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Invited Review

COX-2 Inhibitors and Cardiovascular Risk

Funk, Colin D PhD*; FitzGerald, Garret A MD

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Journal of Cardiovascular Pharmacology: November 2007 - Volume 50 - Issue 5 - p 470-479
doi: 10.1097/FJC.0b013e318157f72d
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Prostaglandins (PGs) are a family of bioactive lipid mediators that are formed from arachidonic acid contained in membranes in virtually all cells of the body.1-4 They are involved in numerous physiological and pathophysiological activities in humans and animals. Prostaglandin H synthase (PGHS), with 2 distinct catalytic activities referred to as cyclooxygenase (COX) and hydroperoxidase, respectively,1-4 carries out the initial transformation of fatty acid. COX is a common way of referring to this prostaglandin-synthesizing enzyme. About 10 years after the initial isolation of PGD2, PGE2, and PGF (the classical prostaglandins), between 1962 and 1964 an unstable compound, TxA2 was identified by Samuelsson and co workers.5-8 Moncada, Bunting, and Vane were the first to describe the formation of a prostaglandin substance, first called PGX, in 1976 as an unstable substance that inhibits platelet aggregation.9,10 Its chemical structure was quickly elucidated and was termed prostacyclin by the end of the same year.11 During the early 1970s, the research of Vane, Flower, and Moncada's group led to the understanding that nonsteroidal antiinflammatory drugs (NSAIDs) can block the synthesis of prostaglandins that represented a monumental advance, explaining the mechanism of action of these most commonly consumed of all drugs.12-14 There are additional enzymes required to transform the PGH2 intermediate derived from COX (PGD, PGE, PGF, PGI, and Tx synthases) into the 5 prostaglandins. Until 1989, researchers were under the distinct impression that there was only 1 COX enzyme or isoform, and this was reinforced by the cloning of only 1 DNA sequence for a COX gene from sheep15,16 and 1 from humans17 during 1988 and 1989.


During the late eighties, 3 separate research groups independently described experiments suggesting that a second form of COX, one that is inducible, might exist.18-21 Using sheep trachea epithelial cultured cells, Holtzman's group18 identified increased COX activity that did not correlate with the only known COX gene known at the time (a 2.8 kb mRNA). Needleman's team19 found that using human dermal fibroblast interleukin-1, an inflammatory cytokine, could potently induce COX activity and that this induction could be regulated by glucocorticoids. They also found that a COX activity in human monocytes could be controlled by the bacterial substance lipopolysaccharide (LPS), and they postulated that “cells may contain two pools of COX, each with a differential sensitivity to LPS or dexamethasone.”20 Han et al21 detected persistent induction of a COX in viral-transformed mouse 3T3 fibroblasts. However, it took a couple of years before other investigators could connect these early results to a second COX enzyme. Thus, 2 research groups working in completely different experimental systems cloned the gene for a second COX isoform.22,23 Simmon's group22 was working with chicken embryo fibroblasts that were infected with a Rous sarcoma virus and identified a COX-encoding gene with about 60% sequence identity to the original COX isoform. Herschmann's group,23-25 on the other hand, identified a COX gene, originally called TIS-10, that was induced by the tumor- promoting substance 12-O-tetradecanoyl phorbol 13-acetate (TPA) in mouse fibroblasts. Around 1992, investigators started referring to the 2 prostaglandin synthase enzymes as PGHS-1 and PGHS-2 or COX-1 and COX-2 when both the human platelet PGHS-1 (COX-1) and human endothelial cell PGHS-2 (COX-2) sequences were cloned and expressed.26-28 In most experimental systems the COX-1 gene does not undergo large changes in gene expression, and this enzyme isoform is often referred to as the housekeeping or constitutive COX, whereas the more recently discovered COX-2 isoform was found to undergo profound regulation by many factors, including cytokines, tumor promoters, inflammatory stimuli, glucocorticoids, etc. As a result, COX-2 is often called the “inducible” or “regulatable” COX isoform.1-4 However, it is now well appreciated that COX-1 may be regulated in its expression while COX-2 expression is constitutive in several organs, including kidney and brain.


