Diethylstilbestrol can effectively accelerate estradiol-17-O-glucuronidation, while potently inhibiting...

This in vitro study investigates the effects of diethylstilbestrol (DES), a widely used toxic synthetic estrogen, on estradiol-3- and 17-O- (E2-3/17-O) glucuronidation, via culturing human liver microsomes (HLMs) or recombinant UDP-glucuronosyltransferases (UGTs) with DES and E2. DES can potently inhibit E2-3-O-glucuronidation in HLM, a probe reaction for UGT1A1. Kinetic assays indicate that the inhibition follows a competitive inhibition mechanism, with the Ki value of 2.1±0.3μM, which is less than the possible in vivo level. In contrast to the inhibition on E2-3-O-glucuronidation, the acceleration is observed on E2-17-O-glucuronidation in HLM, in which cholestatic E2-17-O-glucuronide is generated. In the presence of DES (0-6.25μM), Km values for E2-17-O-glucuronidation are located in the range of 7.2-7.4μM, while Vmax values range from 0.38 to 1.54nmol/min/mg. The mechanism behind the activation in HLM is further demonstrated by the fact that DES can efficiently elevate the activity of UGT1A4 in catalyzing E2-17-O-glucuronidation. The presence of DES (2μM) can elevate Vmax from 0.016 to 0.81nmol/min/mg, while lifting Km in a much lesser extent from 4.4 to 11μM. Activation of E2-17-O-glucuronidation is well described by a two binding site model, with KA, α, and β values of 0.077±0.18μM, 3.3±1.1 and 104±56, respectively. However, diverse effects of DES towards E2-3/17-O-glucuronidation are not observed in liver microsomes from several common experimental animals. In summary, this study issues new potential toxic mechanisms for DES: potently inhibiting the activity of UGT1A1 and powerfully accelerating the formation of cholestatic E2-17-O-glucuronide by UGT1A4.