Inhibition of the polyprenol kinase enzyme of malaria parasites

• Grant number: 22/09526-4
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: June 01, 2023
• End date: May 31, 2026
• Field of knowledge: Biological Sciences - Biochemistry - Biochemistry of Microorganisms
• Principal Investigator: Alejandro Miguel Katzin
• Grantee: Ignasi Bofill Verdaguer
• Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Plasmodium falciparum is the etiologic agent of human falciform malaria, one of the most prevalent diseases in tropical and subtropical regions of the world. One of the largest problems for controlling the disease is the emergence of drug resistance, which leads to the need for discovering new antimalarial compounds and creating new effective pharmaceutical combinations. One of the most studied metabolic pathways for developing antimalarials is the methylerythritol 4-phosphate (MEP) pathway, which is absent in humans. The MEP pathway can be inhibited directly with fosmidomycin or indirectly with ribosomal inhibitors (e.g., clindamycin, chloramphenicol, etc.). However, both types of compounds show little effectiveness and high recrudescence rates in treating human malaria. Therefore, it is necessary to find a way to potentiate the effect of both types of drugs. The MEP pathway's products, isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), are condensed to form longer isoprenoids, such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP). These isoprenoids are the initial substrates for the biosynthesis of several metabolites, such as dolichols and ubiquinone, and for the isoprenylation of proteins, a post-translational modification that occurs mainly in proteins that regulate intracellular traffic, such as Ras and Rap proteins. Several studies suggest that the loss of the protein isoprenylation process is the proximal cause of death of parasites whose isoprenoid biosynthesis has been pharmacologically inhibited. Although the natural substrates of protein isoprenylation are FPP or GGPP, studies have already shown that isoprenoid alcohols, such as farnesol (FOH) and geranylgeraniol (GGOH) that are present in human food and blood plasma, can rescue parasites from the effect of MEP pathway inhibitors. These data suggest that the parasite has a salvage pathway of polyprenols via phosphorylation, whose pharmacological inhibition could enable the use of fosmidomycin and ribosomal inhibitors. Genes involved in this pathway have been identified only in plants, but biochemical evidence exists to be also active in archaea and animals in which the pathway would strongly relate to different types of cancer and dyslipidemias. In any case, previous evidence in the literature indicates that the polyprenol recycling pathway is universally carried out by two enzymes, a polyprenol kinase and a polyprenyl phosphate kinase. Previous studies led to identifying the gene encoding the P. falciparum polyprenol kinase. The catalytic activity was evaluated using [1-(n)-3H] GGOH and [1-(n)-3H] FOH. Furthermore, knockout parasites for this gene demonstrated to be still viable but more sensitive to MEP pathway inhibitors and were unable to metabolize polyprenols. In the present project, we also propose to create P. berghei (the etiologic agent of murine malaria) knockout parasites for the locus that encodes polyprenol kinase to assess whether the parasite may rely on exogenous polyprenols during the treatment of murine malaria with MEP pathway inhibitors. Finally, we also propose to look for inhibitors of this enzyme and test it in vitro and in vivo alone or with other drugs. We believe that the present project can lead to discovering synergistic molecules with MEP pathway inhibitors and elucidate one of the less studied metabolic pathways in biology. Furthermore, it could set an important precedent to be explored for treating other infectious diseases and various types of cancer.