Pharmacological validation of N-myristoyltransferase as a drug target in Leishmania donovani

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Abstract

Visceral leishmaniasis (VL), caused by the protozoan parasites Leishmania donovani and L. infantum, is responsible for ~30,000 deaths annually. Available treatments are inadequate and there is a pressing need for new therapeutics. N-Myristoyltransferase (NMT) remains one of the few genetically validated drug targets in these parasites. Here, we sought to pharmacologically validate this enzyme in Leishmania. A focused set of 1,600 pyrazolyl sulfonamide compounds was screened against L. major NMT in a robust high-throughput biochemical assay. Several potent inhibitors were identified with marginal selectivity over the human enzyme. There was little correlation between the enzyme potency of these inhibitors and their cellular activity against L. donovani axenic amastigotes and this discrepancy could be due to poor cellular uptake due to the basicity of these compounds. Thus, a series of analogues were synthesised with less basic centres. Although most of these compounds continued to suffer from relatively poor anti-leishmanial activity, our most potent inhibitor of LmNMT (DDD100097, Ki 0.34 nM), showed modest activity against L. donovani intracellular amastigotes (EC50 2.4 µM) and maintained a modest therapeutic window over the human enzyme. Two un-biased approaches, namely screening against our cosmid-based overexpression library and thermal proteome profiling (TPP), confirm that DDD100097 (compound 2) acts on-target within parasites. Oral dosing with compound 2 resulted in a 52% reduction in parasite burden in our mouse model of VL. Thus, NMT is now a pharmacologically validated target in Leishmania. The challenge in finding drug candidates remains to identify alternative strategies to address the drop-off in activity between enzyme inhibition and in vitro activity while maintaining sufficient selectivity over the human enzyme, both issues that continue to plague studies in this area.
Original languageEnglish
Pages (from-to)111-122
Number of pages12
JournalACS Infectious Diseases
Volume5
Issue number1
Early online date1 Nov 2018
DOIs
Publication statusPublished - 11 Jan 2019

Fingerprint

Leishmania donovani
Pharmacology
Parasites
Enzymes
Pharmaceutical Preparations
Visceral Leishmaniasis
Leishmania
Leishmania infantum
Cosmids
Plague
Sulfonamides
Enzyme Inhibitors
Proteome
Libraries
Hot Temperature
glycylpeptide N-tetradecanoyltransferase
Therapeutics

Keywords

  • Leishmania
  • N-myristoyltransferase
  • drug discovery
  • target validation
  • thermal proteome profiling (TPP)

Cite this

@article{ac573eb7715346d7bdde1ab085c2cf7b,
title = "Pharmacological validation of N-myristoyltransferase as a drug target in Leishmania donovani",
abstract = "Visceral leishmaniasis (VL), caused by the protozoan parasites Leishmania donovani and L. infantum, is responsible for ~30,000 deaths annually. Available treatments are inadequate and there is a pressing need for new therapeutics. N-Myristoyltransferase (NMT) remains one of the few genetically validated drug targets in these parasites. Here, we sought to pharmacologically validate this enzyme in Leishmania. A focused set of 1,600 pyrazolyl sulfonamide compounds was screened against L. major NMT in a robust high-throughput biochemical assay. Several potent inhibitors were identified with marginal selectivity over the human enzyme. There was little correlation between the enzyme potency of these inhibitors and their cellular activity against L. donovani axenic amastigotes and this discrepancy could be due to poor cellular uptake due to the basicity of these compounds. Thus, a series of analogues were synthesised with less basic centres. Although most of these compounds continued to suffer from relatively poor anti-leishmanial activity, our most potent inhibitor of LmNMT (DDD100097, Ki 0.34 nM), showed modest activity against L. donovani intracellular amastigotes (EC50 2.4 µM) and maintained a modest therapeutic window over the human enzyme. Two un-biased approaches, namely screening against our cosmid-based overexpression library and thermal proteome profiling (TPP), confirm that DDD100097 (compound 2) acts on-target within parasites. Oral dosing with compound 2 resulted in a 52{\%} reduction in parasite burden in our mouse model of VL. Thus, NMT is now a pharmacologically validated target in Leishmania. The challenge in finding drug candidates remains to identify alternative strategies to address the drop-off in activity between enzyme inhibition and in vitro activity while maintaining sufficient selectivity over the human enzyme, both issues that continue to plague studies in this area.",
keywords = "Leishmania, N-myristoyltransferase, drug discovery, target validation, thermal proteome profiling (TPP)",
author = "{Corpas Lopez}, Victoriano and Sonia Moniz and Michael Thomas and Richard Wall and Leah Torrie and Dorothea Zander-Dinse and Michele Tinti and Stephen Brand and Laste Stojanovski and Sujatha Manthri and Irene Hallyburton and Fabio Zuccotto and Paul Wyatt and {De Rycker}, Manu and David Horn and Michael Ferguson and Joachim Clos and Kevin Read and Alan Fairlamb and Ian Gilbert and Susan Wyllie",
note = "Funding: Wellcome Trust (Grants 105021, 101842, 203134, 092340 and 100476); Drugs for Neglected Diseases Initiative.",
year = "2019",
month = "1",
day = "11",
doi = "10.1021/acsinfecdis.8b00226",
language = "English",
volume = "5",
pages = "111--122",
journal = "ACS Infectious Diseases",
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publisher = "American Chemical Society",
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T1 - Pharmacological validation of N-myristoyltransferase as a drug target in Leishmania donovani

