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. 2004 Apr 1;379(Pt 1):99–106. doi: 10.1042/BJ20031695

N-cyclohexanecarbonylpentadecylamine: a selective inhibitor of the acid amidase hydrolysing N-acylethanolamines, as a tool to distinguish acid amidase from fatty acid amide hydrolase.

Kazuhito Tsuboi 1, Christine Hilligsmann 1, Séverine Vandevoorde 1, Didier M Lambert 1, Natsuo Ueda 1
PMCID: PMC1224050  PMID: 14686878

Abstract

Anandamide ( N-arachidonoylethanolamine) and other bioactive N-acylethanolamines are degraded to their corresponding fatty acids and ethanolamine. This hydrolysis is mostly attributed to catalysis by FAAH (fatty acid amide hydrolase), which exhibits an alkaline pH optimum. In addition, we have identified another amidase which catalyses the same reaction exclusively at acidic pH values [Ueda, Yamanaka and Yamamoto (2001) J. Biol. Chem. 276, 35552-35557]. In attempts to find selective inhibitors of this acid amidase, we screened various derivatives of palmitic acid, 1-hexadecanol, and 1-pentadecylamine with N-palmitoylethanolamine as substrate. Here we show that N-cyclohexanecarbonylpentadecylamine inhibits the acid amidase from rat lung with an IC50 of 4.5 microM, without inhibiting FAAH at concentrations up to 100 microM. The inhibition was reversible and non-competitive. This compound also inhibited the acid amidase in intact alveolar macrophages. With the aid of this inhibitor, it was revealed that rat basophilic leukaemia cells possess the acid amidase as well as FAAH. Thus the inhibitor may be a useful tool to distinguish the acid amidase from FAAH in various tissues and cells and to elucidate the physiological role of the enzyme.

