Evidence from 32P-postlabeling and the use of pentachlorophenol for a novel metabolic activation pathway of diethylstilbestrol and its dimethyl ether in mouse live: likely alpha-hydroxylation of ethyl group(s) followed by sulfate conjugation.

Abstract

Diethylstilbestrol (DES), a synthetic stilbene estrogen, is a potent development toxin and carcinogen in humans and rodents. A number of 32P-postlabeling studies suggest that genotoxic effects of DES substantially contribute to these biological effects. The mechanisms involved in DES-mediated genotoxicity are not completely understood, however. As reported here, the structural resemblance of tamoxifen to DES led to the hypothesis that DES may be hydroxylated and sulfated at the allylic C2 and/or C5 of the ethyl side chains in analogy to alpha-hydroxylation and sulfation of and DNA adduct formation by tamoxifen. Female ICR mice were administered 500 mumol/kg DES or its dimethyl ether derivative (DiMeDES), either alone or in combination with the sulfotransferase inhibitor pentachlorophenol (PCP) (75 mumol/kg), once daily for 4 days. Liver DNA adducts were measured 24 h after the last dose by dinucleotide/monophosphate 32P-postlabeling. Administration of DES or DiMeDES led to the formation of a unique and novel pattern of several major DNA adducts which were absent in vehicle controls. With minor exceptions the pattern was qualitatively similar for the two compounds, suggesting rapid O-demethylation of DiMeDES to DES in vivo followed by metabolic activation. Adducts formed in vivo did not chromatographically match DES quinone adducts synthesized in vitro. Co-administration of PCP with DES or DiMeDES significantly decreased adduct formation from either compound, by 33-61%. Taken together, these results are consistent with a hitherto unrecognized pathway of metabolic activation and DNA adduct formation by DES involving the putative hydroxylation of the allylic alpha-carbon of the ethyl side chain(s), followed by formation of DNA-reactive sulfuric acid esters. DES is now known to induce DNA damage in vivo by at least four different mechanisms. It is postulated that this multiplicity of mechanisms in itself explains why this drug elicits such a plethora of unique and complex pathophysiological effects in adults and off-spring of different species.