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In vitro assessment of time-dependent inhibitory effects on CYP2C8 and CYP3A activity by fourteen protein kinase inhibitors.

Drug metabolism and disposition: the biological fate of chemicals (2014-04-10)
Anne M Filppula, Pertti J Neuvonen, Janne T Backman
ABSTRACT

Previous studies have shown that several protein kinase inhibitors are time-dependent inhibitors of cytochrome P450 (CYP) 3A. We screened 14 kinase inhibitors for time-dependent inhibition of CYP2C8 and CYP3A. Amodiaquine N-deethylation and midazolam 1'-hydroxylation were used as marker reactions for CYP2C8 and CYP3A activity, respectively. A screening, IC50 shift, and mechanism-based inhibition were assessed with human liver microsomes. In the screening, bosutinib isomer 1, crizotinib, dasatinib, erlotinib, gefitinib, lestaurtinib, nilotinib, pazopanib, saracatinib, sorafenib, and sunitinib exhibited an increased inhibition of CYP3A after a 30-min preincubation with NADPH, as compared with no preincubation. Axitinib and vandetanib tested negative for time-dependent inhibition of CYP3A and CYP2C8, and bosutinib was the only inhibitor causing time-dependent inhibition of CYP2C8. The inhibitory mechanism by bosutinib was consistent with weak mechanism-based inhibition, and its inactivation variables, inhibitor concentration that supports half-maximal rate of inactivation (KI) and maximal inactivation rate (kinact), were 54.8 µM and 0.018 1/min. As several of the tested inhibitors were reported to cause mechanism-based inactivation of CYP3A4 during the progress of this work, detailed experiments with these were not completed. However, lestaurtinib and saracatinib were identified as mechanism-based inhibitors of CYP3A. The KI and kinact of lestaurtinib and saracatinib were 30.7 µM and 0.040 1/min, and 12.6 µM and 0.096 1/min, respectively. Inhibition of CYP2C8 by bosutinib was predicted to have no clinical relevance, whereas therapeutic lestaurtinib and saracatinib concentrations were predicted to increase the plasma exposure to CYP3A-dependent substrates by ≥2.7-fold. The liability of kinase inhibitors to affect CYP enzymes by time-dependent inhibition may have long-lasting consequences and result in clinically relevant drug-drug interactions.

MATERIALS
Product Number
Brand
Product Description

Supelco
Ammonium formate, eluent additive for LC-MS, LiChropur, ≥99.0%
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Ammonium formate, BioUltra, ≥99.0% (calc. based on dry substance, NT)
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Ketoconazole, 99.0-101.0% (EP, titration)
Supelco
Ketoconazole, Pharmaceutical Secondary Standard; Certified Reference Material
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Ammonium formate solution, 10 mM in H2O, suitable for HPLC
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Ketoconazole
USP
Ketoconazole, United States Pharmacopeia (USP) Reference Standard
Ketoconazole, European Pharmacopoeia (EP) Reference Standard

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