Focused structure-activity-relationship studies of MMV020136, 1, a whole-cell phenotypic screening hit originating from the Pathogen Box against the human malaria parasite, Plasmodium falciparum, led to the identification of compound 15 (MMV1581361). Compound 15 retained nanomolar activity against drug-sensitive, drug-resistant, and field isolates of Plasmodium falciparum blood-stage parasites. It was found to display a moderate rate-of-kill profile similar to pyrimethamine in vitro and efficacy in the humanized P. falciparum Pf3D70087/N9 NODscidIL2Rγnull mouse model at a single dose of 25 mg/kg, with similar kinetics to chloroquine. Furthermore, compound 15 demonstrated significant transmission-blocking potential, achieving 90% inhibition of male gamete exflagellation, a 98% reduction in mean oocyst intensity, and an 88% block in transmission. Mechanism-of-action studies revealed that compound 15 disrupts Na+ ion homeostasis in P. falciparum, indicating that it functions as a PfATP4 inhibitor. However, due to the resistance risk associated with the PfATP4 target and the presence of more advanced clinical candidates, further development of 15 into a viable antimalarial drug is currently deprioritised.

A new class of benzoxaboroles with a phenylglycine appendage was found to display in vitro blood stage activity against the human malaria parasite Plasmodium falciparum (Pf). Structure-activity relationship studies of the starting hit compound 3 resulted in compounds active against PfNF54 drug-sensitive and PfK1 drug-resistant strains with an in vitro antiplasmodium IC50 < 0.4 μM, selectivity over mammalian cell-lines (selectivity index > 47) and high aqueous solubility (160 to >200 μM). Selected compounds showed good in vitro metabolic stability when incubated with human, rat, and mouse liver microsomes and showed no cross-resistance against barcoded mutant lines. Two frontrunner compounds, 6 and 7, were dosed orally at 50 mg·kg-1 using a standard quadrupole dosing regimen in a P. berghei mouse infection model and showed encouraging in vivo efficacy. This work identifies a promising new class of phenylglycine-based benzoxaboroles, which warrants further medicinal chemistry optimization.

Negamycin is a natural product antibiotic discovered in 1970 and shown to have a Gram-negative spectrum of activity. It has served as the starting point in drug discovery efforts due in large part to its structural simplicity and novel mode of inhibition of the bacterial ribosome. It follows that negamycin does not show cross-resistance with other antibacterial agents that operate on the ribosome, whether this would be due to target modification, drug efflux, or drug metabolism. Because of the deficiencies of current drug regimens for the treatment of infections caused by Gram-negative pathogens, having a new agent brought to the infectious disease formulary represents a critical medical need, as has been promoted by the World Health Organization and other entities. Negamycin has been the subject of over 20 total syntheses, often highlighting stereoselective chemistry toward installing its two chiral centers on an acyclic chain. Novel synthetic methodologies thereby developed can stimulate the synthesis of novel analogs. With this, progress has been made in devising more potent analogs than negamycin. Structural work has determined that negamycin binds to the A-site of the 30S ribosome encounter complex with tRNA. Advancements have been made to understand the mechanism of transport of negamycin to the bacterial cytoplasm to enable engagement of the ribosome. This review surveys much of what has been published around negamycin and its analogs, including aspects of the biological spectrum of activity and mode of action as well as limitations that have held back clinical development.

In the search for novel chemotypes with high sp3 character and activity against the human malaria parasite Plasmodium falciparum, a spiroindoline series was identified from a phenotypic high-throughput screening campaign. The spiroindoline hit 2 displayed good activity against both drug-sensitive and multidrug-resistant Plasmodium strains, making it an attractive starting point for hit-to-lead progression. Structure-activity relationship studies led to the identification of a novel pyridylspiroindoline frontrunner (50) with improved antiplasmodial activity, aqueous solubility, and microsomal metabolic stability. Data from additional parasitological profiling suggested that 50 likely has a mode of action differing from that of the original spiroindoline hit. Compound 50 showed excellent in vivo pharmacokinetics with efficacy being achieved in a humanized immunodeficient NSG mouse P. falciparum infection model. This provided a pharmacological proof-of-concept for this series, making it a valuable starting point for further optimization in the quest for novel antimalarial therapeutics.

