The enoyl-acyl carrier protein reductase InhA of M. tuberculosis is an attractive, validated target for anti-tuberculosis drug development. Moreover, direct inhibitors of InhA remain effective against InhA variants with mutations associated with isoniazid resistance, offering the potential for activity against MDR isolates. Here, structure based virtual screening supported by biological assays was applied to identify novel InhA inhibitors as potential antituberculosis agents. High-speed Glide SP docking was initially performed against two conformations of InhA differing in the orientation of the active site Tyr158. The resulting hits were filtered for drug-likeness based on Lipinski's rule and avoidance of PAINS-like properties, and finally subjected to Glide XP docking to improve accuracy. Sixteen compounds were identified and selected for in vitro biological assays, of which two (compounds 1 and 7) showed MIC of 12.5 and 25 µg/ml against M. tuberculosis H37Rv, respectively. Inhibition assays against purified recombinant InhA determined IC50 values for these compounds of 0.38 and 0.22 µM, respectively. A crystal structure of the most potent compound, compound 7, bound to InhA revealed the inhibitor to occupy a hydrophobic pocket implicated in binding the aliphatic portions of InhA substrates but distant from the NADH cofactor, i.e. in a site distinct from those occupied by the great majority of known InhA inhibitors. This compound provides an attractive starting template for ligand optimization aimed at discovery of new and effective compounds against M. tuberculosis that act by targeting InhA.
Mycobacterium tuberculosis DNA gyrase
manipulates the DNA topology using controlled breakage and religation
of DNA driven by ATP hydrolysis. DNA gyrase has been validated as
the enzyme target of fluoroquinolones (FQs), second-line antibiotics
used for the treatment of multidrug-resistant tuberculosis. Mutations
around the DNA gyrase DNA-binding site result in the emergence of
FQ resistance in M. tuberculosis; inhibition
of DNA gyrase ATPase activity is one strategy to overcome this. Here,
virtual screening, subsequently validated by biological assays, was
applied to select candidate inhibitors of the M. tuberculosis DNA gyrase ATPase activity from the Specs compound library (). Thirty compounds
were identified and selected as hits for in vitro biological assays,
of which two compounds, G24 and G26, inhibited
the growth of M. tuberculosis H37Rv
with a minimal inhibitory concentration of 12.5 μg/mL. The two
compounds inhibited DNA gyrase ATPase activity with IC50 values of 2.69 and 2.46 μM, respectively, suggesting this
to be the likely basis of their antitubercular activity. Models of
complexes of compounds G24 and G26 bound
to the M. tuberculosis DNA gyrase ATP-binding
site, generated by molecular dynamics simulations followed by pharmacophore
mapping analysis, showed hydrophobic interactions of inhibitor hydrophobic
headgroups and electrostatic and hydrogen bond interactions of the
polar tails, which are likely to be important for their inhibition.
Decreasing compound lipophilicity by increasing the polarity of these
tails then presents a likely route to improving the solubility and
activity. Thus, compounds G24 and G26 provide
attractive starting templates for the optimization of antitubercular
agents that act by targeting DNA gyrase.
The Mycobacterium tuberculosis protein kinase B (PknB) is critical for growth and survival of M. tuberculosis within the host. The series of aminopyrimidine derivatives show impressive activity against PknB (IC50 < .5 μM). However, most of them show weak or no cellular activity against M. tuberculosis (MIC > 63 μM). Consequently, the key structural features related to activity against of both PknB and M. tuberculosis need to be investigated. Here, two- and three-dimensional quantitative structure-activity relationship (2D and 3D QSAR) analyses combined with molecular dynamics (MD) simulations were employed with the aim to evaluate these key structural features of aminopyrimidine derivatives. Hologram quantitative structure-activity relationship (HQSAR) and CoMSIA models constructed from IC50 and MIC values of aminopyrimidine compounds could establish the structural requirements for better activity against of both PknB and M. tuberculosis. The NH linker and the R1 substituent of the template compound are not only crucial for the biological activity against PknB but also for the biological activity against M. tuberculosis. Moreover, the results obtained from MD simulations show that these moieties are the key fragments for binding of aminopyrimidine compounds in PknB. The combination of QSAR analysis and MD simulations helps us to provide a structural concept that could guide future design of PknB inhibitors with improved potency against both the purified enzyme and whole M. tuberculosis cells.
A series of diphenyl ether derivatives were developed and showed promising potency for inhibiting InhA, an essential enoyl acyl carrier protein reductase involved in mycolic acid biosynthesis, leading to the lysis of Mycobacterium tuberculosis. To understand the structural basis of diphenyl ether derivatives for designing more potent inhibitors, molecular dynamics (MD) simulations were performed. Based on the obtained results, the dynamic behaviour in terms of flexibility, binding free energy, binding energy decomposition, conformation, and the inhibitor-enzyme interaction of diphenyl ether inhibitors were elucidated. Phe149, Tyr158, Met161, Met199, Val203 and NAD+ are the key residues for binding of diphenyl ether inhibitors in the InhA binding pocket. Our results could provide the structural concept to design new diphenyl ether inhibitors with better enzyme inhibitory activity against M. tuberculosis InhA. The present work facilitates the design of new and potentially more effective anti-tuberculosis agents.
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