Design, synthesis and evaluation of small molecule inhibitors of human N-myristoyltransferase
Author(s)
Shen, Mengjie
Type
Thesis
Abstract
N-myristoyltransferase (NMT) is an enzyme that catalyses the transfer of myristate to the N-terminal glycine of protein substrates. In human, two NMT proteins are expressed, playing crucial roles in protein membrane binding and targeting. They are involved in many diseases like cancer and common cold. This project aims to develop human NMT inhibitors as tools for NMT functional study and as potential treatments of N-myristoylation dependent diseases.
In subchapter 2.1, five previously developed inhibitors were re-purified and tested for enzyme activity by the CMP assay. All compounds show inhibitions against both human NMT1/2 with IC50s between 1.3-25 µM, but no inhibition against Lm NMT. The reason for the abnormal curves found in the CPM assay was systematically explored. Inhibitor-CPM adducts were identified and a likely side reaction mechanism was proposed.
In subchapter 2.2, ten inhibitors with a new linker were synthesized based on a developing series. The new linker is generally tolerant, brings better EC50/IC50 ratios and the best compound shows an IC50 of 0.31 µM. Crystallography structures of the best compound and its old-linker analogue were solved and used to explore SARs. Tagging assay showed dose-dependent labelling, providing direct evidence of target engagement.
In subchapter 2.3, a target repurposing approach was used to explore nematode NMT inhibitors from existing NMT inhibitors. Eight plasmodium or malaria NMT inhibitors were tested in the enzyme activity study and all showed inhibition. Their SARs were analysed using human NMT structures. Seven compounds were further tested for in vivo activities and one showed a higher potency than the positive control.
In Chapter 3, eleven ligand-protein structures of the imidazo[1,2-a]pyridin-6-phenoxy series and nine structures of azetidinopyrimidines series were solved by X-ray crystallography. Extensive binding modes analysis has revealed their key SARs. Multi-conformations are identified and investigated. Insights of crystal obtainment were summarized and discussed.
In subchapter 2.1, five previously developed inhibitors were re-purified and tested for enzyme activity by the CMP assay. All compounds show inhibitions against both human NMT1/2 with IC50s between 1.3-25 µM, but no inhibition against Lm NMT. The reason for the abnormal curves found in the CPM assay was systematically explored. Inhibitor-CPM adducts were identified and a likely side reaction mechanism was proposed.
In subchapter 2.2, ten inhibitors with a new linker were synthesized based on a developing series. The new linker is generally tolerant, brings better EC50/IC50 ratios and the best compound shows an IC50 of 0.31 µM. Crystallography structures of the best compound and its old-linker analogue were solved and used to explore SARs. Tagging assay showed dose-dependent labelling, providing direct evidence of target engagement.
In subchapter 2.3, a target repurposing approach was used to explore nematode NMT inhibitors from existing NMT inhibitors. Eight plasmodium or malaria NMT inhibitors were tested in the enzyme activity study and all showed inhibition. Their SARs were analysed using human NMT structures. Seven compounds were further tested for in vivo activities and one showed a higher potency than the positive control.
In Chapter 3, eleven ligand-protein structures of the imidazo[1,2-a]pyridin-6-phenoxy series and nine structures of azetidinopyrimidines series were solved by X-ray crystallography. Extensive binding modes analysis has revealed their key SARs. Multi-conformations are identified and investigated. Insights of crystal obtainment were summarized and discussed.
Version
Open Access
Date Issued
2018-12
Date Awarded
2019-07
Copyright Statement
Creative Commons Attribution NonCommercial NoDerivatives Licence
Advisor
Tate, Edward
Sponsor
Lee family scholarship
Publisher Department
Chemistry
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)