Integrated in vitro and in silico strategies in the search for lead compounds against trypanosomatid-related diseases based on the selective inhibition of dihydroorotate dehydrogenases

Abstract

The Dihydroorotate dehydrogenase enzyme has been investigated as a therapeutic target for the development of drugs with trypanocidal activity. DHODHs from trypanosomatids are attractive targets since (i) they participate in an important biochemical pathway, (ii) there are important structural differences between human and trypanosomatid enzymes (iii) trypanosomatid DHODH enzymes have different validated pockets, to be exploited in drug design.Extensive studies on trypanosomatid DHODH enzymes (TrypDHODHs) have been performed in our laboratory. From the development of a reproducible protocol for expression and purification, kinetic characterization, crystallization, structure determination, identification of the first generation of ligands, validation of potential sites for ligand docking, and more recently the construction of different mutants, characterization of pre-steady state kinetic by stopped-flow and molecular dynamics simulations have been used to fully characterize TrypDHODHs. The next desirable step is to proceed with new enzyme inhibition studies, taking advantage not only of our deeper understanding of both catalytic mechanism and structural data, but also of in silico and in vitro methodologies available in the field of drug discovery, yet not implemented in our laboratory. With this in mind, we established collaboration with Prof. William Hunter, from the Division of Biological Chemistry and Drug Discovery (BCDD), College of Life Sciences - University of Dundee. BCDD is a multi-disciplinary and highly collaborative research division, encompassing both fundamental research and translational research. BCDD is composed by a large team working on the kinetoplastid parasites who invest efforts in the design of new potential therapeutic agents taking advantage of bioinformatics and chemoinformatics tools, a variety of biophysical methods and structural biology techniques. The main goal of this project is to offer the student training in the state-of-art in structure-based drug discovery, focusing on the integration of computational (molecular docking and chemical design based on structure activity relationships analyses) and experimental methods (biolayer interferometry technique, x-ray crystallography, enzymology). This approach will be applied in the search of selective ligands against the enzyme dihydroorotate dehydrogenase from Trypanosoma cruzi and Leishmania major, potential targets for the development of new therapies against Chagas' disease and Leishmaniasis, respectively. (AU)