Studies in the early 1990s indicated that the inducible COX-2 was largely responsible for prostaglandin formation associated with inflammation, while COX-1 was important for prostaglandins of housekeeping function (eg, gastric cytoprotection).29-31 Thus, the pharmaceutical industry devoted immediate attention to develop selective COX-2 inhibitors (known as coxibs) on the basis of this COX-2 hypothesis. To do this, experimental assays had to be established to discern the relative potencies of inhibition by drugs of COX-1 and COX-2.32-36 Two leading drug candidates for human development were selected by Pfizer (formerly Monsanto Searle, Pharmacia) and Merck and became known as celecoxib and rofecoxib, respectively.37-40

Celecoxib is a sulfonamide-containing 1,5-diarylpyrazole compound that was identified through in vitro screening and in vivo testing from a structural lead compound SC-236 and was referred to as SC-58635 during development.37 Using in vitro assays, celecoxib was found to have 155-fold to 3200-fold selectivity for COX-2 compared to COX-1.41 The sulfonamide group in celecoxib binds within a side-pocket of the COX-2 active site that is not present in COX-1 and confers the selectivity of the drug. The drug binds in a competitive manner at first and then becomes irreversible.41 Selectivity is greatly influenced by the type of assay and preincubation time period with the inhibitor. Using a whole blood assay (WBA), the selectivity ratio for celecoxib is between 8 and 30.36,42-44

Rofecoxib, 4-(4′-methylsulfonylphenyl)-3-phenyl-2-(5H)-furanone, also known as Vioxx, was developed as a highly selective and potent COX-2 inhibitor that showed over 800-fold selectivity in a cell assay system39,43,44 and 35-fold to 272-fold selectivity using the WBA.36,42-44 Various animal studies showed efficacy of rofecoxib in models of inflammation and pain and without GI toxicity.3 Rofecoxib provided dental pain relief on par with NSAIDs (eg, ibuprofen) and significantly greater than placebo.40 Various clinical trials showed efficacy in managing pain in osteoarthritis (OA) and rheumatoid arthritis (RA) without upper gastrointestinal (UGI) complications. The United States Food and Drug Administration approved the therapeutic use of rofecoxib in 1999 at the recommended dose of 25 to 50 mg. The VIoxx Gastrointestinal Outcomes Research study (VIGOR) evaluated about 8000 RA patients in a randomized trial comparing rofecoxib (50 mg/d) to naproxen (500 mg bid) over 9 months.45 This drug was in use for about 5 years when it was removed from the market for reasons mentioned below.

Several coxibs were developed with improved COX-2 selectivity after celecoxib and rofecoxib.3,46-49 These include valdecoxib (Bextra), etoricoxib (Arcoxia), parecoxib and lumiracoxib (Prexige). Parecoxib is a prodrug of valdecoxib that is administered by intravenous or intramuscular injection routes. This drug is converted to the active compound valdecoxib by deoxymethylation.46 Valdecoxib showed efficacy for management of OA, RA, and menstrual pain3 and was approved for use in the United States in 2003, but it has since been withdrawn from the market. The most selective coxib is lumiracoxib and is followed by etoricoxib. The FDA recently failed to approve etoricoxib for sale in the United State, and lumiracoxib is yet to be approved for the US market.



Prostacyclin (PGI2) is a potent inhibitor of platelet aggregation9,10,50 synthesized by human arterial and venous tissues/cells from arachidonic acid, with highest capacity to synthesize prostacyclin in the arterial endothelium.50-52 Prostacyclin can also be made in other tissues of the body both in the blood vessels within these tissues and the tissues themselves as detected by the mRNA for the main prostacyclin-synthesizing enzyme, prostacyclin synthase.50,53 Prostacyclin can both inhibit platelet clumping and disrupt clumps of previously aggregated platelets.54 It can also promote blood vessel wall relaxation (vasodilatation) and modulate vascular tone50,55 and inhibit proliferation of vascular smooth muscle cells.56 Clinically, PGI2 (epoprostanol) and its analogs are used in the treatment of primary pulmonary hypertension and in treatment of portopulmonary hypertension that arises secondary to liver disease.46,55 In the cardiovascular system, this prostaglandin is generally regarded as an important, beneficial homeostatic regulator to limit thrombosis.


