Signaling and Antiproliferative Effects Mediated by GnRH Receptors After Expression in Breast Cancer Cells Using Recombinant Adenovirus

• Published in: Endocrinology (Philadelphia) 2001-11, Vol.142 (11), p.4663-4672
• Main Authors: Everest, Helen M, Hislop, James N, Harding, Tom, Uney, James B, Flynn, Andrea, Millar, Robert P, McArdle, Craig A
• Format: Article
• Language: English
• Subjects: Accumulation; Adenoviridae; Adenoviruses; Affinity; Agonists; Analogs; Antiproliferatives; Binding, Competitive; Breast cancer; Breast Neoplasms - pathology; Breast Neoplasms - physiopathology; Cell Division - physiology; Female; Gene Transfer Techniques; Gonadotropin-releasing hormone; Gonadotropin-Releasing Hormone - pharmacology; Humans; Infections; Inositol Phosphates - metabolism; Mitogen-Activated Protein Kinase 1 - metabolism; Multiplicity of infection; Phosphorylation; Phosphorylation - drug effects; Pituitary; Pituitary (anterior); Receptors; Receptors, LHRH - physiology; Recombination, Genetic; Signal Transduction - physiology; Stimulation; Thymidine; Thymidine - antagonists & inhibitors; Tumor Cells, Cultured
• ISSN: 0013-7227; 1945-7170
• DOI: 10.1210/endo.142.11.8503

GnRH receptors (GnRH-Rs) are found in human cancers, including those of the breast, and GnRH can inhibit the growth of cell lines derived from such cancers. Although pituitary and extrapituitary GnRH-R transcripts appear identical, their functional characteristics may differ. Most extrapituitary GnRH-Rs have low affinity for GnRH analogs and may not activate PLC or discriminate between agonists and antagonists in the same way as pituitary GnRH-Rs. Here we have assessed whether GnRH-Rs expressed exogenously in breast cancer cells differ from those in gonadotropes. We found no evidence for endogenous GnRH-Rs in MCF7 cells, but after infection with adenovirus expressing the GnRH-R (Ad GnRH-R) at a multiplicity of infection of 10 or greater, at least 80% expressed GnRH-Rs. These had high affinity (Kd for [125I]buserelin, 1.4 nm) and specificity (rank order of potency, buserelin>GnRH≫chicken GnRH-II) and mediated stimulation of [3H]IP accumulation. Increasing viral titer [from multiplicity of infection, 3–300] increased receptor number (10,000–225,000 sites/cell) and [3H]IP responses. GnRH stimulated ERK2 phosphorylation in Ad GnRH-R-infected cells, and this effect, like stimulation of [3H]IP accumulation, was blocked by GnRH-R antagonists. GnRH also inhibited[ 3H]thymidine incorporation into Ad GnRH-R-infected cells (but not control cells). This effect was mimicked by agonist analogs and inhibited by two antagonists. Thus, when exogenous GnRH-Rs are expressed at density comparable to that in gonadotropes, they are functionally indistinguishable from the endogenous GnRH-Rs in gonadotropes, and increasing expression of high affinity GnRH-Rs can dramatically enhance the direct antiproliferative effect of GnRH agonists on breast cancer cells.