Biochemically based design of cyclooxygenase-2 (COX-2) inhibitors: Facile conversion of nonsteroidal antiinflammatory drugs to potent and highly selective COX-2 inhibitors

Abstract

All nonsteroidal antiinflammatory drugs (NSAIDs) inhibit the cyclooxygenase (COX) isozymes to different extents, which accounts for their anti-inflammatory and analgesic activities and their gastrointestinal side effects. We have exploited biochemical differences between the two COX enzymes to identify a strategy for converting carboxylate-containing NSAIDs into selective COX-2 inhibitors. Derivatization of the carboxylate moiety in moderately selective COX-1 inhibitors, such as 5,8,11,14-eicosatetraynoic acid (ETYA) and arylacetic and fenamic acid NSAIDs, exemplified by indomethacin and meclofenamic acid, respectively, generated potent and selective COX-2 inhibitors. In the indomethacin series, esters and primary and secondary amides are superior to tertiary amides as selective inhibitors. Only the amide derivatives of ETYA and meclofenamic acid inhibit COX-2; the esters are either inactive or nonselective. Inhibition kinetics reveal that indomethacin amides behave as slow, tight-binding inhibitors of COX-2 and that selectivity is a function of the time-dependent step. Site-directed mutagenesis of murine COX-2 indicates that the molecular basis for selectivity differs from the parent NSAIDs and from diarylheterocycles. Selectivity arises from novel interactions at the opening and at the apex of the substrate-binding site. Lead compounds in the present study are potent inhibitors of COX-2 activity in cultured inflammatory cells. Furthermore, indomethacin amides are orally active, nonulcerogenic, anti-inflammatory agents in an in vivo model of acute inflammation. Expansion of this approach can be envisioned for the modification of all carboxylic acid-containing NSAIDs into selective COX-2 inhibitors.

Structural and Functional Basis of Cyclooxygenase Inhibition

“…For example, the discovery that amide derivatives of indomethacin and several other NSAIDs bind to COX-2 but not COX-1 provides a potential strategy for incorporating additional functionality into COX-2 inhibitors. 73 Our laboratory has had success in tethering fluorophores to indomethacin to generate compounds that retain inhibitory potency against COX-2 in intact cells and selectively image COX-2-expressing cells (but not COX-2-negative cells) (M. J. Uddin and L. J. Marnett, unpublished results). A modification of this strategy would allow the introduction of functionality, such as the inhibition of thromboxane synthesis or antagonism of the thromboxane receptor, induction of Nrf2, or activation of PPARγ, all of which might be anticipated to introduce cardiovascular protective activity.…”

“…84 Modification of the Nsubstituted indole-3-acetic acid framework of indomethacin is also an effective strategy. For example, conversion of the carboxylic acid group to the corresponding ester or amide 73,85,86 as well as to the reverse esters/amides 87 results in selective COX-2 inhibition (Figure 12). Interestingly, the 2′-methyl group of indomethacin is critical for inhibitory potency in the neutral amide and ester series, as the des-methyl derivatives are extremely poor inhibitors of the COX enzymes.…”

“…Interestingly, the 2′-methyl group of indomethacin is critical for inhibitory potency in the neutral amide and ester series, as the des-methyl derivatives are extremely poor inhibitors of the COX enzymes. 73,86 Structureactivity studies with diclofenac analogues indicate that methyl or chlorine substituents on the lower aniline ring in the ortho position are necessary to achieve potent inhibition of COX. 88 An o,o-difluoro substituted analogue is considerably less active, and compounds that are synthesized to resemble hydroxylated metabolites of diclofenac show inhibitory potencies that are 100 times lower than that of diclofenac.…”

Systematic discovery of multicomponent therapeutics

“…The refinement of this approach has resulted in the ability to discover compounds with great selectivity for a chosen protein target. The recent success of Gleevec (imatinib mesylate), an inhibitor of the breakpoint cluster regionabelson (BCR-ABL) kinase, and of selective cyclooxygenase-2 (COX-2) inhibitors Vioxx (rofecoxib) and Celebrex (celecoxib) are evidence that the target-based approach can be successful (5,6).…”

Series Introduction: COX in a crystal ball: current status and future promise of prostaglandin research

“…While the tertiary structures of both COX isozymes are remarkably similar, COX-2 is characterized by a side pocket extension to the hydrophobic channel. Although the initial selective COX-2 inhibitors were discovered with the tools of classical biochemical pharmacology, structural studies reveal their localization in the side pocket (Figure 3), where they interact with slow, tight-binding kinetics (13).…”