1COX-2 inhibitors:A cautionary taleOctober 1, 2007Molecular interventions in human disease...With whom the herbs have come togetherLike kingly chiefs unto the gathering,That Brahman is called a “healer” (bhisaj),A demon-killer, a plague-dispeller.From the Rg Veda (1500-900 BCE)Datura stramoniumVol. 2, plate 28 from theHortus Indicus MalabaricusPublished 1679Source of scopolamine and atropineAn approach as old as human civilization.2Aspirin from willow barkWritten records of willow bark use in ancient Greek and Arabic medical documentsTraditional medication for pain and fever throughout much of the world“Ethnobotany” - study of the role of plants in human societies“Zoopharmacognosy” - self-medication in animals (best documented in primates)Development of aspirin1830s: Salicin purified from willow bark - too irritating forhuman use1850s: Acetylsalicylic acid synthesized - analgesic,antipyretic, anti-inflammatory1897: Felix Hoffmann synthesizes acetylsalicylic acid forhis arthritic father; Bayer gets the patent1971: John Vane finds that aspirin inhibits prostaglandinsynthesis (Nobel Prize 1982)1970s: Aspirin shown to act by acetylating Serine 530 ofcyclooxygenase (prostaglandin synthase)1988: Physician's Health Study shows daily aspirin reducesincidence of a first heart attack by 40%3Eicosanoid = 20 carbons (compare icosahedron)Various prostaglandins, cell types, cell responses...Regulate smooth muscle contraction (lowering blood pressure, stimulating labor)Mediators of pain and inflammationInhibit gastric acid secretion, protect against gastric ulcersOther GPCRs forprostaglandins signalthrough calciumSome prostaglandinreceptors are nuclearhormone receptors (e.g.PPAR gamma) thatdirectly activate genetranscription4COX reactionsrequires heme, tyrosyl radical formedsecond active site on same polypeptideCOX = cyclooxygenase(aka prostaglandin synthase)Aspirin mechanismNote COVALENT attachment to enzyme, so irreversible inhibitionImportant in platelets: no new gene expression (no nucleus), so onedose of aspirin is effective for the life of the plateletDrawback: can contribute to serious bleedingLow-dose aspirin regimens recommended for decreasing likelihoodof stroke and myocardial infarction in at-risk patientsSerine 530:Near active site, NOT involved in catalysisContrast penicillin, protease inhibitors (via acyl-enzyme intermediate)In vitro, aspirin acetylates many targets5COX structure with indomethacinNSAID = non-steroidal anti-inflammatory drugIbuprofen, naproxen, sulindac, others.NONCOVALENT, competitive enzyme inhibition HemeCyclooxygenase active siteA role for COX inhibitors intreatment of colorectal cancer1983: Polyps regress in patients taking NSAIDS1991-1993: Large scale studies show prophylactic effect ofNSAIDS (40-50% risk reduction) and effectiveness in patientswith APC deficiency (familial adenomatous polyposis)6Why does this work?Maybe targets other than COX?Prostaglandin E2via Wnt/beta-catenin?Castellone et al.Science 2005 310:1504But wait, there’s more...1991: Two different COX enzymesCOX-1 expressed constitutively in many tissuesGenerates protective prostaglandins in the GI tractStimulates platelet activation via thromboxane synthesisCOX-2 induced at sites of injury and inflammation2002: COX-3 (splice variant of COX-1)expressed in CNS, selectively inhibited byacetaminophen (works for pain and fever, no effecton inflammation)7COX-2-specific inhibitorsidentified by drug screeningWork poorly in standard assay(rat kidney or testis extracts)Work great in brain extracts(Vioxx)(Celebrex)combined sales worldwide >$4 billion per year (through 2004)8Change in colorectal polyposis forFAP patients on COX-2 inhibitorsPatients receiving 400 mg of Celecoxib2x daily show improvement in allareas of colonPatients receiving 100 mg of Celecoxib2x daily show trend towardsimprovement in rectum, ascendingcolon, and cecumAre these two areas the first targets ofCOX-2 inhibitor-mediated polypregression? Is the mechanismdifferent for different dosages?Steinbach et al., 2000.N Engl J Med. 342(26):1946-52.News Flash9What happened? Why?Why does COX-2 inhibition carry cardiovascular risk, whileinhibition of both COX-1 and COX-2 does not?Hypothesis: A COX-2 product (prostaglandin I2) is atheroprotectiveSupported in female but not male miceEgan et al., 2004,Science 306: 1964Other possibilities: Rofecoxib-specific effects?Increases susceptibility of LDL and membrane lipids to oxidative modificationWhat happened? Why?Medical vs. policy questionsCOX-2 inhibitors have a clear clinical advantage for the subset ofarthritis patients likely to experience gastric bleedingVioxx and Celebrex were direct-marketed to all arthritis patientswithout clear explanation of the relevance only to patients at riskfor bleeding…millions of otherwise healthy people took anexpensive prescription drug instead of an OTC NSAIDSeveral clinical trials excluded patients with known cardiovascularrisk factors; the study patients were not representative of thetarget populationShould the cardiovascular risk have been clear earlier thanSeptember 2004?10Cumulative meta-analysis of published trialsA large Merck-sponsored trial in 2000 (Bombadier et al., “VIGOR” Vioxxgastrointestinal outcomes) showed definitive evidence for increased risk ofMI; attributed to a “cardioprotective effect” of naproxen.Vioxx was pulled from the market only after data from the much smallerMerck-sponsored APPROVe trial (Adenomatous Polyp Prevention onVioxx) confirmed increased risk relative to placebo.Juni et al. (2004) Lancet 364(9450):2021-9.How can we predict how different people are going torespond to “selective” enzyme inhibitors?Fries S, Grosser T, Price TS, Lawson JA, Kapoor S, DeMarcoS, Pletcher MT, Wiltshire T, FitzGerald, GAMarked interindividual variability in the response to selectiveinhibitors of cyclooxygenase-2.Gastroenterology. 2006 Jan;130(1):55-64.How does altering COX activity affect other importantlife processes?North TC, Goessling W, Walklev CR, Lengerke C, KopaniKR, Lord AM, Weber GJ, Bowman TV, Jang IH, GrosserT, FitzGerald GA, Daley GQ, Orkin SH, Zon LIProstaglandin E2 regulates vertebrate haematopoietic stemcell homeostasis.Nature. 2007 Jun