The Roles of CYP3A and CYP2B Isoforms in Hepatic Bioactivation and Detoxification of the Pyrrolizidine Alkaloid Senecionine in Sheep and Hamsters

The roles of cytochrome CYP3A and CYP2B isozymes in the bioactivation and detoxification of the pyrrolizidine alkaloid (PA) senecionine (SN) have been investigatedin vitrowith sheep and hamster hepatic microsomes. Our results show that the rate of SN activation measured by (±)-6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolizine (DHP) formation greatly exceeded the rate of SNN-oxide formation (detoxification) in hamsters. In contrast, SNN-oxide, a detoxification product, was the major metabolite in sheep with much lower DHP production. Immunoinhibition studies with anti-sheep CYP3A and CYP2B antibodies show that members of CYP3A subfamily play the major role in the conversion of PA to pyrrolic metabolites in both species (over 90% in sheep; 68% in hamster). These enzymes also contribute 38.8 and 41.3% of SN N-oxidation in sheep and hamsters, respectively. In contrast, CYP2B isoforms have a limited capacity toward DHP formation in both species (47% in sheep; 32% in hamster), while these enzymes catalyzed only 24.6 and 35.4% SN N-oxidation in sheep and hamster, respectively. Using triacetyloleandomycin (TAO) and gestodene, two highly selective chemical inhibitors of CYP3A isoforms, our data show that 90% of DHP formation was inhibited by either inhibitor in sheep. Gestodene appeared to be more efficient than TAO in the inhibition of DHP production in hamsters. Testosterone 6β-hydroxylase activity, a functional marker of CYP3A, was significantly inhibited by TAO and gestodene in sheep liver microsomes and by gestodene (100 μM) in hamster liver microsomes. These results suggest that CYP3A isozymes have important roles in bioactivation and detoxification of PA in both species, whereas CYP2B subfamily members are less efficient in biotransformation of PA.