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J Pharmacol Exp Ther. 2018 Jul 5. pii: jpet.118.249557. doi: 10.1124/jpet.118.249557. [Epub ahead of print]

A Specific Probe Substrate for Evaluation of CYP4A11 Activity in Human Tissue Microsomes and a Highly Selective CYP4A11 Inhibitor: Luciferin-4A and Epalrestat.

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Department of Pharmacy, Shinshu University Hospital;
Faculty of Pharmaceutical Sciences, Hokuriku University.
Graduate School of Medicine, Shinshu University.
Department of Pharmacy, Shinshu University Hospital.


The specificity of cytochrome P450 4A11 (CYP4A11) against luciferin-4A O-demethylation in human liver and renal microsomes (HLMs and HRMs) and selectivity of CYP4A11 inhibition by epalrestat were investigated. Kinetic analysis of luciferin-4A O-demethylation yielded Vmax and S50 values of 39.7 pmol/min/mg protein and 43.2 μM for HLMs (Hill coefficient 1.24) and 39.4 pmol/min/mg protein and 33.8 μM for HRMs (Hill coefficient 1.34), respectively. Among the selective CYP inhibitors tested, HET0016 (CYP4 inhibitor) exclusively inhibited luciferin-4A O-demethylation by HLMs and HRMs. Furthermore, anti-CYP4A11 antibody nearly abolished the activity of both tissue microsomes. Luciferin-4A O-demethylase activity of HLMs was significantly correlated with lauric acid ω-hydroxylase activity, a marker of CYP4A11 activity (r = 0.904, P < 0.0001). Next, effects of epalrestat on CYP-mediated drug oxidations were examined. Epalrestat showed the most potent inhibition against CYP4A11 (IC50 = 1.82 μM) among the 17 recombinant enzymes tested. The inhibitory effect of epalrestat on CYP4A11 was at least 10-fold stronger than those on CYP4F2, CYP4F3B, and CYP4F12. For known CYP4 inhibitors, in contrast, HET0016 inhibited the activities of CYP4A11 and CYP4F2 (IC50 = 0.0137 - 0.0182 μM); 17-octadecynoic acid reduced activities of CYP4A11, CYP4F2, CYP4F3B, and CYP4F12 to a similar extent (IC50 = 5.70 - 17.7 μM). Epalrestat selectively and effectively inhibited the CYP4A11 activity of HLMs (IC50 = 0.913 μM) and HRMs (IC50 = 0.659 μM). These results indicated that luciferin-4A O-demethylase activity is a good CYP4A11 marker of HLMs and HRMs, and that epalrestat is a more selective CYP4A11 inhibitor compared to known CYP4 inhibitors.


CYP inhibition; CYP4; drug metabolism

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