Correlation between in vitro assays and in vivo assays: search for approaches in the study of snake venoms that dispense with the use of animals following the 3R's principle

• Grant number: 24/00382-5
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: May 01, 2024
• End date: July 31, 2024
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Anita Mitico Tanaka-Azevedo
• Grantee: Lidia Jorge Tasima
• Supervisor: Juan Calvete
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Institution abroad: Instituto de Biomedicina de Valencia (IBV), Spain
• Associated to the scholarship: 21/04287-9 - Correlation between in vitro assays and in vivo assays: search for approaches in the study of snake venoms that dispense with the use of animals following the 3R's principle, BP.DR

Abstract

The study of variation and quality control of snake venoms has been the subject of intense investigation, given their use in the production of antivenoms and in the search for new products of biotechnological interest. To this end, various biological activities of venoms are studied using both in vivo and in vitro methods. However, it is recognized that have some problems related of current in vivo tests: a large number of animals are required, time-consuming, expensive, no correlation with envenoming and therapy in human and the ethical point of view. Given the importance of these issues, the development of alternative methods for studies and quality control of venoms and antivenoms has a strong incentive and there is an effort by the scientific community to follow the concept of the 3 R's (Reduce, Refine and Replace) in relation to assays with animals. Although the use of animals to evaluate the effectiveness of antivenom serums is still essential, methodologies that can replace in vivo assays in research related to the effects of venom would not only reduce the use of animals, but also enable standardization and better comparison between results obtained in different research. With this in mind, the aim of this project is to search methodologies for analyzing the venom activities of different snakes in vitro that present results comparable to the in vivo tests ones in order to find alternatives to the use of animals in laboratory methodologies. For this, we used six venom pools from snakes of the genus Bothrops, Crotalus, Lachesis, B. erythromelas, B. leucurus and B. atrox. The results obtained so far have shown strong correlations between the lethal dose 50% results and Phospholipase A2 activity and caseinolytic activities. Strong correlations were found between myotoxic activity and LAAO, hyaluronidase, antibacterial, cytotoxic and coagulant activity. Edematogenic activity showed good correlations with LAAO activities and cytotoxicity in two lineages. When analyzing only with bothropic venoms, some correlations are added to the list of experiments with good correlation. Knowing the amount of each protein family present in the venom of each group of snakes can be a way of predicting the clinical manifestations present in the case of snakebite envenoming and also the intensity of these symptoms in a specific snake species. Therefore, the use of methodologies to analyze the composition of these venoms, such as proteomics, can present interesting results for the proposed project.