Research paper
Novel propanamides as fatty acid amide hydrolase inhibitors

https://doi.org/10.1016/j.ejmech.2017.05.033Get rights and content

Highlights

  • An efficient synthesis afforded new heteroarylpropanamides.

  • Synthesized compounds were assayed in vitro as FAAH inhibitors.

  • Enzymatic assays showed potencies toward FAAH in the nanomolar to micromolar range.

  • The binding mode for the lead compound of the series was proposed and discussed.

Abstract

Fatty acid amide hydrolase (FAAH) has a key role in the control of the cannabinoid signaling, through the hydrolysis of the endocannabinoids anandamide and in some tissues 2-arachidonoylglycerol. FAAH inhibition represents a promising strategy to activate the cannabinoid system, since it does not result in the psychotropic and peripheral side effects characterizing the agonists of the cannabinoid receptors. Here we present the discovery of a novel class of profen derivatives, the N-(heteroaryl)-2-(4-((2-(trifluoromethyl)pyridin-4-yl)amino)phenyl)propanamides, as FAAH inhibitors. Enzymatic assays showed potencies toward FAAH ranging from nanomolar to micromolar range, and the most compounds lack activity toward the two isoforms of cyclooxygenase. Extensive structure-activity studies and the definition of the binding mode for the lead compound of the series are also presented. Kinetic assays in rat and mouse FAAH on selected compounds of the series demonstrated that slight modifications of the chemical structure could influence the binding mode and give rise to competitive (TPA1) or non-competitive (TPA14) inhibition modes.

Introduction

Anandamide (AEA), 2-acylethanolamines and 2-arachidonoyl glycerol are signaling lipids belonging to the class of endocannabinoids, endogenous agonists of the cannabinoid receptor type-1 (CB1) and type-2 (CB2). The cannabinoid system regulates many physiological functions, both in the central and peripheral nervous systems and in peripheral organs. In order to maintain the normal functionality of nerve systems, many fatty acid amides, including AEA, are degraded by fatty acid amide hydrolase (FAAH), a member of the serine hydrolase enzyme family, characterized by the unusual catalytic triad Ser217-Lys142-Ser241.

Blockade of FAAH activity is a suitable strategy to modulate the endocannabinoid system compared to exogenous cannabinoid receptor agonists, whose therapeutic profile is heavily limited by CB1-mediated psychotropic and peripheral side effects such as impairment in cognition, motor coordination, and psychoses. It is now well established that the endocannabinoid system is involved in the regulation of mood, and in animal models of anxiety, and that FAAH inhibitors show a potentially useful profile [1]. Additionally, FAAH inhibitors may be useful in disorders ranging from cannabinoid dependence [2] to preventing or reducing the inflammatory process associated with Aβ deposition in Alzheimer's Disease [3] and in the treatment of comorbidity between psychological disorders and cardiac disease [4]. The principal classes of covalent and non-covalent inhibitors as well as the FAAH inhibitors entered into clinical studies have been recently reviewed [5]. Clinical studies so far undertaken indicate that FAAH inhibitors are generally well tolerated [6], [7]. A serious brain injury was reported in a phase I study with the FAAH inhibitor BIA 10-2474, but this appears to be an off-target effect of the drug, not correlated to FAAH inhibition [8]. Indeed, a computer-based proteome-docking approach suggested that BIA 10-2474 could interact with several targets in a manner likely to cause brain haemorrhaging [9].

We previously reported that some NSAIDs as ibuprofen display modest FAAH inhibitory activity. The conversion of the carboxylic group of ibuprofen into heterocyclic amide namely 2-(4-isobutylphenyl)-N-(3-methylpyridin-2-yl)propanamide (Ibu-AM5) that showed high FAAH inhibitory activity, while retaining COX inhibitory properties of the parent ibuprofen [10], [11], [12]. Encouraged by these finding, we have been prompted to investigate further new Ibu-AM5 analogs as potential FAAH inhibitors. Since superimposition of Ibu-AM5 with FAAH co-crystallized inhibitors indicated that the isobutyl chain of Ibu-AM5 overlap aromatic hydrophilic moieties, as a rational development we designed and synthesized a new series of compounds where the isobutyl group is replaced by a 2-(trifluoromethyl)pyridin-4-ylamino group (Fig. 1). In the present study we report the discovery, synthesis, pharmacological and biochemical characterization of these compounds, as well as their putative binding mode of the lead and finally the efforts to identify key structural features of this novel series of N-(heteroaryl)-2-(4-((2-(trifluoromethyl)pyridin-4-yl)amino)phenyl)propanamides as FAAH inhibitors. With the aim to define the critical requirements for FAAH inhibitory activity we have explored the effects of structural modification in the heteroaryl group, in the linker between the heteroaryl group and the phenylcarbonyl moiety, the effect of replacement of the methyl on the C-2 with smaller and larger substituents, and finally the effect of introducing different polar rings in the (trifluoromethyl)pyridine region.

Section snippets

Chemistry

The synthesis of the target amides was undertaken as outlined in Scheme 1, Scheme 2, Scheme 3, Scheme 4, Scheme 5, Scheme 6. The first step of the synthetic approach to the new series of TPA amides was based on a previously described procedure that allows the construction of the trifluoromethylpyridine ring linked by an amino nitrogen to an aromatic ring [13]. This synthetic approach was modified and successfully applied for the preparation of

Conclusions

In the present study by replacement of Ibu-AM5 isobutyl chain with a trifluoromethylamino moiety led to a novel class of FAAH inhibitors. The N-(heteroaryl)-2-(4-((2-(trifluoromethyl)pyridin-4-yl)amino)phenyl)propanamides were found to act as reversible inhibitors. The 2-amino-3-methylpyridine amide (TPA1) retained the FAAH inhibitory potency of Ibu-AM5 and behaved essentially as competitive inhibitor in the rat enzyme. Docking and MD studies indicated the acyl chain binding (ABP) channel in

General methods

Commercially available solvents and reagents were used without further purification unless otherwise stated. Tetrahydrofuran (THF) was freshly distilled from sodium/benzophenone. Reactions requiring anhydrous conditions were performed in oven-dried glassware under argon atmosphere. 1H NMR spectra were recorded on a Varian Inova 500 spectrometer. The chemical shifts (δ) are reported in part per million downfield from tetramethylsilane (TMS), which was used as internal standard, and the spectra

Acknowledgments

This work was supported by grants from the Regione Autonoma della Sardegna (Project L.R. 7/2007, no. 2012_CRP-59473) to VO; the Swedish Research Council (Grant no. 12158, medicine) and the Research Funds of the Medical Faculty, Umeå University to CJF; the Generalitat de Catalunya (2014SGR1189), ICREA Academia, and Consorci de Serveis Universitaris de Catalunya to FJL. CMM and BC acknowledge the CINECA (ISCRA C projects ID HP10C34LJ7, HP10CPFIRC and HP10CSJ2H8) for the access to high performance

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    These authors equally contribute to the present paper.

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