Dual angiotensin-converting enzyme (ACE) and neprilysin (NEP) inhibitors such as omapatrilat showed promise as potent treatments for hypertension but produced adverse effects due to their high affinity for both domains of ACE (nACE and cACE). This led to the search for compounds that retained NEP potency but selectively inhibit cACE, leaving nACE active to degrade other peptides such as bradykinin. Lisinopril-tryptophan (LisW) has previously been reported to have cACE selectivity. Three mercapto-3-phenylpropanoyl inhibitors were synthesized, combining features of omapatrilat and LisW to probe structural characteristics required for potent dual cACE/NEP inhibition. We report the synthesis of these inhibitors, enzyme inhibition data, and high-resolution crystal structures in complex with nACE and cACE. This provides valuable insight into factors driving potency and selectivity and shows that the mercapto-3-phenylpropanoyl backbone is significantly better for NEP potency than a P1 carboxylate. Future chemistry efforts could be directed at identifying alternative chemotypes for optimization of cACE/NEP inhibitors.

Previously reported antimalarial Plasmodium phosphatidylinositol 4-kinase IIIβ 2,8-diaryl-1,5-naphthyridine inhibitors have shown suboptimal physicochemical and pharmacokinetic properties. A focused target-based structure-activity relationship and structure-property optimization studies identified several compounds with good target and whole-cell activities and improved physicochemical properties. A new frontrunner compound 27 showed an improved pharmacokinetic profile and reduced parasitaemia (91% at 4 × 50 mg/kg QD doses) in the humanized NOD-scid IL-2Rγnull mouse model of Plasmodium falciparum malaria. Compound 27 poses no hERG channel inhibition at high concentrations or mammalian cytotoxicity but shows low selectivity against related human lipid kinases (PI3Kα and PI4Kβ); however, significantly higher selectivity margins were observed against the human MINK1 and MAP4K4 kinases.

Structure-activity relationship studies of 2,8-disubstituted-1,5-naphthyridines, previously reported as potent inhibitors of Plasmodium falciparum (Pf) phosphatidylinositol-4-kinase β (PI4K), identified 1,5-naphthyridines with basic groups at 8-position, which retained Plasmodium PI4K inhibitory activity but switched primary mode of action to the host hemoglobin degradation pathway through inhibition of hemozoin formation. These compounds showed minimal off-target inhibitory activity against the human phosphoinositide kinases and MINK1 and MAP4K kinases, which were associated with the teratogenicity and testicular toxicity observed in rats for the PfPI4K inhibitor clinical candidate MMV390048. A representative compound from the series retained activity against field isolates and lab-raised drug-resistant strains of Pf. It was efficacious in the humanized NSG mouse malaria infection model at a single oral dose of 32 mg/kg. This compound was nonteratogenic in the zebrafish embryo model of teratogenicity and has a low predicted human dose, indicating that this series has the potential to deliver a preclinical candidate for malaria.

Gyrase and topoisomerase IV are the cellular targets for fluoroquinolones, a critically important class of antibacterial agents used to treat a broad spectrum of human infections. Unfortunately, the clinical efficacy of the fluoroquinolones has been curtailed by the emergence of target-mediated resistance. This is especially true for Neisseria gonorrhoeae, the causative pathogen of the sexually transmitted infection gonorrhea. Spiropyrimidinetriones (SPTs), a new class of antibacterials, were developed to combat the growing antibacterial resistance crisis. Zoliflodacin is the most clinically advanced SPT and displays efficacy against uncomplicated urogenital gonorrhea in human trials. Like fluoroquinolones, the primary target of zoliflodacin in N. gonorrhoeae is gyrase, and topoisomerase IV is a secondary target. Because unbalanced gyrase/topoisomerase IV targeting has facilitated the evolution of fluoroquinolone-resistant bacteria, it is important to understand the underlying basis for the differential targeting of zoliflodacin in N. gonorrhoeae. Therefore, we assessed the effects of this SPT on the catalytic and DNA cleavage activities of N. gonorrhoeae gyrase and topoisomerase IV. In all reactions examined, zoliflodacin displayed higher potency against gyrase than topoisomerase IV. Moreover, zoliflodacin generated more DNA cleavage and formed more stable enzyme-cleaved DNA-SPT complexes with gyrase. The SPT also maintained higher activity against fluoroquinolone-resistant gyrase than topoisomerase IV. Finally, when compared to zoliflodacin, the novel SPT H3D-005722 induced more balanced double-stranded DNA cleavage with gyrase and topoisomerase IV from N. gonorrhoeae, Escherichia coli, and Bacillus anthracis. This finding suggests that further development of the SPT class could yield compounds with a more balanced targeting against clinically important bacterial infections.

