Contributed equally to this study
Characterization of an anandamide degradation system in prostate epithelial PC-3 cells: synthesis of new transporter inhibitors as tools for this study
Article first published online: 30 JAN 2009
2004 British Pharmacological Society
British Journal of Pharmacology
Volume 141, Issue 3, pages 457–467, February 2004
How to Cite
Ruiz-Llorente, L., Ortega-Gutiérrez, S., Viso, A., Sánchez, M. G., Sánchez, A. M., Fernández, C., Ramos, J. A., Hillard, C., Lasunción, M. A., López-Rodríguez, M. L. and Díaz-Laviada, I. (2004), Characterization of an anandamide degradation system in prostate epithelial PC-3 cells: synthesis of new transporter inhibitors as tools for this study. British Journal of Pharmacology, 141: 457–467. doi: 10.1038/sj.bjp.0705628
- Issue published online: 30 JAN 2009
- Article first published online: 30 JAN 2009
- (Received September 21, 2003, Revised October 15, 2003, Accepted November 11, 2003)
- Anandamide degradation;
- anandamide uptake inhibitors;
- prostate PC-3 cells;
- fatty acid amidohydrolase;
- endogenous cannabinoid system
The response of anandamide is terminated by a carrier-mediated transport followed by degradation catalyzed by the cloned enzyme fatty acid amidohydrolase (FAAH). In this study, we provide biochemical data showing an anandamide uptake process and the expression of FAAH in human prostate.
Anandamide was accumulated in PC-3 cells by a saturable and temperature-dependent process. Kinetic studies of anandamide uptake, determined in the presence of cannabinoid and vanilloid antagonists, revealed apparent parameters of KM=4.7±0.2 μM and Vmax=3.3±0.3 pmol min−1 (106 cells)−1.
The accumulation of anandamide was moderately inhibited by previously characterized anandamide transporter inhibitors (AM404, UCM707 and VDM11) but was unaffected by inhibitors of other lipid transport systems (phloretin or verapamil) and moderately affected by the FAAH inhibitor methyl arachidonyl fluorophosphonate.
The presence of FAAH in human prostate epithelial PC-3 cells was confirmed by analyzing its expression by Western blot and measuring FAAH activity.
To further study the structural requirements of the putative carrier, we synthesized a series of structurally different compounds 1–8 and evaluated their capacity as uptake inhibitors. They showed different inhibitory capacity in PC-3 cells, with (9Z,12Z)-N-(fur-3-ylmethyl)octadeca-9,12-dienamide (4, UCM119) being the most efficacious, with maximal inhibition and IC50 values of 49% and 11.3±0.5 μM, respectively.
In conclusion, PC-3 cells possess a complete inactivation system for anandamide formed by an uptake process and the enzyme FAAH. These results suggest a possible physiological function of anandamide in the prostate, reinforcing the role of endocannabinoid system as a neuroendocrine modulator.
British Journal of Pharmacology (2004) 141, 457–467. doi:10.1038/sj.bjp.0705628