catechol-O-methyltransferase Activity Research Articles

Amelioration of intracerebroventricular streptozotocin induced cognitive impairment by Evolvulus alsinoides in rats: In vitro and in vivo evidence

Evolvulus alsinoides, also known as Shankpushapi, is a commonly used traditional medicine for enhancing memory. We evaluated the in vitro free radical scavenging and enzymes [acetylcholinesterase, butyrylcholinestrase, glycogen synthase kinase-3-β (GSK-3-β), rho kinase (ROCK II), prolyl endopeptidase (PEP), catechol-O-methyl transferase (COMT) and lipoxygenase (LOX)] inhibitory activities of aqueous and hydro-alcoholic extracts of E. alsinoides. Hydro-alcoholic extract of E. alsinoides demonstrated more free radical scavenging activity as compared to aqueous extract. Hydro-alcoholic extract also showed higher cholinesterase, GSK-3-β, ROCK II, PEP, COMT and LOX enzyme inhibitory activities as compared to aqueous extract. Phytochemical analysis revealed more flavanoids in hydro-alcoholic extract as compared to aqueous extract but no significant difference in phenolic content of the two extracts was observed. Based on in vitro data, hydro-alcoholic extract (100, 300 and 500mg/kg, p.o.) was selected for in vivo study in intracerebroventricularly injected streptozotocin (STZ) induced cognitive impairment in male Wistar rats. Elevated plus maze, passive avoidance and Morris water maze were used for assessment of cognitive function on 14th, 21st and 28th day after STZ injection. Oxidative stress parameters (malondialdehyde, reduced glutathione, nitric oxide levels and superoxide dismutase activity), cholinergic dysfunction and rho kinase (ROCK II) expression were studied in cerebral cortex and hippocampus of rat brain at the end of the study. Hydro-alcoholic extract of E. alsinoides dose dependently prevented STZ induced cognitive impairment by reducing the oxidative stress, improving cholinergic function and preventing the increase in rho kinase expression. The results suggest an anti-Alzheimer potential of hydro-alcoholic extract of E. alsinoides.

Influence of catechol-O-methyltransferase Val158Met polymorphism on startle response in the presence of high estradiol levels

AbstractIntroduction: both human and animal studies have shown a somewhat complex COMT-by-sex interaction effect on brain function and dysfunction. A functional variation in the gene coding for the catechol-O-methyltransferase (COMT) enzyme, which metabolizes dopamine and noradrenaline, has been related to executive and emotional functions, and to sex dimorphism. Aim: to investigate if COMT Val158Met genotype influences startle response in pregnant women, given their physiologically elevated estradiol levels. Methods: seventy-three pregnant women were assessed in gestational week 38 for acoustic startle response, measured by electromyography of the blink reflex, during control condition, positive and negative anticipation stimuli, and pleasant and unpleasant image stimuli. A blood sample was taken for measurement of estradiol levels and genetic analysis. Results: the results indicated a COMT Val158Met effect on startle response across all conditions (main effect of genotype, F(2,70=3.58), p=0.033), where Val/Val women displayed higher startle magnitudes than Val/Met carriers (Cohen's d=0.71). No significant difference by genotype was found in affective modulation. The findings also suggested an estrogen dose-dependent effect of COMT Val158Met on startle reflex. Among women with higher pregnancy-induced estradiol levels, Val/Val carriers had markedly higher startle response across conditions than heterozygotes (Cohen's d=1.36; F(4,21=11.07); p=0.003), while this effect was not present in women with estradiol levels under the median concentration. Conclusions: the observed effect of COMT Val158Met by estradiol on overall startle response is likely to be due to a variable noradrenergic transmission depending on COMT activity in a possible interaction with estradiol.

