Cloning, expression, and selective inhibition of canine cyclooxygenase-1 and cyclooxygenase-2.
Cyclooxygenase (COX) performs the critical initial reaction in the arachidonic metabolic cascade, leading to formation of proinflammatory prostaglandins, thromboxanes, and prostacyclins. The discovery of a second COX isoform (COX-2) associated with inflammation led to agents that selectively inhibit COX-2. Cyclooxygenase-2 inhibitors are also being developed for canine applications. To assess the compound potency on canine enzymes, canine COX-1 and COX-2 were cloned, expressed, and purified. Cyclooxygenase-1 was cloned from a canine kidney complementary DNA (cDNA) library, with 96 % sequence homology to human COX-1. Cyclooxygenase-2 was cloned from canine kidney and lipopolysaccharide-stimulated macrophage cDNA libraries, with a 93 % sequence homology to human COX-2. The arachidonic acid Michaelis constants for canine COX-1 and COX-2 were 4.8 and 6.6 micrometer, respectively, compared with 9.6 and 10.2 micrometer for ovine. Inhibition results indicated that, for all compounds tested, there was no significant difference between potencies determined for canine enzymes and those for human enzymes.
Gierse
James K
JK
Pharmacia Discovery Research, Chesterfield, MO 63198, USA.
Staten
Nicholas R
NR
Casperson
Gerald F
GF
Koboldt
Carol M
CM
Trigg
Jennifer S
JS
Reitz
Beverly A
BA
Pierce
Jennifer L
JL
Seibert
Karen
K
D000595:Amino Acid Sequence; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D003001:Cloning, Molecular; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology*; D004285:Dogs / Q000235:genetics*; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D015723:Gene Library; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D007668:Kidney / Q000201:enzymology; D008565:Membrane Proteins; D016133:Polymerase Chain Reaction; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity
D000894:Anti-Inflammatory Agents, Non-Steroidal; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D007527:Isoenzymes; D008565:Membrane Proteins; D012333:RNA, Messenger; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases