Enhancing the fight against malaria : from genome to structure and activity of a G-protein coupled receptor from the mosquito, Anopheles Gambiae

Includes abstract.Includes bibliographical references (leaves 183-184).G-proton coupled receptors (GPCRs) are excellent drug targets that occupy a central position in the physiology of insects and are involved in transmission of signal from the extracellular to the intracellular side of the cell. Adipokinetic hormone receptors (AKHRs) are GPCRs that mediate physiological functions of the neurohormones, adipokinetic hormones (AKHs) that regulate mobilisation of energy reserves during mosquito flight. Ligand binding to GPCRs depends on the three dimensional (3D) structures of the receptors but to date no crystal structures of insect GPCRs are available. This work focused on building molecular models of AKHR from the genome of the malaria mosquito, identifying its binding site and studying the conformational and structural changes during molecular dynamics of the active and inactive receptor

Computational de-orphaning of Mycobacterium tuberculosis targets

Tuberculosis (TB) continues to be a major health hazard worldwide due to the resurgence of drug discovery strains of Mycobacterium tuberculosis (Mtb) and co-infection. For decades drug discovery has concentrated on identifying ligands for ~10 Mtb targets, hence most of the identified essential proteins are not utilised in TB chemotherapy. Here computational techniques were used to identify ligands for the orphan Mtb proteins. These range from ligand-based and structure-based virtual screening modelling the proteome of the bacterium. Identification of ligands for most of the Mtb Proteins will provide novel TB drugs and targets and hence address drug resistance, toxicity and the duration of TB treatment

Computational Deorphaning of <em>Mycobacterium tuberculosis</em> Targets

Tuberculosis (TB) continues to be a major health hazard worldwide due to the resurgence of drug discovery strains of Mycobacterium tuberculosis (Mtb) and co-infection. For decades drug discovery has concentrated on identifying ligands for ~10 Mtb targets, hence most of the identified essential proteins are not utilised in TB chemotherapy. Here computational techniques were used to identify ligands for the orphan Mtb proteins. These range from ligand-based and structure-based virtual screening modelling the proteome of the bacterium. Identification of ligands for most of the Mtb proteins will provide novel TB drugs and targets and hence address drug resistance, toxicity and the duration of TB treatment

Novel inhibitors of Mycobacterium tuberculosis GuaB2 identified by a target based high-throughput phenotypic screen

High-throughput phenotypic screens have re-emerged as screening tools in antibiotic discovery. The advent of such technologies has rapidly accelerated the identification of ‘hit’ compounds. A pre-requisite to medicinal chemistry optimisation programmes required to improve the drug-like properties of a ‘hit’ molecule is identification of its mode of action. Herein, we have combined phenotypic screening with a biased target-specific screen. The inosine monophosphate dehydrogenase (IMPDH) protein GuaB2 has been identified as a drugable target in Mycobacterium tuberculosis, however previously identified compounds lack the desired characteristics necessary for further development into lead-like molecules. This study has identified 7 new chemical series from a high-throughput resistance-based phenotypic screen using Mycobacterium bovis BCG over-expressing GuaB2. Hit compounds were identified in a single shot high-throughput screen, validated by dose response and subjected to further biochemical analysis. The compounds were also assessed using molecular docking experiments, providing a platform for their further optimisation using medicinal chemistry. This work demonstrates the versatility and potential of GuaB2 as an anti-tubercular drug target

Mycobacterial dihydrofolate reductase inhibitors identified using chemogenomic methods and in vitro validation.

The lack of success in target-based screening approaches to the discovery of antibacterial agents has led to reemergence of phenotypic screening as a successful approach of identifying bioactive, antibacterial compounds. A challenge though with this route is then to identify the molecular target(s) and mechanism of action of the hits. This target identification, or deorphanization step, is often essential in further optimization and validation studies. Direct experimental identification of the molecular target of a screening hit is often complex, precisely because the properties and specificity of the hit are not yet optimized against that target, and so many false positives are often obtained. An alternative is to use computational, predictive, approaches to hypothesize a mechanism of action, which can then be validated in a more directed and efficient manner. Specifically here we present experimental validation of an in silico prediction from a large-scale screen performed against Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis. The two potent anti-tubercular compounds studied in this case, belonging to the tetrahydro-1,3,5-triazin-2-amine (THT) family, were predicted and confirmed to be an inhibitor of dihydrofolate reductase (DHFR), a known essential Mtb gene, and already clinically validated as a drug target. Given the large number of similar screening data sets shared amongst the community, this in vitro validation of these target predictions gives weight to computational approaches to establish the mechanism of action (MoA) of novel screening hit

