It has been suggested that binding of 11β-chloromethyl estradiol (11β-CME2) to the estrogen receptor is irreversible, since its complex with receptor fails to undergo exchange with estradiol (E2). To investigate this behavior directly, 11β-CME2 was prepared in high specific activity, tritium-labeled form: The binding of [3H]11β-CME2, to the estrogen receptor from lamb and rat uterus and MCF-7 human breast cancer cells was shown to be fully reversible; the 11β-CME2 complex with receptor, as well as that of a structural analog 11β-ethyl estradiol, however, do not dissociate or exchange with [3h]e2over a 22 h period at 25°C. By competitive or direct binding assays, the affinity of 11β-CME2for the estrogen receptor can be estimated to be as much as 10- to 30-fold higher than that of E2. The complexes of estrogen receptor from MCF-7 cells with [3H]11β-CME2 and [3H]E2 show identical velocity sedimentation profiles on sucrose gradients, under conditions when the receptor is either a monomer of a dimer. Because of its very high affinity and unusual dissociation kinetics, [3H]11β-CME2 should be a very useful ligand for studies of estrogen receptor dynamics and in the assay of estrogen receptor concentrations in tumors and tissues.
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