UCT594 is a 2-aminopyrazine carboxylic acid Plasmodium phosphatidylinositol 4-kinase inhibitor with potent asexual blood-stage activity, the potential for interrupting transmission, as well as liver-stage activities. Herein, we investigated pharmacokinetic/pharmacodynamic (PK/PD) relationships relative to blood-stage activity toward predicting the human dose. Dose-fractionation studies were conducted in the Plasmodium falciparum NSG mouse model to determine the PK/PD indices of UCT594, using the in vivo minimum parasiticidal concentration as a threshold. UCT594 demonstrated concentration-dependent killing in the P. falciparum-infected NSG mouse model. Using this data and the preclinical pharmacokinetic data led to a low predicted human dose of <50 mg. This makes UCT594 an attractive potential antimalarial drug.

This viewpoint outlines the case for developing new chemical entities (NCEs) as racemates in infectious diseases and where both enantiomers and racemate retain similar on- and off-target activities as well as similar PK profiles. There are not major regulatory impediments for the development of a racemic drug, and minimizing the manufacturing costs becomes a particularly important objective when bringing an anti-infective therapeutic to the marketplace in the endemic settings of infectious diseases.

ADVERTISEMENT RETURN TO ISSUEViewpointNEXTThe 2022 H3D Symposium: Celebrating over a Decade of African-Led Infectious Disease Drug Discovery to Enhance Global HealthJohn G. WoodlandJohn G. WoodlandHolistic Drug Discovery and Development (H3D) Centre, University of Cape Town, Rondebosch 7700, South AfricaSouth African Medical Research Council Drug Discovery and Development Research Unit, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town 7925, South AfricaMore by John G. Woodlandhttps://orcid.org/0000-0001-6273-5596, Gregory S. BasarabGregory S. BasarabHolistic Drug Discovery and Development (H3D) Centre, University of Cape Town, Rondebosch 7700, South AfricaMore by Gregory S. Basarabhttps://orcid.org/0000-0001-5684-6046, Koketso MogweraKoketso MogweraHolistic Drug Discovery and Development (H3D) Centre, University of Cape Town, Rondebosch 7700, South AfricaMore by Koketso Mogwera, Susan WinksSusan WinksHolistic Drug Discovery and Development (H3D) Centre, University of Cape Town, Rondebosch 7700, South AfricaMore by Susan Winks, and Kelly Chibale*Kelly ChibaleHolistic Drug Discovery and Development (H3D) Centre, University of Cape Town, Rondebosch 7700, South AfricaSouth African Medical Research Council Drug Discovery and Development Research Unit, Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town 7925, South Africa*[email protected]More by Kelly Chibalehttps://orcid.org/0000-0002-1327-4727Cite this: ACS Infect. Dis. 2023, 9, 3, 389–393Publication Date (Web):February 10, 2023Publication History Received24 January 2023Published online10 February 2023Published inissue 10 March 2023https://pubs.acs.org/doi/10.1021/acsinfecdis.3c00041https://doi.org/10.1021/acsinfecdis.3c00041article-commentaryACS PublicationsCopyright © 2023 American Chemical Society. This publication is available under these Terms of Use. Request reuse permissions This publication is free to access through this site. Learn MoreArticle Views1226Altmetric-Citations-LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail PDF (1 MB) Get e-Alertsclose Get e-Alerts

The rise in drug-resistant tuberculosis has necessitated the search for alternative antibacterial treatments. Spiropyrimidinetriones (SPTs) represent an important new class of compounds that work through gyrase, the cytotoxic target of fluoroquinolone antibacterials. The present study analyzed the effects of a novel series of SPTs on the DNA cleavage activity of Mycobacterium tuberculosis gyrase. H3D-005722 and related SPTs displayed high activity against gyrase and increased levels of enzyme-mediated double-stranded DNA breaks. The activities of these compounds were similar to those of the fluoroquinolones, moxifloxacin, and ciprofloxacin and greater than that of zoliflodacin, the most clinically advanced SPT. All the SPTs overcame the most common mutations in gyrase associated with fluoroquinolone resistance and, in most cases, were more active against the mutant enzymes than wild-type gyrase. Finally, the compounds displayed low activity against human topoisomerase IIα. These findings support the potential of novel SPT analogues as antitubercular drugs.

