Norethisterone-4 beta,5-oxide and laevonorgestrel-4 beta,5-oxide: formation in rat liver microsomal incubations and interference with microsomal epoxide hydrolase and cytoplasmic glutathione S-transferase.

Abstract

Abstract When norethisterone is incubated with NADPH and liver microsomes from rats pretreated with phenobarbital or 3-methyl-cholanthrene, hydroxylated metabolites, ring-A-reduced metabolites and norethisterone-4 β,5-oxide are formed. Pretreatment with 3-methylcholanthrene especially favours formation of ring-A-reduced metabolites. Laevonorgestrel, when incubated under similar conditions was also metabolized to hydroxylated metabolites, to reduced metabolites, and to its epoxide, laevonorgestrel-4β,5-oxide. Under the conditions used the epoxides were quantitatively important metabolites (15–25% of ethyl acetate extractable metabolites) of either of the two steroids. Neither the parent steroids (norethisterone, laevonorgestrel) nor their epoxides (norethisterone-4β,5-oxide, laevonorgestrel-4β,5-oxide) were mutagenic in the bacterial system according to Ames. Norethisterone-4β,5-oxide inhibits purified rat liver microsomal epoxide hydrolase competitively with a K i of 1.9 × 10 −4 M. Laevonorgestrel-4β,5-oxide does not show this effect. Similar data were obtained with microsomes from human liver and from human placenta. Norethisterone-4β,5-oxide and laevonorgestrel-4β,5-oxide inhibit glutathione S-transferases in 100,000 g supernatant of homogenate. With purified glutathione S-transferase B the inhibition by norethisterone-4 β,5-oxide was competitive with respect to the substrate 2,4-dinitrochlorobenzene and of a mixed-type with respect to the cosubstrate glutathione. The data suggest that, at high doses of norethisterone, sufficient concentrations of the metabolite norethisterone-4β,5-oxide may be reached to inhibit enzymes that are involved in the control of reactive metabolites of environmental carcinogens.