Prostaglandins and glucocorticoids are potent mediators of inflammation. Non-steroidal anti-inflammatory drugs (NSAIDs) exert their effects by inhibition of prostaglandin production. The pharmacological target of NSAIDs is cyclooxygenase (COX, also known as PGH synthase), which catalyses the first committed step in arachidonic-acid metabolism. Two isoforms of the membrane protein COX are known: COX-1, which is constitutively expressed in most tissues, is responsible for the physiological production of prostaglandins; and COX-2, which is induced by cytokines, mitogens and endotoxins in inflammatory cells, is responsible for the elevated production of prostaglandins during inflammation. The structure of ovine COX-1 complexed with several NSAIDs has been determined. Here we report the structures of unliganded murine COX-2 and complexes with flurbiprofen, indomethacin and SC-558, a selective COX-2 inhibitor, determined at 3.0 to 2.5 A resolution. These structures explain the structural basis for the selective inhibition of COX-2, and demonstrate some of the conformational changes associated with time-dependent inhibition.

Nonsteroidal anti-inflammatory drugs (NSAIDs) currently available for clinical use inhibit both COX-1 and COX-2. This suggests that clinically useful NSAIDs inhibit pro-inflammatory prostaglandins (PGs) derived from the activity of COX-2, as well as PGs in tissues like the stomach and kidney (via COX-1). A new class of compounds has recently been developed (SC-58125) that have a high degree of selectivity for the inducible form of cyxlooxygenase (COX-2) over the constitutive form (COX-1). This unique class of compounds exhibit a timedependent irreversible inhibition of COX-2, while reversibly inhibiting COX-1. The molecular basis of this selectivity was probed by site-directed mutagenesis of the active site of COX-2. The sequence differences in the active site were determined by amino acid replacement of the COX-2 sequences based on the known crystal structure of COX-1, which revealed a single amino acid difference in the active site (valine 509 to isoleucine) and a series of differences at the mouth of the active site. Mutants with the single amino acid substitution in the active site and a combination of three changes in the mouth of the active site were made in human COX-2, expressed in insect cells and purified. The single amino acid change of valine 509 to isoleucine confers selectivity of COX-2 inhibitors in the class of SC-58125 and others of the same class (SC-236, NS-398), while commonly used NSAIDs such as indomethacin showed no change in selectivity. Substitutions of COX-1 sequences in COX-2 at the mouth of the active site of COX-2 did not change the selectivity of SC-58125. This indicates that the single amino acid substitution of isoleucine at position 509 for a valine is sufficient to confer COX-2 selectivity in this example of a diaryl-heterocycle COX inhibitor.Prostaglandin synthase catalyzes two separate reactions; the first being the cyclooxygenase function, which is the addition of molecular oxygen to arachidonic acid to form the unstable PGG 2 , 1 and the second the further conversion of PGG 2 to the more stable PGH 2 by a peroxidase function. Hence, this "cyclooxygenase" (COX) enzyme performs the critical initial reaction in the arachidonic metabolic cascade leading to the formation of the prostaglandins, thromboxane, and prostacyclin (1).Recently, a second form of the COX enzyme was isolated whose expression is inducible by cytokines and growth factors (COX-2) (2-6). This inducible COX-2 is linked to inflammatory cell types and tissues and is believed to be the target enzyme for the anti-inflammatory activity of nonsteroidal anti-inflammatory drugs (NSAIDs) (7-12). NSAIDs currently available for clinical use inhibit both COX-1 and COX-2 (13, 14). This suggests that clinically useful NSAIDs inhibit pro-inflammatory PGs derived from the activity of COX-2, as well as PGs in tissues like the stomach and kidney (via COX-1). These homeostatic PGs are linked to normal gastric and renal function (15). It is possible that a selective COX-2 inhibitor may eliminate the side effects associated with COX...

Six docking programs (FlexX, GOLD, ICM, LigandFit, the Northwestern University version of DOCK, and QXP) were evaluated in terms of their ability to reproduce experimentally observed binding modes (poses) of small-molecule ligands to macromolecular targets. The accuracy of a pose was assessed in two ways: First, the RMS deviation of the predicted pose from the crystal structure was calculated. Second, the predicted pose was compared to the experimentally observed one regarding the presence of key interactions with the protein. The latter assessment is referred to as interactions-based accuracy classification (IBAC). In a number of cases significant discrepancies were found between IBAC and RMSD-based classifications. Despite being more subjective, the IBAC proved to be a more meaningful measure of docking accuracy in all these cases.