Host-Derived Geranylgeraniol Shields Intraerythrocytic Stages of Malaria Parasites from Fosmidomycin

Fosmidomycin was proposed as an antimalarial drug but failed in clinical trials due to recrudescence, a phenomenon whose causes remain poorly understood. The mechanism of action of fosmidomycin is the inhibition of the methylerythritol 4-phosphate (MEP) pathway, essential for producing isoprenoids in Plasmodium parasites. The key isoprenoids produced by the MEP pathway are farnesyl and geranylgeranyl pyrophosphates (FPP and GGPP), vital for protein isoprenylation, ubiquinone, and dolichol biosynthesis. In vitro studies have demonstrated that prenols, like farnesol (FOH) and geranylgeraniol (GGOH), can temporarily circumvent the MEP pathway, rescuing parasites from fosmidomycin effects. Our group identified a parasitic prenol kinase (PolK), responsible for converting FOH and GGOH into their active pyrophosphate forms. Additionally, GGOH’s human plasma concentration is sufficient to affect MEP inhibitors. This suggests that the parasite's uptake of host prenols could diminish fosmidomycin effectiveness against malaria. To test this hypothesis, we generated Pb PolK knockout P. berghei parasites (Δ Pb PolK). These transgenic parasites were viable but could not utilize exogenous prenols for protein prenylation and caused a form of murine malaria that responded more effectively to fosmidomycin therapy compared to parasites preserving PbPolK. Consequently, we explored compounds that could inhibit the parasite utilization of exogenous prenols, using biochemical and bioinformatics approaches, as well as in vitro assays in wild type and ΔPfPolK P. falciparum strains. Geraniol inhibits PolK activity and the incorporation of GGOH into P. falciparum. Moreover, geraniol enhanced fosmidomycin antimalarial effect in P. falciparum in vitro , even in the presence of GGOH. Δ Pf PolK P. falciparum strains exhibited profound metabolic dysregulation in carbon metabolism, as assessed by proteomics. Taking all together, findings here presented demonstrate that the prenol salvage pathway is a modulatory mechanism of metabolic homeostasis, facilitates prenol utilization from the host, and contributes to the limited efficacy of fosmidomycin in malaria treatment.