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Inhibition of monoacylglycerol lipase by troglitazone, N-arachidonoyl dopamine and the irreversible inhibitor JZL184: comparison of two different assays
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience.
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience.
Umeå University, Faculty of Medicine, Department of Pharmacology and Clinical Neuroscience, Pharmacology.
2010 (English)In: British Journal of Pharmacology, ISSN 0007-1188, E-ISSN 1476-5381, Vol. 161, no 7, 1512-1526 p.Article in journal (Refereed) Published
Abstract [en]

BACKGROUND AND PURPOSE: Drugs used clinically usually have a primary mechanism of action, but additional effects on other biological targets can contribute to their effects. A potentially useful additional target is the endocannabinoid metabolizing enzyme monoacylglycerol lipase (MGL). We have screened a range of drugs for inhibition of MGL and compared the observed potencies using different MGL enzyme assays.

EXPERIMENTAL APPROACH: MGL activity was screened using recombinant human MGL (cell lysates and purified enzyme) with 4-nitrophenyl acetate (NPA) as substrate. 2-Oleolyglycerol metabolism by rat cerebellar cytosolic MGL and by recombinant MGL was also investigated.

KEY RESULTS: Among the 96 compounds screened in the NPA assay, troglitazone, CP55,940, N-arachidonoyl dopamine and AM404 inhibited NPA hydrolysis by the lysates with IC(50) values of 1.1, 4.9, 0.78 and 3.1µM, respectively. The potency for troglitazone is in the same range as its primary pharmacological activity, activation of peroxisome proliferator-activated receptor (PPAR) γ. Among PPARγ ligands, the potency order towards human MGL was troglitazone > ciglitazone > rosiglitazone > 15-deoxy-Δ(12,14) -prostaglandin J(2) ≈ CAY 10415 > CAY 10514. In contrast to the time-dependent inhibitor JZL184, the potency of troglitazone was dependent upon the enzyme assay system used. Thus, troglitazone inhibited rat cytosolic 2-oleoylglycerol hydrolysis less potently (IC(50) 41µM) than hydrolysis of NPA by the human MGL lysates.

CONCLUSIONS AND IMPLICATIONS: 'Hits' in screening programmes for MGL inhibitors should be assessed in different MGL assays. Troglitazone may be a useful lead for the design of novel, dual action MGL inhibitors/PPARγ activators.

Place, publisher, year, edition, pages
2010. Vol. 161, no 7, 1512-1526 p.
Keyword [en]
monoacylglycerol lipase; cannabinoid; troglitazone; rosiglitazone; peroxisome proliferator-activated receptor γ
URN: urn:nbn:se:umu:diva-41591DOI: 10.1111/j.1476-5381.2010.00974.xISI: 000283948800008PubMedID: 20735405OAI: diva2:407126
Available from: 2011-03-29 Created: 2011-03-29 Last updated: 2014-08-13Bibliographically approved
In thesis
1. The endocannabinoid system: a translational study from Achilles tendinosis to cyclooxygenase
Open this publication in new window or tab >>The endocannabinoid system: a translational study from Achilles tendinosis to cyclooxygenase
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The endogenous cannabinoids anandamide (arachidonoyl ethanolamide, AEA) and 2-arachidonoyl glycerol (2-AG) exert their effect by activating cannabinoid receptors (CB). These receptors mediate a broad range of physiological functions such as beneficial effects in pain and inflammation, although little is known about the expression of CB receptors in human pain conditions. AEA and 2-AG are short- lived molecules due to their rapid cellular accumulation and metabolism. The enzymes primarily responsible for their degradation are fatty acid amide hydrolase (FAAH) for AEA and monoacylglycerol lipase (MGL) for 2-AG. Inhibition of endocannabinoid metabolism is a potential approach for drug development, and there is a need for the identification of novel compounds with inhibitory effects upon FAAH and MGL.

In Paper I of this thesis, the expression of CB1 receptors in human Achilles tendon was examined. We found expression of CB1 receptors in tenocytes, blood vessel wall as well as in the perineurium of the nerve. A semi-quantitative analysis showed an increase of CB1 receptors in painful human Achilles tendinosis.

In papers II and III, termination of AEA signalling was investigated via inhibition of FAAH. In Paper II, Flu-AM1, an analogue of flurbiprofen, was investigated. The compound inhibited both FAAH and the oxygenation of 2-AG by cyclooxygenase-2. In Paper III the antifungal compound ketoconazole was shown to inhibit the cellular uptake of AEA in HepG2, CaCo-2 and C6 cell lines in a manner consistent with inhibition of FAAH.

The role of FAAH in gating the cellular accumulation of AEA was investigated in Paper IV. FAAH has been shown to control the concentration gradient of AEA across the plasmamembrane in RBL2H3 cells, whereas no such effect is seen in other FAAH-expressing cell lines. To determine whether this effect is assay dependent or due to intrinsic differences between the cell lines, we assayed four cell lines with different levels of FAAH expression using the same methodology. We found that the sensitivity of FAAH uptake inhibition was not dependent on the expression level of FAAH, suggesting that factors other than FAAH are important for uptake.

Paper V is focused on the inhibition of MGL. Prior to this study no selective inhibitors of the enzyme had been described. Thus, we screened a number of compounds for their inhibitory effect on MGL. Troglitazone was found to be an inhibitor of MGL, although its potency was dependent upon the enzyme assay used. 

Place, publisher, year, edition, pages
Umeå: Umeå universitet, 2014. 72 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1663
National Category
Pharmacology and Toxicology
urn:nbn:se:umu:diva-91573 (URN)978-91-7601-089-1 (ISBN)
Public defence
2014-09-05, Hörsal E04 Unod R1, Byggnad 6E, Norrlands universitetssjukhus, Umeå, 09:00 (Swedish)
Available from: 2014-08-15 Created: 2014-08-11 Last updated: 2014-08-14Bibliographically approved

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Björklund, EmmelieNorén, ENilsson, JFowler, Christopher J
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