Plasmodium spp. kinase inhibitors as potential antimalarial candidates: virtual screening and experimental validation on yeast target-based system

Abstract

Malaria is among the three infectious human diseases causing the greatest number of deaths worldwide, together with AIDS and tuberculosis. The use of antimalarials is essential for the containment of the disease and treatment of those infected. The frequent parasitic resistance to current antimalarials claims for the constant study and development of new chemical compounds capable of killing the parasite by new alternative modes of action. With the success of kinases inhibitors for cancer treatment in mind, the kinome of Plasmodium spp. started to be investigated, in order to find potential therapeutic targets for the treatment of malaria. Through gene ontology analysis, we chose the protein Casein Kinase 1 (CK1) from Plasmodium spp. as a target for the screen of chemical compounds for further parasite inhibition testing. CK1 is conserved among Plasmodium species, and highly expressed throughout the parasite's blood cycle. It has a physical interaction with the Rab-5B protein, thus having a metabolic control role on the parasite. In addition, reverse genetics studies have shown that this protein is essential for the development of the blood stage of the parasite. In the course of the studies, it was found that the casein kinase 2 alpha domain (CK2±) may also be a promising target. Studying these two complementary proteins in parallel could be advantageous. Although both kinases belong to the same family, serine-threonine kinases are divergent in structure, function and response to effector molecules. With the increasing computational capacity and improvement of in silico screening, we performed virtual screening with the purpose of reducing experimental tests and increasing assertive capacity, gaining time and reducing costs. We build the target protein structures by homology, after which we use two screening methods. The first was based on Structure-Based Virtual Screening (SBVS) and the second based on active molecules in Plasmodium spp., Quantitative Structure-Activity Relationship (QSAR). We also performed virtual screenings with phosphatidyl inositol-4-kinase (PI4K), another protein resulting from our ontogenetic screen, which is already described in the literature as a possible therapeutic target for malaria. We started the experimental tests of 205 compounds resulting from the virtual screening of the 3 target proteins of this project. As a result, we have been able (to date) to identify 23 compounds with e90% parasite growth inhibition rates. Aiming at understanding the mode of action of these compounds towards our predicted kinase targets, we intend through this project to construct tester yeast strains expressing our heterologous potential target P. vivax kinases CK1 and PI4K, as well as their human orthologs for cytotoxicity determination. We will also use yeast chemical genomics in yeast, to validate our screen predictions against our P. vivax CK1 and CK2 alpha kinases. (AU)