Explore the vast range of titles available.

The proteasome of the malaria parasite Plasmodium falciparum (Pf20S) is an advantageous drug target because its inhibition kills P. falciparum in multiple stages of its life cycle and synergizes with artemisinins. We recently developed a macrocyclic peptide, TDI-8304, that is highly selective for Pf20S over human proteasomes and is potent in vitro and in vivo against P. falciparum . A mutation in the Pf20S β6 subunit, A117D, confers resistance to TDI-8304, yet enhances both enzyme inhibition and anti-parasite activity of a tripeptide vinyl sulfone β2 inhibitor, WLW-vs. Here we present the high-resolution cryo-EM structures of Pf20S with TDI-8304, of human constitutive proteasome with TDI-8304, and of Pf20Sβ6 A117D with WLW-vs that give insights into the species selectivity of TDI-8304, resistance to it, and the collateral sensitivity associated with resistance, including that TDI-8304 binds β2 and β5 in wild type Pf20S as well as WLW-vs binds β2 and β5 in Pf20Sβ6 A117D . We further show that TDI-8304 kills P. falciparum as quickly as chloroquine and artemisinin and is active against P. cynomolgi at the liver stage. This increases interest in using these structures to facilitate the development of Pf20S inhibitors that target multiple proteasome subunits and limit the emergence of resistance. The proteasome of Plasmodium falciparum (Pf20S) is a promising multi-stage drug target. Using CryoEM Hsu et al. report the structures of parasite and human proteasome in complex with the recently developed inhibitor TDI-8304 to gain insights into its species selectivity.

Malaria is a devastating infection caused by protozoa of the genus Plasmodium . Drug resistance is widespread, no new chemical class of antimalarials has been introduced into clinical practice since 1996 and there is a recent rise of parasite strains with reduced sensitivity to the newest drugs. We screened nearly 2 million compounds in GlaxoSmithKline’s chemical library for inhibitors of P. falciparum , of which 13,533 were confirmed to inhibit parasite growth by at least 80% at 2 µM concentration. More than 8,000 also showed potent activity against the multidrug resistant strain Dd2. Most (82%) compounds originate from internal company projects and are new to the malaria community. Analyses using historic assay data suggest several novel mechanisms of antimalarial action, such as inhibition of protein kinases and host–pathogen interaction related targets. Chemical structures and associated data are hereby made public to encourage additional drug lead identification efforts and further research into this disease. Antimalarial arsenal There are still nearly 250 million malaria cases reported annually, over 800,000 fatal, with most deaths being children under 5. The malaria parasite Plasmodium falciparum is notoriously adept at developing drug resistance, and new drugs are urgently needed. Two reports raise hopes that alternatives to artemisinins might be found, by identifying thousands of compounds inhibiting the growth of P. falciparum asexual-stage parasites in red blood cells, many distinct in structure and mechanism from current drugs. Guiguemde et al . present a chemical genomics screen of over 300,000 compounds: the 1,300 'hits' include 561 with good potency and broad therapeutic windows. Gamo et al . screened nearly 2 million compounds from GlaxoSmithKline's chemicals library, finding over 13,500 hits, many active against multidrug-resistant isolates. These studies provide a rich source of potential leads, freely available to academic and industry labs looking for new antimalarials. Here, nearly 2 million compounds from GlaxoSmithKline's chemical library were screened for inhibitors of the malaria parasite Plasmodium falciparum , grown in red blood cells. Of these compounds, some 13,500 inhibited parasite growth, and more than 8,000 also showed potent activity against a multidrug resistant strain. The targets of these compounds were inferred through bioinformatic analysis, revealing several new mechanisms of antimalarial action.

Benzoxaboroles are effective against bacterial, fungal and protozoan pathogens. We report potent activity of the benzoxaborole AN3661 against Plasmodium falciparum laboratory-adapted strains (mean IC 50 32 nM), Ugandan field isolates (mean ex vivo IC 50 64 nM), and murine P. berghei and P. falciparum infections (day 4 ED 90 0.34 and 0.57 mg kg −1 , respectively). Multiple P. falciparum lines selected in vitro for resistance to AN3661 harboured point mutations in pfcpsf3 , which encodes a homologue of mammalian cleavage and polyadenylation specificity factor subunit 3 (CPSF-73 or CPSF3). CRISPR-Cas9-mediated introduction of pfcpsf3 mutations into parental lines recapitulated AN3661 resistance. PfCPSF3 homology models placed these mutations in the active site, where AN3661 is predicted to bind. Transcripts for three trophozoite-expressed genes were lost in AN3661-treated trophozoites, which was not observed in parasites selected or engineered for AN3661 resistance. Our results identify the pre-mRNA processing factor PfCPSF3 as a promising antimalarial drug target. Benzoxaboroles have been shown to be active against different pathogens. Here, the authors show that the benzoxaborole AN3661 inhibits Plasmodium falciparum in vitro and in mouse models, and identify a homologue of a mammalian cleavage and polyadenylation specificity factor as a drug target.

