Cytosol from human breast cancer was used in saturation analysis over a wide range of [3H]-estradiol concentrations. Hydroxylapatite was used to measure specific ligand binding. This type of saturation analysis demonstrated the presence of a second, specific estrogen binding site, named type II estrogen binding site, in human breast cancer cytosol. Type II estrogen binding sites have a low affinity (Kd = 80 nM vs 0.31 nM) but a higher concentration (200 fmol/mg cytosol protein vs 30.3fmol/mg cytosol protein) than the classical cytoplasmic estrogen receptor. The presence of type II estrogen binding site in human breast cancer cytosol was confirmed by a "post-labeled" sucrose gradient method. Type II estrogen binding site was shown to bind only to estrogens and was found only in estrogen target tissue. Individual tumors were examined for the presence of type II estrogen binding sites and 23 25 (92%) contained significant amounts of these sites, including six which lacked estrogen receptor. The concentration of type II estrogen binding site was strongly correlated to the concentration of cytoplasmic estrogen receptor. We conclude that human breast cancer cytosol contains a specific, secondary estrogen binding site, similar to that previously described in the rat uterus. Knowledge about this type II estrogen binding site may provide us with additional information on the mechanisms of hormonally-dependent growth in breast cancer.
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