Sexual dimorphisms in the immune system of catechol-O-methyltransferase knockout mice

The enzyme catechol-O-methyltransferase (COMT) is part of the metabolic pathway of 17β-estradiol, converting 2-hydroxyestradiol to 2-methoxyestradiol. We recently showed that administration of the COMT product 2-methoxyestradiol has anti-inflammatory and anti-osteoporotic effects. We have now investigated whether COMT affects the immune system, by immunologically phenotyping COMT deficient (COMT−/−) mice.Immunoglobulin production, T lymphocyte proliferation, NK cell cytotoxicity and oxygen radical production were assessed.In male COMT−/−-mice, the total number of T-, and B-lymphocytes from spleen increased but the T-cell proliferative response decreased. The NK cell population shifted toward less mature cells, leaving cytotoxic capacity unaffected. In COMT−/−-females, a higher frequency of neutrophils was found but the oxygen radical production was unaltered.In conclusion, only minor changes of the immune system were seen in COMT deficient mice, and the changes were usually seen in males. This study provides clues into how COMT activity, and hence gender differences, affects the immune system.

Ultraviolet B radiation differentially modifies catechol‐O‐methyltransferase activity in keratinocytes and melanoma cells

Catechol-O-methyltransferase (COMT) is a ubiquitous enzyme inactivating catecholic compounds. COMT is expressed also in human skin samples, and in melanoma cells it may be cytoprotective. A role of COMT in keratinocytes (HaCat) is unknown. The objective of this study is: to investigate whether ultraviolet-B (UVB) radiation modifies COMT activity in melanocytes and HaCat and whether COMT inhibition plays a role in UVB-induced cell death. Human cell lines of melanotic melanoma (SK-mel-1) and HaCat were used. COMT activity was evaluated under basal conditions and after UVB irradiation (311 nm) at a low (8 mJ/cm(2)) and a high dose (60 mJ/cm(2)). Tolcapone 1 μM was used to inhibit COMT. Both SK-mel-1 and Ha-Cat cells express COMT activity. In SK-mel-1, COMT activity is reduced nearly 50% both 24 h and 48 h after a high dose UVB. In Ha-Cat cells, COMT activity increased 24 h after a high dose UVB but decreased at 48 h. Tolcapone increases significantly the cytotoxic effect of high dose UVB irradiation only in HaCat. High concentrations of tolcapone reduced melanin levels in melanoma cells parallel to reduced cell numbers. Ultraviolet radiation differentially modifies COMT activity in melanoma cells and HaCat. Furthermore, tolcapone increased death of HaCat after irradiation but did not affect melanoma cells.

Identification of the ortho-Benzoquinone Intermediate of 5-O-Caffeoylquinic Acid In Vitro and In Vivo: Comparison of Bioactivation under Normal and Pathological Situations

5-O-Caffeoylquinic acid (5-CQA) is one of the major bioactive ingredients in some Chinese herbal injections. Occasional anaphylaxis has been reported for these injections during their clinical use, possibly caused by reactive metabolites of 5-CQA. This study aimed at characterizing the bioactivation pathway(s) of 5-CQA and the metabolic enzyme(s) involved. After incubating 5-CQA with GSH and NADPH-supplemented human liver microsomes, two types of GSH conjugates were characterized: one was M1-1 from the 1,4-addition of GSH to ortho-benzoquinone intermediate; the other was M2-1 and M2-2 from the 1,4-addition of GSH directly to the α,β-unsaturated carbonyl group of the parent. The formation of M1-1 was cytochrome P450 (P450)-mediated, with 3A4 and 2E1 as the principal catalyzing enzymes, whereas the formation of M2-1 and M2-2 was independent of NADPH and could be accelerated by cytosolic glutathione transferase. In the presence of cumene hydroperoxide, M1-1 formation increased 6-fold, indicating that 5-CQA can also be bioactivated by P450 peroxidase under oxidizing conditions. Furthermore, M1-1 could be formed by myeloperoxidase in activated human leukocytes, implying that 5-CQA bioactivation is more likely to occur under inflammatory conditions. This finding was supported by experiments on lipopolysaccharide-induced inflammatory rats, where a greater amount of M1-1 was detected. In S-adenosyl methionine- and GSH-supplemented human S9 incubations, M1-1 formation decreased by 80% but increased after tolcapone-inhibited catechol-O-methyltransferase (COMT) activity. In summary, the high reactivities of the ortho-benzoquinone metabolite and α,β-unsaturated carbonyl group of 5-CQA to nucleophiles have been demonstrated. Different pathological situations and COMT activities in patients may alter the bioactivation extent of 5-CQA.

