168 results found
Bell AS, Yu Z, Hutton JA, et al., 2020, Novel thienopyrimidine inhibitors of Leishmania N-myristoyltransferase with on-target activity in intracellular amastigotes, Journal of Medicinal Chemistry, Vol: 14, Pages: 7740-7765, ISSN: 0022-2623
The leishmaniases, caused by Leishmania species of protozoan parasites, are neglected tropical diseases with 12-15 million cases worldwide. Current therapeutic approaches are limited by toxicity, resistance and cost. N-Myristoyltransferase (NMT), an enzyme ubiquitous and essential in all eukaryotes, has been validated via genetic and pharmacological methods as a promising antileishmanial target. Here we describe a comprehensive structure activity relationship study of a thienopyrimidine series previously identified in a high throughput screen against Leishmania NMT, across 68 compounds in enzyme- and cell-based assay formats. Using a chemical tagging target engagement biomarker assay we identify the first inhibitor in this series with on-target NMT activity in leishmania parasites. Furthermore, crystal structure analyses of 12 derivatives in complex with Leishmania major NMT revealed key factors important for future structure-guided optimization delivering IMP-105 (43), a compound with modest activity against L. donovani intracellular amastigotes and excellent selectivity (>660-fold) for Leishmania NMT over human NMTs.
Mousnier A, Bell AS, Swieboda DP, et al., 2018, Fragment-derived inhibitors of human N-myristoyltransferase block capsid assembly and replication of the common cold virus, Nature Chemistry, Vol: 10, Pages: 599-606, ISSN: 1755-4330
Rhinoviruses are the pathogens most often responsible for the common cold, and are a frequent cause of exacerbations in asthma, chronic obstructive pulmonary disease and cystic fibrosis. Here we report discovery of IMP-1088, a picomolar dual inhibitor of the human N-myristoyltransferases NMT1 and NMT2, and use it to demonstrate that pharmacological inhibition of host cell N-myristoylation rapidly and completely prevents rhinoviral replication without inducing cytotoxicity. Identification of cooperative binding between weak-binding fragments led to rapid inhibitor optimization through fragment reconstruction, structure-guided fragment linking, and conformational control over linker geometry. We show that inhibition of co-translational myristoylation of a specific virus-encoded protein (VP0) by IMP-1088 potently blocks a key step in viral capsid assembly, delivering low nanomolar antiviral activity against multiple rhinovirus strains, poliovirus and foot-and-mouth disease virus, and protection of cells against virus-induced killing, highlighting the potential of host myristoylation as a drug target in picornaviral infections.
Cilibrizzi A, Fedorova M, Collins J, et al., 2017, A tri-functional vanadium(IV) complex to detect cysteine oxidation, DALTON TRANSACTIONS, Vol: 46, Pages: 6994-7004, ISSN: 1477-9226
The development of effective molecular probes to detect and image the levels of oxidative stress in cells remains a challenge. Herein we report the design, synthesis and preliminary biological evaluation of a novel optical probe to monitor oxidation of thiol groups in cysteine-based phosphatases (CBPs). Following orthogonal protecting approaches we synthesised a new vanadyl complex designed to bind to CBPs. This complex is functionalised with a well-known dimedone derivative (to covalently trap sulfenic acids, SOHs) and a coumarin-based fluorophore for optical visualization. We show that this new probe efficiently binds to a range of phosphatases in vitro with nanomolar affinity. Moreover, preliminary flow cytometry and microscopy studies in live HCT116 cells show that this probe can successfully image cellular levels of sulfenic acids – one of the species resulting from protein oxidative damage.
Goncalves V, Brannigan JA, Laporte A, et al., 2016, Structure-guided optimization of quinoline inhibitors of Plasmodium N-myristoyltransferase, MedChemComm, Vol: 8, Pages: 191-197, ISSN: 2040-2511
The parasite Plasmodium vivax is the most widely distributed cause of recurring malaria. N-myristoyltransferase (NMT), an enzyme that catalyses the covalent attachment of myristate to the N-terminal glycine of substrate proteins, has been described as a potential target for the treatment of this disease. Herein, we report the synthesis and the structure-guided optimization of a series of quinolines with balanced activity against both Plasmodium vivax and Plasmodium falciparum N-myristoyltransferase (NMT).
Aronica PG, Verma C, Popovic B, et al., 2016, The Parasol Protocol for computational mutagenesis, Protein Engineering Design & Selection, Vol: 29, Pages: 253-261, ISSN: 1741-0134
To aid in the discovery and development of peptides and proteins as therapeutic agents, a virtual screen can be used to predict trends and direct workflow. We have developed the Parasol Protocol, a dynamic method implemented using the AMBER MD package, for computational site-directed mutagenesis. This tool can mutate between any pair of amino acids in a computationally expedient, automated manner. To demonstrate the potential of this methodology, we have employed the protocol to investigate a test case involving stapled peptides, and have demonstrated good agreement with experiment.
