Human Fatty Acid Amide Hydrolase Research Articles

Discovery of PF-04457845: A Highly Potent, Orally Bioavailable, and Selective Urea FAAH Inhibitor

Fatty acid amide hydrolase (FAAH) is an integral membrane serine hydrolase that degrades the fatty acid amide family of signaling lipids, including the endocannabinoid anandamide. Genetic or pharmacological inactivation of FAAH leads to analgesic and anti-inflammatory 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 the treatment of inflammatory pain and other nervous system disorders. Herein, we report the discovery and characterization of a highly efficacious and selective FAAH inhibitor PF-04457845 (23). Compound 23 inhibits FAAH by a covalent, irreversible mechanism involving carbamylation of the active-site serine nucleophile of FAAH with high in vitro potency (k(inact)/K(i) and IC(50) values of 40300 M(-1) s(-1) and 7.2 nM, respectively, for human FAAH). Compound 23 has exquisite selectivity for FAAH relative to other members of the serine hydrolase superfamily as demonstrated by competitive activity-based protein profiling. Oral administration of 23 at 0.1 mg/kg results in efficacy comparable to that of naproxen at 10 mg/kg in a rat model of inflammatory pain. Compound 23 is being evaluated in human clinical trials.

Biochanin A, a naturally occurring inhibitor of fatty acid amide hydrolase

Inhibitors of fatty acid amide hydrolase (FAAH), the enzyme responsible for the metabolism of the endogenous cannabinoid (CB) receptor ligand anandamide (AEA), are effective in a number of animal models of pain. Here, we investigated a series of isoflavones with respect to their abilities to inhibit FAAH. In vitro assays of FAAH activity and affinity for CB receptors were used to characterize key compounds. In vivo assays used were biochemical responses to formalin in anaesthetized mice and the 'tetrad' test for central CB receptor activation. Of the compounds tested, biochanin A was adjudged to be the most promising. Biochanin A inhibited the hydrolysis of 0.5 microM AEA by mouse, rat and human FAAH with IC(50) values of 1.8, 1.4 and 2.4 microM respectively. The compound did not interact to any major extent with CB(1) or CB(2) receptors, nor with FAAH-2. In anaesthetized mice, URB597 (30 microg i.pl.) and biochanin A (100 microg i.pl.) both inhibited the spinal phosphorylation of extracellular signal-regulated kinase produced by the intraplantar injection of formalin. The effects of both compounds were significantly reduced by the CB(1) receptor antagonist/inverse agonist AM251 (30 microg i.pl.). Biochanin A (15 mg.kg(-1) i.v.) did not increase brain AEA concentrations, but produced a modest potentiation of the effects of 10 mg.kg(-1) i.v. AEA in the tetrad test. It is concluded that biochanin A, in addition to its other biochemical properties, inhibits FAAH both in vitro and peripherally in vivo.

Biomarkers of Endocannabinoid System Activation in Severe Obesity

BackgroundObesity is a worldwide epidemic, and severe obesity is a risk factor for many diseases, including diabetes, heart disease, stroke, and some cancers. Endocannabinoid system (ECS) signaling in the brain and peripheral tissues is activated in obesity and plays a role in the regulation of body weight. The main research question here was whether quantitative measurement of plasma endocannabinoids, anandamide, and related N-acylethanolamines (NAEs), combined with genotyping for mutations in fatty acid amide hydrolase (FAAH) would identify circulating biomarkers of ECS activation in severe obesity.Methodology/Principal FindingsPlasma samples were obtained from 96 severely obese subjects with body mass index (BMI) of ≥40 kg/m2, and 48 normal weight subjects with BMI of ≤26 kg/m2. Triple-quadrupole mass spectroscopy methods were used to measure plasma ECS analogs. Subjects were genotyped for human FAAH gene mutations. The principal analysis focused on the FAAH 385 C→A (P129T) mutation by comparing plasma ECS metabolite levels in the FAAH 385 minor A allele carriers versus wild-type C/C carriers in both groups. The main finding was significantly elevated mean plasma levels of anandamide (15.1±1.4 pmol/ml) and related NAEs in study subjects that carried the FAAH 385 A mutant alleles versus normal subjects (13.3±1.0 pmol/ml) with wild-type FAAH genotype (p = 0.04), and significance was maintained after controlling for BMI.Conclusions/SignificanceSignificantly increased levels of the endocannabinoid anandamide and related NAEs were found in carriers of the FAAH 385 A mutant alleles compared with wild-type FAAH controls. This evidence supports endocannabinoid system activation due to the effect of FAAH 385 mutant A genotype on plasma AEA and related NAE analogs. This is the first study to document that FAAH 385 A mutant alleles have a direct effect on elevated plasma levels of anandamide and related NAEs in humans. These biomarkers may indicate risk for severe obesity and may suggest novel ECS obesity treatment strategies.

