Selective Biotransformation of Taxol to 6α-Hydroxytaxol by Human Cytochrome P450 2C8

Abstract

The principal taxol biotransformation reaction of humans and of human hepatic in vitro preparations is 6α-hydroxylation of the taxane ring, but a separate, minor hydroxylation pathway (metabolite B formation) also exists. Taxol metabolism was studied using membrane fractions from Hep G₂ cells infected with recombinant vaccinia viruses that contain complementary DNAs encoding several human cytochrome P450 enzymes. Only P450 2C8 formed detectable 6α-hydroxytaxol. Metabolite B formation was catalyzed by complementary DNA-expressed 3A3 and 3A4, but not by 3A5. Each P450 3A preparation catalyzed felodipine oxidation. The apparent K_m and V_max values for taxol 6α-hydroxylation were 5.4 ± 1.0 µm and 30 ± 1.5 nmol/min/nmol P450, respectively, for complementary DNA-expressed P450 2C8; the values were 4.0 ± 1.0 µm and 0.87 ± 0.06 nmol/min/mg protein, respectively, for human hepatic microsomes. The inhibition of 6α-hydroxytaxol formation by quercetin was competitive with an apparent K_i of 1.3 or 1.1 µm with 2C8 or hepatic microsomes, respectively; retinoic acid was inhibitory, showing an apparent K_i of 27 µm with hepatic microsomes; inhibition by tolbutamide was complex, weak, and unlikely to be clinically relevant. The correlation between hepatic 2C8 protein content and 6α-hydroxytaxol formation was high (r² = 0.82), while the correlation with 2C9 content was low (r² = 0.38).

These data show that human biotransformation routes of taxol result from catalysis by specific enzymes of two P450 families and that taxol 6α-hydroxylation is a useful indicator of P450 2C8 activity in human hepatic microsomes.