Janus Molecule I: Dichotomous Effects of COMT in Neuropathic vs Nociceptive Pain Modalities

The enzyme catechol-O-methyltransferase (COMT) has been shown to play a critical role in pain perception by regulating levels of epinephrine (Epi) and norepinephrine (NE). Although the key contribution of catecholamines to the perception of pain has been recognized for a long time, there is a clear dichotomy of observations. More than a century of research has demonstrated that increasing adrenergic transmission in the spinal cord decreases pain sensitivity in animals. Equally abundant evidence demonstrates the opposite effect of adrenergic signaling in the peripheral nervous system, where adrenergic signaling increases pain sensitivity. Viewing pain processing within spinal and peripheral compartments and determining the directionality of adrenergic signaling helps clarify the seemingly contradictory findings of the pain modulatory properties of adrenergic receptor agonists and antagonists presented in other reviews. Available evidence suggests that adrenergic signaling contributes to pain phenotypes through α(1/2) and β(2/3) receptors. While stimulation of α(2) adrenergic receptors seems to uniformly produce analgesia, stimulation of α(1) or β receptors produces either analgesic or hyperalgesic effects. Establishing the directionality of adrenergic receptor modulation of pain processing, and related COMT activity in different pain models are needed to bring meaning to recent human molecular genetic findings. This will enable the translation of current findings into meaningful clinical applications such as diagnostic markers and novel therapeutic targets for complex human pain conditions.

Abstract 1860: Protective effect of catechol-O-methyltransferase-mediated 4-hydroxyestradiol metabolism in human renal cell cancers

Abstract Long-term exposure to estrogen and its metabolites may play an important role in renal cell carcinogenesis. Catechol-O-methyltransferase (COMT) participates in the estrogen metabolism pathway by neutralizing toxic substances. Although reduced COMT activity has been suggested to be a risk factor for estrogen-associated cancers, no studies have investigated the biological significance of COMT in the pathogenesis of human renal cell cancers (RCC). Using quantitative PCR, Western blotting and immunohistochemical staining, we initially found that COMT levels are significantly decreased in human RCC tissues and cells suggesting a suppressive role of COMT in tumor development. However, transient over-expression of COMT has no effect on the proliferation, apoptosis and cell cycle progression of RCC cell lines. In contrast, when cells over-expressing COMT are treated with its substrate 4-hydroxyestradiol (4-OHE2), growth is inhibited by apoptotic cell death. We also found that COMT over-expression combined with 4-OHE2 induces up-regulation of growth arrest- and DNA damage-inducible protein ≤ (GADD45α). We further show that down-regulation of GADD45α by a siRNA-mediated approach inhibits cell death, indicating the essential role of GADD45α in the underlying mechanism of COMT action in response to 4-OHE2. Finally, 4-methoxyestradiol (4-ME) fully reproduces the anti-proliferative function of COMT with 4-OHE2 by promoting GADD45α induction. Together, our study shows that COMT in the presence of 4-OHE2 prevents RCC cell proliferation by enhancing apoptosis, and that GADD45α plays a critical role in the COMT-mediated inhibition of RCC. These findings will provide new direction for the biomarker detection and corresponding treatment in the prevention of human RCC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1860. doi:1538-7445.AM2012-1860

Serotonin-Induced Hypersensitivity via Inhibition of Catechol O-Methyltransferase Activity

Serotonin-Induced Hypersensitivity via Inhibition of Catechol O-Methyltransferase Activity

144 INHIBITORY EFFECT OF CATECHOL-O-METHYLTRANSFERASE-MEDIATED 4-HYDROXYESTRADIOL METABOLISM ON THE DEVELOPMENT OF HUMAN RENAL CELL CARCERS

