Study conducted by researchers at the Federal University of São Paulo (Unifesp) and European collaborators reveal a possible new mechanism of action of the drug heparin in the treatment of Covid-19. In addition to combating coagulation disorders that can affect lung vessels and impair oxygenation, the drug also appears to be able to hinder the entry of the new coronavirus (SARS-CoV-2) into cells.

Samples of tests for coronavirus in a laboratory in Bern, Switzerland 22/04/2020 REUTERS / Arnd Wiegmann

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In laboratory tests, carried out on cell lines from the kidney of the African green monkey (Cercopithecus aethiops), heparin reduced the invasion of cells by the new coronavirus by 70%. The results of the study, supported by Fapesp (São Paulo State Research Support Foundation) under a project selected in the so-called Fapesp “Quick implementation supplements against covid-19”, were described in an article published on the bioRxiv platform, still in pre-print version (without peer review). The research was attended by scientists from England and Italy.

“There was evidence that heparin, which is a drug that performs several pharmacological functions, was also able to prevent viral infections, including by coronavirus, but the evidence was not very robust. We were able to prove this property of the drug in in vitro tests”, tells Agência Fapesp Helena Bonciani Nader, professor at Unifesp and coordinator of the project on the Brazilian side.

Nader’s group has studied glycosaminoglycans for more than 40 years – the class of complex carbohydrates to which heparin belongs – and developed the first low molecular weight heparins, used clinically as anticoagulant and antithrombotic agents, including in patients with covid-19.

One of the discoveries made by the group over this period was that heparin is a multi-target drug, because in addition to its effect in preventing blood clotting, it can bind to various proteins. Among them, growth factors and cytokines that bind to specific receptors on the surface of target cells.

In recent years, studies by other groups have suggested that the surface proteins of other coronaviruses previously reported could bind to a glycosaminoglycan in mammalian cells, called heparam sulfate, to infect them.

With the emergence of SARS-CoV-2, Unifesp researchers, in collaboration with English and Italian colleagues, had the idea of ​​evaluating whether the surface protein of the new coronavirus responsible for the infection of cells – called spike protein – binds to heparin, since the drug molecule has a structure very similar to that of heparan sulfate.

The experiments confirmed the hypothesis. Through techniques of surface plasmon resonance and circular dichroism spectroscopy, it was observed that heparin, when bound to SARS-CoV-2 spike proteins, causes a conformational change in these molecules. In this way, the “lock” for the virus to enter the cells will malfunction.

“If it does not enter the cell, the virus cannot multiply and is not successful in infection,” explains Nader.

Structural shape

The researchers also evaluated which structural forms of heparin have the best interaction and are able to change the conformation of the new coronavirus spike proteins, based on a library of derivatives and different fragments of the molecule, defined by size.

“The results of the analyzes indicated that the heparin that presents the best interaction and conformational alteration activity of the SARS-CoV-2 spike protein is with eight polysaccharides, that is, an octosaccharide”, says Nader.

Researchers are now making structural changes to heparins to identify a molecule that has the same binding effect and conformational change as the new coronavirus spike protein, but that causes less bleeding – a potential side effect of the drug.

In addition, they are also testing other compounds called mimetic heparins – which mimic the action of heparin.

“The idea is to arrive at a molecule with a better antiviral effect”, says Nader, who is also a member of Fapesp’s Superior Council.

According to the researcher, the ongoing studies will be carried out with structural biology technologies that involve techniques of nuclear magnetic resonance, of quick interaction kinetics by stop-flow, confocal microscopy and flow cytometry, among others, using different cell models.

The article Heparin inhibitis cellular invasion by SARS-CoV-2: structural dependence of the interaction of the surface protein (spike) S1 receptor binding domain with heparin (DOI: 10.1101 / 2020.04.28.066761), by Courtney J. Mycroft-West, Dunhao Su, Isabel Pagani, Timothy R. Rudd, Stefano Elli, Scott E. Guimond, Gavin Miller, Maria CZ Meneghetti, Helena B. Nader, Yong Li, Quentin M. Nunes, Patricia Procter, Nicasio Mancini, Massimo Clementi, Nicholas R. Forsyth, Jeremy E. Turnbull, Marco Guerrini, David G. Fernig, Elisa Vicenzi, Edwin A. Yates, Marcelo A. Lima and Mark A. Skidmore, can be read in the bioRxiv.

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