Biological characterization of chemically diverse compounds targeting the Plasmodium falciparum coenzyme A synthesis pathway

Abstract

BackgroundIn the fight against malaria, the discovery of chemical compounds with a novel mode of action and/or chemistry distinct from currently used drugs is vital to counteract the parasite’s known ability to develop drug resistance. Another desirable aspect is efficacy against gametocytes, the sexual developmental stage of the parasite which enables the transmission through Anopheles vectors. Using a chemical rescue approach, we previously identified compounds targeting Plasmodium falciparum coenzyme A (CoA) synthesis or utilization, a promising target that has not yet been exploited in anti-malarial drug development.ResultsWe report on the outcomes of a series of biological tests that help to define the species- and stage-specificity, as well as the potential targets of these chemically diverse compounds. Compound activity against P. falciparum gametocytes was determined to assess stage-specificity and transmission-reducing potential. Against early stage gametocytes IC50 values ranging between 60 nM and 7.5 μM were obtained. With the exception of two compounds with sub-micromolar potencies across all intra-erythrocytic stages, activity against late stage gametocytes was lower. None of the compounds were specific pantothenate kinase inhibitors. Chemical rescue profiling with CoA pathway intermediates demonstrated that most compounds acted on either of the two final P. falciparum CoA synthesis enzymes, phosphopantetheine adenylyltransferase (PPAT) or dephospho CoA kinase (DPCK). The most active compound targeted either phosphopantothenoylcysteine synthetase (PPCS) or phosphopantothenoylcysteine decarboxylase (PPCDC). Species-specificity was evaluated against Trypanosoma cruzi and Trypanosoma brucei brucei. No specific activity against T. cruzi amastigotes was observed; however three compounds inhibited the viability of trypomastigotes with sub-micromolar potencies and were confirmed to act on T. b. brucei CoA synthesis. ConclusionsUtilizing the compounds we previously identified as effective against asexual P. falciparum, we demonstrate for the first time that gametocytes, like the asexual stages, depend on CoA, with two compounds exhibiting sub-micromolar potencies across asexual forms and all gametocytes stages tested. Furthermore, three compounds inhibited the viability of T. cruzi and T. b. brucei trypomastigotes with sub-micromolar potencies and were confirmed to act on T. b. brucei CoA synthesis, indicating that the CoA synthesis pathway might represent a valuable new drug target in these parasite species.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1860-3) contains supplementary material, which is available to authorized users.

Characterization of Plasmodium falciparum Pantothenate Kinase and Identification of Its Inhibitors From Natural Products

“…Many potential inhibitors that interfere with the CoA biosynthetic pathway in Plasmodium have been reported ( Fletcher and Avery, 2014 ; Fletcher et al., 2016 ; Weidner et al., 2017 ). Although pantothenate analogs such as pantothenol (PanOH), CJ-15,801 ( Saliba and Kirk, 2005 ; Saliba et al., 2005 ; Spry et al., 2005 ), N5-trz-C1-Pan, and N -PE-αMe-PanAm ( Macuamule et al., 2015 ; Howieson et al., 2016 ) were expected to target PanK, they appeared to inhibit other enzymes than PanK.…”

“…Coenzyme A (CoA) is an essential cofactor in many metabolic processes, involving more than 9% of approximately 3,500 cellular activities ( ). CoA biosynthesis has been attracting attention as a promising drug target ( Fletcher et al., 2016 ). In P. falciparum , CoA is generated by four enzymatic reactions initiated by the conversion of pantothenate (Vitamin B5) to 4-phosphopantothenate, catalyzed by pantothenate kinase ( Pf PanK) ( Saliba and Kirk, 2001 ).…”

“…It was previously shown that in P. falciparum blood stage parasites, either pantothenate or pantetheine must be incorporated from the host cytoplasm because the biosynthetic pathways for pantothenate and pantetheine are lacking (Saliba et al, 1998). Incorporation of pantetheine allows a bypass of pantothenate to 4 ' -p h o s p h o p a n t e t h e i n e , b y d i r e c t l y y i e l d i n g 4 'phosphopantetheine from pantetheine by PfPanK, as demonstrated by the growth rescue of both sexual and asexual stages by pantethine supplementation in the culture (Fletcher et al, 2016). 4'-Phosphopantetheine subsequently generates CoA by the last two enzymes, phosphopantetheine adenylyltransferase (PfPPAT) and dephospho-CoA kinase (PfDPCK).…”

Structure-activity analysis of CJ-15,801 analogues that interact with Plasmodium falciparum pantothenate kinase and inhibit parasite proliferation

“…As the CoA-dependent fatty acid synthesis pathway is required for survival of liver stage parasites [26,27], inhibitors of CoA biosynthesis (pending the ability to permeate liver cells) are anticipated to be active against liver stage parasites. Fletcher et al [28] have identified compounds structurally distinct from pantothenate analogues that inhibit growth of P. falciparum via an effect on CoA biosynthesis. Some of these molecules also inhibit early and late stage gametocytes, suggesting that other molecules that target CoA biosynthesis might also be effective against these forms of the parasite.…”

Antiplasmodial dihetarylthioethers target the coenzyme A synthesis pathway in Plasmodium falciparum erythrocytic stages

“…For further structure-guided optimization of this compound class, it will be necessary to identify which of the enzymes involved in CoA synthesis constitutes the eventual target. A recent study using the same rescue approach used here, but with additional CoA pathway intermediates found that addition of pantethine, dephospho CoA and CoA all rescued the activity of Amb180780 to more than 75%, pointing at the enzymes phosphopantothenoylcysteine synthetase or phosphopantothenoylcysteine decarboxylase as the putative targets for Amb180780 [ 29 ]. For similar studies aiming at corroborating the putative targets of the new compound class presented here, the readily soluble and potent inhibitor KuWei173 ( 8e ) is now available.…”