You have accessJournal of UrologyKidney Cancer: Basic Research I1 Apr 2012144 INHIBITORY EFFECT OF CATECHOL-O-METHYLTRANSFERASE-MEDIATED 4-HYDROXYESTRADIOL METABOLISM ON THE DEVELOPMENT OF HUMAN RENAL CELL CARCERS Inik Chang, Jan Liu, Shahana Majid, Sharanjot Saini, Mohd Zaman, Soichiro Yamamura, Vary Shahryari, Takeshi Chiyomaru, Guoren Deng, Rajvir Dahiya, and Yuichiro Tanaka Inik ChangInik Chang San Francisco, CA More articles by this author , Jan LiuJan Liu San Francisco, CA More articles by this author , Shahana MajidShahana Majid San Francisco, CA More articles by this author , Sharanjot SainiSharanjot Saini San Francisco, CA More articles by this author , Mohd ZamanMohd Zaman San Francisco, CA More articles by this author , Soichiro YamamuraSoichiro Yamamura San Francisco, CA More articles by this author , Vary ShahryariVary Shahryari San Francisco, CA More articles by this author , Takeshi ChiyomaruTakeshi Chiyomaru San Francisco, CA More articles by this author , Guoren DengGuoren Deng San Francisco, CA More articles by this author , Rajvir DahiyaRajvir Dahiya San Francisco, CA More articles by this author , and Yuichiro TanakaYuichiro Tanaka San Francisco, CA More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2012.02.194AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Long-term exposure to estrogen and its metabolites may play an important role in renal cell carcinogenesis. Catechol-O-methyltransferase (COMT) participates in the estrogen metabolism pathway by neutralizing toxic substances. Although reduced COMT activity has been suggested to be a risk factor for estrogen-associated cancers, no studies have investigated the biological significance of COMT in the pathogenesis of human renal cell cancers (RCC). METHODS Gene expression was examined by quantitative RT-PCR, Western blotting and immunohistochemical staining. Cell proliferation was determined by MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium)-based assay. Apoptosis was analyzed with an Annexin V-FITC/7-amino-actinomycin D staining system. Gene silencing was achieved by siRNA transfection. Gene expression profiling of selected apoptosis-related genes was investigated by PCR array. RESULTS We initially found that COMT levels are significantly decreased in human RCC tissues and cells suggesting a suppressive role of COMT in tumor development. However, transient over-expression of COMT has no effect on the proliferation, apoptosis and cell cycle progression of RCC cell lines. In contrast, when cells over-expressing COMT are treated with its substrate 4-hydroxyestradiol (4-OHE2), growth is inhibited by apoptotic cell death. We also found that COMT over-expression combined with 4-OHE2 induces up-regulation of growth arrest- and DNA damage-inducible protein α (GADD45α). We further show that down-regulation of GADD45α by a siRNA-mediated approach inhibits cell death, indicating the essential role of GADD45α in the underlying mechanism of COMT action in response to 4-OHE2. Finally, 4-methoxyestradiol fully reproduces the anti-proliferative function of COMT with 4-OHE2 by promoting GADD45α induction. CONCLUSIONS Our study shows that COMT in the presence of 4-OHE2 prevents RCC cell proliferation by enhancing apoptosis, and that GADD45α plays a critical role in the COMT-mediated inhibition of RCC. These findings will provide new direction for the biomarker detection and corresponding treatment in the prevention of human RCC. © 2012 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 187Issue 4SApril 2012Page: e60 Advertisement Copyright & Permissions© 2012 by American Urological Association Education and Research, Inc.MetricsAuthor Information Inik Chang San Francisco, CA More articles by this author Jan Liu San Francisco, CA More articles by this author Shahana Majid San Francisco, CA More articles by this author Sharanjot Saini San Francisco, CA More articles by this author Mohd Zaman San Francisco, CA More articles by this author Soichiro Yamamura San Francisco, CA More articles by this author Vary Shahryari San Francisco, CA More articles by this author Takeshi Chiyomaru San Francisco, CA More articles by this author Guoren Deng San Francisco, CA More articles by this author Rajvir Dahiya San Francisco, CA More articles by this author Yuichiro Tanaka San Francisco, CA More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Anti-inflammatory functions of purpurogallin in LPS-activated human endothelial cells

