Identification of New Inhibitors of Plasmodium falciparum Enoyl- ACP Reductase Symposium on: Advances in Parasitology “Education and Research in Parasitology in the service of Mankind “

Malaria, what is it? A parasitic infection caused by the Plasmodium parasite and transmitted by female Anopheles mosquitoes.

Why do we need to focus on malaria? Infects million people every year! Kills one million peoples annually!!! Kills one child every 30 seconds!!!!

Gambian child with severe malaria anemia

DrugIntroduction In vivo resistance confirmed Origin Chloroquine 1945 Early 1960s Southeast Asia, South America Sulfadoxine and pyrimethamine(Fansidar) 1967 Late 1960s Southeast Asia, South America Mefloquine (Lariam) 1985 Early 1990sSoutheast Asia Atovaquone and proguanil (Malarone) Africa Artemether and lumefantrine (Coartem) Southeast Asia The Challenge of Resistance!

How could the problem of resistance be tackled Three approaches are under investigation:  1/ The novel use of older drugs ( use in combination)  2/ The re-design of existing drugs ( develop analogues, discovery of natural products)  3/ Validation of novel parasite- specific drug targets ( benefit from the better understanding of the parasite biology and genomics)

 e.g. targeting a parasitic enzyme, which is not present in mammals, or which has significant structural differences from the corresponding enzyme in mammals

 Example: The chosen target, may over time, lose its sensitivity to the drug  Example: The penicillin-binding-protein (PBP) known to be the primary target of penicillin in the bacterial species Staphylococcus aureus has evolved a mutant form that no longer recognizes penicillin.

Compound library (natural, biotransformed, synthetic ) Medium-throughput screening (MTS) PfENR inhibition assay Active compounds ( ≥ 50% PfENR inhibition) IC50 determinationIn vitro Kinetics studies Molecular docking modeling Cytotoxicity evaluation

PfENR Inhibition assay was established, standardized and optimized for screening

Classification of PfENR inhibitors PfENR inhibitors Source NaturalSynthetic Chemical class Flavonoid s Phenolics Aromatic acids Steroids Coumarin derivatives Miscellane ous

Compound codeConcentration (mM)% inhibition IC 50 (µM) ± SEM ± ± ± ± ± ± ± ± ± ± ± ± ± ± 3.2 % of inhibition and IC 50 values of PfENR inhibitors from natural sources

Gossypol (compound 1) 27 28

Kinetic studies on gossypol (compound 1): IC 50 = 2.0 µM K i = 4.0 Mechanism of Inhibition: competitive with regard to the substrate, crotonyl CoA

Kinetic Behavior of Gossypol

Schematic representation of the binding pocket of Gossypol on the surface of PfENR.

Binding models of PfENR in complex with compound 4 Score value = 10.34

Representation of compound 21 docked to the active site of PfENR. Score value =7.20/ IC 50 = 100 µM

CONCLUSION AND FUTURE DIRECTIONS  The current study has resulted in the identification of new PfENR inhibitors with chemical diversity and broad range of potency  We have identified here for the first time the well know natural compound ‘Gossypol’ as a potent PfENR inhibitor with IC 50 and Ki values of 2.0 µM and 4 respectively  Gossypol was know for its antimalarial activity but through the inhibition of Plasmodium falciparum Lactate Dehydrogenase (PfLDH). If we consider its newly identified PfENR inhibitory activity, gossypol could be described as dual-target inhibitor, a critical feature for a hit to be developed into potential antimalarial lead  The kinetic behaviour and the analysis of the inhibitors-PfENR complexes would pave the way for the development of these inhibitors into antiplasmodial agents

Sudan's diverse and virgin flora Rich traditional medicine knowledge Well-equiped drug discovery facility

Enzymology and enzyme inhibition Rsearch Antioxidant discovery research Structural computational chemistry research.

Medical Biochemistry Research Lab. (

ACKNOWLEDGEMENTS Many people have contributed to this work Prof Asaad Khalid Members of ICCBS Prof Mohammed Galal Prof Hassan Alsubki Members of lab # 404, HEJ Prof Mohamed Iqbal Members of MAPRI Dr. Mohamed A. Mesaik Financial fund of the IDB Dr. Omer M. Abdellah Financial fund of the Ministry Dr. Talal A.Awad of Higher Education Mr. Elteab Fadul Omdurman Islamic University Miss Fatima Elfatih Miss Ghada

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