Homology modelling and molecular dynamics study of human fatty acid amide hydrolase

The three-dimensional (3D) model of the human fatty acid amide hydrolase (hFAAH) was constructed based on the crystal structure of the rat FAAH (PDB code 1MT5) in complex with a substrate using Modeller9v2 program. With the aid of molecular mechanics and molecular dynamics method, the last model was obtained and further assessed by Profile-3D, Prosa2003 and Procheck, which confirms that the refined model is reliable. Furthermore, the docking results of propofol and its structural analogue into the active site of hFAAH indicate that 2,6-di-sec-butyl phenol is a more preferred ligand than others, which is in good agreement with the experimental results. From the docking studies, we also suggest that Phe192, Ile238, Thr377, Leu380, Phe381, Phe388 and Leu404 in the hFAAH are seven important determinant residues in binding as they have strong van der Waal interactions with the ligand.

Mining biologically-active molecules for inhibitors of fatty acid amide hydrolase (FAAH): Identification of phenmedipham and amperozide as FAAH inhibitors

The screening of known medicinal agents against new biological targets has been shown to be a valuable approach for revealing new pharmacology of marketed compounds. Recently, carbamate, urea and ketone inhibitors of fatty acid amide hydrolase (FAAH) have been described as promising treatments for pain, anxiety, depression and other CNS-related conditions. In order to find novel FAAH inhibitors, a focused screen of molecules containing potentially reactive moieties or having in vivo effects that are possibly relevant to the biology of FAAH was conducted. These studies revealed phenmedipham 13 and amperozide 14 to be inhibitors of human FAAH, with an IC 50 of 377 nM and 1.34 μM, respectively.

Synthesis and in Vitro Evaluation of N-Substituted Maleimide Derivatives as Selective Monoglyceride Lipase Inhibitors

The endocannabinoid 2-arachidonoylglycerol (2-AG) plays a major role in many physiological processes, and its action is quickly terminated via enzymatic hydrolysis catalyzed by monoglyceride lipase (MGL). Regulating its endogenous level could offer therapeutic opportunities; however, few selective MGL inhibitors have been described so far. Here, we describe the synthesis of N-substituted maleimides and their pharmacological evaluation on the recombinant human fatty acid amide hydrolase (FAAH) and on the purified human MGL. A few N-arylmaleimides were previously described ( Saario , S. M. ; Salo , O. M. ; Nevalainen , T. ; Poso , A. ; Laitinen , J. T. ; Jarvinen , T. ; Niemi , R. Characterization of the Sulfhydryl-Sensitive Site in the Enzyme Responsible for Hydrolysis of 2-Arachidonoylglycerol in Rat Cerebellar Membranes . Chem. Biol. 2005 , 12 , 649 - 656 ) as MGL inhibitors, and along these lines, we present a new set of maleimide derivatives that showed low micromolar IC(50) and high selectivity toward MGL vs FAAH. Then, structure-activity relationships have been investigated and, for instance, 1-biphenyl-4-ylmethylmaleimide inhibits MGL with an IC(50) value of 790 nM. Furthermore, rapid dilution experiments reveal that these compounds act as irreversible inhibitors. In conclusion, N-substituted maleimides constitute a promising class of potent and selective MGL inhibitors.