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Selected References

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  1. Bisogno T., Maurelli S., Melck D., De Petrocellis L., Di Marzo V. Biosynthesis, uptake, and degradation of anandamide and palmitoylethanolamide in leukocytes. J Biol Chem. 1997 Feb 7;272(6):3315–3323. doi: 10.1074/jbc.272.6.3315. [DOI] [PubMed] [Google Scholar]
  2. Bisogno Tiziana, De Petrocellis Luciano, Di Marzo Vincenzo. Fatty acid amide hydrolase, an enzyme with many bioactive substrates. Possible therapeutic implications. Curr Pharm Des. 2002;8(7):533–547. doi: 10.2174/1381612023395655. [DOI] [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Calignano A., La Rana G., Giuffrida A., Piomelli D. Control of pain initiation by endogenous cannabinoids. Nature. 1998 Jul 16;394(6690):277–281. doi: 10.1038/28393. [DOI] [PubMed] [Google Scholar]
  5. Clement Angela B., Hawkins E. Gregory, Lichtman Aron H., Cravatt Benjamin F. Increased seizure susceptibility and proconvulsant activity of anandamide in mice lacking fatty acid amide hydrolase. J Neurosci. 2003 May 1;23(9):3916–3923. doi: 10.1523/JNEUROSCI.23-09-03916.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cravatt B. F., Demarest K., Patricelli M. P., Bracey M. H., Giang D. K., Martin B. R., Lichtman A. H. Supersensitivity to anandamide and enhanced endogenous cannabinoid signaling in mice lacking fatty acid amide hydrolase. Proc Natl Acad Sci U S A. 2001 Jul 24;98(16):9371–9376. doi: 10.1073/pnas.161191698. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cravatt Benjamin F., Lichtman Aron H. The enzymatic inactivation of the fatty acid amide class of signaling lipids. Chem Phys Lipids. 2002 Dec 31;121(1-2):135–148. doi: 10.1016/s0009-3084(02)00147-0. [DOI] [PubMed] [Google Scholar]
  8. De Petrocellis L., Melck D., Ueda N., Maurelli S., Kurahashi Y., Yamamoto S., Marino G., Di Marzo V. Novel inhibitors of brain, neuronal, and basophilic anandamide amidohydrolase. Biochem Biophys Res Commun. 1997 Feb 3;231(1):82–88. doi: 10.1006/bbrc.1997.6000. [DOI] [PubMed] [Google Scholar]
  9. Deutsch D. G., Omeir R., Arreaza G., Salehani D., Prestwich G. D., Huang Z., Howlett A. Methyl arachidonyl fluorophosphonate: a potent irreversible inhibitor of anandamide amidase. Biochem Pharmacol. 1997 Feb 7;53(3):255–260. doi: 10.1016/s0006-2952(96)00830-1. [DOI] [PubMed] [Google Scholar]
  10. Deutsch D. G., Ueda N., Yamamoto S. The fatty acid amide hydrolase (FAAH). Prostaglandins Leukot Essent Fatty Acids. 2002 Feb-Mar;66(2-3):201–210. doi: 10.1054/plef.2001.0358. [DOI] [PubMed] [Google Scholar]
  11. Devane W. A., Hanus L., Breuer A., Pertwee R. G., Stevenson L. A., Griffin G., Gibson D., Mandelbaum A., Etinger A., Mechoulam R. Isolation and structure of a brain constituent that binds to the cannabinoid receptor. Science. 1992 Dec 18;258(5090):1946–1949. doi: 10.1126/science.1470919. [DOI] [PubMed] [Google Scholar]
  12. Di Marzo Vincenzo, De Petrocellis L., Fezza F., Ligresti A., Bisogno T. Anandamide receptors. Prostaglandins Leukot Essent Fatty Acids. 2002 Feb-Mar;66(2-3):377–391. doi: 10.1054/plef.2001.0349. [DOI] [PubMed] [Google Scholar]
  13. Fowler C. J., Jacobsson S. O. P. Cellular transport of anandamide, 2-arachidonoylglycerol and palmitoylethanolamide--targets for drug development? Prostaglandins Leukot Essent Fatty Acids. 2002 Feb-Mar;66(2-3):193–200. doi: 10.1054/plef.2001.0357. [DOI] [PubMed] [Google Scholar]
  14. Fowler C. J., Jonsson K. O., Tiger G. Fatty acid amide hydrolase: biochemistry, pharmacology, and therapeutic possibilities for an enzyme hydrolyzing anandamide, 2-arachidonoylglycerol, palmitoylethanolamide, and oleamide. Biochem Pharmacol. 2001 Sep 1;62(5):517–526. doi: 10.1016/s0006-2952(01)00712-2. [DOI] [PubMed] [Google Scholar]
  15. Fu Jin, Gaetani Silvana, Oveisi Fariba, Lo Verme Jesse, Serrano Antonia, Rodríguez De Fonseca Fernando, Rosengarth Anja, Luecke Hartmut, Di Giacomo Barbara, Tarzia Giorgio. Oleylethanolamide regulates feeding and body weight through activation of the nuclear receptor PPAR-alpha. Nature. 2003 Sep 4;425(6953):90–93. doi: 10.1038/nature01921. [DOI] [PubMed] [Google Scholar]
  16. Glaser Sherrye T., Abumrad Nada A., Fatade Folayan, Kaczocha Martin, Studholme Keith M., Deutsch Dale G. Evidence against the presence of an anandamide transporter. Proc Natl Acad Sci U S A. 2003 Mar 24;100(7):4269–4274. doi: 10.1073/pnas.0730816100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hansen H. S., Moesgaard B., Hansen H. H., Petersen G. N-Acylethanolamines and precursor phospholipids - relation to cell injury. Chem Phys Lipids. 2000 Nov;108(1-2):135–150. doi: 10.1016/s0009-3084(00)00192-4. [DOI] [PubMed] [Google Scholar]
