Leung K.

Publication Details



In vitro Rodents



Fatty acid amide hydrolase (FAAH) is an integral membrane-bound serine hydrolase and a part of the endocannabinoid system (ECS), which comprises the cannabinoid receptors (CB1 and CB2), endogenous ligands termed endocannabinoids (anandamide and oleamide), transporters, and enzymes (1, 2). FAAH plays a key role in the hydrolysis of a number of primary and secondary fatty acid amides, controlling the levels of the neuromodulatory endocannabinoids in the ECS (3, 4). FAAH is widely expressed in many tissues, with the highest levels in the liver and brain (5). Genetic or pharmacological inactivation of FAAH in the brain leads to analgesic, anti-inflammatory, anxiolytic, and anti-depression effects in animal models (6-8).

Cyclohexylcarbamic acid 3'-carbamoylbiphenyl-3-yl ester (URB597) is an irreversible, substrate-like inhibitor of FAAH involving carbamylation of the catalytic nucleophilic Ser241 and the O-biaryl group as the leaving group (9-11). On the basis of the structure of URB597, Wyffels et al. (11) prepared biphenyl-3-yl-4-[11C]methoxyphenylcarbamate ([11C]-1) for in vivo positron emission tomography (PET) imaging studies of the brain FAAH in mice. It was proposed that the carbamylation of Ser241 would leave the [11C]-methoxyanilino group bound to the FAAH for visualization of FAAH in the brain. However, the results of in vitro and ex vivo studies indicated that [11C]-1 is a reversible inhibitor of FAAH, and the rapid brain washout of the tracer limits its utility as a PET agent for in vivo measurements of FAAH. Wilson et al. (10) reported the 11C-radiolabeling of a close analog of URB597, 6-hydroxy-[1,1'-biphenyl]-3-yl-cyclohexylcarbamate (URB694), yielding [11C-carbonyl]URB694 ([11C]CURB) for PET imaging of FAAH in the brain. [11C]CURB showed good brain accumulation with regional heterogeneity, irreversibility, and specific binding to FAAH in vivo in rats.

Skaddan et al. (9) prepared 4-(3-((5-(2-[18F]fluoroethoxy)pyridine-2-yl)oxy)benzylidene)-N-(pyridazin-3-yl)piperidine-1-carboxamide ([18F]PF-9811) for use with in vivo PET imaging of FAAH. PF-9811 is a close analog of 4-(3-((5-(2-trifluoromethyl)pyridine-2-yl)oxy)benzylidene)-N-(pyridazin-3-yl)piperidine-1-carboxamide (PF-04457845), which is a clinical candidate for treatment of inflammatory and non-inflammatory pain. [18F]PF-9811 showed good brain accumulation, with regional heterogeneity and specific binding to FAAH in vivo in rats.



Skaddan et al. (9) synthesized [18F]PF-9811 via a 3-step, one-pot reaction sequence in 87 min from the end of bombardment. [18F]PF-9811 was purified with high-performance liquid chromatography with >98% radiochemical purity. The specific activity of [18F]PF-9811 was >444 GBq/μmol (12 Ci/µmol) at the end of synthesis. The LogP7.4 value of [18F]PF-9811 was 3.14 ± 0.02.

In Vitro Studies: Testing in Cells and Tissues


Skaddan et al. (9) showed that unlabeled PF-04457845 and PF-9811 inhibited FAAH activity in mouse brain homogenates with IC50 values of 3.2 and 16 nM, respectively.

Animal Studies



Ex vivo biodistribution studies in normal male rats (n = 4/group) were performed at 10, 60, and 90 min after intravenous injection of ~70 MBq (2.1 mCi) [18F]PF-9811 (0.4 nmol/kg) (9). The accumulation level of radioactivity was the highest in the cortex and cerebellum, followed by the hippocampus, striatum, hypothalamus, thalamus, and brain stem, with standard uptake values (SUVs) of 0.32–0.67 at 10 min, and the accumulation gradually increased to 0.39–0.86 SUV at 90 min. The distribution of radioactivity in the brain regions is in agreement with the known distribution of FAAH in the rat brain. The SUVs in the liver and bone at 90 min were 16.1 and 0.16 (indicating minimal defluorination), respectively. Radioactivity in the blood was lower than in the brain. The brain/blood ratios increased from 2.7 at 10 min to 12.9 at 90 min after injection. Pretreatment (60 min) with PF-04457845 (1 mg/kg, oral) inhibited the radioactivity levels by 37%–73%, depending on the brain regions, and 74% in the liver at 90 min. The specific/non-specific ratios were 2.3, 2.4, and 2.6 for the hippocampus, cerebellum, and cortex, respectively.