Fatty acid amide hydrolase as a potential therapeutic target for the treatment of pain and CNS disorders

Background: Fatty acid amide hydrolase (FAAH) is an integral membrane enzyme that hydrolyzes the endocannabinoid anandamide and related amidated signaling lipids. Genetic or pharmacological inactivation of FAAH produces analgesic, anti-inflammatory, anxiolytic and antidepressant phenotypes without showing the undesirable side effects of direct cannabinoid receptor agonists, indicating that FAAH may be a promising therapeutic target. Objectives: This review highlights advances in the development of FAAH inhibitors of different mechanistic classes and their in vivo efficacy. Also highlighted are advances in technology for the in vitro and in vivo selectivity assessment of FAAH inhibitors using activity-based protein profiling and click chemistry-activity-based protein profiling, respectively. Recent reports on structure-based drug design for human FAAH generated by protein engineering using interspecies active site conversion are also discussed. Methods: The literature searches of Medline and SciFinder databases were used. Conclusions: There has been tremendous progress in our understanding of FAAH and development of FAAH inhibitors with in vivo efficacy, selectivity and drug-like pharmacokinetic properties.

Biochemical and Biological Properties of 4-(3-phenyl-[1,2,4] thiadiazol-5-yl)-piperazine-1-carboxylic acid phenylamide, a Mechanism-Based Inhibitor of Fatty Acid Amide Hydrolase

Fatty acid amide hydrolase (FAAH) is an integral membrane enzyme within the amidase-signature family. It catalyzes the hydrolysis of several endogenous biologically active lipids, including anandamide (arachidonoyl ethanolamide), oleoyl ethanolamide, and palmitoyl ethanolamide. These endogenous FAAH substrates have been shown to be involved in a variety of physiological and pathological processes, including synaptic regulation, regulation of sleep and feeding, locomotor activity, pain and inflammation. Here we describe the biochemical and biological properties of a potent and selective FAAH inhibitor, 4-(3-phenyl-[1,2,4]thiadiazol-5-yl)-piperazine-1-carboxylic acid phenylamide (JNJ-1661010). The time-dependence of apparent IC(50) values at rat and human recombinant FAAH, dialysis and mass spectrometry data indicate that the acyl piperazinyl fragment of JNJ-1661010 forms a covalent bond with the enzyme. This bond is slowly hydrolyzed, with release of the piperazinyl fragment and recovery of enzyme activity. The lack of inhibition observed in a rat liver esterase assay suggests that JNJ-1661010 is not a general esterase inhibitor. JNJ-1661010 is >100-fold preferentially selective for FAAH-1 when compared to FAAH-2. JNJ-1661010 dose-dependently increases arachidonoyl ethanolamide, oleoyl ethanolamide, and palmitoyl ethanolamide in the rat brain. The compound attenuates tactile allodynia in the rat mild thermal injury model of acute tissue damage and in the rat spinal nerve ligation (Chung) model of neuropathic pain. JNJ-1661010 also diminishes thermal hyperalgesia in the inflammatory rat carrageenan paw model. These data suggest that FAAH inhibitors with modes of action similar to JNJ-1661010 may be useful clinically as broad-spectrum analgesics.