  18. Hillard C. J., Jarrahian A. The movement of N-arachidonoylethanolamine (anandamide) across cellular membranes. Chem Phys Lipids. 2000 Nov;108(1-2):123–134. doi: 10.1016/s0009-3084(00)00191-2. [DOI] [PubMed] [Google Scholar]
  19. Jacobsson S. O., Fowler C. J. Characterization of palmitoylethanolamide transport in mouse Neuro-2a neuroblastoma and rat RBL-2H3 basophilic leukaemia cells: comparison with anandamide. Br J Pharmacol. 2001 Apr;132(8):1743–1754. doi: 10.1038/sj.bjp.0704029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lambert D. M., DiPaolo F. G., Sonveaux P., Kanyonyo M., Govaerts S. J., Hermans E., Bueb J., Delzenne N. M., Tschirhart E. J. Analogues and homologues of N-palmitoylethanolamide, a putative endogenous CB(2) cannabinoid, as potential ligands for the cannabinoid receptors. Biochim Biophys Acta. 1999 Sep 22;1440(2-3):266–274. doi: 10.1016/s1388-1981(99)00132-8. [DOI] [PubMed] [Google Scholar]
  21. Lambert Didier M., Vandevoorde Severine, Jonsson Kent-Olov, Fowler Christopher J. The palmitoylethanolamide family: a new class of anti-inflammatory agents? Curr Med Chem. 2002 Mar;9(6):663–674. doi: 10.2174/0929867023370707. [DOI] [PubMed] [Google Scholar]
  22. Lichtman Aron H., Hawkins E. Gregory, Griffin Graeme, Cravatt Benjamin F. Pharmacological activity of fatty acid amides is regulated, but not mediated, by fatty acid amide hydrolase in vivo. J Pharmacol Exp Ther. 2002 Jul;302(1):73–79. doi: 10.1124/jpet.302.1.73. [DOI] [PubMed] [Google Scholar]
  23. Maccarrone Mauro, Pauselli Riccardo, Di Rienzo Marianna, Finazzi-Agrò Alessandro. Binding, degradation and apoptotic activity of stearoylethanolamide in rat C6 glioma cells. Biochem J. 2002 Aug 15;366(Pt 1):137–144. doi: 10.1042/BJ20020438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Patricelli M. P., Cravatt B. F. Proteins regulating the biosynthesis and inactivation of neuromodulatory fatty acid amides. Vitam Horm. 2001;62:95–131. doi: 10.1016/s0083-6729(01)62002-8. [DOI] [PubMed] [Google Scholar]
  25. Pertwee R. G. Pharmacological, physiological and clinical implications of the discovery of cannabinoid receptors. Biochem Soc Trans. 1998 May;26(2):267–272. doi: 10.1042/bst0260267. [DOI] [PubMed] [Google Scholar]
  26. Rodríguez de Fonseca F., Navarro M., Gómez R., Escuredo L., Nava F., Fu J., Murillo-Rodríguez E., Giuffrida A., LoVerme J., Gaetani S. An anorexic lipid mediator regulated by feeding. Nature. 2001 Nov 8;414(6860):209–212. doi: 10.1038/35102582. [DOI] [PubMed] [Google Scholar]
  27. Schmid Harald H. O., Schmid Patricia C., Berdyshev Evgueni V. Cell signaling by endocannabinoids and their congeners: questions of selectivity and other challenges. Chem Phys Lipids. 2002 Dec 31;121(1-2):111–134. doi: 10.1016/s0009-3084(02)00157-3. [DOI] [PubMed] [Google Scholar]
  28. Schmid P. C., Zuzarte-Augustin M. L., Schmid H. H. Properties of rat liver N-acylethanolamine amidohydrolase. J Biol Chem. 1985 Nov 15;260(26):14145–14149. [PubMed] [Google Scholar]
  29. Sugiura T., Kobayashi Y., Oka S., Waku K. Biosynthesis and degradation of anandamide and 2-arachidonoylglycerol and their possible physiological significance. Prostaglandins Leukot Essent Fatty Acids. 2002 Feb-Mar;66(2-3):173–192. doi: 10.1054/plef.2001.0356. [DOI] [PubMed] [Google Scholar]
  30. Ueda N., Puffenbarger R. A., Yamamoto S., Deutsch D. G. The fatty acid amide hydrolase (FAAH). Chem Phys Lipids. 2000 Nov;108(1-2):107–121. doi: 10.1016/s0009-3084(00)00190-0. [DOI] [PubMed] [Google Scholar]
  31. Ueda N., Yamamoto K., Yamamoto S., Tokunaga T., Shirakawa E., Shinkai H., Ogawa M., Sato T., Kudo I., Inoue K. Lipoxygenase-catalyzed oxygenation of arachidonylethanolamide, a cannabinoid receptor agonist. Biochim Biophys Acta. 1995 Jan 20;1254(2):127–134. doi: 10.1016/0005-2760(94)00170-4. [DOI] [PubMed] [Google Scholar]
  32. Ueda N., Yamanaka K., Terasawa Y., Yamamoto S. An acid amidase hydrolyzing anandamide as an endogenous ligand for cannabinoid receptors. FEBS Lett. 1999 Jul 9;454(3):267–270. doi: 10.1016/s0014-5793(99)00820-0. [DOI] [PubMed] [Google Scholar]
  33. Ueda N., Yamanaka K., Yamamoto S. Purification and characterization of an acid amidase selective for N-palmitoylethanolamine, a putative endogenous anti-inflammatory substance. J Biol Chem. 2001 Jul 19;276(38):35552–35557. doi: 10.1074/jbc.M106261200. [DOI] [PubMed] [Google Scholar]
  34. Vandevoorde Séverine, Tsuboi Kazuhito, Ueda Natsuo, Jonsson Kent-Olov, Fowler Christopher J., Lambert Didier M. Esters, retroesters, and a retroamide of palmitic acid: pool for the first selective inhibitors of N-palmitoylethanolamine-selective acid amidase. J Med Chem. 2003 Oct 9;46(21):4373–4376. doi: 10.1021/jm0340795. [DOI] [PubMed] [Google Scholar]

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