Assessment of NSAIDs as potential inhibitors of the fatty acid amide hydrolase I (FAAH-1) using three different primary fatty acid amide substrates in vitro

Background Pain relief remains a major subject of inadequately met need of patients. Therapeutic agents designed to treat pain and inflammation so far have low to moderate efficiencies with significant untoward side effects. FAAH-1 has been proposed as a promising target for the discovery of drugs to treat pain and inflammation without significant adverse effects. FAAH-1 is the primary enzyme accountable for the degradation of AEA and related fatty acid amides. Studies have revealed that the simultaneous inhibition of COX and FAAH-1 activities produce greater pharmacological efficiency with significantly lowered toxicity and ulcerogenic activity. Recently, the metabolism of endocannabinoids by COX-2 was suggested to be differentially regulated by NSAIDs. Methods We analysed the affinity of oleamide, arachidonamide and stearoylamide at the FAAH-1 in vitro and investigated the potency of selected NSAIDs on the hydrolysis of endocannabinoid-like molecules (oleamide, arachidonamide and stearoylamide) by FAAH-1 from rat liver. NSAIDs were initially screened at 500 μM after which those that exhibited greater potency were further analysed over a range of inhibitor concentrations. Results The substrate affinity of FAAH-1 obtained, increased in a rank order of oleamide < arachidonamide < stearoylamide with resultant Vmax values in a rank order of arachidonamide > oleamide > stearoylamide. The selected NSAIDs caused a concentration-dependent inhibition of FAAH-1 activity with sulindac, carprofen and meclofenamate exhibiting the greatest potency. Michaelis-Menten analysis suggested the mode of inhibition of FAAH-1 hydrolysis of both oleamide and arachidonamide by meclofenamate and indomethacin to be non-competitive in nature. Conclusion Our data therefore suggest potential for study of these compounds as combined FAAH-1-COX inhibitors.

The Biosynthesis and Metabolism of the N-Acylated Aromatic Amino Acids: N-Acylphenylalanine, N-Acyltyrosine, N-Acyltryptophan, and N-Acylhistidine

The fatty acid amides are a family of lipids composed of two chemical moieties, a fatty acid and a biogenic amine linked together in an amide bond. This lipid family is structurally related to the endocannabinoid anandamide (N-arachidonoylethanolamine) and, thus, is frequently referred to as a family of endocannabinoid-related lipids. The fatty acid amide family is divided into different classes based on the conjugate amine; anandamide being a member of the N-acylethanolamine class (NAE). Another class within the fatty acid amide family is the N-acyl amino acids (NA-AAs). The focus of this review is a sub-class of the NA-AAs, the N-acyl aromatic amino acids (NA-ArAAs). The NA-ArAAs are not broadly recognized, even by those interested in the endocannabinoids and endocannabinoid-related lipids. Herein, the NA-ArAAs that have been identified from a biological source will be highlighted and pathways for their biosynthesis, degradation, enzymatic modification, and transport will be presented. Also, information about the cellular functions of the NA-ArAAs will be placed in context with the data regarding the identification and metabolism of these N-acylated amino acids. A review of the current state-of-knowledge about the NA-ArAAs is to stimulate future research about this underappreciated sub-class of the fatty acid amide family.

Identifying FAAH Inhibitors as New Therapeutic Options for the Treatment of Chronic Pain through Drug Repurposing

Chronic pain determines a substantial burden on individuals, employers, healthcare systems, and society. Most of the affected patients report dissatisfaction with currently available treatments. There are only a few and poor therapeutic options—some therapeutic agents are an outgrowth of drugs targeting acute pain, while others have several serious side effects. One of the primary degradative enzymes for endocannabinoids, fatty acid amide hydrolase (FAAH) attracted attention as a significant molecular target for developing new therapies for neuropsychiatric and neurological diseases, including chronic pain. Using chemical graph mining, quantitative structure–activity relationship (QSAR) modeling, and molecular docking techniques we developed a multi-step screening protocol to identify repurposable drugs as FAAH inhibitors. After screening the DrugBank database using our protocol, 273 structures were selected, with five already approved drugs, montelukast, repaglinide, revefenacin, raloxifene, and buclizine emerging as the most promising repurposable agents for treating chronic pain. Molecular docking studies indicated that the selected compounds interact with the enzyme mostly non-covalently (except for revefenacin) through shape complementarity to the large substrate-binding pocket in the active site. A molecular dynamics simulation was employed for montelukast and revealed stable interactions with the enzyme. The biological activity of the selected compounds should be further confirmed by employing in vitro and in vivo studies.

