Cellular mechanisms of growth inhibition of human epithelial ovarian cancer cell line by LH-releasing hormone antagonist Cetrorelix

We investigated the direct effects of LH-releasing hormone (LH-RH) antagonist, Cetrorelix, on the growth of HTOA human epithelial ovarian cancer cell line. RT-PCR revealed the expression of mRNA for LH-RH and its receptor in HTOA cells. Cetrorelix, at concentrations between 10^-9 and 10^-5 M, exerted a dose-dependent antiproliferative action on HTOA cells, as measured by 5-bromo-2′-deoxyuridine incorporation into DNA. Flow cytometric analysis indicated that Cetrorelix, at 10^-5 M, arrested cell cycle in HTOA cells, at G1 phase, after 24 h of treatment. Western blot analysis of cell cycle-regulatory proteins demonstrated that treatment with Cetrorelix (10^-5 M) for 24 h did not change the steady-state levels of cyclin D1, cyclin E, and cyclin-dependent kinase (Cdk)4 but decreased the levels of cyclin A and Cdk2. The protein levels of p21 (a Cdk inhibitor) and p53 (a suppressor of tumor cell growth and a positive regulator for p21 expression) were increased by Cetrorelix, but the levels of p27 (a Cdk inhibitor) did not change significantly. Flow cytometric analysis and terminal deoxynucleotidyltransferase-mediated deoxyuridine 5-triphosphate nick end labeling staining demonstrated that Cetrorelix (10^-5 M) induced apoptosis in HTOA cells. In conclusion, Cetrorelix directly inhibits the proliferation of human epithelial ovarian cancer cells through mechanisms mediated by LH-RH receptor and involving multiple events in cell cycle progression, including G1 phase cell cycle arrest coupled with down-regulation of cyclin A-Cdk2 complex levels, presumably attributable to an up-regulation of p53 and p21 protein levels and apoptosis.