The fatty acid ethanolamides (FAEs) are a family of bioactive lipid mediators that include the endogenous agonist of peroxisome proliferator-activated receptor-alpha, palmitoylethanolamide (PEA). FAEs are hydrolyzed intracellularly by either fatty acid amide hydrolase or N-acylethanolamine-hydrolyzing acid amidase (NAAA). Selective inhibition of NAAA by (S)-N-(2-oxo-3-oxetanyl)-3-phenylpropionamide [(S)-OOPP, 7a] prevents PEA degradation in mouse leukocytes and attenuates responses to proinflammatory stimuli. Starting from the structure of 7a, a series of beta-lactones was prepared and tested on recombinant rat NAAA to explore structure-activity relationships (SARs) for this class of inhibitors and improve their in vitro potency. Following the hypothesis that these compounds inhibit NAAA by acylation of the catalytic cysteine, we identified several requirements for recognition at the active site and obtained new potent inhibitors. In particular, (S)-N-(2-oxo-3-oxetanyl)biphenyl-4-carboxamide (7h) was more potent than 7a at inhibiting recombinant rat NAAA activity (7a, IC(50) = 420 nM; 7h, IC(50) = 115 nM) in vitro and at reducing carrageenan-induced leukocyte infiltration in vivo.

The compound URB754 was recently identified as a potent inhibitor of the endocannabinoid-deactivating enzyme monoacylglycerol lipase (MGL) by screening of a commercial chemical library. Based on HPLC/MS, NMR and EI/MS analyses, the present paper shows that the MGL-inhibitory activity attributed to URB754 is in fact due to a chemical impurity present in the commercial sample, identified as bis(methylthio)mercurane. Although this organomercurial compound is highly potent at inhibiting MGL (IC50 = 11.9 +/- 1.1 nM), its biological use is prohibited by its toxicity and target promiscuity.

The β-lactone ring of N-(2-oxo-3-oxetanyl)amides, a class of N-acylethanolamine acid amidase (NAAA) inhibitors endowed with anti-inflammatory properties, is responsible for both NAAA inhibition and low compound stability. Here, we investigate the structure-activity and structure-property relationships for a set of known and new β-lactone derivatives, focusing on the new class of N-(2-oxo-3-oxetanyl)carbamates. Replacement of the amide group with a carbamate one led to different stereoselectivity for NAAA inhibition and higher intrinsic stability, because of the reduced level of intramolecular attack at the lactone ring. The introduction of a syn methyl at the β-position of the lactone further improved chemical stability. A tert-butyl substituent in the side chain reduced the reactivity with bovine serum albumin. (2S,3R)-2-Methyl-4-oxo-3-oxetanylcarbamic acid 5-phenylpentyl ester (27, URB913/ARN077) inhibited NAAA with good in vitro potency (IC(50) = 127 nM) and showed improved stability. It is rapidly cleaved in plasma, which supports its use for topical applications.

Identifying points of control in inflammation is essential to discovering safe and effective antiinflammatory medicines. Palmitoylethanolamide (PEA) is a naturally occurring lipid amide that, when administered as a drug, inhibits inflammatory responses by engaging peroxisome proliferator-activated receptor-alpha (PPAR-alpha). PEA is preferentially hydrolyzed by the cysteine amidase N-acylethanolamine-hydrolyzing acid amidase (NAAA), which is highly expressed in macrophages. Here we report the discovery of a potent and selective NAAA inhibitor, N-[(3S)-2-oxo-3-oxetanyl]-3-phenylpropanamide [(S)-OOPP], and show that this inhibitor increases PEA levels in activated leukocytes and blunts responses induced by inflammatory stimuli both in vitro and in vivo. These effects are stereoselective, mimicked by exogenous PEA, and abolished by PPAR-alpha deletion. (S)-OOPP also attenuates inflammation and tissue damage and improves recovery of motor function in mice subjected to spinal cord trauma. The results suggest that PEA activation of PPAR-alpha in leukocytes serves as an early stop signal that contrasts the progress of inflammation. The PEA-hydrolyzing amidase NAAA may provide a previously undescribed target for antiinflammatory medicines.