Identification of fungal natural products with potent inhibition in Toxoplasma gondii.

In an effort to identify novel compounds with potent inhibition against Toxoplasma gondii, a phenotypic screen was performed utilizing a library of 683 pure compounds derived primarily from terrestrial and marine fungi. An initial screen with a fixed concentration of 5 µM yielded 91 hits with inhibition comparable to an equal concentration of artemisinin. These compounds were then triaged based on known biological and chemical concerns and liabilities. From these, 49 prioritized compounds were tested in a dose response format with T. gondii and human foreskin fibroblasts (HFFs) for cytotoxicity. Ten compounds were identified with an IC50 less than 150 nM and a selectivity index (SI) greater than 100. An additional eight compounds demonstrated submicromolar IC50 and SI values equal to or greater than 35. While the majority of these scaffolds have been previously implicated against apicomplexan parasites, their activities in T. gondii were largely unknown. Herein, we report the T. gondii activity of these compounds with chemotypes including xanthoquinodins, peptaibols, heptelidic acid analogs, and fumagillin analogs, with multiple compounds demonstrating exceptional potency in T. gondii and limited toxicity to HFFs at the highest concentrations tested. IMPORTANCE: Current therapeutics for treating toxoplasmosis remain insufficient, demonstrating high cytotoxicity, poor bioavailability, limited efficacy, and drug resistance. Additional research is needed to develop novel compounds with high efficacy and low cytotoxicity. The success of artemisinin and other natural products in treating malaria highlights the potential of natural products as anti-protozoan therapeutics. However, the exploration of natural products in T. gondii drug discovery has been less comprehensive, leaving untapped potential. By leveraging the resources available for the malaria drug discovery campaign, we conducted a phenotypic screen utilizing a set of natural products previously screened against Plasmodium falciparum. Our study revealed 18 compounds with high potency and low cytotoxicity in T. gondii, including four novel scaffolds with no previously reported activity in T. gondii. These new scaffolds may serve as starting points for the development of toxoplasmosis therapeutics but could also serve as tool compounds for target identification studies using chemogenomic approach.

Human nuclear hormone receptor activity contributes to malaria parasite liver stage development.

Chemical genetic approaches have had a transformative impact on discovery of drug targets for malaria but have primarily been used for parasite targets. To identify human pathways required for intrahepatic development of parasite, we implemented multiplex cytological profiling of malaria infected hepatocytes treated with liver stage active compounds. Some compounds, including MMV1088447 and MMV1346624, exhibited profiles similar to cells treated with nuclear hormone receptor (NHR) agonist/antagonists. siRNAs targeting human NHRs, or their signaling partners identified eight genes that were critical for Plasmodium berghei infection. Knockdown of NR1D2, a host NHR, significantly impaired parasite growth by downregulation of host lipid metabolism. Importantly, treatment with MMV1088447 and MMV1346624 but not other antimalarials, phenocopied the lipid metabolism defect of NR1D2 knockdown. Our data underlines the use of high-content imaging for host-cellular pathway deconvolution, highlights host lipid metabolism as a drug-able human pathway and provides new chemical biology tools for studying host-parasite interactions.

Advances in malaria pharmacology and the online guide to MALARIA PHARMACOLOGY: IUPHAR review 38.

Antimalarial drug discovery has until recently been driven by high-throughput phenotypic cellular screening, allowing millions of compounds to be assayed and delivering clinical drug candidates. In this review, we will focus on target-based approaches, describing recent advances in our understanding of druggable targets in the malaria parasite. Targeting multiple stages of the Plasmodium lifecycle, rather than just the clinically symptomatic asexual blood stage, has become a requirement for new antimalarial medicines, and we link pharmacological data clearly to the parasite stages to which it applies. Finally, we highlight the IUPHAR/MMV Guide to MALARIA PHARMACOLOGY, a web resource developed for the malaria research community that provides open and optimized access to published data on malaria pharmacology.

Adaptive laboratory evolution in S. cerevisiae highlights role of transcription factors in fungal xenobiotic resistance.