Endocannabinoid System as a Promising Therapeutic Target in Inflammatory Bowel Disease – A Systematic Review

Inflammatory bowel disease (IBD) is a general term used to describe a group of chronic inflammatory conditions of the gastrointestinal tract of unknown etiology, including two primary forms: Crohn’s disease (CD) and ulcerative colitis (UC). The endocannabinoid system (ECS) plays an important role in modulating many physiological processes including intestinal homeostasis, modulation of gastrointestinal motility, visceral sensation, or immunomodulation of inflammation in IBD. It consists of cannabinoid receptors (CB1 and CB2), transporters for cellular uptake of endocannabinoid ligands, endogenous bioactive lipids (Anandamide and 2-arachidonoylglycerol), and the enzymes responsible for their synthesis and degradation (fatty acid amide hydrolase and monoacylglycerol lipase), the manipulation of which through agonists and antagonists of the system, shows a potential therapeutic role for ECS in inflammatory bowel disease. This review summarizes the role of ECS components on intestinal inflammation, suggesting the advantages of cannabinoid-based therapies in inflammatory bowel disease.

Potential of Fatty Acid Amide Hydrolase (FAAH), Monoacylglycerol Lipase (MAGL), and Diacylglycerol Lipase (DAGL) Enzymes as Targets for Obesity Treatment: A Narrative Review

The endocannabinoid system (ECS) plays an integral role in maintaining metabolic homeostasis and may affect hunger, caloric intake, and nutrient absorption. Obesity has been associated with higher levels of the endogenous cannabinoid transmitters (endocannabinoids). Therefore, the ECS is an important target in obesity treatment. Modulating the enzymes that synthesize and degrade endocannabinoids, namely fatty acid amide hydrolase (FAAH), monoacylglycerol lipase (MAGL), and diacylglycerol lipase (DAGL), may be a promising strategy to treat obesity. This review aims to synthesize all studies investigating pharmacological or genetic manipulation of FAAH, MAGL, or DAGL enzymes in association with obesity-related measures. Pharmacological inhibition or genetic deletion of FAAH tended to promote an obesogenic state in animal models, though the relationships between human FAAH polymorphisms and obesity-related outcomes were heterogeneous, which could be due to FAAH having both pro-appetitive and anti-appetitive substrates. Genetic deletion of Mgll and Dagla as well as pharmacological inhibition of DAGL tended to reduce body weight and improve metabolic state in animal studies, though the effects of Mgll manipulation were tissue-dependent. Monitoring changes in body weight in ongoing clinical trials of FAAH inhibitors may clarify whether FAAH inhibition is a potential therapeutic strategy for treatment obesity. More preclinical work is needed to characterize the role of MAGL and DAGL modulation in obesity-related outcomes.

FAAH polymorphism (rs324420) modulates extinction recall in healthy humans: an fMRI study

Gold standard treatments for anxiety- and trauma-related disorders focus on exposure therapy promoting extinction learning and extinction retention. However, its efficacy is limited. Preclinical and particularly animal research has been able to demonstrate that homozygosity for the fatty acid amide hydrolase (FAAH) C385A allele, similar to FAAH inhibition, is associated with elevated concentrations of anandamide (AEA) and facilitates extinction learning and extinction recall. However, in humans, the underlying neurobiological processes are less well understood, and further knowledge might enhance the development of more effective therapies. In this functional magnetic resonance imaging (fMRI) study, a fear conditioning, fear extinction and extinction recall paradigm was conducted with 55 healthy male adults. They were genotyped for the FAAH single-nucleotide polymorphism (SNP) rs324420 to investigate differences related to extinction recall in neural activation and State–Trait Anxiety Inventory (STAI) ratings between AC heterozygotes and CC homozygotes (FAAH C385A SNP). Differential brain activation upon an unextinguished relative to an extinguished stimulus, was greater in AC heterozygotes as compared to CC homozygotes in core neural structures previously related to extinction recall, such as the medial superior frontal gyrus, the dorsal anterior cingulate and the anterior and middle insular cortex. Furthermore, AC heterozygotes displayed higher AEA levels and lower STAI-state ratings. Our data can be interpreted in line with previous suggestions of more successful extinction recall in A-allele carriers with elevated AEA levels. Data corroborate the hypothesis that the endocannabinoid system, particularly AEA, plays a modulatory role in the extinction of aversive memory.

