Researchers investigating ways to develop a novel class of antiviral drugs to treat coronaviruses, including the SARS-CoV-2 virus that causes COVID-19, have developed a platform that can rapidly screen thousands of compounds to identify potential candidates. The team, led by researchers at the Johns Hopkins Bloomberg School of Public Health, used their new high-throughput platform to screen drug compounds that inhibit the macrodomain—a protein fold that plays a critical role in the coronavirus life cycle.
Prior studies suggest that some coronaviruses as well as alphaviruses largely lose their ability to replicate in cells and cause disease when their macrodomain's enzymatic activity is disrupted. In this study, researchers targeted the macrodomain of SARS-CoV-2 called Mac1.
This is thought to be the first high-throughput system that can screen for compounds that block macrodomain activity. In a proof-of-principle demonstration, they used the screening platform to identify existing drugs that block the coronavirus Mac1 activity. The researchers then showed that one of these drugs did not disrupt the enzymatic activity of the human macrodomain that is most similar to Mac1. Compounds that affect human macrodomains would be more likely to cause unwanted side effects in patients.
The new screening platform may enable the development of broad-spectrum drugs that treat existing coronaviruses, including SARS-CoV-2 and the virus that causes Middle East Respiratory Syndrome, and potentially new coronaviruses that could emerge from animal reservoirs such as bats.
"A viral-specific macrodomain inhibitor drug could be very useful in the treatment of COVID-19, MERS, and in a possible future pandemic caused by a novel coronavirus," says study senior author Anthony K. L. Leung, associate professor in the Bloomberg School's Department of Biochemistry and Molecular Biology. "While developing new drugs takes time, our versatile screening platform gives us hope that we can one day find one."
The researchers note that Mac1 is also a promising drug target because it is preserved in SARS-CoV-2 variants, including delta and omicron.
Mac1 is found in SARS-CoV-2 on a protein called nsp3 (non-structural protein 3) and has a type of enzyme activity called ADP-ribosylhydrolase activity.
In the study, Leung and colleagues developed a simple assay called ADPr-Glo that registers, with luminescence, the degree of enzymatic activity of a macrodomain. In the presence of compounds that inhibit the macrodomain, luminescence will be reduced. For this demonstration, they used the assay to rapidly screen two small libraries of existing drug compounds—a total of 3,233 compounds—for their ability to inhibit Mac1.
The researchers also measured those Mac1-inhibiting compounds' ability to inhibit human MacroD2, an ADP-ribosylhydrolase enzyme that is the closest human counterpart to Mac1. MacroD2 mutation or deletion is implicated in cancer formation and neurological disorders. Since compounds that affect MacroD2 could cause unwanted side effects in patients, the aim was to demonstrate that this new method could identify compounds that inhibit Mac1 without affecting MacroD2.
The experiments uncovered one existing drug, dasatinib, that does inhibit Mac1 with moderate potency, yet does not measurably inhibit the human MacroD2. Notably, it also inhibits Mac1 from another highly-pathogenic coronavirus that causes MERS. Dasatinib is a leukemia drug that was developed to target another class of enzymes and thus hits other targets in humans beyond Mac1. Dasatinib is known to be toxic to cells at the concentrations that inhibited Mac1. It would need modification to become an antiviral against coronavirus infections.
Identifying this drug demonstrates that viral Mac1's unique structure can be targeted by small-molecule drugs without affecting their closest human counterpart.
The ongoing pandemic has renewed urgency around identifying and developing antiviral treatments in the near term and for future pandemics. Earlier this year, the U.S. National Institute of Allergy and Infectious Diseases launched an Antiviral Program for Pandemics to promote the development of new drugs to combat viruses with pandemic potential.
The researchers plan to continue screening compounds to identify new Mac1 inhibitors.