Vilar Compte R, Collins J, Woscholski R, et al., 2016, Vanadyl complexes with dansyl-labelled di-picolinic acid ligands: synthesis, phosphatase inhibition activity and cellular uptake studies, Dalton Transactions, Vol: 45, Pages: 7104-7113, ISSN: 1477-9226
Vanadium complexes have been previously utilised as potent inhibitors of cysteine based phosphatases (CBPs) . Herein, we present the synthesis and characterisation of two new fluorescently labelled vanadyl complexes (14 and 15 ) with bridged dipicolinic acid ligand. These compounds differ significantly from previous vanadyl complexes with phosphatase inhibition properties in that the metal-chelating part is a single tetradentate unit, which should afford greater stability and scope for synthetic elaboration then the earlier complexes. These new complexes inhibit a selection of cysteine based phosphatases (CBPs) in the nM range with some selectivity. Fluorescence spectroscopic studies (including fluorescence anisotropy) were carried out to demonstrate that the complexes are not simply acting as vanadyl delivery vehicles but they interact with the proteins. Finally, we present preliminary fluorescence microscopy studies to demonstrate that the complexes are cell permeable and localise throughout the cytoplasm of NIH3T3 cells.
Rackham MD, Yu Z, Brannigan JA, et al., 2015, Discovery of high affinity inhibitors of Leishmania donovani N-myristoyltransferase, MedChemComm, Vol: 6, Pages: 1761-1766, ISSN: 2040-2511
N-Myristoyltransferase (NMT) is a potential drug target in Leishmania parasites. Scaffold-hopping from published inhibitors yielded the serendipitous discovery of a chemotype selective for Leishmania donovani NMT; development led to high affinity inhibitors with excellent ligand efficiency. The binding mode was characterised by crystallography and provides a structural rationale for selectivity.
Pitchford WH, Kim H-J, Ivanov AP, et al., 2015, Synchronized Optical and Electronic Detection of Biomolecules Using a Low Noise Nanopore Platform, ACS NANO, Vol: 9, Pages: 1740-1748, ISSN: 1936-0851
Yu Z, Brannigan JA, Rangachari K, et al., 2015, Discovery of pyridyl-based inhibitors of Plasmodium falciparum N-myristoyltransferase, MEDCHEMCOMM, Vol: 6, Pages: 1767-1772, ISSN: 2040-2503
Paape D, Bell AS, Heal WP, et al., 2014, Using a Non-Image-Based Medium-Throughput Assay for Screening Compounds Targeting N-myristoylation in Intracellular Leishmania Amastigotes, PLOS NEGLECTED TROPICAL DISEASES, Vol: 8, ISSN: 1935-2735
Hutton JA, Goncalves V, Brannigan JA, et al., 2014, Structure-Based Design of Potent and Selective Leishmania N-Myristoyltransferase Inhibitors, JOURNAL OF MEDICINAL CHEMISTRY, Vol: 57, Pages: 8664-8670, ISSN: 0022-2623
Olaleye TO, Brannigan JA, Roberts SM, et al., 2014, Peptidomimetic inhibitors of N-myristoyltransferase from human malaria and leishmaniasis parasites, Organic & Biomolecular Chemistry, Vol: 12, Pages: 8132-8137, ISSN: 1477-0539
N-Myristoyltransferase (NMT) has been shown to be essential in Leishmania and subsequently validated as a drug target in Plasmodium. Herein, we discuss the use of antifungal NMT inhibitors as a basis for inhibitor development resulting in the first sub-micromolar peptidomimetic inhibitors of Plasmodium and Leishmania NMTs. High-resolution structures of these inhibitors with Plasmodium and Leishmania NMTs permit a comparative analysis of binding modes, and provide the first crystal structure evidence for a ternary NMT-Coenzyme A/myristoylated peptide product complex.
