The cyclooxygenase enzymes (COX-1 and COX-2) catalyze the conversion arachidonic acid (AA) to prostaglandin H2 (PGH2), which is the precursor to biologically active prostanoids. The primary mechanism of action of non-steroidal anti-inflammatory drugs (NSAIDs) is the inhibition of prostaglandin biosynthesis by binding within the active site of the COX enzymes. Naproxen, a non-selective NSAID, has been marketed as analgesic and anti-inflammatory agent for over thirty years. In the work presented herein, the structure and dynamics of naproxen binding to COX is elucidated. We determined a 1.7 Å crystal structure of naproxen complexed to COX-2, which indicates that naproxen is bound similarly to other arylpropionic acid inhibitors with the carboxylate moiety making critical interactions at the base of the active site. Interestingly, we identified a novel interaction at the top of the active site between Trp-387 and the p-methoxy moiety of naproxen. Each of the major functional groups of naproxen is required for inhibitory activity suggesting that the development of more potent and/or COX-2 preferring naproxen analogs may be difficult. However, we have synthesized derivatives of naproxen with single atom modifications at the 6-position resulting in COX-2-preferring inhibitors. The crystal structure of one of these analogs, p-methylthio naproxen, bound to COX-2 suggests that the analogs occupy relatively the same conformation as naproxen within the active site.
COX-2 has the ability to metabolize alternative fatty acid substrates in addition to AA including the endocannabinoids, 2-arachidonoylglycerol (2-AG) and anandamide (AEA). Previous studies have shown that ibuprofen and mefenamic acid are weak, competitive inhibitors of COX-2 mediated AA metabolism, but potent, non-competitive inhibitors of 2-AG oxygenation; this phenomenon was dubbed substrate-selective inhibition. In the present work, we demonstrate that a series of reversible inhibitors are significantly more potent inhibitors of 2-AG oxygenation compared to AA whereas a series of tight-binding inhibitors block the oxygenation of both substrates by COX-2 with comparable potency. Furthermore, (R)-arylpropionates, which were previously thought to lack COX inhibitory activity, are potent inhibitors of COX-2-mediated 2-AG oxygenation. A highly substrate-selective inhibitor may represent a novel analgesic agent that lacks the deleterious side effects associated with the use of traditional NSAIDs.
| Identifer | oai:union.ndltd.org:VANDERBILT/oai:VANDERBILTETD:etd-01142011-132702 |
| Date | 14 January 2011 |
| Creators | Duggan, Kelsey Constance |
| Contributors | Alan R. Brash, Richard N. Armstrong, John. A. Oates, Lawrence J. Marnett, F. Peter Guengerich |
| Publisher | VANDERBILT |
| Source Sets | Vanderbilt University Theses |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.vanderbilt.edu/available/etd-01142011-132702/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to Vanderbilt University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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