Amino Acid Residues Affecting the Activities of Human Cytochrome P450 2C9 and 2C19

Niwa, Toshiyuki; Kageyama, Akira; Kishimoto, Kae; Yabusaki, Yoshiyasu; Ishibashi, Fumihide; Katagiri, Masatoshi

doi:10.1124/dmd.30.8.931

Cited by 17 publications

(17 citation statements)

References 22 publications

(38 reference statements)

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“…Interestingly, CYP2C19 and CYP2C9 have very distinctive substrate specificities, although they are the most highly conserved (91% amino acid sequence identity) among human CYP2C enzymes (Romkes et al 1991;Goldstein and de Morais 1994;Miners et al 2000;Tsao et al 2001). For example, His99 of CYP2C19 and Ile99 of CYP2C9 in SRS-1 have been speculated to be important residues for substrate binding (Ibeanu et al 1996;Niwa et al 2002). The amino acids of interest in this study, Arg329 and Ser51, are located in the J and A helices, respectively.…”

Section: Discussionmentioning

confidence: 97%

Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population

Hanioka¹,

Tsuneto²,

Saito³

et al. 2007

Xenobiotica

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Cytochrome P450 2C19 (CYP2C19) plays an important role in the metabolism of a wide range of therapeutic drugs and exhibits genetic polymorphism with interindividual differences in metabolic activity. We have previously described two CYP2C19 allelic variants, namely CYP2C19*18 and CYP2C19*19 with Arg329His/Ile331Val and Ser51Gly/Ile331Val substitutions, respectively. In order to investigate precisely the effect of amino acid substitutions on CYP2C19 function, CYP2C19 proteins of the wild-type (CYP2C19.1B having Ile331Val) and variants (CYP2C19.18 and CYP2C19.19) were heterologously expressed in yeast cells, and their S-mephenytoin 4'-hydroxylation activities were determined. The K(m) value of CYP2C19.19 for S-mephenytoin 4'-hydroxylation was significantly higher (3.0-fold) than that of CYP2C19.1B. Although no significant differences in V(max) values on the basis of microsomal and functional CYP protein levels were observed between CYP2C19.1B and CYP2C19.19, the V(max)/K(m) values of CYP2C19.19 were significantly reduced to 29-47% of CYP2C19.1B. By contrast, the K(m), V(max) or V(max)/K(m) values of CYP2C19.18 were similar to those of CYP2C19.1B. These results suggest that Ser51Gly substitution in CYP2C19.19 decreases the affinity toward S-mephenytoin of CYP2C19 enzyme, and imply that the genetic polymorphism of CYP2C19*19 also causes variations in the clinical response to drugs metabolized by CYP2C19.

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Section: Discussionmentioning

confidence: 97%

Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population

Hanioka¹,

Tsuneto²,

Saito³

et al. 2007

Xenobiotica

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“…11) Further, residues 237 and 286 were suggested to play important roles in (S)-mephenytoin 4′-hydroxylation, 12,14) and replacement of asparagine 286 with serine (N286S) enhanced tolbutamide p-methyl hydroxylase, diclofenac 4′-hydroxylase, and ibuprofen hydroxylase activities. 16) Moreover, the CYP2C19 KSN triple mutant (E241K/N286S/I289N) was reported to metabolize both (R)-and (S)-warfarin with a broadened regioselectivity.…”

Section: Discussionmentioning

confidence: 99%

“…# These authors contributed equally to this work. residues as key determinants for omeprazole 5-hydroxylase activity, 11,12) (S)-mephenytoin 4′-hydroxylation, 13,14) and binding and metabolism of warfarin, 15) diclofenac, and ibuprofen. 16,17) It is noteworthy that the role of CYP2C19 residues glutamic acid 72 (Glu72) and glutamic acid 241 (Glu241), and CYP2C9 residues lysine 72 (Lys72) and lysine 241 (Lys241) in metabolic activity and binding affinity for TCA drugs, as well as basic drugs, has not been investigated.…”