Aronica PGA, Gould IR, Leatherbarrow RJ, 2014, Development of a new protocol for computational site-directed mutagenesis, 248th National Meeting of the American-Chemical-Society (ACS), Publisher: AMER CHEMICAL SOC, ISSN: 0065-7727
Cilibrizzi A, Collins J, Woscholski R, et al., 2014, New vanadium complexes as optical probes to detect Cys sulfenic modifications in PTEN, 12th European Biological Inorganic Chemistry Conference (EuroBIC), Publisher: Springer Verlag (Germany), Pages: S873-S873, ISSN: 1432-1327
Brannigan JA, Roberts SM, Bell AS, et al., 2014, Diverse modes of binding in structures of Leishmania major N-myristoyltransferase with selective inhibitors, IUCrJ, Vol: 1, Pages: 250-260, ISSN: 2052-2525
The leishmaniases are a spectrum of global diseases of poverty associated withimmune dysfunction and are the cause of high morbidity. Despite the longhistory of these diseases, no effective vaccine is available and the currently useddrugs are variously compromised by moderate efficacy, complex side effects andthe emergence of resistance. It is therefore widely accepted that new therapiesare needed. N-Myristoyltransferase (NMT) has been validated pre-clinically asa target for the treatment of fungal and parasitic infections. In a previouslyreported high-throughput screening program, a number of hit compounds withactivity against NMT from Leishmania donovani have been identified. Here,high-resolution crystal structures of representative compounds from four hitseries in ternary complexes with myristoyl-CoA and NMT from the closelyrelated L. major are reported. The structures reveal that the inhibitors associatewith the peptide-binding groove at a site adjacent to the bound myristoyl-CoAand the catalytic -carboxylate of Leu421. Each inhibitor makes extensiveapolar contacts as well as a small number of polar contacts with the protein.Remarkably, the compounds exploit different features of the peptide-bindinggroove and collectively occupy a substantial volume of this pocket, suggestingthat there is potential for the design of chimaeric inhibitors with significantlyenhanced binding. Despite the high conservation of the active sites of theparasite and human NMTs, the inhibitors act selectively over the host enzyme.The role of conformational flexibility in the side chain of Tyr217 in conferringselectivity is discussed.
Gray K, Elghadban S, Thongyoo P, et al., 2014, Potent and specific inhibition of the biological activity of the type-II transmembrane serine protease matriptase by the cyclic microprotein MCoTI-II, Thrombosis and Haemostasis, Vol: 112, Pages: 402-411, ISSN: 0340-6245
Matriptase is a type-II transmembrane serine protease involved in epithelial homeostasis in both health and disease, and is implicated in the development and progression of a variety of cancers. Matriptase mediates its biological effects both via as yet undefined substrates and pathways, and also by proteolytic cleavage of a variety of well-defined protein substrates, several of which it shares with the closely-related protease hepsin. Development of targeted therapeutic strategies will require discrimination between these proteases. Here we have investigated cyclic microproteins of the squash Momordica cochinchinensis trypsin-inhibitor family (generated by total chemical synthesis) and found MCoTI-II to be a high-affinity (Ki 9 nM) and highly selective (> 1,000-fold) inhibitor of matriptase. MCoTI-II efficiently inhibited the proteolytic activation of pro-hepatocyte growth factor (HGF) by matriptase but not by hepsin, in both purified and cell-based systems, and inhibited HGF-dependent cell scattering. MCoTI-II also selectively inhibited the invasion of matriptase-expressing prostate cancer cells. Using a model of epithelial cell tight junction assembly, we also found that MCoTI-II could effectively inhibit the re-establishment of tight junctions and epithelial barrier function in MDCK-I cells after disruption, consistent with the role of matriptase in regulating epithelial integrity. Surprisingly, MCoTI-II was unable to inhibit matriptase-dependent proteolytic activation of prostasin, a GPI-anchored serine protease also implicated in epithelial homeostasis. These observations suggest that the unusually high selectivity afforded by MCoTI-II and its biological effectiveness might represent a useful starting point for the development of therapeutic inhibitors, and further highlight the role of matriptase in epithelial maintenance.
Bell AS, Goncalves V, Hutton JA, et al., 2014, N-Myristoyltransferase inhibitors as anti-leishmanial agents, 247th National Spring Meeting of the American-Chemical-Society (ACS), Publisher: AMER CHEMICAL SOC, ISSN: 0065-7727
Rackham MD, Brannigan JA, Rangachari K, et al., 2014, Design and Synthesis of High Affinity Inhibitors of Plasmodium falciparum and Plasmodium vivax N-Myristoyltransferases Directed by Ligand Efficiency Dependent Lipophilicity (LELP), Journal of Medicinal Chemistry, Vol: 57, Pages: 2773-2788, ISSN: 0022-2623
N-Myristoyltransferase (NMT) is an essential eukaryotic enzyme and an attractive drug target in parasiticinfections such as malaria. We have previously reported that 2-(3-(piperidin-4-yloxy)benzo[b]thiophen-2-yl)-5-((1,3,5-trimethyl-1H-pyrazol-4-yl)methyl)-1,3,4-oxadiazole (34c) is a high affinity inhibitor of both Plasmodium falciparum and P. vivax NMT anddisplays activity in vivo against a rodent malaria model. Here we describe the discovery of 34c through optimization of apreviously described series. Development, guided by targeting a ligand efficiency dependent lipophilicity (LELP) score of lessthan 10, yielded a 100-fold increase in enzyme affinity and a 100-fold drop in lipophilicity with the addition of only two heavyatoms. 34c was found to be equipotent on chloroquine-sensitive and -resistant cell lines and on both blood and liver stage formsof the parasite. These data further validate NMT as an exciting drug target in malaria and support 34c as an attractive tool forfurther optimization.
