Sabine Fletcher

Thiaplakortones A–D: Antimalarial Thiazine Alkaloids from the Australian Marine Sponge Plakortis lita

A high-throughput screening campaign using a prefractionated natural product library and an in vitro antimalarial assay identified active fractions derived from the Australian marine sponge Plakortis lita . Bioassay-guided fractionation of the CH2Cl2/CH3OH extract from P. lita resulted in the purification of four novel thiazine-derived alkaloids, thiaplakortones A-D (1-4). The chemical structures of 1-4 were determined following analysis of 1D/2D NMR and MS data. Comparison of the chiro-optical data for 3 and 4 with literature values of related N-methyltryptophan natural products was used to determine the absolute configuration for both thiaplakortones C and D as 11S. Compounds 1-4 displayed significant growth inhibition against chloroquine-sensitive (3D7) and chloroquine-resistant (Dd2) Plasmodium falciparum (IC50 values <651 nM) and only moderate cytotoxicity against HEK293 cells (IC50 values >3.9 μM). Thiaplakortone A (1) was the most active natural product, with IC50 values of 51 and 6.6 nM against 3D7 and Dd2 lines, respectively.

Two-pronged attack: dual inhibition of Plasmodium falciparum M1 and M17 metalloaminopeptidases by a novel series of hydroxamic acid-based inhibitors

Plasmodium parasites, the causative agents of malaria, have developed resistance to most of our current antimalarial therapies, including artemisinin combination therapies which are widely described as our last line of defense. Antimalarial agents with a novel mode of action are urgently required. Two Plasmodium falciparum aminopeptidases, PfA-M1 and PfA-M17, play crucial roles in the erythrocytic stage of infection and have been validated as potential antimalarial targets. Using compound-bound crystal structures of both enzymes, we have used a structure-guided approach to develop a novel series of inhibitors capable of potent inhibition of both PfA-M1 and PfA-M17 activity and parasite growth in culture. Herein we describe the design, synthesis, and evaluation of a series of hydroxamic acid-based inhibitors and demonstrate the compounds to be exciting new leads for the development of novel antimalarial therapeutics.

A novel approach for the discovery of chemically diverse anti-malarial compounds targeting the Plasmodium falciparum Coenzyme A synthesis pathway

BackgroundMalaria is a devastating parasitic disease, causing more than 600,000 deaths annually. Drug resistance has rendered previous generation anti-malarials ineffective and is also rapidly emerging against the current therapeutics of choice, artemisinin and its derivatives, making the discovery of new anti-malarials with novel mechanisms of action a priority. The Coenzyme A (CoA) synthesis pathway, a well-known anti-microbial drug target that is also essential for the malaria parasite Plasmodium falciparum, has not yet been exploited in anti-malarial drug development. A novel high throughput approach for the identification of chemically diverse inhibitors of the CoA synthesis pathway is reported.MethodsTo identify novel CoA synthesis pathway inhibitors, a chemical rescue screening approach was developed. In short, a test compound was considered likely to inhibit the P. falciparum CoA synthesis pathway, if addition of the end product of the pathway, CoA, was able to negate the growth-inhibitory action of the compound on P. falciparum parasites.ResultsThe chemical rescue approach was employed to screen the Medicines for Malaria Venture malaria box and a small focussed compound library. This resulted in the identification of 12 chemically diverse potential inhibitors of the CoA pathway. To ascertain accurate potency and selectivity, the half-maximal inhibitory concentration (IC50 value) of these compounds was determined for both P. falciparum and a human cell line. Seven compounds showed submicromolar activity against the parasite, with selectivity indices ranging between six and greater than 300. CoA supplementation was confirmed to alleviate the effects on parasite growth and cell viability in a dose dependent manner. Microscopic investigation into the stage of effect and phenotype of treated parasites was performed on a selection of the active compounds.ConclusionsThe chemical rescue approach described resulted in the identification of a set of chemically diverse CoA synthesis pathway inhibitors with IC50 values ranging between 120 nM and 6 μM. The identified compounds will be utilized as tools for further investigating the parasite CoA synthesis pathway to define their exact mechanism of action. Furthermore, the chemical diversity of the compounds identified substantiates the suitability of this approach to identify novel starting points for future anti-malarial drug development.

Total synthesis and antiplasmodial activity of pohlianin C and analogues.

The first synthesis of the glycine-rich cyclic octapeptide pohlianin C is reported, confirming the structure of this natural product. Screening against Plasmodium falciparum reveals moderate antiplasmodial activity, consistent with data obtained from the natural sample. In addition, the synthesis of three analogues reveals that the antiplasmodial activity of pohlianin C can be preserved or increased with simplified structures.

Discovery of chemically diverse compounds targeting the Plasmodium falciparum coenzyme A pathway

Background Drug resistance has rendered previous generation antimalarials ineffective and is also rapidly emerging against the current therapeutics of choice, artemisinin and its derivatives, making the discovery of new anti-malarials with novel mechanisms of action a priority. The Coenzyme A (CoA) synthesis pathway is a well-known anti-microbial drug target that is also essential for the malaria parasite Plasmodium falciparum. To date, this pathway has not been exploited in anti-malarial drug development. Herein a novel high throughput approach for the identification of chemically diverse inhibitors of the CoA synthesis pathway from chemical compound libraries is reported. Materials and methods To identify novel CoA synthesis pathway inhibitors, a chemical rescue screening approach was developed by modifying a well-established P. falciparum imaging assay. In short, a test compound was considered likely to inhibit the P. falciparum CoA synthesis pathway if addition of the pathway’ se nd product, CoA, was able to negate the compound’s growth-inhibitory action on P. falciparum asexual parasites. Results The chemical rescue approach was employed to screen the Medicines for Malaria Venture (MMV) malaria box and another small focussed compound library. This resulted in the identification of 12 chemically diverse inhibitors. To ascertain accurate potency and selectivity, the half maximal inhibitory concentration (IC50 )o f these compounds was determined for both P. falciparum and human embryonic kidney cells. Seven compounds showed sub-micromolar activity against the parasite, with selectivity indices ranging between 6 and over 300 times. CoA supplementation was confirmed to alleviate the effects on parasite growth and cell viability in a dose dependant manner. Microscopic investigation of the stage of effect and phenotype of treated parasites was performed on a selection of the active compounds. Conclusions