Specific low affinity binding sites for gonadotropin-releasing hormone in human endometrial carcinomata

Abstract

Specific low-affinity high-capacity binding sites for gonadotropin-releasing hormone (GnRH) have recently been discovered in human breast and ovarian carcinomata. We checked whether similar binding sites are present in human endometrial cancer. Plasma membrane preparations were incubated with [ 125I, d-Ala 6-desGly 10]-GnRH-ethylamide in the presence or absence of unlabelled GnRH agonists or other peptides. GnRH-binding could be demonstrated in all 12 tumor samples tested. The mathematical analysis of the binding data was consistent with a single class of low affinity ( K a = (0.8−1.4) × 10 5 M −1) and high-capacity ( B max = (134–142) × 10 −12 M/mg membrane protein) binding sites. Native GnRH had a similar affinity to the binding sites as the GnRH agonist used. Other peptides such as oxytocin, somatostatin and thyrotropin-releasing hormone did not crossreact with the binding sites. A photolabelled derivative of [ d-Lys 6]-GnRH was prepared with the bifunctional photolabile reagent (4-azidobenzyl)- N-hydroxysuccinimide. Photoaffinity labelling of endometrial carcinoma membranes and subsequent sodium dodecyl sulfate electrophoresis in 10% polyacrylamide gel revealed the presence of a single molecular mass component of 62 ± 1.9 kDa. The appearance of this photolabelled binding site could be largely suppressed by the addition of unlabelled GnRH-agonist (10 −4 M) and thus represents the specific binding site for GnRH in endometrial cancer.