Mechanism-based inactivation of cytochrome P450 2C9 by tienilic acid and (+/-)-suprofen: a comparison of kinetics and probe substrate selection.

Article Details

Citation

Hutzler JM, Balogh LM, Zientek M, Kumar V, Tracy TS

Mechanism-based inactivation of cytochrome P450 2C9 by tienilic acid and (+/-)-suprofen: a comparison of kinetics and probe substrate selection.

Drug Metab Dispos. 2009 Jan;37(1):59-65. doi: 10.1124/dmd.108.023358. Epub 2008 Oct 6.

PubMed ID
18838506 [ View in PubMed
]
Abstract

In vitro experiments were conducted to compare k(inact), K(I) and inactivation efficiency (k(inact)/K(I)) of cytochrome P450 (P450) 2C9 by tienilic acid and (+/-)-suprofen using (S)-flurbiprofen, diclofenac, and (S)-warfarin as reporter substrates. Although the inactivation of P450 2C9 by tienilic acid when (S)-flurbiprofen and diclofenac were used as substrates was similar (efficiency of approximately 9 ml/min/micromol), the inactivation kinetics were characterized by a sigmoidal profile. (+/-)-Suprofen inactivation of (S)-flurbiprofen and diclofenac hydroxylation was also described by a sigmoidal profile, although inactivation was markedly less efficient (approximately 1 ml/min/micromol). In contrast, inactivation of P450 2C9-mediated (S)-warfarin 7-hydroxylation by tienilic acid and (+/-)-suprofen was best fit to a hyperbolic equation, where inactivation efficiency was moderately higher (10 ml/min/micromol) and approximately 3-fold higher (3 ml/min/micromol), respectively, relative to that of the other probe substrates, which argues for careful consideration of reporter substrate when mechanism-based inactivation of P450 2C9 is assessed in vitro. Further investigations into the increased inactivation seen with tienilic acid relative to that with (+/-)-suprofen revealed that tienilic acid is a higher affinity substrate with a spectral binding affinity constant (K(s)) of 2 microM and an in vitro half-life of 5 min compared with a K(s) of 21 microM and a 50 min in vitro half-life for (+/-)-suprofen. Lastly, a close analog of tienilic acid with the carboxylate functionality replaced by an oxirane ring was devoid of inactivation properties, which suggests that an ionic binding interaction with a positively charged residue in the P450 2C9 active site is critical for recognition and mechanism-based inactivation by these close structural analogs.

DrugBank Data that Cites this Article

Drug Enzymes
DrugEnzymeKindOrganismPharmacological ActionActions
SuprofenCytochrome P450 2C9ProteinHumans
Unknown
Substrate
Inhibitor
Inactivator
Details
Tienilic acidCytochrome P450 2C9ProteinHumans
Unknown
Inhibitor
Details