Divergent Effects of Genetic Variation in Endocannabinoid Signaling on Human Threat- and Reward-Related Brain Function

Fatty acid amide hydrolase (FAAH) is a key enzyme in regulating endocannabinoid (eCB) signaling. A common single nucleotide polymorphism (C385A) in the human FAAH gene has been associated with increased risk for addiction and obesity. Using imaging genetics in 82 healthy adult volunteers, we examined the effects of FAAH C385A on threat- and reward-related human brain function. Carriers of FAAH 385A, associated with reduced enzyme and possibly increased eCB signaling, had decreased threat-related amygdala reactivity but increased reward-related ventral striatal reactivity in comparison with C385 homozygotes. Similarly divergent effects of FAAH C385A genotype were manifest at the level of brain-behavior relationships. The 385A carriers showed decreased correlation between amygdala reactivity and trait anxiety but increased correlation between ventral striatal reactivity and delay discounting, an index of impulsivity. Our results parallel pharmacologic and genetic dissection of eCB signaling, are consistent with the psychotropic effects of Delta(9)-tetrahydrocannabinol, and highlight specific neural mechanisms through which variability in eCB signaling impacts complex behavioral processes related to risk for addiction and obesity.

Search for monoglyceride lipase inhibitors: synthesis and screening of arylthioamides derivatives

Monoglyceride lipase (MGL) is the enzyme responsible for the termination of 2-arachidonoylglycerol (2-AG) signalling, an endogenous ligand for the G-protein coupled cannabinoid receptors CB1 and CB2. Its known abundance and physiological roles emphasize the interest of MGL as an attractive therapeutic target. Search for MGL inhibitors was undertaken by screening an arylthioamide series. The evaluation of arylthioamides derivatives activity as MGL inhibitors measured by the hydrolysis of [H-3]-2-oleoylglycerol by human purified MGL led to the identification of (2-chloro-phenyl)-morpholin-4-yl-methanethione (2) and (3-nitro-phenyl) morpholin-4-yl-methanethione (12), which moreover exhibit good selectivity compared with human fatty acid amide hydrolase inhibition.

Structure-guided inhibitor design for human FAAH by interspecies active site conversion

The integral membrane enzyme fatty acid amide hydrolase (FAAH) hydrolyzes the endocannabinoid anandamide and related amidated signaling lipids. Genetic or pharmacological inactivation of FAAH produces analgesic, anxiolytic, and antiinflammatory phenotypes but not the undesirable side effects of direct cannabinoid receptor agonists, indicating that FAAH may be a promising therapeutic target. Structure-based inhibitor design has, however, been hampered by difficulties in expressing the human FAAH enzyme. Here, we address this problem by interconverting the active sites of rat and human FAAH using site-directed mutagenesis. The resulting humanized rat (h/r) FAAH protein exhibits the inhibitor sensitivity profiles of human FAAH but maintains the high-expression yield of the rat enzyme. We report a 2.75-A crystal structure of h/rFAAH complexed with an inhibitor, N-phenyl-4-(quinolin-3-ylmethyl)piperidine-1-carboxamide (PF-750), that shows strong preference for human FAAH. This structure offers compelling insights to explain the species selectivity of FAAH inhibitors, which should guide future drug design programs.

9.22 A Human Fatty Acid Amide Hydrolase (FAAH) Functional Gene Variant Is Associated with Lower Blood Pressure in Young Males

9.22 A Human Fatty Acid Amide Hydrolase (FAAH) Functional Gene Variant Is Associated with Lower Blood Pressure in Young Males

A Human Fatty Acid Amide Hydrolase (FAAH) Functional Gene Variant Is Associated With Lower Blood Pressure in Young Males

