COVALENT MODIFICATION OF CYCLOOXYGENASE-2 (COX-2) BY 2-ACETOXYPHENYL ALKYL SULFIDES, A NEW CLASS OF SELECTIVE COX-2 INACTIVATORS

All of the selective COX-2 inhibitors described to date inhibit the is oform by binding tightly but noncovalently at the substrate binding si te. Recently, we reported the first account of selective covalent modi fication of COX-2 by a novel inactivator, 2-acetoxyphenyl hept-2-ynyl sulfide (70) (Science 1998, 280, 1268-1270). Compound 70 selectively i nactivates COX-2 by acetylating the same serine residue that aspirin a cetylates. This paper describes the extensive structure-activity relat ionship (SAR) studies on the initial lead compound 2-acetoxyphenyl met hyl sulfide (36) that led to the discovery of 70. Extension of the S-a lkyl chain in 36 with higher alkyl homologues led to significant incre ases in inhibitory potency. The heptyl chain in 2-acetoxyphenyl heptyl sulfide (46) was optimum for COX-2 inhibitory potency, and introducti on of a triple bond in the heptyl chain (compound 70) led to further i ncrements in potency and selectivity. The alkynyl analogues were more potent and selective COX-2 inhibitors than the corresponding alkyl hom ologues. Sulfides were more potent and selective COX-2 inhibitors than the corresponding sulfoxides or sulfones or other heteroatom-containi ng compounds. In addition to inhibiting purified COX-2, 36, 46, and 70 also inhibited COX-2 activity in murine macrophages. Analogue 36 whic h displayed moderate potency and selectivity against purified human CO X-2 was a potent inhibitor of COX-2 activity in the mouse macrophages. Tryptic digestion and peptide mapping of COX-2 reacted with [1-C-14-a cetyl]-36 indicated that selective COX-2 inhibition by 36 also resulte d in the acetylation of Ser516. That COX-2 inhibition by aspirin resul ted from the acetylation of Ser516 was confirmed by tryptic digestion and peptide mapping of COX-2 labeled with [1-C-14-acetyl]salicyclic ac id. The efficacy of the sulfides in inhibiting COX-2 activity in infla mmatory cells, our recent results on the selectivity of 70 in attenuat ing growth of COX-2-expressing colon cancer cells, and its selectivity for inhibition of COX-2 over COX-1 in vivo indicate that this novel c lass of covalent modifiers may serve as potential therapeutic agents i n inflammatory and proliferative disorders.