A New Class of Cyp2c9 Inhibitors: Probing 2c9 Specificity With High-Affinity Benzbromarone Derivatives

Locuson, Charles W.; Wahlstrom, Jan L.; Rock, Denise A.; Rock, Dan A.; Jones, Jeffrey P.

doi:10.1124/dmd.31.7.967

Cited by 31 publications

(38 citation statements)

References 28 publications

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“…Arg108 was also reported to form ionic and hydrogen bonds with aspartic acid 293 (Asp293) and Asn289, respectively. 28) Moreover, the Arg= 108 residue was shown to be critical for substrate binding. 29) This suggests that mutations at position 241 can cause conformational changes in B'C-loop through Arg108, resulting in re-orientation of Phe100 and Phe114 and leading to alterations in drug binding.…”

Section: Discussionmentioning

confidence: 99%

Effect of Cytochrome P450 2C19 and 2C9 Amino Acid Residues 72 and 241 on Metabolism of Tricyclic Antidepressant Drugs

et al. 2014

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Although cytochromes P450 2C9 (CYP2C9) and 2C19 (CYP2C19) have 91% amino acid identity, they have different substrate specificities. Previous studies have suggested that several amino acid residues may be involved in substrate specificity. In this study, we focused on the roles of two amino acids, residues 72 and 241. The amino acids in these positions have opposite charges in CYP2C9 and 2C19; the former has lysines in both positions (Lys72 and Lys241), and the latter has glutamic acids (Glu72 and Glu241). Reciprocal mutants for both CYP2C19 and 2C9 were produced, and their metabolic activities and spectroscopic properties were examined using three tricyclic antidepressant (TCA) drugs: amitriptyline, imipramine, and dothiepin. Although CYP2C19 wild-type (WT) had a high metabolic activity for all three drugs, the E72K mutation decreased enzymatic activity by 29-37%, while binding affinities were diminished 2.5-to 20-fold. On the other hand, low activity and low affinity of CYP2C9 WT were recovered notably by K72E mutation. The metabolic activities and binding affinities were minimally affected by CYP2C19 E241K and CYP2C9 K241E mutations. We could also show linear correlations between metabolic activities and binding affinities, and hence we conclude that amino acid residue 72 plays a key role in TCA drug metabolism by limiting the binding affinities of CYP2C19 and CYP2C9.Key words cytochrome P450 2C19; cytochrome P450 2C9; drug metabolism; antidepressant; dissociation constant Depression is a psychiatric condition that affects 120 million people worldwide, and can interfere with independence and productivity in essentially all aspects of daily life. Depression is also associated with a high risk of self-harm, and ultimately suicide. Despite the considerable advances in depression treatments, tricyclic antidepressants (TCAs) remain an important therapeutic option for depression. TCAs such as amitriptyline (AMI), imipramine (IMI), and dothiepin (DOT) are heterocyclic chemical compounds (Fig. 1). These drugs are characterized by substantial pre-systemic first-pass metabolism and their clearance is dependent primarily on hepatic cytochrome P450 (CYP) oxidative enzymes, in particular CYP2C19.1) Because the activity of some CYP isoforms is individually influenced by genetic variations, there is wide inter-individual variability in TCA pharmacokinetics and active metabolite production, 2,3) which complicates the clinical use of these drugs. Therefore, the understanding of TCA drug metabolism at the molecular level is essential for the safe use of these drugs.CYPs are a superfamily of heme-containing enzymes that are mainly responsible for oxidative transformation of xenobiotics in humans, animals, and plants. The CYP1, 2, and 3 families in mammals have a wide range of overlapping substrates and play a role in the metabolism of both drug and endogenous substrates through N-, O-, and S-dealkylation, aromatic and aliphatic hydroxylation, epoxidation; oxidative desulfurization, and sulfoxidation. 4,5) The CYP2C subfamily in...

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Section: Discussionmentioning

confidence: 99%

Effect of Cytochrome P450 2C19 and 2C9 Amino Acid Residues 72 and 241 on Metabolism of Tricyclic Antidepressant Drugs

et al. 2014

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“…4B). The presence of Arg-108 in the active site of P450 2C9 is likely to contribute to the high binding affinity observed for benzbromarone and other ionized phenols (40). Mutagenesis studies indicate that substitution of alanine for Arg-108 of P450 2C9 dramatically reduces diclofenac 4-hydroxylase activity (41).…”

Section: Comparison Of the Four Molecules In The Asymmetric Unit-mentioning

confidence: 99%

Structural Characterization of Human Cytochrome P450 2C19

Reynald¹,

Sansen²,

Stout

et al. 2012

Journal of Biological Chemistry

114

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Background: Knowledge of the structural features of P450 2C19 that underlie its distinct roles in human drug metabolism is lacking. Results: The structure of P450 2C19 was determined by x-ray crystallography. Conclusion:The structure of the enzyme exhibits features that distinguish it from closely related P450s 2C8 and 2C9. Significance: Structural characterization of P450 2C19 contributes to a better understanding of its role in drug clearance.

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“…Interestingly, benzbromarone and its analogs are the most potent class of CYP2C9 inhibitors identified to date (data based on S-warfarin inhibition). 12 These data suggest that CYP2C9 may possess characteristics similar to CYP3A4 and that more detailed analysis of the active site may be necessary to fully understand CYP2C9 inhibition. Furthermore, ibuprofen and naproxen showed no inhibition of MFC metabolism.…”

Section: Resultsmentioning

confidence: 99%

Influence of Fluorescent Probe Size and Cytochrome b5 on Drug-Drug Interactions in CYP2C9

Hummel

Tracy

Hutzler³

et al. 2006

SLAS Discovery

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7-Methoxy-4-trifluoromethylcoumarin (MFC) has been used extensively in high-throughput screens for the identification of potential CYP2C9 interactions. More recently, additional probes from Invitrogen have been used. Vivid 2C9 Green is the largest of the probes and has had limited prior characterization. The new series of probes differ significantly from MFC and were examined for their ability to identify interactions with 19 CYP2C9 substrates/inhibitors. The inhibition profiles depend largely on the physical differences between the fluorescent probe substrates. Cytochrome b5 (cyt b5) was also investigated for the ability to alter the inhibition profile of a given compound. The stoichiometric addition of cyt b5 caused an increase in Vmax of MFC and Vivid 2C9 Green 4.4 and 1.7 times, respectively. Furthermore, cyt b5 imposes a steric component to the active site as the inhibition profiles were significantly affected in incubations with MFC. The addition of cyt b5 had limited impact on the inhibition profiles generated with Vivid 2C9 Green. The K m of Vivid 2C9 Green increased from 1.2 ± 0.2 µM to 4.8 ± 0.3 µM as a result of cyt b5 addition. These results illustrate that multiple substrate probes may be necessary for screening drug-drug interaction in CYP2C9 and that cyt b5 effects can impart steric restraints on the CYP2C9 active site. (Journal of Biomolecular Screening 2006:303-309)

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A New Class of Cyp2c9 Inhibitors: Probing 2c9 Specificity With High-Affinity Benzbromarone Derivatives

Cited by 31 publications

References 28 publications

Effect of Cytochrome P450 2C19 and 2C9 Amino Acid Residues 72 and 241 on Metabolism of Tricyclic Antidepressant Drugs

Effect of Cytochrome P450 2C19 and 2C9 Amino Acid Residues 72 and 241 on Metabolism of Tricyclic Antidepressant Drugs

Structural Characterization of Human Cytochrome P450 2C19

Influence of Fluorescent Probe Size and Cytochrome b5 on Drug-Drug Interactions in CYP2C9

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