Evidence for hormone-responsive cleavage of gonadotropin-releasing hormone by the plasma membrane of ovarian cancer cells.

Gonadotropin-releasing hormone (GnRH) produced by ovarian cancer cells acts as a negative autocrine regulator in the ovarian cancer cell proliferation. GnRH and its analogs retard the proliferation of ovarian cancers bearing GnRH receptor, and thus the GnRH hydrolysis may be responsible for their growth. We determined the principal mechanism of GnRH hydrolysis by ovarian cancer cells and its coupling to GnRH receptor. GnRH cleaving activity was assessed by measuring the degradation of exogenous [3H]GnRH labeled at the N-terminal residue and the appearance of labeled degradation product. Approximately 98% of GnRH hydrolysis by plasma membrane from ovarian cancers surgically removed was accounted for by the cleavage at Tyr5-Gly6 to yield (1-5)GnRH. The GnRH degrading activity in the plasma membrane was approximately 6-fold higher than those in homogenate fraction. The membrane GnRH cleaving activity was metal ion-independent and sensitive to inhibition by N-ethymaleimide and endopeptidase inhibitors; characteristics suggestive of membrane-bound endopeptidase activity. No effects of buserelin and leuprolide on the [3H]GnRH hydrolysis at 4-fold higher concentration of the [3H]GnRH substrate suggested great resistance of the GnRH analogs toward the endopeptidase. GnRH degrading activity increased by 1.5-fold when the ovarian cancer Caov-3 cells were exposed to buserelin in intact cells prior to assay of cell membranes. These data demonstrate that GnRH is hydrolysed in the plasma membrane of ovarian cancer cells by the action of endopeptidase and provide the initial evidence for the functional coupling of the GnRH degradation to GnRH receptor activation.