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Fragment-Based Approaches to the Development of Mycobacterium tuberculosis CYP121 Inhibitors.

Kavanagh ME, Coyne AG, McLean KJ, James GG, Levy CW, Marino LB, de Carvalho LP, Chan DS, Hudson SA, Surade S, Leys D, Munro AW, Abell C - J. Med. Chem. (2016)

Bottom Line: Synthetic merging and optimization of 1 produced a 100-fold improvement in binding affinity, yielding lead compound 2 (KD = 15 μM).Structure-guided addition of a metal-binding pharmacophore onto LE retrofragment scaffolds produced low nanomolar (KD = 15 nM) CYP121 ligands.Analysis of the factors governing ligand potency and selectivity using X-ray crystallography, UV-vis spectroscopy, and native mass spectrometry provides insight for subsequent drug development.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.

ABSTRACT
The essential enzyme CYP121 is a target for drug development against antibiotic resistant strains of Mycobacterium tuberculosis. A triazol-1-yl phenol fragment 1 was identified to bind to CYP121 using a cascade of biophysical assays. Synthetic merging and optimization of 1 produced a 100-fold improvement in binding affinity, yielding lead compound 2 (KD = 15 μM). Deconstruction of 2 into its component retrofragments allowed the group efficiency of structural motifs to be assessed, the identification of more LE scaffolds for optimization and highlighted binding affinity hotspots. Structure-guided addition of a metal-binding pharmacophore onto LE retrofragment scaffolds produced low nanomolar (KD = 15 nM) CYP121 ligands. Elaboration of these compounds to target binding hotspots in the distal active site afforded compounds with excellent selectivity against human drug-metabolizing P450s. Analysis of the factors governing ligand potency and selectivity using X-ray crystallography, UV-vis spectroscopy, and native mass spectrometry provides insight for subsequent drug development.

No MeSH data available.


Related in: MedlinePlus

X-ray crystal structures of Ar3 analogues(a) 26a (PDB 5IBI) and (b) 26h (PDB 5IBH)in complex with CYP121. Both compounds closely recapitulated the bindingmode of lead 2. Ar3 substituents remained flexible, interactingonly with active site water molecules for 26a or formingnovel interactions with active site residues Ala75 and Asn74 for 26h. The omit Fo – Fc electron density maps of ligands 26a and 26h contoured to 3σ have been provided inSupporting Information Figure S5. Figuresprepared using PyMOL v1.7.4 (Schrödinger, LLC).
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fig8: X-ray crystal structures of Ar3 analogues(a) 26a (PDB 5IBI) and (b) 26h (PDB 5IBH)in complex with CYP121. Both compounds closely recapitulated the bindingmode of lead 2. Ar3 substituents remained flexible, interactingonly with active site water molecules for 26a or formingnovel interactions with active site residues Ala75 and Asn74 for 26h. The omit Fo – Fc electron density maps of ligands 26a and 26h contoured to 3σ have been provided inSupporting Information Figure S5. Figuresprepared using PyMOL v1.7.4 (Schrödinger, LLC).

Mentions: The only analogue in the Ar1 or Ar3 series of compounds toperturbthe Soret band was the 3(4)-imidazole-methylamine substituted compound 26h. The type II shift observed was small (Δλmax26h (100 μM) = +1.5 nm) compared tothat observed for the 3-aminophenyl substituted Ar2 analogues (Δλmax25a [100 μM] = +5.0 nm), which was consistentwith results observed for heteroaromatic nitrogen substituents duringour initial selection of a heme binding functional group for Ar2 analogues(Table 1). A 2.0 ÅX-ray crystal structure of 26h in complex with CYP121indicated that the compound must adopt multiple different bindingmodes, as the 3(4)-imidazole substituent of the crystallized compoundresided in the Ar3 pocket, distal to the heme cofactor (Figure 8b). These competing bindingmodes might account for the weak affinity of 26h (KD = 274 μM) compared to the structurallysimilar 3-thiophenyl 26c or 2-furanyl 26d methylamine analogues, which did not perturb the optical spectrumof CYP121 (Table 2).Owing to a lack of spectral perturbation, the binding affinity ofall other Ar1 and Ar3 analogues was assumed to result from bindinginteractions made with residues in the distal active site pocket andprovided support that these analogues retained a similar binding modeto lead 2.


