Novel Mechanistic Class of Fatty Acid Amide Hydrolase Inhibitors with Remarkable Selectivity

Abstract

Fatty acid amide hydrolase (FAAH) is an integral membrane enzyme that degrades the fatty acid amide family of signaling lipids, including the endocannabinoid anandamide. Genetic or pharmacological inactivation of FAAH leads to analgesic, anti-inflammatory, anxiolytic, and antidepressant phenotypes in rodents without showing the undesirable side effects observed with direct cannabinoid receptor agonists, indicating that FAAH may represent an attractive therapeutic target for treatment of pain, inflammation, and other central nervous system disorders. However, the FAAH inhibitors reported to date lack druglike pharmacokinetic properties and/or selectivity. Herein we describe piperidine/piperazine ureas represented by and N-phenyl-4-(quinolin-2-ylmethyl)piperazine-1-carboxamide (PF-622) as a novel mechanistic class of FAAH inhibitors. PF-750 and PF-622 show higher in vitro potencies than previously established classes of FAAH inhibitors. Rather unexpectedly based on the high chemical stability of the urea functional group, PF-750 and PF-622 were found to inhibit FAAH in a time-dependent manner by covalently modifying the enzyme's active site serine nucleophile. Activity-based proteomic profiling revealed that PF-750 and PF-622 were completely selective for FAAH relative to other mammalian serine hydrolases. We hypothesize that this remarkable specificity derives, at least in part, from FAAH's special ability to function as a C(O)-N bond hydrolase, which distinguishes it from the vast majority of metabolic serine hydrolases in mammals that are restricted to hydrolyzing esters and/or thioesters. The piperidine/piperazine urea may thus represent a privileged chemical scaffold for the synthesis of FAAH inhibitors that display an unprecedented combination of potency and selectivity for use as potential analgesic and anxiolytic/antidepressant agents.Fatty acid amide hydrolase (FAAH 1 ) is an integral membrane enzyme (1, 2) that regulates the fatty acid amide family of lipid transmitters, which includes the endogenous cannabinoid N-arachidonyl ethanolamine (anandamide) (3), the anti-inflammatory factor N-palmitoyl ethanolamine (PEA) (4), the sleep-inducing substance 9(Z)-octadecenamide (oleamide) (5), and the satiating signal N-oleoyl ethanolamine (OEA) (6).FAAH inactivation by either genetic deletion of the FAAH gene in mice (7) or by chemical inhibitors (8) leads to elevated endogenous levels of fatty acid amides with concomitant analgesic (9-13), anxiolytic (8), antidepressant (14, 15), sleep-enhancing (16), and anti-inflammatory (9,17,18) phenotypes. Notably the behavioral phenotypes observed in FAAH knockout (-/-) mice (7, 9) or in rodents treated with FAAH inhibitors (8,(11)(12)(13)(14)18) occur in the absence of alterations in motility, weight gain, or body temperature that are typically observed with direct cannabinoid receptor 1 (CB1) agonists. These findings suggest that FAAH may represent an attractive therapeutic target for treatment of pain, inflammation, and other central nervous system (CNS) diso...

The synthesis and biological evaluation of para-substituted phenolic N-alkyl carbamates as endocannabinoid hydrolyzing enzyme inhibitors

“…Not surprisingly, in this series of 1,2,3-thiadiazol-4-yl substituted derivatives (8,(25)(26)(27)(28)(29), the length and the shape of the N-alkyl group had a similar effect on the inhibitory potencies against both enzymes, FAAH and MGL, as it had in the previous series. Thus, by replacing the N-n-butyl chain of 8 by N-n-hexyl (25) the inhibitory potencies toward FAAH and MGL were improved (IC 50 s; 0.019 and 11 mM, respectively).…”

“…Furthermore, the most potent inhibitor in this series with N-dodecyl moiety (26) inhibited FAAH with an IC 50 value of 0.012 mM. Moreover, hydrolysis of 2-AG in rat brain membranes as well as hrMGL mediated 2-OG hydrolysis However, the compounds with N-cyclopentyl (27), N-cyclohexyl (28) and N-benzyl (29) moieties were more potent FAAH inhibitors than the corresponding dihydrothiazoline derivatives 14-16, indicating that FAAH inhibitory activity could be affected by the type of heterocycle. This effect was not seen for MGL.…”

“…Therefore, several FAAH inhibitors have been developed (see for review Ref. [12]), including various fatty acid derivatives [13][14][15][16][17], and non-lipid inhibitors such as a-keto heterocycles [18][19][20], carbamate derivatives [21][22][23][24][25], (thio)-hydantoins [26] and most recently, selective piperidine/piperazine ureas [27,28]. Probably, the most well-studied FAAH inhibitors are the carbamate-based 3 0 -carbamoylbiphenyl-3-yl ester (4, URB597) ( Fig.…”

Avibactam is a covalent, reversible, non–β-lactam β-lactamase inhibitor

“…Compared with the ester linkage, the carbamoyl acyl-enzyme linkage is generally more stable to enzyme-assisted hydrolysis (27,28), and inhibitors that acylate through a carbamate have been reported with extremely long residence times, indicating stability to hydrolytic deacylation (27,29). In this light, the hydrolytic stability of the avibactam acylenzyme is not without precedent.…”

Chiral 3-(4,5-dihydrooxazol-2-yl)phenyl alkylcarbamates as novel FAAH inhibitors: Insight into FAAH enantioselectivity by molecular docking and interaction fields

“…Inhibition by N-alkylcarbamates is based on irreversible acylation of the active site serine [25]. Recently reported other structure families with inhibitory activity against FAAH include (thio)hydantoins [26], piperidine-and piperazine ureas [27,28], sulfonyl derivatives [29] and boronic acid derivatives [30].…”