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Kidnapping the mailman

By interrupting cell communication, artificial RNA kills the parasite that causes schistosomiasis

RAFAELA OLIVEIRAHazards in water: larvae from parasites penetrate the body through the skinRAFAELA OLIVEIRA

In the manner of a game of spies whose objective is to intercept communication with the enemy and prevent messages from arriving at their destination, researchers from São Paulo blocked the cell mechanism responsible for the production of a protein that is essential for the survival of the Schistosoma mansoni parasite, which causes schistosomiasis. In one treatment session, they eliminated one fourth of the parasites that infested mice and thus may have stumbled upon a new way of dealing with one of the most serious known parasite-caused diseases, which affects 200 million people around the world and has become resistant to the drugs currently available.

The main difference between the schistosomiasis drugs and the experimental therapy developed at the State University of Campinas (Unicamp) and at Paulista State University (Unesp) is how the parasite is annihilated. Praziquantel and oxamniquine open up holes in their cells, whereas the treatment proposed by the team from São Paulo is similar to the intelligence service: just like a postman who is kidnapped and the letters he is carrying are torn up, the treatment captures and destroys the mechanism that guides the generation of the energy necessary for the survival of the Schistosoma and for the multiplication of its cells. Thus, the parasite is unable to replace the cells that deteriorated in the course of time and it dies.

In 2002, geneticist Iscia Lopes Cendes’s idea was to adopt this strategy to attack the Schistosoma, when she started working with a molecular biology technique developed by the Andrew Fire and Craig Mello, from the United States. By studying the Caenorhab­ditis elegans parasite, they found that it was possible to interfere with the cells’ chain of command and thus prevent the production of a specific protein by using laboratory produced RNA molecules.

In the cells of most living things, including parasites, the information on how to manufacture the protein is stored in a gene, a tiny segment of the DNA molecule, comprised of two nitrogenous-based parallel strands in the form of a spiral. Whenever the cell needs a protein, its formula is copied by a simpler molecule – the messenger RNA, comprised of a single line of nitrogenous bases and transported to the region where the proteins are manufactured.

Silent Genes
In 1998, Fire and Mello identified a way of preventing the messenger RNA from doing its work. They fed the parasites with artificial RNA molecules, which were comprised of two strands instead of one. When penetrating the cells, the double-strand RNA joins a protein complex and intercepts the messenger RNA. As a result, the protein complex is destroyed and the gene is silenced. The discovery of this phenomenon, which Craig and Mello named RNA interference, or RNAi, meant that they won the Nobel Prize for Medicine and Physiology in 2006.

“This worked with a free parasite, which absorbs RNA molecules through a resistant cuticle. So I thought that this would probably work with parasites with a thinner cuticle, that live in the organism of the hosts, like the Schistosoma,” says Iscia. Together with biologists Tiago Campos Pereira, Vinícius Bittencourt and Rafael Marchesini, she looked for a vital protein for the Schistosoma, but one whose messenger RNA would differ from that of mice. She identified hypoxantine-guanine-phosphorribosyl-transferase (HGPRTase), which is essential to split the cell and for the production of energy. A computer program developed by Pereira and Ivan de Godoy Maia, from Unesp at Botucatu, helped the team from Campinas design and produce specific double-strand RNA molecules to prevent the production of this protein. The next step was to test it on Schistosoma.

Iscia wanted to verify the effect of RNA interference on the schistosomiasis parasite in its natural environment, and on the blood vessels of the animals that host the parasite, instead of doing so under artificial lab-created conditions. As she herself does not work with animal testing, she contacted parasitology experts Eliana and Luiz Augusto Magalhães, who had developed a schistosomiasis model in mice years ago.

In this experiment, the researchers separated the Schistosoma-infected mice into four groups. The first group was given an injection in the vein that consisted of five micrograms of RNA molecules, designed to block the production of HGPRTase. Another group was injected with RNA molecules similar to those injected into the first group, but which had some slight modifications. The third group was treated with RNA unable to identify the formula of any of its genes, and the last group was injected with saline solution.

As expected, only the first treatment was successful against the Schistosoma: 27% of the adult parasites were killed. But it was necessary to discover whether the death of the parasites had really been caused by the RNA designed by Iscia’s team. Iscia applied the molecules that the team had produced on parasites kept under glass plates and observed a 60% reduction in the production of HGPRTase, as explained in the article published this month in Experimental Parasitology. “This might seem insignificant, but it really is not,” says the geneticist from Unicamp. “In this initial test, we used a lower dose, capable of having some effect on the parasite.” According to Iscia, it is possible to improve this performance with repeated applications or by increasing the dose up to ten times.

Although the results are promising, many more tests are still necessary, along with years of work, until it is finally proven that this is a feasible and safe strategy to treat schistosomiasis. “We didn’t identify any undesired effects in the mice,” says Iscia, “but nobody knows the long-term consequences of RNAi therapy.” So far, no results of clinical tests on humans have been reported.

In view of all these doubts, is it worthwhile to invest in this path” Iscia believes it is, because the therapeutic potential of this strategy goes beyond treatment of schistosomiasis. In principle, RNAi can be used to treat any illness caused by a hyperactive gene or by a defective gene. In addition, it is easier and faster to design RNA molecules to silence a gene and prevent the production of a protein instead of looking for them in nature or designing molecules that block the action of such a protein after it is ready. But can this therapy with RNA molecules cause resistance? – “It can”, says Iscia, “but in less than two days we are able to design and produce RNA molecules that prevent the production of another protein.”