AU - Corpas Lopez, Victoriano

AU - Moniz, Sonia

AU - Thomas, Michael

AU - Wall, Richard

AU - Torrie, Leah

AU - Zander-Dinse, Dorothea

AU - Tinti, Michele

AU - Brand, Stephen

AU - Stojanovski, Laste

AU - Manthri, Sujatha

AU - Hallyburton, Irene

AU - Zuccotto, Fabio

AU - Wyatt, Paul

AU - De Rycker, Manu

AU - Horn, David

AU - Ferguson, Michael

AU - Clos, Joachim

AU - Read, Kevin

AU - Fairlamb, Alan

AU - Gilbert, Ian

AU - Wyllie, Susan

N1 - Funding: Wellcome Trust (Grants 105021, 101842, 203134, 092340 and 100476); Drugs for Neglected Diseases Initiative.

PY - 2019/1/11

Y1 - 2019/1/11

N2 - Visceral leishmaniasis (VL), caused by the protozoan parasites Leishmania donovani and L. infantum, is responsible for ~30,000 deaths annually. Available treatments are inadequate and there is a pressing need for new therapeutics. N-Myristoyltransferase (NMT) remains one of the few genetically validated drug targets in these parasites. Here, we sought to pharmacologically validate this enzyme in Leishmania. A focused set of 1,600 pyrazolyl sulfonamide compounds was screened against L. major NMT in a robust high-throughput biochemical assay. Several potent inhibitors were identified with marginal selectivity over the human enzyme. There was little correlation between the enzyme potency of these inhibitors and their cellular activity against L. donovani axenic amastigotes and this discrepancy could be due to poor cellular uptake due to the basicity of these compounds. Thus, a series of analogues were synthesised with less basic centres. Although most of these compounds continued to suffer from relatively poor anti-leishmanial activity, our most potent inhibitor of LmNMT (DDD100097, Ki 0.34 nM), showed modest activity against L. donovani intracellular amastigotes (EC50 2.4 µM) and maintained a modest therapeutic window over the human enzyme. Two un-biased approaches, namely screening against our cosmid-based overexpression library and thermal proteome profiling (TPP), confirm that DDD100097 (compound 2) acts on-target within parasites. Oral dosing with compound 2 resulted in a 52% reduction in parasite burden in our mouse model of VL. Thus, NMT is now a pharmacologically validated target in Leishmania. The challenge in finding drug candidates remains to identify alternative strategies to address the drop-off in activity between enzyme inhibition and in vitro activity while maintaining sufficient selectivity over the human enzyme, both issues that continue to plague studies in this area.

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KW - Leishmania

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KW - drug discovery

KW - target validation

KW - thermal proteome profiling (TPP)

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