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Title page for ETD etd-01142011-132702

Type of Document Dissertation
Author Duggan, Kelsey Constance
Author's Email Address kelsey.c.duggan@vanderbilt.edu
URN etd-01142011-132702
Title Structural and Functional Analysis of Cyclooxygenase-2 Inhibition by Non-Steroidal Anti-Inflammatory Drugs
Degree PhD
Department Biochemistry
Advisory Committee
Advisor Name Title
Lawrence J. Marnett Committee Chair
Alan R. Brash Committee Member
F. Peter Guengerich Committee Member
John. A. Oates Committee Member
Richard N. Armstrong Committee Member
  • naproxen
  • prostaglandins
  • cyclooxygenase
  • non-steroidal anti-inflammatory drugs
Date of Defense 2010-12-06
Availability unrestricted
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.

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