Preventing parasite transmission from humans to mosquitoes is recognised to be critical for achieving elimination and eradication of malaria. Consequently developing new antimalarial drugs with transmission-blocking properties is a priority. Large screening campaigns have identified many new transmission-blocking molecules, however little is known about how they target the mosquito-transmissible Plasmodium falciparum stage V gametocytes, or how they affect their underlying cell biology. To respond to this knowledge gap, we have developed a machine learning image analysis pipeline to characterise and compare the cellular phenotypes generated by transmission-blocking molecules during male gametogenesis. Using this approach, we studied 40 molecules, categorising their activity based upon timing of action and visual effects on the organisation of tubulin and DNA within the cell. Our data both proposes new modes of action and corroborates existing modes of action of identified transmission-blocking molecules. Furthermore, the characterised molecules provide a new armoury of tool compounds to probe gametocyte cell biology and the generated imaging dataset provides a new reference for researchers to correlate molecular target or gene deletion to specific cellular phenotype. Our analysis pipeline is not optimised for a specific organism and could be applied to any fluorescence microscopy dataset containing cells delineated by bounding boxes, and so is potentially extendible to any disease model.

Identifying how small molecules act to kill malaria parasites can lead to new “chemically validated” targets. By pressuring Plasmodium falciparum asexual blood stage parasites with three novel structurally-unrelated antimalarial compounds (MMV665924, MMV019719 and MMV897615), and performing whole-genome sequence analysis on resistant parasite lines, we identify multiple mutations in the P. falciparum acyl-CoA synthetase (ACS) genes Pf ACS10 (PF3D7_0525100, M300I, A268D/V, F427L) and Pf ACS11 (PF3D7_1238800, F387V, D648Y, and E668K). Allelic replacement and thermal proteome profiling validates Pf ACS10 as a target of these compounds. We demonstrate that this protein is essential for parasite growth by conditional knockdown and observe increased compound susceptibility upon reduced expression. Inhibition of Pf ACS10 leads to a reduction in triacylglycerols and a buildup of its lipid precursors, providing key insights into its function. Analysis of the Pf ACS11 gene and its mutations point to a role in mediating resistance via decreased protein stability. Drug resistance to current antimalarials is rising and new drugs and targets are urgently needed. Here the authors identify Plasmodium falciparum acyl-CoA synthetase 10 as a new target whose inhibition leads to a decrease in triacylglycerols.

Chemotherapy is still the cornerstone for malaria control. Developing drugs against Plasmodium parasites and monitoring their efficacy requires methods to accurately determine the parasite killing rate in response to treatment. Commonly used techniques essentially measure metabolic activity as a proxy for parasite viability. However, these approaches are susceptible to artefacts, as viability and metabolism are two parameters that are coupled during the parasite life cycle but can be differentially affected in response to drug actions. Moreover, traditional techniques do not allow to measure the speed-of-action of compounds on parasite viability, which is an essential efficacy determinant. We present here a comprehensive methodology to measure in vitro the direct effect of antimalarial compounds over the parasite viability, which is based on limiting serial dilution of treated parasites and re-growth monitoring. This methodology allows to precisely determine the killing rate of antimalarial compounds, which can be quantified by the parasite reduction ratio and parasite clearance time, which are key mode-of-action parameters. Importantly, we demonstrate that this technique readily permits to determine compound killing activities that might be otherwise missed by traditional, metabolism-based techniques. The analysis of a large set of antimalarial drugs reveals that this viability-based assay allows to discriminate compounds based on their antimalarial mode-of-action. This approach has been adapted to perform medium throughput screening, facilitating the identification of fast-acting antimalarial compounds, which are crucially needed for the control and possibly the eradication of malaria.