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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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“…We previously demonstrated that several human hepatic P450s, including CYP2C8, CYP2C9, and CYP2C19, exhibit aminopyrine Ndemethylation activity (16). Tolbutamide is a typical substrate of CYP2C9 in vitro and a probe drug used for in vivo drug interaction studies as recommended in the guidance by EMA (19) and by FDA (20); however, tolbutamide methylhydroxylation was catalyzed by not only CYP2C9 but also CYP2C19 (8). In the present study, CYP2C19 showed a higher Km and Vmax than that shown by CYP2C9 and the Vmax/Km value for CYP2C19 was twice that of CYP2C9, whereas methylhydroxylated tolbutamide catalyzed by CYP2C8 was not detected.…”

Section: Discussionmentioning

confidence: 99%

“…The CYP2C subfamily in human liver comprises three members: CYP2C8, CYP2C9, and CYP2C19. Although the three CYP2C enzymes share more than 82% amino acid sequence identity, they have rather distinct substrate specificities (6)(7)(8)(9).…”

Section: Introductionmentioning

confidence: 99%

Substrate Specificity of Human Cytochrome P450 (CYP) 2C Subfamily and Effect of Azole Antifungal Agents on CYP2C8

Niwa¹,

Imagawa²

2016

J Pharm Pharm Sci

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-PURPOSE: The metabolic activities of aminopyrine N-demethylation and tolbutamide methylhydroxylation by the human hepatic cytochrome P450 (P450 or CYP) 2C subfamily were compared and the effects of azole antifungal agent on the drug-metabolizing activity of CYP2C8 were investigated. METHODS: Aminopyrine N-demethylation and tolbutamide methylhydroxylation by CYP2C8, CYP2C9, and CYP2C19 were determined by the previous reported methods. The effects of five azole antifungal agents, fluconazole, itraconazole, ketoconazole, miconazole, and voriconazole, on the aminopyrine N-demethylation activity by CYP2C8 were investigated. RESULTS: With regard to aminopyrine N-demethylation, CYP2C19 had the lowest Michaelis constant (Km) and CYP2C8 had the highest maximal velocity (Vmax) among the CYP2C subfamily members. The Vmax/Km values for CYP2C8 were the highest, followed by CYP2C19. For tolbutamide methylhydroxylation, the Km and Vmax for CYP2C19 were three and six times higher than the corresponding values for CYP2C9, and the Vmax/Km value for CYP2C19 was twice that for CYP2C9, whereas hydroxylated tolbutamide formed by CYP2C8 was not detected. Fluconazole, itraconazole, and voriconazole at a concentration of 2 or 10 µM neither inhibited nor stimulated CYP2C8-mediated aminopyrine Ndemethylation activity at substrate concentrations around the Km (5 mM). However, ketoconazole and miconazole noncompetitively inhibited CYP2C8-mediated aminopyrine N-demethylation with the inhibitory constant values of 1.98 and 0.86 µM, respectively. CONCLUSION: These results suggest that ketoconazole and miconazole might inhibit CYP2C8 clinically.

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Amino Acid Residues Affecting the Activities of Human Cytochrome P450 2C9 and 2C19

Cited by 17 publications

References 22 publications

Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population

Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population

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

Substrate Specificity of Human Cytochrome P450 (CYP) 2C Subfamily and Effect of Azole Antifungal Agents on CYP2C8

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Amino Acid Residues Affecting the Activities of Human Cytochrome P450 2C9 and 2C19

Cited by 17 publications

References 22 publications

Functional characterization of two novel CYP2C19 variants (CYP2C19*18andCYP2C19*19) found in a Japanese population

Functional characterization of two novel CYP2C19 variants (CYP2C19*18andCYP2C19*19) found in a Japanese population

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

Substrate Specificity of Human Cytochrome P450 (CYP) 2C Subfamily and Effect of Azole Antifungal Agents on CYP2C8

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Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population

Functional characterization of two novel CYP2C19 variants (***CYP2C19*18***andCYP2C19*19) found in a Japanese population