Roqué Rosell NR, Mokhlesi L, Milton NE, et al., 2014, Design and synthesis of irreversible inhibitors of foot-and-mouth disease virus 3C protease., Bioorg Med Chem Lett, Vol: 24, Pages: 490-494
Foot-and-mouth disease virus (FMDV) causes a highly infectious and economically devastating disease of livestock. The FMDV genome is translated as a single polypeptide precursor that is cleaved into functional proteins predominantly by the highly conserved viral 3C protease, making this enzyme an attractive target for antiviral drugs. A peptide corresponding to an optimal substrate has been modified at the C-terminus, by the addition of a warhead, to produce irreversible inhibitors that react as Michael acceptors with the enzyme active site. Further investigation highlighted key structural determinants for inhibition, with a positively charged P2 being particularly important for potency.
Wright MH, Clough B, Rackham MD, et al., 2013, Validation of N-myristoyltransferase as an antimalarial drug target using an integrated chemical biology approach, Nature Chemistry, Vol: 6, Pages: 112-121, ISSN: 1755-4349
Malaria is an infectious disease caused by parasites of the genus Plasmodium, which leads to approximately one million deaths per annum worldwide. Chemical validation of new antimalarial targets is urgently required in view of rising resistance to current drugs. One such putative target is the enzyme N-myristoyltransferase, which catalyses the attachment of the fatty acid myristate to protein substrates (N-myristoylation). Here, we report an integrated chemical biology approach to explore protein myristoylation in the major human parasite P. falciparum, combining chemical proteomic tools for identification of the myristoylated and glycosylphosphatidylinositol-anchored proteome with selective small-molecule N-myristoyltransferase inhibitors. We demonstrate that N-myristoyltransferase is an essential and chemically tractable target in malaria parasites both in vitro and in vivo, and show that selective inhibition of N-myristoylation leads to catastrophic and irreversible failure to assemble the inner membrane complex, a critical subcellular organelle in the parasite life cycle. Our studies provide the basis for the development of new antimalarials targeting N-myristoyltransferase.
Smith LD, Leatherbarrow RJ, Spivey AC, 2013, Development of small molecules to target the IgE:Fc epsilon RI protein-protein interaction in allergies, FUTURE MEDICINAL CHEMISTRY, Vol: 5, Pages: 1423-1435, ISSN: 1756-8919
Rackham MD, Brannigan JA, Moss DK, et al., 2013, Discovery of Novel and Ligand-Efficient Inhibitors of Plasmodium falciparum and Plasmodium vivax N-Myristoyltransferase, JOURNAL OF MEDICINAL CHEMISTRY, Vol: 56, Pages: 371-375, ISSN: 0022-2623
Yu Z, Brannigan JA, Moss DK, et al., 2012, Design and Synthesis of Inhibitors of Plasmodium falciparum N-Myristoyltransferase, A Promising Target for Antimalarial Drug Discovery, Journal of Medicinal Chemistry, Vol: 55, Pages: 8879-8890, ISSN: 0022-2623
Design of inhibitors for N-myristoyltransferase (NMT), an enzyme responsible for protein trafficking in Plasmodium falciparum, the most lethal species of parasites that cause malaria, is described. Chemistry-driven optimization of compound 1 from a focused NMT inhibitor library led to the identification of two early lead compounds 4 and 25, which showed good enzyme and cellular potency and excellent selectivity over human NMT. These molecules provide a valuable starting point for further development.
