Pharmacoperone IN3 enhances the apoptotic effect of leuprolide in prostate cancer cells by increasing the gonadotropin-releasing hormone receptor in the cell membrane.
Gonadotropin-releasing hormone (GnRH) agonists are widely used for the treatment of advanced prostate cancer (PCa). Agonists activate the GnRH receptor (GnRH-R), triggering apoptosis in PCa cells. In gonadotropes, the amount of GnRH-R in the plasma membrane is regulated by protein folding and endoplasmic reticulum retention, mechanisms that can be overcome by the pharmacoperone IN3. Our aim was to describe the intracellular distribution of GnRH-R in PCa cells and its relation to response to GnRH analog treatments. The expressions of GnRH-R in PCa biopsies were evaluated by immunohistochemistry and the intracellular distribution was determined by immunofluorescence in primary cell cultures from human PCa samples. Cultured cells were pretreated with IN3 and then with leuprolide. Cell survival was evaluated by 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT) thiazolyl blue formazan and cell cycle and apoptosis by flow cytometry. We observed that the expression of GnRH-R decreased according to malignant progression. Most GnRH-R are located inside the cell, colocalizing with endoplasmic reticulum markers. The treatment with IN3 decreased cellular GnRH-R retention, increasing plasma membrane expression in approximately 60%. Pretreatment with IN3 decreased PCa cell survival compared with leuprolide-alone treatment, primarily because of an increase in apoptosis. We conclude that the response of PCa cells to leuprolide is related to the amount of GnRH-R in the plasma membrane. Therefore, pretreatment evaluation of the amount of these receptors may be a predictor of the outcome of leuprolide treatment in PCa patients. Assessment of systemic IN3 effect would be necessary to determine its utility as an adjuvant treatment in hormone-resistant tumors.
Laboratory of Molecular and Cellular Andrology, Physiology and Biophysics Program, ICBM, Faculty of Medicine, University of Chile, Independencia 1027, PO Box 70005, 838 0453 Santiago, Chile. csanchezn@clc.cl
Catherine A
CA
Sánchez
Alejandro J
AJ
Mercado
Héctor R
HR
Contreras
Juan C
JC
Cabezas
Christian C
CC
Huidobro
Enrique A
EA
Castellón
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology*; D018929:Cell Culture Techniques; D002453:Cell Cycle / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D004357:Drug Synergism; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007211:Indoles / Q000494:pharmacology*; D016729:Leuprolide / Q000494:pharmacology*; D008297:Male; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D017510:Protein Folding; D011725:Pyridines / Q000494:pharmacology*; D011966:Receptors, LHRH / Q000819:agonists* / Q000096:biosynthesis; D014407:Tumor Cells, Cultured
D019086:Bridged Bicyclo Compounds, Heterocyclic; C461300:IN3 compound; D007211:Indoles; D011725:Pyridines; D011966:Receptors, LHRH; D016729:Leuprolide