, and their classic property of enabling the transfer of metal cations in return for protons should lead to altered lysosomal pH. This is similar to the mechanism of cationic amphiphiles like HCQ, which could indicate that related mechanisms are at work.
The researchers undertook morphological profiling to compare the compounds on cultured cells without any viral infection. They found that HCQ showed a different pattern of bioactivity from the PEIs, or at least a subset of them. The bioactivity was broadly corresponding to the concentrations associated with antiviral activity. The conclusion was that PEIs “mediate their antiviral effects through a mechanism that is different from that of the lysosomotropic, cationic amphiphiles” like HCQ.
Unexpectedly Potent Inhibition of SARS-CoV-2
No human safety data on PEIs are known, and some animals find them toxic, but they are used in the agricultural industry, which implies they are produced on an industrial scale, with safety. Thus, the current study looked at whether PEIs have broad-spectrum antiviral activity, and specifically against SARS-CoV-2.
The researchers found that the commonly used PEIs like salinomycin, monensin, and lasalocid are effective against a range of viruses. In addition, these are also effective against SARS-CoV-2. In particular, X-206 is strikingly powerful and selective as an antiviral with the above spectrum of action.
The study concludes, “Our future efforts will be focused on understanding the precise origin of the strong antiviral activity of X-206, which may also help to shed light on the possibilities for further pre-clinical development.”
bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.