Controlling drug resistance by targeting Plasmodium falciparum heat shock protein 70-1, a chaperone at the centre of protein quality control mechanism: a review

The survival of the malaria parasite is highly dependent on withstanding physiological stresses, which are a proportional response to parasite invasion within the hostile human host environment. Plasmodium falciparum heat shock protein 70-1 (PfHSP70-1) plays a significant role through its network of interactions with the substrate scanners PfHSP40s, the functional maturation protein PfHSP90, the nucleotide exchange factor PfHSP110c/PfHSP70-z, apoptosis and parasite homeostasis agent PfHSP60, also the Clp machinery for the unfolding and degrading of misfolded proteins PfHSP100. These proteins work together to maintain the health and function of the parasite, but also possess individual functionalities. Here, we review the functional interplay between these heat shock proteins (HSPs), highlighting the central role of PfHSP70-1, its prospects in antimalarial drug discovery and possible implications in drug resistance

Natural products : a potential source of malaria transmission blocking drugs?

The ability to block human-to-mosquito and mosquito-to-human transmission of Plasmodium parasites is fundamental to accomplish the ambitious goal of malaria elimination. The WHO currently recommends only primaquine as a transmission-blocking drug but its use is severely restricted by toxicity in some populations. New, safe and clinically effective transmission-blocking drugs therefore need to be discovered. While natural products have been extensively investigated for the development of chemotherapeutic antimalarial agents, their potential use as transmission-blocking drugs is comparatively poorly explored. Here, we provide a comprehensive summary of the activities of natural products (and their derivatives) of plant and microbial origins against sexual stages of Plasmodium parasites and the Anopheles mosquito vector. We identify the prevailing challenges and opportunities and suggest how these can be mitigated and/or exploited in an endeavor to expedite transmission-blocking drug discovery efforts from natural products.The South African Research Chairs Initiative of the Department of Science and Technology, administered through the South African National Research Foundation and the NRF.http://www.mdpi.com/journal/pharmaceuticalsam2021BiochemistryChemistryGeneticsMicrobiology and Plant PathologyParaclinical Science

In silico Comparison of Antimycobacterial Natural Products with Known Antituberculosis Drugs

The chemical space based on physicochemical properties and structural features of a diverse group of natural products with reported in vitro activity against different Mycobacterium tuberculosis strains is investigated using in silico tools. This is compared to the chemical space occupied by drugs currently recommended for the treatment of various forms of tuberculosis as well as compounds in preclinical and clinical development. Docking studies exploring possible binding affinities and modes of two main clusters of natural products on two different mycobacterial targets are also reported. Our docking results suggest that scytoscalarol, an antibacterial and antifungal guanidine-bearing sesterterpene, can inhibit arabinosyltransferase Mtb EmbC, and the β-carboline alkaloids 8-hydroxymanzamine A and manzamine A can bind to the oxidoreductase of Mtb INHA. On this basis, these products showing high binding affinities to the two targets in silico could be rationally selected for in vitro testing against these targets and/or semisynthetic modification

Tetrazole-based deoxyamodiaquines: synthesis, ADME/PK profiling and pharmacological evaluation as potential antimalarial and antituberculosis agents

A series of new deoxyamodiaquine-based compounds was synthesized via the modified TMSN3-Ugi multi-component reaction and evaluated in vitro for antiplasmodial and antimycobacterial activity. The most potent compounds, 6b, 6c and 6j, showed IC50 values in the range of 6 to 77 nM against chloroquine-resistant K1- and W2-strain P. falciparum and MABA MIC90 values in the range of 7.6 to 59.3 µM against M. tuberculosis H37Rv. In vitro ADME characterization of compounds 6b and 6c indicates that these two compounds are rapidly metabolized and have a high clearance rate in human and rat liver microsomes. This result correlated well with an in vivo pharmacokinetics study, which showed low bioavailability of 6c in rats. Tentative metabolite identification was determined by LC-MS and suggested metabolic lability of groups attached to the tertiary nitrogen. Preliminary studies on 6b and 6c suggested strong inhibitory activity against the major CYP450 enzymes. In silico docking studies were used to rationalize strong inhibition of CYP3A4 by 6c