Interaction of the radiolabelled high-affinity anti-oestrogen [3H]H1285 with the cytoplasmic oestrogen receptor

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Abstract
The high-affinity triarylethylene anti-estrogen H1285 [4-(N,N-diethylaminoethoxy)-.beta.-ethyl-.alpha.-(p-hydroxyphenyl)-4''-methoxystilbene] was tritiated to high specific radioactivity (35 Ci/mmol). Competition experiments between [3H]H1285 and H1285 or estradiol demonstrated that both compounds would compete with [3H]H1285 for estrogen-specific binding sites in rate uterine cytosol. [3H]H1285 had at least 10 times the affinity for the receptor compared with estradiol at the 50% competition level. [3H]H1285 appeared to have at least twice the association rate for the estrogen receptor compared with [3H]estradiol. The dissociation half-life (t1/2) of specific binding of [3H]H1285 to estrogen receptors at 0.degree. C was about 220 h compared with a value of 60 h for [3H]estradiol. Because of the extremely slow dissociation of [3H]H1285 from the estrogen receptor, the sedimentation profiles of [3H]H1285-receptor complexes were compared with those of [3H]-estradiol-receptor complexes in the presence of 0.4 M-KCl on 5-20% sucrose density gradients. [3H]estradiol-receptor complexes had a major peak at 4.4 S with a smaller peak at 5.6 S, whereas with [3H]H1285-receptor complexes the 5.6S peak was always higher than the 4.4S peak. There was significant variation between the dissociation behavior at 20.degree. C of [3H]H1285-receptor complexes and [3H]estradiol-receptor complexes pre-activated at 25.degree. C for 30 min in the presence and in the absence of 10 mM-sodium molybdate. The dissociation t1/2 of [3H]estradiol-receptor complexes at 20.degree. C decreased from 1.5 h to 0.5 h when molybdate was present during heat treatment whereas the dissociation t1/2 for [3H]H1285-receptor complexes was 5 h for both conditions. There evidently are fundamental differences in the initial interaction of H1285 and estradiol with the estrogen receptor. [Clinical implications for breast tumor treatment are considered.].