Fatty acid amide hydrolase (FAAH) inhibitors, preventing endocannabinoid (EC) degradation, reduce blood pressure (BP) and heart rate in young male (YM) hypertensive rodents. The functional human FAAH 129T gene variant results in reduced protein level and enzymatic activity but its relationship with BP is unknown. This study investigates the relationship among FAAH P129T alleles and cardiovascular features in YMs at baseline and after 9-year follow-up, and in older male obese hypertensive (OH) patients, in whom the EC system (ECS) is overactive. Genotype analysis was performed in 215 Caucasian male students (24 (0.2) years old) and in 185 older OH patients (50 (0.2) years old). YMs were also followed up for 9 years. Clinical and anthropometric variables, BP, cardiac and carotid artery echographic measurements were evaluated. YMs with the FAAH 129T allele had lower systolic (P = 0.042) and mean BP (P = 0.022), and a trend toward lower diastolic BP (P = 0.06). Such significant association was maintained at follow-up. In contrast, the same allele was not associated with BP in older OH. No association was found with other cardiac and vascular variables. An FAAH defective gene variant results in lower BP in YMs, similar to the findings in young rodents. This effect is lost in older OH patients. Because cannabinoid CB1 receptor blockade is associated with BP reduction in OH patients, EC effects and the use of ECS-interfering drugs is likely to be age and clinical-condition dependent.

Influence of sulfur oxidation state and steric bulk upon trifluoromethyl ketone (TFK) binding kinetics to carboxylesterases and fatty acid amide hydrolase (FAAH)

Carboxylesterases metabolize numerous exogenous and endogenous ester-containing compounds including the chemotherapeutic agent CPT-11, anti-influenza viral agent oseltamivir, and many agrochemicals. Trifluoromethyl ketone (TFK)-containing compounds with a sulfur atom β to the ketone moiety are some of the most potent carboxylesterase and amidase inhibitors identified to date. This study examined the effects of alkyl chain length (i.e., steric effects) and sulfur oxidation state upon TFK inhibitor potency (IC 50) and binding kinetics ( k i). The selective carboxylesterase inhibitor benzil was used as a non-TFK containing control. These effects were examined using two commercial esterases (porcine and rabbit liver esterase) and two human recombinant esterases (hCE-1 and hCE-2) as well as human recombinant fatty acid amide hydrolase (FAAH). In addition, the inhibition mechanism was examined using a combination of 1H NMR, X-ray crystallography, and ab initio calculations. Overall, the data show that while sulfur oxidation state profoundly affects both inhibitor potency and binding kinetics, the steric effects dominate and override the contributions of sulfur oxidation. In addition, the data suggest that inclusion of a sulfur atom β to the ketone contributes an increase (∼5-fold) in inhibitor potency due to effects upon ketone hydration and/or intramolecular hydrogen bond formation. These results provide further information on the nature of the TFK binding interaction and will be useful in increasing our understanding of this basic biochemical process.

Genetic variation in endocannabinoid metabolism, gastrointestinal motility, and sensation

Cannabinoid agonist inhibits gastrointestinal motility. The endocannabinoid, anandamide, is inactivated by fatty acid amide hydrolase (FAAH). A single nucleotide polymorphism in the human FAAH gene (C385A) reduces FAAH expression. Our aim was to evaluate associations between FAAH genotype variation and symptom phenotype, gastric emptying and volume, colonic transit, and rectal sensation in patients with functional gastrointestinal disorders (FGID). 482 FGID patients [Rome II positive, 159 constipation disorders, 184 diarrhea disorders (D-IBS), 86 mixed bowel function (M-IBS), 20 chronic abdominal pain (CAP), 33 functional dyspepsia], and 252 healthy volunteers (HV) underwent questionnaires and studies of phenotype and genotype from 2000 to 2007: 250 gastric emptying, 210 fasting and postprandial gastric volume, 152 colonic transit, and 123 rectal sensation. All had FAAH genotype [CC vs. polymorphic (CA/AA)] determined by TaqMan. FAAH genotype distribution of FGID patients and HV did not deviate from Hardy-Weinberg equilibrium. There was a significant association of FAAH genotype with FGID phenotype (overall chi(2), P = 0.011) and with specific individual phenotypes (P = 0.048). Thus FAAH CA/AA increases the odds (relative to HV) for D-IBS (P = 0.008), M-IBS (P = 0.012), and, possibly, CAP (P = 0.055). There was a significant association of FAAH CA/AA genotype with accelerated colonic transit in D-IBS (P = 0.037). There was no association of FAAH genotype with rectal sensation thresholds or ratings. The association of genetic variation in metabolism of endocannabinoids with symptom phenotype in D-IBS and M-IBS and with faster colonic transit in D-IBS supports the hypothesis that cannabinoid mechanisms may play a role in the control of colonic motility in humans and deserve further study.

