Estrogen replacement therapy has been correlated with an increased risk for developing breast and
endometrial cancers. One potential mechanism of
estrogen carcinogenesis involves metabolism of
estrogens to 2- and 4-hydroxylated
catechols, which are further oxidized to electrophilic/redox active o-
quinones that have the potential to both initiate and promote the carcinogenic process. Previously, we showed that the equine
estrogens,
equilin and
equilenin, which are major components of the
estrogen replacement formulation
Premarin (Wyeth-Ayerst), are primarily metabolized to the
catechol,
4-hydroxyequilenin. This
catechol was found to autoxidize to an o-
quinone causing oxidation and alkylation of
DNA in vitro and in vivo. To block
catechol formation from
equilenin, 4-halogenated
equilenin derivatives were synthesized. These derivatives were tested for their ability to bind to the
estrogen receptor, induce
estrogen sensitive genes, and their potential to form
catechol metabolites. We found that the 4-fluoro derivatives were more estrogenic than the 4-chloro and 4-bromo derivatives as demonstrated by a higher binding affinity for
estrogen receptors alpha and beta, an enhanced induction of
alkaline phosphatase activity in Ishikawa cells, pS2 expression in S30 cells, and PR expression in Ishikawa cells. Incubation of these compounds with
tyrosinase in the presence of GSH showed that the halogenated
equilenin compounds formed less
catechol GSH conjugates than the parent compounds,
equilenin and 17beta-hydroxyequilenin. In addition, these halogenated compounds showed less cytotoxicity in the presence of
tyrosinase than the parent compounds in S30 cells. Also, as stated above, the 4-fluoro derivatives showed similar
estrogenic effects as compared with parent compounds; however, they were less toxic in S30 cells as compared to
equilenin and 17beta-equilenin. Because 17beta-hydroxy-4-halogenated
equilenin derivatives showed higher
estrogenic effects than the halogenated
equilenin derivatives in vitro, we studied the relative ability of the 17beta-hydroxy-4-halogenated
equilenin derivatives to induce
estrogenic effects in the ovariectomized rat model. The 4-fluoro derivative showed higher activity than 4-chloro and 4-bromo derivatives as demonstrated by inducing higher vaginal cellular differentiation, uterine growth, and mammary gland branching. However, 17beta-hydroxy-4-fluoroequilenin showed a lower estrogenic activity than 17beta-hydroxyequilenin and
estradiol, which could be due to alternative pharmacokinetic properties for these compounds. These data suggest that the 4-fluoroequilenin derivatives have promise as alternatives to traditional
estrogen replacement therapy due to their similar estrogenic properties with less overall toxicity.