Notícia

News Bulletin 247

Research points to new targets for creating male contraceptives (11 notícias)

Publicado em 12 de fevereiro de 2022

Researchers from the Universidade Estadual Paulista (Unesp) studied a protein existing in sperm and discovered two new targets that, combined, can be used for the development of male contraceptives. The study also demonstrates the feasibility of using mice as a model for in vivo testing. So far, scientists interested in this protein have been experimenting with primates, which makes the research more complex, time-consuming and expensive.

The focus of the project is EPPIN, an acronym for epididymal protease inhibitor, whose main function is to modulate sperm motility, that is, its ability to swim to the egg. Scientists and the pharmaceutical industry are looking to develop male contraceptives that act on sperm motility, as it is more difficult to come up with a drug capable of preventing the production of the male gamete.

“The degree of complexity of sperm production is greater than that of female egg production. The spermatogenesis process lasts about two months and occurs continuously”, explains Erick José Ramo da Silva, professor at the Department of Biophysics and Pharmacology at the Instituto de Biociências de Botucatu (IBB-Unesp) and one of the authors of the article Dissecting EPPIN protease inhibitor domains in sperm motility and fertilizing ability: repercussions for male contraceptive development, published in the journal Molecular Human Reproduction. “If a male contraceptive was produced that prevented sperm production, the drug would take three to four months to take effect from the moment a man started using it,” he adds.

authorization to swim

To understand how EPPIN works, it is first necessary to know some details about the human fertilization process. When the man ejaculates, the sperm is expelled from the epididymis, where it is stored, and travels through the urethra, being bathed in this way by various fluids coming from organs such as the seminal gland, the prostate and the epididymis.

In the case of mammals, especially primates, newly ejaculated semen has a gelatinous appearance and is quite viscous, as it is composed of several proteins that form the so-called semen clot. One of them is semenogelin, with which EPPIN interacts, blocking sperm locomotion. “Until ejaculation, the sperm does not swim, despite already having the machinery for it”, explains Ramo da Silva.

To continue on its way to the egg, the sperm must be released from the semen clot, which is gel-like because of the presence of proteins secreted by the seminal vesicles. In this step, the protease PSA, or prostate-specific antigen, comes into action, known as a marker for the diagnosis of prostate cancer. PSA cleaves several proteins that form this clot, liquefying the semen. Among the “broken” proteins is semenogelin.

“With the cleavage carried out by the PSA protease, the sperm can swim, which we call progressive motility, and penetrate the outermost layers of the egg, in a movement known as hyperactivated motility”, explains the researcher.

This process of protein cleavage by PSA occurs in the female reproductive tract, between five and ten minutes after ejaculation. “Until the moment when PSA acts, the only mechanism that is taking the sperm towards the egg is ejaculation. It doesn’t need motility before this stage and so it saves energy to go all the way to the uterus” , adds.

In previous research, monkeys were given recombinant human EPPIN vaccines. [produzidas em laboratório por microrganismos geneticamente modificados] and developed antibodies capable of binding to this protein, which caused infertility by blocking semenogelin breakdown processes and delaying semen liquefaction. This proved that EPPIN is related to the control of sperm motility.

As there are many differences between rodents and primates, the scientists’ initial option was to work with the animals of the second group. In the research conducted by Ramo da Silva’s team, the option was to use mice as an experimental model. That’s because they have a protein, SVS2, which plays the same role as semenogelin in humans: it joins EPPIN and blocks sperm movement.

promising results

In the research conducted at Unesp, the mice received three types of antibodies to verify if they bound to EPPIN and blocked sperm motility. By binding to the target molecule, the antibodies showed in which domains of the protein there should be an intervention to reduce or prevent sperm motility.

“As antibodies are made to act against a piece of the protein, they do not bind to other portions of EPPIN”, explains the scientist.

The antibodies that inhibited sperm motility bound to an initial region of the peptide chain called the “C-terminal Kunitz”, which was expected. But other antibodies that bound to the “N-terminal WFDC” domain also showed the ability to inhibit sperm motility – a novelty for scientists. Furthermore, both antibodies against the C-terminal and N-terminal regions promoted inhibition of the rate of in vitro fertilization, confirming that their binding to EPPIN affects the fertile potential of the sperm.

When verifying the motility of spermatozoa, its reduction was observed, which proves that both regions have protease inhibitors that regulate motility and that can be the target of new drugs. In other words, the research showed that it is possible to design molecules that bind to these two regions and not just to the C-terminus, preventing sperm motility.

The research also points out which sequences of the 133 amino acid chain that form EPPIN should be the target of anyone thinking of developing a male contraceptive that acts on sperm motility. Finally, it proves that it is possible to use mice as models for in vivo tests, which can make pre-clinical research simpler, faster and cheaper.

The article in Molecular Human Reproduction that describes these results has as its main authors Alan Andrew dos Santos Silva and Tamiris Rocha Fanti Raimundo, who developed the research during their master’s degree at the Department of Biophysics and Pharmacology at IBB-Unesp. The study was developed in partnership with the Department of Pharmacology and the Department of Biological Sciences at the Federal University of São Paulo (Unifesp), in addition to the Instituto de Biología y Medicina Experimental of the Consejo Nacional de Investigaciones Científicas y Técnicas, in Argentina.

The research was supported by FAPESP through three projects (15/08227-0, 17/11363-8 and 19/13661-1) and can be accessed in full at: https://doi.org/10.1093/molehr/ gaab066.

As next steps, Ramo da Silva will coordinate a team that will seek to design small sequences of amino acids capable of binding to EPPIN in a similar way to semenogelin and test them to see if they interrupt sperm motility. There is already a partnership with researchers from Portugal and the United Kingdom with this objective.

The idea is to test it in mice in Brazil and use human semen in the tests in Portugal, where the procedures to obtain this biological material are faster. “We now want to study the mechanism of action, how inhibition occurs and which steps take place to interrupt motility. With this, we can even identify other targets, other specific sperm proteins that are involved in this process”, he concludes.