Uncomplicated malaria is effectively treated with oral artemisinin-based combination therapy (ACT). Yet, there is an unmet clinical need for the intravenous treatment of the more fatal severe malaria. There is no combination intravenous therapy for uncomplicated due to the nonavailability of a water-soluble partner drug for the artemisinin, artesunate. The currently available treatment is a two-part regimen split into an intravenous artesunate followed by the conventional oral ACT . In a novel application of polymer therapeutics, the aqueous insoluble antimalarial lumefantrine is conjugated to a carrier polymer to create a new water-soluble chemical entity suitable for intravenous administration in a clinically relevant formulation . The conjugate is characterized by spectroscopic and analytical techniques, and the aqueous solubility of lumefantrine is determined to have increased by three orders of magnitude. Pharmacokinetic studies in mice indicate that there is a significant plasma release of lumefantrine and production its metabolite desbutyl-lumefantrine (area under the curve of metabolite is ≈10% that of the parent). In a Plasmodium falciparum malaria mouse model, parasitemia clearance is 50% higher than that of reference unconjugated lumefantrine. The polymer-lumefantrine shows potential for entering the clinic to meet the need for a one-course combination treatment for severe malaria.

New antibiotics with either a novel mode of action or novel mode of inhibition are urgently needed to overcome the threat of drug-resistant tuberculosis (TB). The present study profiles new spiropyrimidinetriones (SPTs), DNA gyrase inhibitors having activity against drug-resistant Mycobacterium tuberculosis (Mtb), the causative agent of TB. While the clinical candidate zoliflodacin has progressed to phase 3 trials for the treatment of gonorrhea, compounds herein demonstrated higher inhibitory potency against Mtb DNA gyrase (e.g., compound 42 with IC50 = 2.0) and lower Mtb minimum inhibitor concentrations (0.49 μM for 42). Notably, 42 and analogues showed selective Mtb activity relative to representative Gram-positive and Gram-negative bacteria. DNA gyrase inhibition was shown to involve stabilization of double-cleaved DNA, while on-target activity was supported by hypersensitivity against a gyrA hypomorph. Finally, a docking model for SPTs with Mtb DNA gyrase was developed, and a structural hypothesis was built for structure-activity relationship expansion.

Selective inhibition of the angiotensin-converting enzyme C-domain (cACE) and neprilysin (NEP), leaving the ACE N-domain (nACE) free to degrade bradykinin and other peptides, has the potential to provide the potent antihypertensive and cardioprotective benefits observed for nonselective dual ACE/NEP inhibitors, such as omapatrilat, without the increased risk of adverse effects. We have synthesized three 1-carboxy-3-phenylpropyl dipeptide inhibitors with nanomolar potency based on the previously reported C-domain selective ACE inhibitor lisinopriltryptophan (LisW) to probe the structural requirements for potent dual cACE/NEP inhibition. Here we report the synthesis, enzyme kinetic data, and high-resolution crystal structures of these inhibitors bound to nACE and cACE, providing valuable insight into the factors driving potency and selectivity. Overall, these results highlight the importance of the interplay between the S-1' and S-2' subsites for ACE domain selectivity, providing guidance for future chemistry efforts toward the development of dual cACE/NEP inhibitors.

