7 alpha-iodine-125-iodo-5 alpha-dihydrotestosterone: a radiolabeled ligand for the androgen receptor

We describe the preparation of 7 alpha-[125I]iodo-5 alpha-dihydrotestosterone (7 alpha-[125I]IDHT) and its characterization as a ligand for the androgen receptor. We designed a route to prepare the radioiodine-labeled androgen on microscale through treatment of the 7 beta-tosylate of 7 beta-hydroxy-5 alpha-dihydrotestosterone-17 beta-p-nitro-benzoate with Na125I, followed by alkaline hydrolysis. The radiolabeled steroid was tested as a ligand for the androgen receptor in cytosol from MCF-7 cells, and for its in vivo tissue distribution in the rat. In addition, we tested 7 alpha-[125I]IDHT as a ligand in a novel assay for the detection and quantification of the ligand activated androgen receptor by in vitro autoradiography. The above synthetic route produced the 17 beta-p-nitrobenzoate of 7 alpha-[125I]IDHT in carrier-free form and in good yield. The 17 beta-ester was removed with alkali and the resulting 7 alpha-[125I]IDHT was purified by HPLC. 7 alpha-[125I]IDHT bound with high affinity, Kd = 0.26 nM, to the androgen receptor and showed low nonspecific binding. Since the ligand was carrier free and thus of very high specific activity, approximately 2,200 Ci/mmole, the sensitivity of the assay was much greater than with [3H]R1881, the classical androgen receptor ligand with which it was compared. When tested as a ligand for in vitro autoradiography, 7 alpha-[125I]IDHT produced excellent autoradiograms of the activated receptor with very low nonspecific binding and with only overnight exposure of the film. These studies demonstrate that 7 alpha-[125I]IDHT is an excellent ligand for the androgen receptor.