Diethylstilboestrol: II, pharmacology, toxicology and carcinogenicity in experimental animals

The diethylstilbestrol (DES) metabolite (β-dienestrol), which had been identified in mouse, rat, monkey, and human urine, and two proposed metabolic intermediates (diethylstilbestrol α,α′-epoxide and α,α′-dihydroxy DES) were synthesized and their estrogenic activities determined. In addition, three DES analogs, α-dienestrol, DES-dihydroxy diethyl phenanthrene (DES-phenanthrene), and 1-(α-ethyl, 4α-hydroxyphenyl)indanyl-5-ol (indanyl-DES), were studied. Estrogenic activities of the compounds in vivo were determined by the immature mouse uterine weight bioassay; in vitro, their estradiol receptor binding activity (competitive equilibrium binding, sucrose gradient analysis, and association rate inhibition assays) was determined. Results of the mouse uterine weight bioassay gave the following order of estrogenicity: DES > α-dienestrol ≥ DES-epoxide > indanyl-DES > dihydroxy DES > β-dienestrol > DES-phenanthrene. Results of competitive equilibrium binding analyses of these compounds with estradiol-17β for the mouse uterine cytosol receptor followed the same order seen for the bioassay, except for indanyl-DES. DES, indanyl-DES, and α-dienestrol had the greatest affinities (Ka values approximately 0.5-19.1 × 1010 M-1), while DES-phenanthrene had the lowest (Ka = 3.5 × 107 M-1 ± 1.2). Sucrose gradient analysis of the above competition preparations illustrated the displacement of [3H]estradiol from the receptor peak. This displacement was receptor specific and concentration dependent and correlated with the equilibrium binding concentrations. In addition, the most hormonally active substances demonstrated the greatest rate inhibition in the estradiol cytosol receptor association rate reaction (V0). The rank order of estrogenicity of the compounds determined in this study should be useful in evaluating alternative metabolic pathways of DES as well as distinguishing biologically active metabolites from relatively inactive ones.

Estrogenic activity in vivo and in vitro of some diethylstilbestrol metabolites and analogs

This literature review presents available data on the metabolism of diethylstilbestrol (DES) to shed light on the fate and the mechanism of toxicity of this compound. Biotransformation effects reviewed include conjugation reactions of DES such as glucuronide formation in vivo and in vitro enterohepatic circulation of DES and its glucuronide and formation of steroid sulfatases and sulfates; oxidative metabolism of DES (aromatic hydroxylation followed by methylation); and species differences in DES metabolism. Excretion curves for 12 animal species show vast differences; whereas urinary excretion predominates in humans chimps rhesus monkeys and guinea pigs; rats hamsters and mice show predominately fecal elimination. The difference among species seems due to capability of biliary excretion. A molecular weight threshold seems to exist and this may account for the varying remnants of DES housed in different species organs. Placental transfer is another major problem. Fetuses of many species seem capable of glucurondizing DES. The formation of reactive metabolites (i.e. affinity for estradiol 17-beta receptor attachment and affinity for other proteins bound by estrogen) through oxidative biotransformation suggests that DES metabolites affect hepatic systems and may activate or transform cells to malignancy. Theories of the organotropism of DES carcinogenicity are presented as well as a discussion of the fate of DES in the environment.

The metabolism of diethylstilbestrol.

Natural and xenobiotic compounds having sex-related actions have long been used for growth promotion and various changes in carcass quality in meat animals. The first compounds used were synthetic estrogens; however, later on a whole battery of compounds having androgenic, and progestogenic actions have also been involved. In surveying the effects of these compounds in meat-producing animals, it became clear that these drugs increase the growth rate of the treated animals and bring about changes in the carcass that are generally characterized by lower fat content and more lean mass. Extensive studies undertaken in various countries, including the European Economic Community (EEC), have shown that if used according to good husbandry practices, the meat from treated animals does not have excessive amounts of residues compared with the endogenous amount of steroid production in the animals in question and also in human beings. The banning of these compounds in the European community brought a new phenomenon of illegal or black market cocktails. These mixtures of anabolic steroids are injected into the body of the animals rather than implanted in the ears, which is the normal practice in countries where they have not yet been banned. Several screening and confirmatory methods are now available for monitoring programs. However, these programs need excessive resources in terms of manpower, funds, and proper legislation, which in underdeveloped countries is questionable, particularly in the absence of strong scientific evidence for the exercise.

Natural sex steroids and their xenobiotic analogs in animal production: Growth, carcass quality, pharmacokinetics, metabolism, mode of action, residues, methods, and epidemiology

Diethylstilbestrol (DES) is considered a teratogen and a transplacental carcinogen in humans. In order to compare its biotransformation in nonhuman primates to that in humans, a metabolic study was carried out in rhesus monkeys and chimpanzees. After an oral dose of 1 mg/kg [14C]DES, approximately 59% of the ingested radioactivity was found in the urine and 28% in the feces of two female rhesus monkeys after 4 days, while in male rhesus monkeys urinary radioactivity accounted for 43% and fecal radioactivity for 35%. In chimpanzees, 63% of a 0.5 mg/kg dose was excreted with the urine in a female and 47% in a male animal. In both species, urinary radioactivity was predominantly (greater than 70%) associated with glucuronides. Besides DES, three metabolites were found in the urinary glucuronide fraction of rhesus monkeys and chimpanzees by radio gas chromatography and were identified as dienestrol and omega-hydroxy derivatives of DES and dienestrol. Fecal radioactivity in rhesus monkeys was shown to consist exclusively of DES. Glucuronidation of DES appears to occur in neonatal and fetal rhesus monkeys.

Biotransformation of diethylstilbestrol in the rhesus monkey and the chimpanzee.

Aerobic incubation of a misture of E[1,1,1-D3]-3,4;bis(p-hydroxyphenyl)-hex-3-ene admixed with an approximately equimolar amount of unlabeled diethylstilbestrol and [ 2-(14)C ] diethylstilbestrol with rat liver homogenates in the presence of NADPH yielded water-soluble metabolites as well as products more and less polar than the starting material Addition of 5-adenosyl-L-methionine increased the quantity of nonpolar metabolites Incubation with rat liver microsomes yielded similar results When polar metabolites from incubation with rat liver microsomes were incubated with catechol O-methyltransferase and S-adenosyl[methyl-3 H]-L-methionine there was conversion to 3H-labeled nonpolar products Examination of reaction products by means of gas chronatography-mass spectrometry gave evidence for the formation of a dihydroxydiethylstilbestrol, a dihydroxydienestrol, and a monomethoxydiethylstilbestrol

Merriam Gr

Papers

Metabolism of diethylstilbestrol by rat liver: a preliminary report.