In vitro evolution and whole genome analysis were used to comprehensively identify the genetic determinants of chemical resistance in Saccharomyces cerevisiae. Sequence analysis identified many genes contributing to the resistance phenotype as well as numerous amino acids in potential targets that may play a role in compound binding. Our work shows that compound-target pairs can be conserved across multiple species. The set of 25 most frequently mutated genes was enriched for transcription factors, and for almost 25 percent of the compounds, resistance was mediated by one of 100 independently derived, gain-of-function SNVs found in a 170 amino acid domain in the two Zn2C6 transcription factors YRR1 and YRM1 (p < 1 × 10-100). This remarkable enrichment for transcription factors as drug resistance genes highlights their important role in the evolution of antifungal xenobiotic resistance and underscores the challenge to develop antifungal treatments that maintain potency.

MalDA, Accelerating Malaria Drug Discovery.

The Malaria Drug Accelerator (MalDA) is a consortium of 15 leading scientific laboratories. The aim of MalDA is to improve and accelerate the early antimalarial drug discovery process by identifying new, essential, druggable targets. In addition, it seeks to produce early lead inhibitors that may be advanced into drug candidates suitable for preclinical development and subsequent clinical testing in humans. By sharing resources, including expertise, knowledge, materials, and reagents, the consortium strives to eliminate the structural barriers often encountered in the drug discovery process. Here we discuss the mission of the consortium and its scientific achievements, including the identification of new chemically and biologically validated targets, as well as future scientific directions.

A consensus-based and readable extension of Linear Code for Reaction Rules (LiCoRR).

Systems glycobiology aims to provide models and analysis tools that account for the biosynthesis, regulation, and interactions with glycoconjugates. To facilitate these methods, there is a need for a clear glycan representation accessible to both computers and humans. Linear Code, a linearized and readily parsable glycan structure representation, is such a language. For this reason, Linear Code was adapted to represent reaction rules, but the syntax has drifted from its original description to accommodate new and originally unforeseen challenges. Here, we delineate the consensuses and inconsistencies that have arisen through this adaptation. We recommend options for a consensus-based extension of Linear Code that can be used for reaction rule specification going forward. Through this extension and specification of Linear Code to reaction rules, we aim to minimize inconsistent symbology thereby making glycan database queries easier. With a clear guide for generating reaction rule descriptions, glycan synthesis models will be more interoperable and reproducible thereby moving glycoinformatics closer to compliance with FAIR standards. Here, we present Linear Code for Reaction Rules (LiCoRR), version 1.0, an unambiguous representation for describing glycosylation reactions in both literature and code.

Probing the Open Global Health Chemical Diversity Library for Multistage-Active Starting Points for Next-Generation Antimalarials.

Most phenotypic screens aiming to discover new antimalarial chemotypes begin with low cost, high-throughput tests against the asexual blood stage (ABS) of the malaria parasite life cycle. Compounds active against the ABS are then sequentially tested in more difficult assays that predict whether a compound has other beneficial attributes. Although applying this strategy to new chemical libraries may yield new leads, repeated iterations may lead to diminishing returns and the rediscovery of chemotypes hitting well-known targets. Here, we adopted a different strategy to find starting points, testing ∼70,000 open source small molecules from the Global Health Chemical Diversity Library for activity against the liver stage, mature sexual stage, and asexual blood stage malaria parasites in parallel. In addition, instead of using an asexual assay that measures accumulated parasite DNA in the presence of compound (SYBR green), a real time luciferase-dependent parasite viability assay was used that distinguishes slow-acting (delayed death) from fast-acting compounds. Among 382 scaffolds with the activity confirmed by dose response (<10 µM), we discovered 68 novel delayed-death, 84 liver stage, and 68 stage V gametocyte inhibitors as well. Although 89% of the evaluated compounds had activity in only a single life cycle stage, we discovered six potent (half-maximal inhibitory concentration of <1 µM) multistage scaffolds, including a novel cytochrome bc1 chemotype. Our data further show the luciferase-based assays have higher sensitivity. Chemoinformatic analysis of positive and negative compounds identified scaffold families with a strong enrichment for activity against specific or multiple stages.