Characterization of steroid receptors in human prostate using mibolerone

Accurate quantitation of androgen receptors requires a radioactive ligand which has affinity and specificity for the receptor and which is stable to metabolic enzymes. In this report, we have characterized the properties of 7 alpha,17 alpha-dimethyl-17 beta-hydroxy-4-estren-3-one (mibolerone) in human benign hyperplastic prostate cytosol and compared them to those of 17 beta-hydroxy-17 alpha-methyl-estra-4,9,11-trien-3-one (R1881). Mibolerone was found to have an affinity (Kd = 1.5 nM) greater than R1881. (Kd = 2.3 nM) for the androgen receptor in human prostate tissue. Surprisingly, mibolerone was found to bind with high affinity to the progesterone receptor in both human prostate (Kd = 5.9 nM) and rabbit uterus (Kd = 1.1 nM). However, binding to this receptor in both species could be blocked with a 500-fold excess of triamcinolone acetonide. [3H]Mibolerone binding to the androgen receptor was competed effectively with unlabeled dihydrotestosterone, R1881, and mibolerone but not by progesterone, diethylstilbestrol or R5020, in the presence of triamcinolone acetonide. Interestingly, mibolerone was more resistant to metabolism than R1881 in prostate cytosol when exposed to elevated temperatures (30 degrees C) for extended periods of time. However, when exposed to high-intensity ultraviolet irradiation, both compounds lost 50% of their binding ability in about 30 minutes. Mibolerone was found to have a very low affinity (Ki = 540 nM) for human sex steroid binding protein. These studies demonstrate that mibolerone is a useful ligand for androgen receptor assays. They also emphasize the need for including competitors of progesterone receptor binding in assays utilizing this steroid for androgen receptor measurements.