Enzymatic oxidation of commercially available pyrogallol was efficiently transformed to an oxidative product, purpurogallin. Purpurogallin plays an important role in inhibiting glutathione S-transferase, xanthine oxidase, catechol O-methyltransferase activities and is effective in the cell protection of several cell types. However, the anti-inflammatory functions of purpurogallin are not well studied. Here, we determined the effects of purpurogallin on lipopolysaccharide (LPS)-mediated proinflammatory responses. The results showed that purpurogallin inhibited LPS-mediated barrier hyper-permeability, monocyte adhesion and migration and such inhibitory effects were significantly correlated with the inhibitory functions of purpurogallin on LPS-mediated cell adhesion molecules (vascular cell adhesion molecules, intracellular cell adhesion molecule, E-selectin). Furthermore, LPS-mediated nuclear factor-κB (NF-κB) and tumor necrosis factor-α (TNF-α) releases from HUVECs were inhibited by purpurogallin. Given these results, purpurogallin showed its anti-inflammatory activities and could be a candidate as a therapeutic agent for various systemic inflammatory diseases. [BMB reports 2012; 45(3): 200-205].

Quercetin Increased the Antiproliferative Activity of Green Tea Polyphenol (-)-Epigallocatechin Gallate in Prostate Cancer Cells

We previously demonstrated that 50% of (-)-epigallocatechin gallate (EGCG) was present in methylated form (4″-MeEGCG) in human prostate tissue, which is less bioactive. We therefore investigated whether quercetin, a natural inhibitor of catechol-O-methyl transferase (COMT), will inhibit EGCG methylation leading to enhanced antiproliferative activity of EGCG in prostate cancer cells. Incubation with both quercetin and EGCG for 2 h increased the cellular concentrations of EGCG by 4- to 8-fold and 6- to 10-fold in androgen-independent PC-3 cells and androgen-dependent LNCaP cells, respectively. Concurrently, the percent of 4″-MeEGCG in the total EGCG was decreased from 39% to 15% in PC-3 cells and from 61% to 38% in LNCaP cells. Quercetin and EGCG in combination synergistically inhibited cell proliferation, caused cell cycle arrest, and induced apoptosis in PC-3 cells. In LNCaP cells, EGCG and quercetin exhibited a stronger antiproliferative activity leading to an additive effect. The synergistic effect of these 2 agents in PC-3 cells could be based on the fact that EGCG primarily inhibited COMT activity, whereas quercetin reduced the amount of COMT protein. In summary, quercetin combined with EGCG in vitro demonstrated enhanced inhibition of cell proliferation by increasing the intracellular concentration of EGCG and decreasing EGCG methylation.

Psychological distress in fibromyalgia patients: A role for catechol-O-methyl-transferase Val158met polymorphism.

Fibromyalgia (FM) has been related to biochemical alterations, central pain sensitization and psychological distress. Among genetic and environmental hypotheses, a role was suggested for catechol-O-methyl-transferase (COMT), a modulator in the metabolism of monoaminergic neurotransmitters. This study compared the COMT Val158Met enzyme polymorphism (rs4680) of 198 FM patients to 99 pain-free controls. Psychological and functional aspects were assessed through investigating anxiety, depression, catastrophizing, perceived health, and functional status. The distribution of the COMT Val158Met polymorphism was similar in FM and controls. Out of 198 patients, 137 were able to stop medication before evaluation. In these patients, the COMT Val158Met genotype was associated with specific psychological profiles. The Met/Met subgroup scored systematically worse on all psychological and functional variables. All variables displayed a "genotype-trend effect" with the Met/Met and Val/Val subgroups at the two ends of the scores. Genotypes distribution in the 61 patients unable to stop medication was significantly different from that of patients able to stop medication and controls (p = .002 and p = .018, respectively) with an increase in the proportion of the Met/Met genotype associated to the lowest COMT activity. These results suggest a possible role of COMT Val158Met polymorphism in the psychological distress observed in FM. The association of COMT genotype with psychological distress may be of importance as identifying subgroups is a challenge in the diagnosis and treatment of fibromyalgia patients. This association may contribute to open new perspectives into the understanding of the pathophysiology of fibromyalgia and stress-related genes.