Bell AS, Mills JE, Williams GP, et al., 2012, Selective Inhibitors of Protozoan Protein N-myristoyltransferases as Starting Points for Tropical Disease Medicinal Chemistry Programs, PLOS Neglected Tropical Diseases, Vol: 6, ISSN: 1935-2735
Inhibition of N-myristoyltransferase has been validated pre-clinically as a target for the treatment of fungal andtrypanosome infections, using species-specific inhibitors. In order to identify inhibitors of protozoan NMTs, we chose toscreen a diverse subset of the Pfizer corporate collection against Plasmodium falciparum and Leishmania donovani NMTs.Primary screening hits against either enzyme were tested for selectivity over both human NMT isoforms (Hs1 and Hs2) andfor broad-spectrum anti-protozoan activity against the NMT from Trypanosoma brucei. Analysis of the screening results hasshown that structure-activity relationships (SAR) for Leishmania NMT are divergent from all other NMTs tested, a finding notpredicted by sequence similarity calculations, resulting in the identification of four novel series of Leishmania-selective NMTinhibitors. We found a strong overlap between the SARs for Plasmodium NMT and both human NMTs, suggesting thatachieving an appropriate selectivity profile will be more challenging. However, we did discover two novel series withselectivity for Plasmodium NMT over the other NMT orthologues in this study, and an additional two structurally distinctseries with selectivity over Leishmania NMT. We believe that release of results from this study into the public domain willaccelerate the discovery of NMT inhibitors to treat malaria and leishmaniasis. Our screening initiative is another example ofhow a tripartite partnership involving pharmaceutical industries, academic institutions and governmental/nongovernmentalorganisations such as Medical Research Council and Wellcome Trust can stimulate research for neglecteddiseases
Goncalves V, Brannigan JA, Whalley D, et al., 2012, Discovery of Plasmodium vivax N-Myristoyltransferase Inhibitors: Screening, Synthesis, and Structural Characterization of their Binding Mode, Journal of Medicinal Chemistry, Vol: 55, Pages: 3578-3582, ISSN: 0022-2623
N-Myristoyltransferase (NMT) is a prospective drug target against parasitic protozoa. Herein we report the successful discovery of a series of Plasmodium vivax NMT inhibitors by high-throughput screening. A high-resolution crystal structure of the hit compound in complex with NMT was obtained, allowing understanding of its novel binding mode. A set of analogues was designed and tested to define the chemical groups relevant for activity and selectivity.
Maskuniitty M, Leatherbarrow R, Ying L, 2012, Fluorescence measurements of catalysis by foot-and-mouth disease virus 3C protease, 11th International Biorelated Polymer Symposium / 243rd National Spring Meeting of the American-Chemical-Society (ACS), Publisher: AMER CHEMICAL SOC, ISSN: 0065-7727
Bradshaw RT, Aronica PGA, Tate EW, et al., 2012, Developing mutational locally enhanced sampling (MULES) for predicting relative binding free energies at protein-protein interfaces, 11th International Biorelated Polymer Symposium / 243rd National Spring Meeting of the American-Chemical-Society (ACS), Publisher: AMER CHEMICAL SOC, ISSN: 0065-7727
Offermann DA, McKendrick JE, Sejberg JJP, et al., 2012, Synthesis and Incorporation into Cyclic Peptides of Tolan Amino Acids and Their Hydrogenated Congeners: Construction of an Array of A-B-loop Mimetics of the Cε3 Domain of Human IgE., J Org Chem
The disruption of the human immunolobulin E-high affinity receptor I (IgE-FcεRI) protein-protein interaction (PPI) is a validated strategy for the development of anti asthma therapeutics. Here, we describe the synthesis of an array of conformationally constrained cyclic peptides based on an epitope of the A-B loop within the Cε3 domain of IgE. The peptides contain various tolan (i.e., 1,2-biarylethyne) amino acids and their fully and partially hydrogenated congeners as conformational constraints. Modest antagonist activity (IC(50) ∼660 μM) is displayed by the peptide containing a 2,2'-tolan, which is the one predicted by molecular modeling to best mimic the conformation of the native A-B loop epitope in IgE.
Goncalves V, Brannigan JA, Thinon E, et al., 2012, A fluorescence-based assay for N-myristoyltransferase activity, Analytical Biochemistry, Vol: 421, Pages: 342-344, ISSN: 1096-0309
N-myristoylation is the irreversible attachment of a C14 fatty acid, myristic acid, to the N-terminal glycine of a protein via formation of an amide bond. This modification is catalyzed by myristoyl–coenzyme A (CoA):protein N-myristoyltransferase (NMT), an enzyme ubiquitous in eukaryotes that is up-regulated in several cancers. Here we report a sensitive fluorescence-based assay to study the enzymatic activity of human NMT1 and NMT2 based on detection of CoA by 7-diethylamino-3-(4-maleimido-phenyl)-4-methylcoumarin. We also describe expression and characterization of NMT1 and NMT2 and assay validation with small molecule inhibitors. This assay should be broadly applicable to NMTs from a range of organisms.
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