Researchers at the Butantan Institute have succeed in isolating and cloning the component responsible for the accidents of hemorrhaging syndrome which occurs in the south of the country due to contact with the hairy caterpillar Lonomia obliqua. “We are dealing with a protein which has received the name of lopap”, reveals the pharmaceutical researcher Ana Marisa Chudzinski-Tavassi, the coordinator of the group. Three scientific papers, signed by the pharmaceutical biochemist Cleyson Valença Reis and other members of the team, and published in the magazines Lancet and Thrombosis Research in 1999 and last June, crown the work of characterizing the protein which began in 1996 as Cleyson’s thesis for his master’s degree.
Once they had identified the protein, their attention was drawn to its possible use as the principal reagent of an anti-coagulant for the treatment of thrombosis. “With the extract from the bristles of the Lonomia, we managed to prepare a very small amount of lopap. However, we are developing a method of producing it on a large scale starting with genetically modified bacteria. All of the molecular biology needed for this was established throughout this year. Once a large quantity of material is available, it will be possible to think about the manufacture of the medicine”, says Ana Marisa.
Epidemic in the south
Since 1989, more than a thousand accidents caused by contact with the Lonomia obliqua have been registered in the south. Characterized by the syndrome of hemorrhaging, accompanied by the drastic lowering of some coagulation factors, the phenomenon has become alarming. Headaches, nausea, stinging dermatitis, bruising, hematomas, the bleeding of recent wounds, of gums and of the nose, are common symptoms. The most serious include blood in the urine and hemorrhaging in the stomach, glands, lungs and brain, which can lead to death. The epidemic made press headlines, but it is estimated that its dimension has been underestimated because, having occurred in rural zones, many cases were not reported to the medical authorities.
Known locally as oruga or ruga (wrinkly), the Lonomia obliqua is a greenish hairy caterpillar, with spots and stripes, and which often mimics the plants which it inhabits. Symmetrically displaced along its back are bristles in the form of branched and pointed thorns, akin to small trees. It is the contact with these bristles, where the lopap protein is located, which causes the accidents. They become more serious because it is difficult for the person to touch only one hairy caterpillar and not more within the colony. Therefore, the consequences depend on the quantity of caterpillars, the intensity of contact and the predisposition of the patient.
However, what has been known since the first steps into medicine, is that what can kill can also save. A thrombosis is the interruption of the flow of blood due to the presence of a coagulant. Since contact with this caterpillar brings on a hemorrhaging syndrome, a substance which impedes the formation of coagulants or even eliminates it, could become the main active ingredient of a medicine against thrombosis. This would be the case of the main reagent of the caterpillar’s venom the protein lopap (acronym: Lonomia oblique prothrombin activator protease).
The researchers obtained the lopap through the purification of the extract from the bristles of the caterpillar. They checked, in vivo, through intraviral microscopy analysis in rats, which it brought on hemorrhaging. “It activates a substance in the blood called prothrombin, producing thrombin, an enzyme that unchains the process of coagulation”, says Marisa. Here there is an apparent contradiction. Exactly by stimulating an exaggerated coagulation, does the lopap rob from circulation a large quantity of the substance which makes the blood coagulate, the fibrinogen.
There settles in the so-called consumption consumption coagulopathy , a reaction of the organism that deprives the blood of that which makes it coagulate. Cleyson Reis, the person mainly responsible for the characterization of lopap, describes the process:
“The lopap activates the prothrombin, forming thrombin. The thrombin requests and breaks down the molecules of fibrinogen, producing fibrin, the raw material of the coagulants. This brings about a dispersed intravascular coagulation (which leads to a thrombosis). The organism reacts against the presence of the coagulants, promoting a secondary fibrinolysis (the destruction of fibrin). Robbed of the components that would allow it to coagulate, the blood can no longer coagulate. There follows from this point, hemorrhaging which, in the accidents with the caterpillars, can even lead to the death of the patient through renal and cerebral complications.” However, with rigorous control of the dosage, the lopap can function against thrombosis, without provoking hemorrhaging reactions.
The group calls attention to the fact that in the accidents with the Lonomia obliqua, the destruction of fibrin is a secondary phenomena, brought about by the reaction of the organism and not by the poison. It is important to differentiate between the cases registered hereand those that happened in Venezuela. There, the poisoning is caused by another species, the Lonomia achelous, in whose venom Venezuelan researchers have affirmed that they have found components that destroy fibrin.
Nevertheless, at the beginning of the 90s, and based on the Venezuelan model, the recommendation was to treat the individuals who were victims in Brazil withanti-fibrinolytes. However, these medicines impede the destruction of fibrin (fibrinolysis) and the researcher energetically contests this directive.”To use anti fibrinolytes is to kill the patient. They stop secondary fibrinolysis from occurring. The coagulants accumulate and the person ends up dying from disseminated intravascular coagulation.” She spoke with the authority of one who had carried out a previous project through FAPESP exactly to study poisoning in patients (Determination of the Parameters of Coagulation and Fibrinolysis in the Plasma of Patients in Contact with Lonomia obliqua).
Furthermore, she adds poisonings in Brazil have been perfectly well treated using a serum that the Butantan Institute has made, even before the group’s research, which now wants to take a further step forward. The current serum is manufactured beginning with the total poison, and the team now wants to produce a new serum based exclusively on lopap.
This serum is already being tested on animals and could be a “cleaner” alternative, since it responds to the specific protein that causes the problem and not to the complex package of proteins of the poison. “After inoculating with the specific serum, we want to challenge the animals with the total poison, as a way of proving that the lopap is really the most important factor”, says Ana Marisa.
The more ambitious goal, nonetheless, continues to be that starting from lopap, to create a disfibrinogent medicine to combat thrombosis. Ana Marisa knows that there is a long way to go between the test of an active source in animals, current phase, and the manufacturing of a medicine for human consumption on a commercial scale, but she is enthusiastic.
1. Determination of the Parameters of Coagulation and Fibrinolysis in the Plasma of Patients in Contact with the Hairy Caterpillar Lonomia obliqua (nº 99/01371-3); Modality Regular line of research assistance; Coordinator Dr. Ana Marisa Chudzinski-Tavassi -Butantan Institute; Investment R$ 125.247,00
2. Proteins of Poisonous Animals that Interfere in the Coagulation (nº 00/11735-1); Modality Regular line of research assistance; Coordinator Dr. Ana Marisa Chudzinski-Tavassi – Butantan Institute; Investment R$ 17.113,00
3. Fibrinolysis Mechanisms: Analysis of Interactions with Cell Receptors (nº 00/14781-4); Modality Regular line of research assistance; Coordinator Dr. Ana Marisa Chudzinski-Tavassi -Butantan Institute; Investment R$ 41.003,90