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Interaction of the N-(3-Methylpyridin-2-yl)amide Derivatives of Flurbiprofen and Ibuprofen with FAAH: Enantiomeric Selectivity and Binding Mode.

Karlsson J, Morgillo CM, Deplano A, Smaldone G, Pedone E, Luque FJ, Svensson M, Novellino E, Congiu C, Onnis V, Catalanotti B, Fowler CJ - PLoS ONE (2015)

Bottom Line: The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH.The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH.This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Clinical Neuroscience, Pharmacology Unit, Umeå University, Umeå, Sweden.

ABSTRACT

Background: Combined fatty acid amide hydrolase (FAAH) and cyclooxygenase (COX) inhibition is a promising approach for pain-relief. The Flu-AM1 and Ibu-AM5 derivatives of flurbiprofen and ibuprofen retain similar COX-inhibitory properties and are more potent inhibitors of FAAH than the parent compounds. However, little is known as to the nature of their interaction with FAAH, or to the importance of their chirality. This has been explored here.

Methodology/principal findings: FAAH inhibitory activity was measured in rat brain homogenates and in lysates expressing either wild-type or FAAH(T488A)-mutated enzyme. Molecular modelling was undertaken using both docking and molecular dynamics. The (R)- and (S)-enantiomers of Flu-AM1 inhibited rat FAAH with similar potencies (IC50 values of 0.74 and 0.99 μM, respectively), whereas the (S)-enantiomer of Ibu-AM5 (IC50 0.59 μM) was more potent than the (R)-enantiomer (IC50 5.7 μM). Multiple inhibition experiments indicated that both (R)-Flu-AM1 and (S)-Ibu-AM5 inhibited FAAH in a manner mutually exclusive to carprofen. Computational studies indicated that the binding site for the Flu-AM1 and Ibu-AM5 enantiomers was located between the acyl chain binding channel and the membrane access channel, in a site overlapping the carprofen binding site, and showed a binding mode in line with that proposed for carprofen and other non-covalent ligands. The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH.

Conclusions/significance: The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH. This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.

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Representation of (R)-Flu-AM1 (A) and (S)-Flu-AM1 (B) in the competitive binding site of the dimeric FAAH as obtained from MD simulations.
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pone.0142711.g006: Representation of (R)-Flu-AM1 (A) and (S)-Flu-AM1 (B) in the competitive binding site of the dimeric FAAH as obtained from MD simulations.

Mentions: Three out of the four ligand-bound systems run for (R)-Flu-AM1 converged to a common binding mode, which is characterized by a stable hydrogen bond between the hydroxyl group of Thr488 and the carbonyl unit of (R)-Flu-AM1 (Fig 6A), as noted in the similar arrangement obtained upon superposition of representative snapshots (see S3 Fig). The ligand is closely packed in the binding site, forming interactions that are preserved along most of the trajectory (Fig 6A). The biphenyl moiety fills a hydrophobic cavity, with the distal ring pointing to the centre of the anionic hole and forming van der Waals contacts with hydrophobic residues lining the ACB channel (Ile491, Phe381, Leu380, Ile238 and Phe194). No specific interactions were observed for the methyl group on the chiral centre. On the other side, transient hydrogen bonds were observed between the pyridine nitrogen and either the backbone of Asp403 or alternatively Gly485 (through a water molecule), and between the amide NH unit and the backbone of Leu401 (S3 Fig). Van der Waals contacts are also formed with Met436 and Ile407. The structural integrity of this binding mode was maintained upon extension of the trajectory up to 100 ns (data not shown).


Interaction of the N-(3-Methylpyridin-2-yl)amide Derivatives of Flurbiprofen and Ibuprofen with FAAH: Enantiomeric Selectivity and Binding Mode.

Karlsson J, Morgillo CM, Deplano A, Smaldone G, Pedone E, Luque FJ, Svensson M, Novellino E, Congiu C, Onnis V, Catalanotti B, Fowler CJ - PLoS ONE (2015)

Representation of (R)-Flu-AM1 (A) and (S)-Flu-AM1 (B) in the competitive binding site of the dimeric FAAH as obtained from MD simulations.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4643906&req=5

pone.0142711.g006: Representation of (R)-Flu-AM1 (A) and (S)-Flu-AM1 (B) in the competitive binding site of the dimeric FAAH as obtained from MD simulations.
Mentions: Three out of the four ligand-bound systems run for (R)-Flu-AM1 converged to a common binding mode, which is characterized by a stable hydrogen bond between the hydroxyl group of Thr488 and the carbonyl unit of (R)-Flu-AM1 (Fig 6A), as noted in the similar arrangement obtained upon superposition of representative snapshots (see S3 Fig). The ligand is closely packed in the binding site, forming interactions that are preserved along most of the trajectory (Fig 6A). The biphenyl moiety fills a hydrophobic cavity, with the distal ring pointing to the centre of the anionic hole and forming van der Waals contacts with hydrophobic residues lining the ACB channel (Ile491, Phe381, Leu380, Ile238 and Phe194). No specific interactions were observed for the methyl group on the chiral centre. On the other side, transient hydrogen bonds were observed between the pyridine nitrogen and either the backbone of Asp403 or alternatively Gly485 (through a water molecule), and between the amide NH unit and the backbone of Leu401 (S3 Fig). Van der Waals contacts are also formed with Met436 and Ile407. The structural integrity of this binding mode was maintained upon extension of the trajectory up to 100 ns (data not shown).

Bottom Line: The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH.The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH.This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Clinical Neuroscience, Pharmacology Unit, Umeå University, Umeå, Sweden.

ABSTRACT

Background: Combined fatty acid amide hydrolase (FAAH) and cyclooxygenase (COX) inhibition is a promising approach for pain-relief. The Flu-AM1 and Ibu-AM5 derivatives of flurbiprofen and ibuprofen retain similar COX-inhibitory properties and are more potent inhibitors of FAAH than the parent compounds. However, little is known as to the nature of their interaction with FAAH, or to the importance of their chirality. This has been explored here.

Methodology/principal findings: FAAH inhibitory activity was measured in rat brain homogenates and in lysates expressing either wild-type or FAAH(T488A)-mutated enzyme. Molecular modelling was undertaken using both docking and molecular dynamics. The (R)- and (S)-enantiomers of Flu-AM1 inhibited rat FAAH with similar potencies (IC50 values of 0.74 and 0.99 μM, respectively), whereas the (S)-enantiomer of Ibu-AM5 (IC50 0.59 μM) was more potent than the (R)-enantiomer (IC50 5.7 μM). Multiple inhibition experiments indicated that both (R)-Flu-AM1 and (S)-Ibu-AM5 inhibited FAAH in a manner mutually exclusive to carprofen. Computational studies indicated that the binding site for the Flu-AM1 and Ibu-AM5 enantiomers was located between the acyl chain binding channel and the membrane access channel, in a site overlapping the carprofen binding site, and showed a binding mode in line with that proposed for carprofen and other non-covalent ligands. The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH.

Conclusions/significance: The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH. This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.

Show MeSH