Microbial resistance to chemotherapy has caused countless deaths where malaria is endemic. Chemotherapy may fail either due to pre-existing resistance or evolution of drug-resistant parasites. Here we use a diverse set of antimalarial compounds to investigate the acquisition of drug resistance and the degree of cross-resistance against common resistance alleles. We assess cross-resistance using a set of 15 parasite lines carrying resistance-conferring alleles in pfatp4 , cytochrome bc 1 , pfcarl , pfdhod, pfcrt , pfmdr, pfdhfr, cytoplasmic prolyl t-RNA synthetase or hsp90 . Subsequently, we assess whether resistant parasites can be obtained after several rounds of drug selection. Twenty-three of the 48 in vitro selections result in resistant parasites, with time to resistance onset ranging from 15 to 300 days. Our data indicate that pre-existing resistance may not be a major hurdle for novel-target antimalarial candidates, and focusing our attention on fast-killing compounds may result in a slower onset of clinical resistance. It is unclear whether new antimalarial compounds may rapidly lose effectiveness in the field because of parasite resistance. Here, Corey et al. investigate the acquisition of drug resistance and the extent to which common resistance mechanisms decrease susceptibility to a diverse set of 50 antimalarial compounds.

Plasmodium parasites are reliant on the Apicomplexan AP2 (ApiAP2) transcription factor family to regulate gene expression programs. AP2 DNA binding domains have no homologs in the human or mosquito host genomes, making them potential antimalarial drug targets. Using an in-silico screen to dock thousands of small molecules into the crystal structure of the AP2-EXP (Pf3D7_1466400) AP2 domain (PDB:3IGM), we identified putative AP2-EXP interacting compounds. Four compounds were found to block DNA binding by AP2-EXP and at least one additional ApiAP2 protein. Our top ApiAP2 competitor compound perturbs the transcriptome of P . falciparum trophozoites and results in a decrease in abundance of log 2 fold change > 2 for 50% (46/93) of AP2-EXP target genes. Additionally, two ApiAP2 competitor compounds have multi-stage anti- Plasmodium activity against blood and mosquito stage parasites. In summary, we describe a novel set of antimalarial compounds that interact with AP2 DNA binding domains. These compounds may be used for future chemical genetic interrogation of ApiAP2 proteins or serve as starting points for a new class of antimalarial therapeutics.

Background Despite the strides made in recent decades, the resistance observed in existing antimalarial drugs, and the intricate life cycle of the Plasmodium parasite underscore the pressing need to develop novel and effective therapeutic interventions. This article provides a comprehensive evaluation of the outcomes stemming from screening a library comprising 48 compounds (TCAMS) against Plasmodium falciparum . Methods This study focused on characterizing the IC 50 values of compounds from the Tres Cantos Antimalarial Set (TCAMS) library via a double-labelling method of P. falciparum parasites with SYBR Green-I and MitoTracker Deep Red, which were evaluated via flow cytometry. Evaluation of the cytotoxicity of the best candidates in human embryonic kidney (HEK293) cells, chemoinformatic analysis, and exploration of the effects of the compounds on the action of serotonin and melatonin in the erythrocytic life cycle of the parasite. Results IC 50 characterization confirmed that 93.75% of the compounds tested exhibited antimalarial activity at concentrations below 2 micromolar (µM), with 5 compounds showing IC 50 values below 50 nM (nM) (15.21 ± 5.97 nM to 45.82 ± 5.11 nM). Furthermore, 12 compounds presented IC 50 values between 50 and 100 nM (57.43 ± 12.25 nM to 100.6 ± 22.89 nM), highlighting their potent in vitro efficacy against P. falciparum . Cytotoxicity evaluation in HEK293 cells revealed that 12 from 17 compounds did not significantly reduce cell viability. Cheminformatics analysis clustered the compounds based on structural and physicochemical similarities, revealing distinct structural patterns. Exploration of hypothetical targets from the TCAMS library identified 27 compounds with potential targets, 15 specifically targeted serotonergic receptors. Subsequent serotonin and melatonin treatment experiments indicated that certain compounds could inhibit both effects on parasitaemia, suggesting a complex interaction with signaling in P. falciparum . Conclusions This study identifies promising antimalarial candidates with low IC 50 values and highlights the significance of targeting serotonin receptors in the development of potential antimalarial drugs.

Chloroquine/hydroxychloroquine have been proposed as potential treatments for COVID-19. These drugs have warning labels for use in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Analysis of whole genome sequence data of 458 individuals from sub-Saharan Africa showed significant G6PD variation across the continent. We identified nine variants, of which four are potentially deleterious to G6PD function, and one (rs1050828) that is known to cause G6PD deficiency. We supplemented data for the rs1050828 variant with genotype array data from over 11,000 Africans. Although this variant is common in Africans overall, large allele frequency differences exist between sub-populations. African sub-populations in the same country can show significant differences in allele frequency (e.g. 16.0% in Tsonga vs 0.8% in Xhosa, both in South Africa, p = 2.4 × 10−3). The high prevalence of variants in the G6PD gene found in this analysis suggests that it may be a significant interaction factor in clinical trials of chloroquine and hydroxychloroquine for treatment of COVID-19 in Africans.