Fragment-Based Approaches to the Development of Mycobacterium tuberculosis CYP121 Inhibitors.

Kavanagh ME, Coyne AG, McLean KJ, James GG, Levy CW, Marino LB, de Carvalho LP, Chan DS, Hudson SA, Surade S, Leys D, Munro AW, Abell C - J. Med. Chem. (2016)

X-ray crystal structures of Ar3 analogues(a) 26a (PDB 5IBI) and (b) 26h (PDB 5IBH)in complex with CYP121. Both compounds closely recapitulated the bindingmode of lead 2. Ar3 substituents remained flexible, interactingonly with active site water molecules for 26a or formingnovel interactions with active site residues Ala75 and Asn74 for 26h. The omit Fo – Fc electron density maps of ligands 26a and 26h contoured to 3σ have been provided inSupporting Information Figure S5. Figuresprepared using PyMOL v1.7.4 (Schrödinger, LLC).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835159&req=5

fig8: X-ray crystal structures of Ar3 analogues(a) 26a (PDB 5IBI) and (b) 26h (PDB 5IBH)in complex with CYP121. Both compounds closely recapitulated the bindingmode of lead 2. Ar3 substituents remained flexible, interactingonly with active site water molecules for 26a or formingnovel interactions with active site residues Ala75 and Asn74 for 26h. The omit Fo – Fc electron density maps of ligands 26a and 26h contoured to 3σ have been provided inSupporting Information Figure S5. Figuresprepared using PyMOL v1.7.4 (Schrödinger, LLC).
Mentions: The only analogue in the Ar1 or Ar3 series of compounds toperturbthe Soret band was the 3(4)-imidazole-methylamine substituted compound 26h. The type II shift observed was small (Δλmax26h (100 μM) = +1.5 nm) compared tothat observed for the 3-aminophenyl substituted Ar2 analogues (Δλmax25a [100 μM] = +5.0 nm), which was consistentwith results observed for heteroaromatic nitrogen substituents duringour initial selection of a heme binding functional group for Ar2 analogues(Table 1). A 2.0 ÅX-ray crystal structure of 26h in complex with CYP121indicated that the compound must adopt multiple different bindingmodes, as the 3(4)-imidazole substituent of the crystallized compoundresided in the Ar3 pocket, distal to the heme cofactor (Figure 8b). These competing bindingmodes might account for the weak affinity of 26h (KD = 274 μM) compared to the structurallysimilar 3-thiophenyl 26c or 2-furanyl 26d methylamine analogues, which did not perturb the optical spectrumof CYP121 (Table 2).Owing to a lack of spectral perturbation, the binding affinity ofall other Ar1 and Ar3 analogues was assumed to result from bindinginteractions made with residues in the distal active site pocket andprovided support that these analogues retained a similar binding modeto lead 2.

Bottom Line: Synthetic merging and optimization of 1 produced a 100-fold improvement in binding affinity, yielding lead compound 2 (KD = 15 μM).Structure-guided addition of a metal-binding pharmacophore onto LE retrofragment scaffolds produced low nanomolar (KD = 15 nM) CYP121 ligands.Analysis of the factors governing ligand potency and selectivity using X-ray crystallography, UV-vis spectroscopy, and native mass spectrometry provides insight for subsequent drug development.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Cambridge , Lensfield Road, Cambridge CB2 1EW, U.K.

ABSTRACT
The essential enzyme CYP121 is a target for drug development against antibiotic resistant strains of Mycobacterium tuberculosis. A triazol-1-yl phenol fragment 1 was identified to bind to CYP121 using a cascade of biophysical assays. Synthetic merging and optimization of 1 produced a 100-fold improvement in binding affinity, yielding lead compound 2 (KD = 15 μM). Deconstruction of 2 into its component retrofragments allowed the group efficiency of structural motifs to be assessed, the identification of more LE scaffolds for optimization and highlighted binding affinity hotspots. Structure-guided addition of a metal-binding pharmacophore onto LE retrofragment scaffolds produced low nanomolar (KD = 15 nM) CYP121 ligands. Elaboration of these compounds to target binding hotspots in the distal active site afforded compounds with excellent selectivity against human drug-metabolizing P450s. Analysis of the factors governing ligand potency and selectivity using X-ray crystallography, UV-vis spectroscopy, and native mass spectrometry provides insight for subsequent drug development.

No MeSH data available.


Related in: MedlinePlus