Rapid screening for potentially relevant polymorphisms in the human fatty acid amide hydrolase gene using Pyrosequencing™

Introduction Fatty acid amides such as the endocannabinoid anandamide serve as mammalian lipid transmitters in various physiological and pathophysiological processes including inflammation. They are rapidly degraded by the fatty acid amide hydrolase (FAAH). Non-functional FAAH resulted in reduced inflammatory and nociceptive responses. Evidence suggests that human genetic FAAH variants modulate pain and addiction but their clinical role is still poorly known. We therefore developed reliable high-throughput screening assays for FAAH polymorphisms to facilitate research of their clinical role. Materials and methods Six simplex Pyrosequencing™ assays were developed for FAAH polymorphisms dbSNP rs932816, rs4141964, rs324420, rs324419, rs2295633 and rs12029329 spanning the whole FAAH gene. They are frequent or have been functionally associated. Assays were established and validated in DNA samples from 350 healthy unrelated Caucasians. Results In all 350 DNA samples the six FAAH polymorphisms were identified correctly as verified by control samples obtained by conventional sequencing. The observed frequencies of homozygous, heterozygous and non-carriers of the minor alleles were in agreement with the Hardy–Weinberg equilibrium. Minor allelic frequencies were: rs932816G > A = 0.26, rs4141964C > T = 0.37, rs324420C > A = 0.20, rs324419C > T = 0.15, rs2295633G > A = 0.35 and rs12029329G > C = 0.25. SNPs were in high linkage except between rs324419 and rs12029329. One single haploblock was identified, spanning either the whole gene range or excluding rs12029329 in the 3′ region, depending on the statistical procedure of haloblock assignment. Conclusion The presently developed Pyrosequencing™ assays allow for quick and reliable detection of FAAH genotypes and may facilitate investigations of FAAH genetic functional associations.

A MBP-FAAH fusion protein as a tool to produce human and rat fatty acid amide hydrolase: expression and pharmacological comparison

Fatty acid amide hydrolase (FAAH), a membrane-anchored enzyme responsible for the termination of endocannabinoid signalling, is an attractive target for treating conditions such as pain and anxiety. Inhibitors of the enzyme, optimized using rodent FAAH, are known but their pharmacology and medicinal chemistry properties on the human FAAH are missing. Therefore recombinant human enzyme would represent a powerful tool to evaluate new drug candidates. However, the production of high amounts of enzyme is hampered by the known refractiveness of FAAH to overexpression. Here, we report the successful overexpression of rat and human FAAH as a fusion to the E. coli maltose-binding protein, retaining catalytic properties of native FAAH. Several known FAAH inhibitors were tested and differences in their potencies toward the human and rat FAAH were found, underscoring the importance of using a human FAAH in the development of inhibitors.

A Second Fatty Acid Amide Hydrolase with Variable Distribution among Placental Mammals