COMT Val158Met Genotype Determines the Direction of Cognitive Effects Produced by Catechol-O-Methyltransferase Inhibition

BackgroundCatechol-O-methyltransferase (COMT) metabolizes dopamine. The COMT Val158Met polymorphism influences its activity, and multiple neural correlates of this genotype on dopaminergic phenotypes, especially working memory, have been reported. COMT activity can also be regulated pharmacologically by COMT inhibitors. The inverted-U relationship between cortical dopamine signaling and working memory predicts that the effects of COMT inhibition will differ according to COMT genotype.MethodsThirty-four COMT Met158Met (Met-COMT) and 33 COMT Val158Val (Val-COMT) men were given a single 200-mg dose of the brain-penetrant COMT inhibitor tolcapone or placebo in a randomized, double-blind, between-subjects design. They completed the N-back task of working memory and a gambling task.ResultsIn the placebo group, Met-COMT subjects outperformed Val-COMT subjects on the 2- back, and they were more risk averse. Tolcapone had opposite effects in the two genotype groups: it worsened N-back performance in Met-COMT subjects but enhanced it in Val-COMT subjects. Tolcapone made Met-COMT subjects less risk averse but Val-COMT subjects more so. In both tasks, tolcapone reversed the baseline genotype differences.ConclusionsDepending on genotype, COMT inhibition can enhance or impair working memory and increase or decrease risky decision making. To our knowledge, the data are the clearest demonstration to date that the direction of effect of a drug can be influenced by a polymorphism in its target gene. The results support the inverted-U model of dopamine function. The findings are of translational relevance, because COMT inhibitors are used in the adjunctive treatment of Parkinson's disease and are under evaluation in schizophrenia and other disorders.

Catechol-O-methyltransferase polymorphisms do not play a significant role in pain perception in male Chinese Han population

Polymorphisms in the human catechol-O-methyltransferase (COMT) gene have been widely studied for their role in pain and analgesia. In this study, sensitivity to potassium iontophoresis, visual analog scale measurements for fixed twofold pain threshold stimulation and pain threshold changes induced by transcutaneous electrical acupoint stimulation (TEAS) were assessed in a population of healthy Chinese males. These results were correlated with the alleles of six single nucleotide polymorphisms (SNP) or diplotypes of common haplotypes designated as low pain sensitive, average pain sensitive, and high pain sensitive in the COMT gene of these subjects. Our results reveal that the alleles of each SNP are not significantly correlated with pain perception except for the rs4633 allele in the 2 Hz TEAS session (P < 0.05). In addition, the six diplotypes of COMT haplotypes, which cover 92.5% of the Chinese population, are also not correlated with pain perception. Moreover, there were no significant differences in pain threshold changes induced by 2 and 100 Hz TEAS among the diplotypes of each SNP or the various haplotypes. These results suggest that COMT activity do not play a significant role in pain perception and TEAS-induced analgesia in the Chinese Han male population.

Quercetin increased bioavailability and decreased methylation of green tea polyphenols in vitro and in vivo

The extensive methylation of green tea polyphenols (GTPs) in vivo may limit their chemopreventive potential. We investigated whether quercetin, a natural inhibitor of catechol-O-methyltransferase (COMT) and multidrug resistance proteins (MRPs), will differentially increase the intracellular concentration and decrease the methylation of GTPs in different cancer cell lines. Intrinsic COMT activity was lowest in lung cancer A549 cells, intermediate in kidney 786-O cells and highest in liver HepG2 cells. Quercetin increased the cellular absorption of epigallocatechin gallate (EGCG) four-fold in A549 cells with a decreased methylation rate from 63 to 19%, 2-fold in 786-O cells with a decreased methylation from 97% to 56%, while no significant effect was observed in HepG2 cells. The combination significantly decreased the activity and protein expression of COMT and decreased the protein expression of MRP1 compared to individual treatments. The combination exhibited the strongest increase in antiproliferation in A549 cells, an intermediate effect in 786-O cells and lowest effect in HepG2 cells. The effect of quercetin on bioavailability and metabolism of GTPs was confirmed in vivo. Severe combined immunodeficiency (SCID) mice were administered brewed green tea (GT) and a diet supplemented with 0.4% quercetin alone or in combination for 2 weeks. We observed a 2- to 3-fold increase of total and non-methylated EGCG in lung and kidney and an increasing trend in liver. In summary, combining quercetin with GT provides a promising approach to enhance the chemoprevention of GT. Responses of different cancers to the combination may vary by tissue depending on the intrinsic COMT and MRP activity.

