Maribor Faculty of Chemistry exploring substances targeting SARS-CoV-2

Researcher Anja Kolarič from the Faculty of Chemistry and Chemical Engineering, where study of substances with potential antiviral effects began when the pandemic started, stresses that the virus is very elusive and that it needs to be made sure that the drug does not have unpredictable effects on human cell.

She told the STA that the development of a vaccine is completely different from that of a drug, so it is difficult to compare the two. The problem that researchers face in developing antiviral agents that constitute a part of the drug is the biology of the virus itself.

Because the virus uses the human cell and its mechanisms to replicate, making sure that the antiviral agent acts only on the virus without side effects for humans is more difficult in drug development than in vaccine development.

A vaccine introduces a viral antigen into the body, leaving the immune system to mount its own defence against the antigen, which is safer than introducing an antiviral agent into the human body.

"On the one hand, it is better to prevent disease than to cure it, but on the other hand, drugs are needed to fight the disease when the virus mutates and evades vaccines. Vaccines are needed for prevention, while medicines are needed to alleviate the symptoms of the disease. Both are very important," Kolarič noted.

A highly effective drug to prevent severe Covid-19 symptoms would eliminate the need for lockdowns. To give doctors more treatment options and to combine more active ingredients in the same drug, researchers at the University of Maribor are developing new drugs, but they need to attack several segments of the virus's life cycle.

Humans as hosts

To replicate, SARS-CoV-2 must attach to the cell's angiotensin-converting enzyme receptor 2, or ACE2, using spike-like proteins called "spike proteins" that gave coronaviruses their name. According to Kolarič, the Maribor researchers want to prevent this process by focusing on two targets.

The first target is the furin protein, which allows the protein to bind to a receptor, and the second is the Neuropilin-1 receptor, which in some tissues facilitates the virus's entry into the cell by binding a spike protein. If researchers are able to find an active ingredient that stops the action of these two targets, they can prevent the virus from entering the cell.

At the same time, the Maribor researchers are working to find agents that would inhibit the action of the two proteases that produce non-structural proteins. These are responsible for the virus' replication by stopping the cell's defence mechanism, thereby delaying the immune response and preventing the translation of human hereditary material.

Compounds that would stop the action of the RNA-dependent RNA polymerase protein, which synthesises viral RNA, are also being investigated. This would prevent viral RNA amplification and the action of the non-structural protein 14 that verifies the correct synthesis of the new viral RNA molecule. This means that mutated versions of the viral RNA would be produced repeatedly, which would not allow the virus to continue to exist outside the cell.

Irreplaceable information technology

Another exceptionally important aspect when designing new antiviral agents is computational support. According to Kolarič, this process often starts with the use of information technology. It is essential to narrow down the broad range of compounds to those that will have the greatest potential for treatment.

Using computational methods, scientists predict which compounds are likely to work best and only then synthesize and experimentally evaluate them. Despite all the computational power, it is ultimately experimentation that produces definitive and reliable results.

Strict security regulations

The Maribor researchers have already identified compounds on certain targets that show the desired antiviral activity, but many other tests are still needed before a drug can reach the market. They are currently working with colleagues from abroad to establish the compounds' efficacy in viral cultures, but face difficulties due to safety restrictions.

Because SARS-CoV-2 is highly transmissible, there are strict rules for working with infected cells. As a result, only a limited number of laboratories are experimenting with the virus, which poses an additional challenge for the Maribor researchers, who have to find external partners to test the active substances on infected cells.

Long road to end result

It takes several years of research to get to a final drug that can be approved for medical use. The active ingredient must have high enough efficacy and low toxicity before preclinical testing can even take place. At this stage, further laboratory research involving animal testing is carried out. Only then do the four phases of human clinical testing and approval of the product follow.

Even more problematic are the financial resources, since the financial burden increases with each successive stage, making the involvement of large pharmaceutical companies essential.

Nevertheless, researchers at the University of Maribor continue to try to contribute to the understanding of the virus and the potential development of a new drug.

Existing treatment options

Two main types of drugs developed for other viral diseases are currently used to treat Covid-19. The first group of drugs acts indirectly on the virus by using monoclonal antibodies that help the immune system to respond quickly. Immunomodulatory drugs are also available to ease the cytokine storm common in Covid-19.

Other drugs have a direct effect on the virus and mostly include active substances that have been developed for the treatment of other viral diseases, but can also be used for SARS-CoV-2 due to similar mechanisms of action.

The European Medicines Agency (EMA) has approved two such drugs. Remdesivir, which was developed for hepatitis C, binds to the viral RNA molecule via RNA polymerase and prevents the virus from replicating further by mutating it. Molnupiravir, an active ingredient developed for influenza, works according to the same principle.

The third active substance, the first to be developed specifically for SARS-CoV-2, is nirmatrelvir, known commercially as Paxlovid. It acts as an inhibitor of the major protease responsible for the formation of non-structural proteins, without which the virus cannot replicate.

While these drugs are already on the market, there remains the problem of viral mutations. Each time a mutation occurs in a virus it allows the virus to evade the effect of a drug, and the process of adaptation can take years.

Nevertheless, researchers have found a way to push the virus into a corner. They can incorporate several different antiviral agents into the drugs, targeting several different targets in the virus's life cycle to ensure that, despite possible mutations and resistance to one agent, the virus is still destroyed by the others.

This has proven very effective in treating HIV infection, Kolarič points out. And that is why it is crucial that research continues.