Fatty acid amides constitute a large and diverse class of lipid transmitters that includes the endogenous cannabinoid anandamide and the sleep-inducing substance oleamide. The magnitude and duration of fatty acid amide signaling are controlled by enzymatic hydrolysis in vivo. Fatty acid amide hydrolase (FAAH) activity in mammals has been primarily attributed to a single integral membrane enzyme of the amidase signature (AS) family. Here, we report the functional proteomic discovery of a second membrane-associated AS enzyme in humans that displays FAAH activity. The gene that encodes this second FAAH enzyme was found in multiple primate genomes, marsupials, and more distantly related vertebrates, but, remarkably, not in a number of lower placental mammals, including mouse and rat. The two human FAAH enzymes, which share 20% sequence identity and are referred to hereafter as FAAH-1 and FAAH-2, hydrolyzed primary fatty acid amide substrates (e.g. oleamide) at equivalent rates, whereas FAAH-1 exhibited much greater activity with N-acyl ethanolamines (e.g. anandamide) and N-acyl taurines. Both enzymes were sensitive to the principal classes of FAAH inhibitors synthesized to date, including O-aryl carbamates and alpha-keto heterocycles. These data coupled with the overlapping, but distinct tissue distributions of FAAH-1 and FAAH-2 suggest that these proteins may collaborate to control fatty acid amide catabolism in primates. The apparent loss of the FAAH-2 gene in some lower mammals should be taken into consideration when extrapolating genetic or pharmacological findings on the fatty acid amide signaling system across species.

A high throughput fluorescent assay for measuring the activity of fatty acid amide hydrolase

Fatty acid amide hydrolase (FAAH) is the enzyme responsible for the rapid degradation of fatty acid amides such as the endocannabinoid anandamide. Inhibition of FAAH activity has been suggested as a therapeutic approach for the treatment of chronic pain, depression and anxiety, through local activation of the cannabinoid receptor CB1. We have developed a high throughput screening assay for identification of FAAH inhibitors using a novel substrate, decanoyl 7-amino-4-methyl coumarin (D-AMC) that is cleaved by FAAH to release decanoic acid and the highly fluorescent molecule 7-amino-4-methyl coumarin (AMC). This assay gives an excellent signal window for measuring FAAH activity and, as a continuous assay, inherently offers improved sensitivity and accuracy over previously reported endpoint assays. The assay was validated using a panel of known FAAH inhibitors and purified recombinant human FAAH, then converted to a 384 well format and used to screen a large library of compounds (>600,000 compounds) to identify FAAH inhibitors. This screen identified numerous novel FAAH inhibitors of diverse chemotypes. These hits confirmed using a native FAAH substrate, anandamide, and had very similar rank order potency to that obtained using the D-AMC substrate. Collectively these data demonstrate that D-AMC can be successfully used to rapidly and effectively identify novel FAAH inhibitors for potential therapeutic use.

The fatty acid amide hydrolase 385 A/A (P129T) variant: haplotype analysis of an ancient missense mutation and validation of risk for drug addiction

The human fatty acid amide hydrolase (FAAH) missense mutation c.385 C-->A, which results in conversion of a conserved proline residue to threonine (P129T), has been associated with street drug use and problem drug abuse. Although a link between the FAAH P129T variant and human drug abuse has been reported, the extent of risk and specific types of substance addiction vulnerability remain to be determined. Here, we investigated the relationship of the FAAH P129T variant to a number of linked single nucleotide polymorphisms to establish a haplotyping system, calculate the estimated age and origin of the FAAH 385 C-->A mutation and evaluate its association with clinically significant drug addiction in a case control study. The results showed a significant over-representation of the FAAH P129T homozygotes in 249 subjects with documented multiple different drug addictions compared to drug free individuals of the same ethnic backgrounds (P = 0.05) using logistic regression analysis controlling for ethnicity. To increase the logistic regression analysis power by increasing the sample size, the data from our previous study (Sipe et al. in Proc Natl Acad Sci USA 99:8394-8399, 2002) were pooled with the present cohort which increased the significance to P = 0.00003. Investigation of the FAAH chromosomal backgrounds of the P129T variant in both multiple different drug addicted and control subjects revealed a common ancestral haplotype, marked population differences in haplotype genetic diversity and an estimated P129T mutation age of 114,425-177,525 years. Collectively, these results show that the P129T mutation is the only common mutation in the FAAH gene and is significantly associated with addictive traits. Moreover, this mutation appears to have arisen early in human evolution and this study validates the previous link between the FAAH P129T variant and vulnerability to addiction of multiple different drugs.