The Proton Pump Inhibitor, Omeprazole, but Not Lansoprazole or Pantoprazole, Is a Metabolism-Dependent Inhibitor of CYP2C19: Implications for Coadministration with Clopidogrel

Published:  27 July 2011

Ogilvie BW, Yerino P, Kazmi F, Buckley DB, Rostami-Hodjegan A, Paris BL, Toren P, Parkinson A
As a direct-acting inhibitor of CYP2C19 in vitro, lansoprazole is more potent than omeprazole and other proton pump inhibitors (PPIs), but lansoprazole does not cause clinically significant inhibition of CYP2C19 whereas omeprazole does. To investigate this apparent paradox, we evaluated omeprazole, esomeprazole, R-omeprazole, lansoprazole, and pantoprazole for their ability to function as direct-acting and metabolism-dependent inhibitors (MDIs) of CYP2C19 in pooled human liver microsomes (HLM) as well as in cryopreserved hepatocytes and recombinant CYP2C19. In HLM, all PPIs were found to be direct-acting inhibitors of CYP2C19 with IC50 values varying from 1.2 μM [lansoprazole; maximum plasma concentration (Cmax) = 2.2 μM] to 93 μM (pantoprazole; Cmax = 6.5 μM). In addition, we identified omeprazole, esomeprazole, R-omeprazole, and omeprazole sulfone as MDIs of CYP2C19 (they caused IC50 shifts after a 30-min preincubation with NADPH-fortified HLM of 4.2-, 10-, 2.5-, and 3.2-fold, respectively), whereas lansoprazole and pantoprazole were not MDIs (IC50 shifts < 1.5-fold). The metabolism-dependent inhibition of CYP2C19 by omeprazole and esomeprazole was not reversed by ultracentrifugation, suggesting that the inhibition was irreversible (or quasi-irreversible), whereas ultracentrifugation largely reversed such effects of R-omeprazole. Under various conditions, omeprazole inactivated CYP2C19 with KI (inhibitor concentration that supports half the maximal rate of inactivation) values of 1.7 to 9.1 μM and kinact (maximal rate of enzyme inactivation) values of 0.041 to 0.046 min−1. This study identified omeprazole, and esomeprazole, but not R-omeprazole, lansoprazole, or pantoprazole, as irreversible (or quasi-irreversible) MDIs of CYP2C19. These results have important implications for the mechanism of the clinical interaction reported between omeprazole and clopidogrel, as well as other CYP2C19 substrates.

Comprehensive Quantitative and Qualitative Liquid Chromatography–Radioisotope–Mass Spectrometry Analysis for Safety Testing of Tolbutamide Metabolites Without Standard Samples

Published:  02 June 2011

Zenzaburo Tozuka, Shinsuke Aoyama, Kohei Nozawa, Shoji Akita, Toshinari Oh-Hara, Yasuhisa Adachi, Shin-Ichi Ninomiya
ADME & Tox. Research Institute, Sekisui Medical Company, Ltd., Tokai, Ibaraki 319-1182, Japan

Liquid chromatography–radioisotope–mass spectrometry (LC–RI–MS) analysis was used to determine the structures of 12 (four previously unknown) 14C–tolbutamide (TB) metabolites in rat biological samples (plasma, urine, bile, feces, and microsomes). The four novel metabolites are ω-carboxy TB, hydroxyl TB (HTB)-O-glucuronide, TB-ortho or meta-glutathion, and tolylsulphoaminocarbo-glutathion. In rat plasma, after oral administration of 14C–TB at therapeutic dose (1 mg/kg) and microdose (1.67 µg/kg), the total RI and six metabolites [HTB, carboxy TB (CTB), M1: desbutyl TB, M2: ω-hydroxyl TB, M3: α-hydroxyl TB, and M4: ω-1-hydroxyl TB] were quantified by LC–RI–MS. The plasma concentrations were calculated using their response factors (MS–RI intensity ratio) without standard samples, and the area under the curve (AUC) of plasma concentration per time for evaluation of Safety Testing of Drug Metabolites (MIST) was calculated using the ratio of TB metabolites AUC/total RI AUC. The ratios were as follows: TB 94.5% and HTB 2.5% for the microdose (1.67 µg/kg) and TB 95.6%, HTB 0.96%, CTB 0.065%, M1 0.62%, M2 0.0035%, M3 0.077%, and M4 0.015% for the therapeutic dose (1 mg/kg). These values were less than 10% of the MIST criteria. © 2011 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 100:4024–4036, 2011

An evaluation of the dilution method for identifying metabolism-dependent inhibitors of cytochrome P450 enzymes

Published:  13 April 2011

Parkinson A, Kazmi F, Buckley DB, Yerino P, Paris BL, Holsapple J, Toren P, Otradovec SM, Ogilvie BW
Metabolism-dependent inhibition (MDI) of cytochrome P450 is usually assessed in vitro by examining whether the inhibitory potency of a drug candidate increases after a 30-min incubation with human liver microsomes (HLMs). To augment the IC50 shift, many researchers incorporate a dilution step whereby the samples, after being preincubated for 30 min with a high concentration of HLMs (with and without NADPH), are diluted before measuring P450 activity. In the present study, we show that the greater IC50 shift associated with the dilution method is a consequence of data processing. With the dilution method, IC50 values for direct-acting inhibitors vary with the dilution factor unless they are based on the final (postdilution) inhibitor concentration, whereas the IC50 values for MDIs vary with the dilution factor unless they are based on the initial (predilution) concentration. When the latter data are processed on the final inhibitor concentration, as is commonly done, the IC50 values for MDI (shifted IC50 values) decrease by the magnitude of the dilution factor. The lower shifted IC50 values are a consequence of data processing, not enhanced P450 inactivation. In fact, for many MDIs, increasing the concentration of HLMs actually leads to considerably less P450 inactivation because of inhibitor depletion and/or binding of the inhibitor to microsomes. A true increase in P450 inactivation and IC50 shift can be achieved by assessing MDI by a nondilution method and by decreasing the concentration of HLMs. These results have consequences for the conduct of MDI studies and the development of cut-off criteria.