Described here is a series of spiropyrimidinetrione (SPT) compounds with activity against Mycobacterium tuberculosis through inhibition of DNA gyrase. The SPT class operates via a novel mode of inhibition, which involves Mg2+-independent stabilization of the DNA cleavage complex with DNA gyrase and is thereby not cross-resistant with other DNA gyrase-inhibiting antibacterials, including fluoroquinolones. Compound 22 from the series was profiled broadly and showed in vitro cidality as well as intracellular activity against M. tuberculosis in macrophages. Evidence for the DNA gyrase mode of action was supported by inhibition of the target in a DNA supercoiling assay and elicitation of an SOS response seen in a recA reporter strain of M. tuberculosis. Pharmacokinetic properties of 22 supported evaluation of efficacy in an acute model of M. tuberculosis infection, where modest reduction in CFU numbers was seen. This work offers promise for deriving a novel drug class of tuberculosis agent without preexisting clinical resistance.

Phenotypic screening of a Medicines for Malaria Venture compound library against Mycobacterium tuberculosis (Mtb) identified a cluster of pan-active 2-pyrazolylpyrimidinones. The biology triage of these actives using various tool strains of Mtb suggested a novel mechanism of action. The compounds were bactericidal against replicating Mtb and retained potency against clinical isolates of Mtb. Although selected MmpL3 mutant strains of Mtb showed resistance to these compounds, there was no shift in the minimum inhibitory concentration (MIC) against a mmpL3 hypomorph, suggesting mutations in MmpL3 as a possible resistance mechanism for the compounds but not necessarily as the target. RNA transcriptional profiling and the checkerboard board 2D-MIC assay in the presence of varying concentrations of ferrous salt indicated perturbation of the Fe-homeostasis by the compounds. Structure-activity relationship studies identified potent compounds with good physicochemical properties and in vitro microsomal metabolic stability with moderate selectivity over cytotoxicity against mammalian cell lines.

Phenotypic whole cell high-throughput screening of a ∼150,000 diverse set of compounds against Mycobacterium tuberculosis (Mtb) in cholesterol-containing media identified 1,3-diarylpyrazolyl-acylsulfonamide 1 as a moderately active hit. Structure-activity relationship (SAR) studies demonstrated a clear scope to improve whole cell potency to MIC values of <0.5 μM, and a plausible pharmacophore model was developed to describe the chemical space of active compounds. Compounds are bactericidal in vitro against replicating Mtb and retained activity against multidrug-resistant clinical isolates. Initial biology triage assays indicated cell wall biosynthesis as a plausible mode-of-action for the series. However, no cross-resistance with known cell wall targets such as MmpL3, DprE1, InhA, and EthA was detected, suggesting a potentially novel mode-of-action or inhibition. The in vitro and in vivo drug metabolism and pharmacokinetics profiles of several active compounds from the series were established leading to the identification of a compound for in vivo efficacy proof-of-concept studies.

A novel diazaspiro[3.4]octane series was identified from a Plasmodium falciparum whole-cell high-throughput screening campaign. Hits displayed activity against multiple stages of the parasite lifecycle, which together with a novel sp3-rich scaffold provided an attractive starting point for a hit-to-lead medicinal chemistry optimization and biological profiling program. Structure-activity-relationship studies led to the identification of compounds that showed low nanomolar asexual blood-stage activity (<50 nM) together with strong gametocyte sterilizing properties that translated to transmission-blocking activity in the standard membrane feeding assay. Mechanistic studies through resistance selection with one of the analogues followed by whole-genome sequencing implicated the P. falciparum cyclic amine resistance locus in the mode of resistance.

Chemical matter is needed to target the divergent biology associated with the different life cycle stages of Plasmodium . Here, we report the parallel de novo screening of the Medicines for Malaria Venture (MMV) Pandemic Response Box against Plasmodium asexual and liver stage parasites, stage IV/V gametocytes, gametes, oocysts and as endectocides. Unique chemotypes were identified with both multistage activity or stage-specific activity, including structurally diverse gametocyte-targeted compounds with potent transmission-blocking activity, such as the JmjC inhibitor ML324 and the antitubercular clinical candidate SQ109. Mechanistic investigations prove that ML324 prevents histone demethylation, resulting in aberrant gene expression and death in gametocytes. Moreover, the selection of parasites resistant to SQ109 implicates the druggable V-type H + -ATPase for the reduced sensitivity. Our data therefore provides an expansive dataset of compounds that could be redirected for antimalarial development and also point towards proteins that can be targeted in multiple parasite life cycle stages.