Serotonin-Induced Hypersensitivity via Inhibition of Catechol O-Methyltransferase Activity

The subcutaneous and systemic injection of serotonin reduces cutaneous and visceral pain thresholds and increases responses to noxious stimuli. Different subtypes of 5-hydroxytryptamine (5-HT) receptors are suggested to be associated with different types of pain responses. Here we show that serotonin also inhibits catechol O-methyltransferase (COMT), an enzyme that contributes to modultion the perception of pain, via non-competitive binding to the site bound by catechol substrates with a binding affinity comparable to the binding affinity of catechol itself (Ki = 44 μM). Using computational modeling, biochemical tests and cellular assays we show that serotonin actively competes with the methyl donor S-adenosyl-L-methionine (SAM) within the catalytic site. Binding of serotonin to the catalytic site inhibits the access of SAM, thus preventing methylation of COMT substrates. The results of in vivo animal studies show that serotonin-induced pain hypersensitivity in mice is reduced by either SAM pretreatment or by the combined administration of selective antagonists for β2- and β3-adrenergic receptors, which have been previously shown to mediate COMT-dependent pain signaling. Our results suggest that inhibition of COMT via serotonin binding contributes to pain hypersensitivity, providing additional strategies for the treatment of clinical pain conditions.

Nicotine normalizes event related potentials in COMT-Val-tg mice and increases gamma and theta spectral density.

Regulation of dopamine neurotransmission is essential for cognitive processes. In humans and rodents, the relationship between dopamine signaling and cognitive performance is described as a dose-dependent, inverted-U curve whereby excess or insufficiency of dopamine in prefrontal cortex has detrimental effects. Previous studies have indicated that prefrontal dopamine levels are associated with genetic variation in catechol-O-methyltransferase (COMT), a regulatory enzyme that controls dopamine availability. Furthermore, smokers who carry the high-activity COMT-Val allele are more prone to cognitive deficits and have an increased risk of smoking relapse. The present study employed transgenic mice expressing the human COMT-Val variant to determine the effects of the high-activity COMT allele on electrophysiological markers, including the P20, N40, and P80 components of the auditory event-related potential, as well as baseline and auditory event-related power and phase-synchrony in theta and gamma ranges. We also examined the effects of nicotine on these measures to investigate the potential effects of smoking on COMT-mediated electrophysiological activity. COMT-Val-tg mice displayed increased N40 latency and decreased P80 amplitude as well as reduced baseline theta and gamma power. Nicotine increased P20 and P80 amplitudes, decreased N40 amplitude, increased P20 and N40 latencies, and reduced P80 latency. Nicotine also increased the event-related power and phase synchrony, yielding an increase in signal-to-noise ratio across theta and gamma ranges. COMT activity specifically alters long-latency components of the event-related response. Nicotine restored normal event-related activity among COMT-Val-tg mice, suggesting one mechanism through which nicotine may normalize cognitive function among people with the high-activity allele.

2.244 PHARMACOKINETICS, PHARMACODYNAMICS AND TOLERABILITY OF OPICAPONE, A NOVEL COMT INHIBITOR, DURING FIRST ADMINISTRATION TO HEALTHY MALE SUBJECTS

Aim: Opicapone was developed to fulfill the need for more potent, safer and longer acting catechol-O-methyltransferase (COMT) inhibitors [1]. This study was aimed to assess the tolerability, pharmacodynamics, and pharmacokinetics of single oral doses of opicapone in healthy male subjects. Design: Single rising oral doses of 10mg to 1200mg of opicapone were studied under a double-blind, randomised, placebo-controlled design. Eight sequential groups of 8 subjects were enrolled. Within each group, 6 subjects were randomised to receive opicapone and 2 subjects to receive placebo. Opicapone/placebo was administered after a 10h overnight fasting. Results: The extent and rate of systemic exposure (AUC and Cmax) to opicapone increased in an approximately dose-proportional manner. Despite the relatively short half-life of opicapone (1 to 4h), inhibition of soluble COMT (S-COMT) activity in erythrocytes was long-lasting, ranging from 6% (10mg) to 55% (1200mg) at 72h postdose. Maximum S-COMT inhibition (Emax) occurred between 1 and 6h post-dose (tEmax), and was 34.5%, 71.7%, 93.8%, 96.3%, 100%, 100%, 100% and 100% with the doses of 10, 25, 50, 100, 200, 400, 800 and 1200mg, respectively. Urine levels of opicapone and its metabolites usually remained below the limit of quantification, showing that kidney is not the primary route of excretion. Opicapone was well tolerated at all doses tested. Conclusion: Opicapone was well tolerated at oral single doses of 10mg to 1200mg, presented a dose-proportional kinetics and effectively inhibited erythrocyte soluble COMT activity.