PET imaging studies were performed in six male rats cannulated in the jugular vein; scans were performed at 1, 2, 5, 10, 15, 20, 30, 45, 60, and 90 min after injection of ~74 MBq (2 mCi) [18F]PF-9811 (9). The highest radioactivity levels were observed in the cortex, cerebellum, and hippocampus. Radioactivity accumulation reached a plateau by ~60 min and gradually increased up to 90 min. Pretreatment (60 min) with PF-04457845 (1 mg/kg, oral) inhibited the radioactivity levels in the cortex, cerebellum, and hippocampus by ~40%–50% at 90 min based on image quantitation.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


No publication is currently available.


Bambico F.R., Duranti A., Tontini A., Tarzia G., Gobbi G. Endocannabinoids in the treatment of mood disorders: evidence from animal models. Curr Pharm Des. 2009;15(14):1623–46. [PubMed: 19442178]
Thomas E.A., Cravatt B.F., Danielson P.E., Gilula N.B., Sutcliffe J.G. Fatty acid amide hydrolase, the degradative enzyme for anandamide and oleamide, has selective distribution in neurons within the rat central nervous system. J Neurosci Res. 1997;50(6):1047–52. [PubMed: 9452020]
Fowler C.J., Naidu P.S., Lichtman A., Onnis V. The case for the development of novel analgesic agents targeting both fatty acid amide hydrolase and either cyclooxygenase or TRPV1. Br J Pharmacol. 2009;156(3):412–9. [PMC free article: PMC2697682] [PubMed: 19226258]
Naidu P.S., Kinsey S.G., Guo T.L., Cravatt B.F., Lichtman A.H. Regulation of inflammatory pain by inhibition of fatty acid amide hydrolase. J Pharmacol Exp Ther. 2010;334(1):182–90. [PMC free article: PMC2912038] [PubMed: 20375198]
Alexander J.P., Cravatt B.F. Mechanism of carbamate inactivation of FAAH: implications for the design of covalent inhibitors and in vivo functional probes for enzymes. Chem Biol. 2005;12(11):1179–87. [PMC free article: PMC1994809] [PubMed: 16298297]
Ahn K., Smith S.E., Liimatta M.B., Beidler D., Sadagopan N., Dudley D.T., Young T., Wren P., Zhang Y., Swaney S., Van Becelaere K., Blankman J.L., Nomura D.K., Bhattachar S.N., Stiff C., Nomanbhoy T.K., Weerapana E., Johnson D.S., Cravatt B.F. Mechanistic and pharmacological characterization of PF-04457845: a highly potent and selective fatty acid amide hydrolase inhibitor that reduces inflammatory and noninflammatory pain. J Pharmacol Exp Ther. 2011;338(1):114–24. [PMC free article: PMC3126636] [PubMed: 21505060]
Basso E., Duranti A., Mor M., Piomelli D., Tontini A., Tarzia G., Traldi P. Tandem mass spectrometric data-FAAH inhibitory activity relationships of some carbamic acid O-aryl esters. J Mass Spectrom. 2004;39(12):1450–5. [PubMed: 15578755]
Mor M., Rivara S., Lodola A., Plazzi P.V., Tarzia G., Duranti A., Tontini A., Piersanti G., Kathuria S., Piomelli D. Cyclohexylcarbamic acid 3'- or 4'-substituted biphenyl-3-yl esters as fatty acid amide hydrolase inhibitors: synthesis, quantitative structure-activity relationships, and molecular modeling studies. J Med Chem. 2004;47(21):4998–5008. [PubMed: 15456244]
Skaddan M.B., Zhang L., Johnson D.S., Zhu A., Zasadny K.R., Coelho R.V., Kuszpit K., Currier G., Fan K.H., Beck E.M., Chen L., Drozda S.E., Balan G., Niphakis M., Cravatt B.F., Ahn K., Bocan T., Villalobos A. The synthesis and in vivo evaluation of [(18)F]PF-9811: a novel PET ligand for imaging brain fatty acid amide hydrolase (FAAH). Nucl Med Biol. 2012;39(7):1058–67. [PMC free article: PMC3611965] [PubMed: 22571907]
Wilson A.A., Garcia A., Parkes J., Houle S., Tong J., Vasdev N. [11C]CURB: Evaluation of a novel radiotracer for imaging fatty acid amide hydrolase by positron emission tomography. Nucl Med Biol. 2011;38(2):247–53. [PubMed: 21315280]
Wyffels L., Muccioli G.G., Kapanda C.N., Labar G., De Bruyne S., De Vos F., Lambert D.M. PET imaging of fatty acid amide hydrolase in the brain: synthesis and biological evaluation of an 11C-labelled URB597 analogue. Nucl Med Biol. 2010;37(5):665–75. [PubMed: 20610171]