Genetic Variants of Fatty Acid Amide Hydrolase Modulate Acute Inflammatory Responses to Colitis in Adult Male Mice

Cannabinoids, including cannabis derived phytocannabinoids and endogenous cannabinoids (endocannabinoids), are typically considered anti-inflammatory. One such endocannabinoid is N-arachidonoylethanolamine (anandamide, AEA), which is metabolized by fatty acid amide hydrolase (FAAH). In humans, there is a loss of function single nucleotide polymorphism (SNP) in the FAAH gene (C385A, rs324420), that leads to increases in the levels of AEA. Using a mouse model with this SNP, we investigated how this SNP affects inflammation in a model of inflammatory bowel disease. We administered 2,4,6-trinitrobenzene sulfonic acid (TNBS) intracolonically, to adult male FAAH SNP mice and examined colonic macroscopic tissue damage and myeloperoxidase activity, as well as levels of plasma and amygdalar cytokines and chemokines 3 days after administration, at the peak of colitis. We found that mice possessing the loss of function alleles (AC and AA), displayed no differences in colonic damage or myeloperoxidase activity compared to mice with wild type alleles (CC). In contrast, in plasma, colitis-induced increases in interleukin (IL)-2, leukemia inhibitory factor (LIF), monocyte chemoattractant protein (MCP)-1, and tumor necrosis factor (TNF) were reduced in animals with an A allele. A similar pattern was observed in the amygdala for granulocyte colony stimulating factor (G-CSF) and MCP-1. In the amygdala, the mutant A allele led to lower levels of IL-1α, IL-9, macrophage inflammatory protein (MIP)-1β, and MIP-2 independent of colitis—providing additional understanding of how FAAH may serve as a regulator of inflammatory responses in the brain. Together, these data provide insights into how FAAH regulates inflammatory processes in disease.

Effects of Fatty Acid Amide Hydroxylase Inhibitor URB597 on the Catecholaminergic Activity of the Adrenal Medulla in Stressed Male and Female Rats

<b><i>Introduction:</i></b> The present study examined the effects of fatty acid amide hydrolase inhibitor URB597 on the level of plasma catecholamine and their content, synthesis, and degradation in the adrenal medulla of male and female rats subjected to chronic unpredictable stress (CUS). <b><i>Material and Methods:</i></b> Male and female Wistar rats were exposed to the 6 weeks of CUS and treated intraperitoneally with either 0.3 mg/kg/day of URB597 or vehicle in the last 2 weeks of stress protocol. Catecholamines’ plasma levels and catecholamines’ levels in adrenal medulla were examined using Elabscience ELISA kits. Western blot analysis was used to detect the protein in the medulla. <b><i>Results:</i></b> The results of our experiment showed that adrenal weights and catecholamine of unstressed control were higher in females and that CUS induced further enlargement of adrenal glands and catecholamine content and its synthesis compared to male rats. CUS caused an increase of plasma norepinephrine and depletion of norepinephrine content as well as unchanged synthesis and degradation of catecholamine in the adrenal medulla of male rats. URB597 reduced enlarged adrenals and catecholamine content and its synthesis in stressed female rats. URB597 reduces increased plasma norepinephrine and restores its content in the adrenal medulla, unchanging the expression of enzyme synthesis, while reduced protein levels of monoamine oxidase A in male rats are exposed to CUS. <b><i>Discussion:</i></b> Our results support the role of endocannabinoids as an antistress mechanism that inhibits elevated adrenomedullary activation and promotes its recovery to baseline in both male and female stressed rats.