2.245 PHARMACOKINETICS, PHARMACODYNAMICS AND TOLERABILITY OF OPICAPONE, A NOVEL COMT INHIBITOR, DURING MULTIPLE RISING DOSE REGIMENS IN HEALTHY MALE SUBJECTS

Aim: Opicapone was developed to fulfill the need for more potent, safer and longer acting catechol-O-methyltransferase (COMT) inhibitors [1]. This study was aimed to assess the tolerability, pharmacodynamics, and pharmacokinetics of single oral doses of opicapone in healthy male subjects. Design: Single rising oral doses of 10mg to 1200mg of opicapone were studied under a double-blind, randomised, placebo-controlled design. Eight sequential groups of 8 subjects were enrolled. Within each group, 6 subjects were randomised to receive opicapone and 2 subjects to receive placebo. Opicapone/placebo was administered after a 10h overnight fasting. Results: The extent and rate of systemic exposure (AUC and Cmax) to opicapone increased in an approximately dose-proportional manner. Despite the relatively short half-life of opicapone (1 to 4h), inhibition of soluble COMT (S-COMT) activity in erythrocytes was long-lasting, ranging from 6% (10mg) to 55% (1200mg) at 72h postdose. Maximum S-COMT inhibition (Emax) occurred between 1 and 6h post-dose (tEmax), and was 34.5%, 71.7%, 93.8%, 96.3%, 100%, 100%, 100% and 100% with the doses of 10, 25, 50, 100, 200, 400, 800 and 1200mg, respectively. Urine levels of opicapone and its metabolites usually remained below the limit of quantification, showing that kidney is not the primary route of excretion. Opicapone was well tolerated at all doses tested. Conclusion: Opicapone was well tolerated at oral single doses of 10mg to 1200mg, presented a dose-proportional kinetics and effectively inhibited erythrocyte soluble COMT activity.

Catechol- O -methyltransferase-mediated metabolism of 4-hydroxyestradiol inhibits the growth of human renal cancer cells through the apoptotic pathway

Long-term exposure to estrogen and its metabolites may play an important role in renal cell carcinogenesis. Catechol-O-methyltransferase (COMT) participates in the estrogen metabolism pathway by neutralizing toxic substances. Although reduced COMT activity has been suggested to be a risk factor for estrogen-associated cancers, no studies have investigated the biological significance of COMT in the pathogenesis of human renal cell cancers (RCCs). We initially found that COMT levels are significantly decreased in human RCC tissues and cells suggesting it plays a suppressive role in tumor development. However, transient overexpression of COMT has no functional effect on RCC cell lines. In contrast, when cells overexpressing COMT are treated with its substrate 4-hydroxyestradiol (4-OHE(2)), growth is inhibited by apoptotic cell death. We also found that COMT overexpression combined with 4-OHE(2) induces upregulation of growth arrest- and DNA damage-inducible protein α (GADD45α). We further show that downregulation of GADD45α by a small interfering RNA-mediated approach inhibits cell death, indicating the essential role of GADD45α in the underlying mechanism of COMT action in response to 4-OHE(2). Finally, 4-methoxyestradiol fully reproduces the antiproliferative function of COMT with 4-OHE(2) by promoting GADD45α induction. Together, these findings show that COMT in the presence of 4-OHE(2) prevents RCC cell proliferation by enhancing apoptosis and that GADD45α plays a critical role in the COMT-mediated inhibition of RCC.