CHARACTERIZATION OF HIGH-AFFINITY RECEPTORS FOR BOMBESIN/GASTRIN RELEASING PEPTIDE ON THE HUMAN PROSTATE-CANCER CELL-LINES PC-3 AND DU-145 - INTERNALIZATION OF RECEPTOR-BOUND (125)I-(TYR(4)) BOMBESIN BY TUMOR-CELLS

Specific receptors for bombesin/gastrin releasing peptide (GRP) on the androgen-independent human prostate cancer cell lines PC-3 and DU-145 were characterized. No specific binding of I-125-[Tyr(4)]-bombesin to the androgen-dependent human prostate cancer fell line LNCaP was detectable. The binding of I-125-[Tyr(4)]-bombesin to PC-3 and DU-145 cells was found to be time- and temperature-dependent, saturable, and reversible. Scatchard analysis revealed a single class of binding sites with high affinity (K-d 9.8 x 10(-11) M for PC-3, and 9.1 x 10(-11) M for DU-145 cells at 25 degrees C) and with a binding capacity of 44,000 binding sites/cell and 19,000 binding sites/cell, respectively. Bound I-125-[Tyr(4)]-bombesin was rapidly internalized by PC-3 cells. The nonhydrolyzable GTP analog GTP-gamma-S caused a dose-dependent inhibition of I-125-[Tyr(4)]-bombesin binding to PC-3 and DU-145 cells, indicating that a G-protein (guanine nucleotide-binding protein) couples the bombesin receptor to intracellular effector systems. Bombesin and GRP(14-27) inhibited the binding of I-125-[Tyr(4)]-bombesin to both cell lines in a dose-dependent manner with inhibition constants (K-i) of 0.5 nM and 0.4 nM, respectively. Both cell lines express the bombesin/GRP preferring bombesin receptor subtype, since, in displacement studies, neuromedin B was more than 200 times less potent than bombesin and GRP(14-27) in inhibiting the binding of I-125-[Tyr(4)]-bombesin. Two synthetic bombesin/GRP antagonists, RC-3095 and RC-3110, powerfully inhibited the specific binding of I-125-[Tyr4]-bombesin with K-i 0.92 nM and 0.26 nM on PC-3 cells, and 3.3 nM and 0.89 nM on DU-145 cells, respectively. These findings indicate that the PC-3 and DU-145 human prostate cancer cell lines possess specific high-affinity receptors for bombesin/GRP, and are suitable models for the evaluation of the antineoplastic activity of new bombesin/GRP antagonists in the treatment of androgen-independent prostate cancer. (C) 1994 Wiley-Liss, Inc.