Thromboxane, an unstable substance derived from arachidonic acid with a half-life of about 30 seconds, is a potent platelet-aggregating agent synthesized within the platelet.8 This powerful platelet-clumping agent synthesized via COX-1 and thromboxane synthase in platelets amplifies the response to other platelet agonists (eg, collagen and ADP).57,58 Besides its effects on platelets, this compound can also constrict blood vessels59 and promote the proliferation of blood vessel smooth muscle cells.60 Activation of this pathway is important in the context of hemostasis.57,61 The importance of this pathway to thrombosis is attested by the findings that a low-dose aspirin regimen has been used by millions as a means to maintain cardiovascular health via its ability to block platelet COX-1-derived thromboxane production. Many clinical trials have shown the benefit of low-dose aspirin in the primary prevention of acute myocardial infarction and in the secondary prevention of myocardial infarction, ischemic stroke, and vascular death.62-68 Dysregulated thromboxane synthesis in the cardiovascular system is regarded as a detrimental event that can lead to thrombosis.

Thromboxane/Prostacyclin “Balance”

Due to the contrasting biological functions of thromboxane and prostacyclin, the concept of balance in the body was first proposed more than 30 years ago10,69,70 for maintenance of healthy cardiovascular status. Vane and colleagues10 stated, “a balance between the amounts of TXA2 formed by platelets and PGX (prostacyclin) formed by vessels might be critical for thrombus formation.” Data from measurements of prostacyclin and thromboxane metabolites (the main urinary metabolites are referred to as PGI-M and Tx-M, respectively) several years later indicated that prostacyclin and thromboxane are formed in small amounts and that they do not act as circulating hormones, but they exert their actions locally in a so-called paracrine/autocrine manner (ie, in the immediate adjacent cells or same cell of synthesis) during platelet-vascular wall interactions.70-75 Thromboxane production by platelets with its pro-platelet activating properties and vascular prostacyclin with its platelet inhibition properties were the key components to this balance. The importance of these 2 eicosanoids has been a reflection of the role thromboxane plays as an amplifying signal for platelet activation by all recognized agonists and the role of prostacyclin as a restraint on all endogenous compounds that promote platelet activation and vasoconstriction.57,76 Indeed, prostacyclin limits platelet activation by thromboxane in vivo and reduces the thrombotic response to vascular injury in mouse models.77 However, this concept of a balance can represent a simplification of the biological context within the vasculature.76,78 These eicosanoids are 1 component of the many facets of an intricately organized and exquisitely regulated vasculature to maintain cardiovascular health. Other cardioprotective mechanisms of the vascular endothelium, in addition to prostacyclin, include nitric oxide (which has similar anti-platelet aggregation and vascular relaxation properties to prostacyclin), CD39 ecto-ADPase [a molecule on the surface of blood vessels that destroys the platelet proaggregating substance adenosine diphosphate (ADP)], thrombomodulin, heparin-like proteoglycan substances, tissue factor pathway inhibitor, and tissue plasminogen activator.76,79,80 Other prothrombotic agents besides thromboxane are thrombin and ADP as well as substances from a perturbed endothelium, which include tissue factor, plasminogen activator inhibitor, von Willebrand factor, collagen, and P-selectin.76,80

Given the important and opposing biological actions of thromboxane and prostacyclin, there have been attempts to define therapies that might depress or block thromboxane while leaving prostacyclin synthesis intact. Such biochemical selectivity could only partially be attained with conventionally formulated low-dose aspirin,81,82 although exploitation of aspirin action on platelets in the presystemic circulation permitted development of a selective formulation.83


COX-1 was generally regarded as the main enzyme capable of forming both thromboxane and prostacyclin in blood platelets and the vascular wall before studies of human pharmacology that ascribed prostacyclin biosynthesis predominantly to COX-2 instead of COX-1.84 In this study and others, urinary PGI-M metabolite, an index of systemic biosynthesis of prostacyclin, derived mainly (about 65% to 80%) from COX-2 and thromboxane synthesis from COX-1.84-86 These conclusions were drawn initially on the basis of administration of celecoxib (a selective COX-2 inhibitor) and ibuprofen (a mixed COX-1/COX-2 inhibitor). Another parallel study revealed that rofecoxib (Vioxx; MK-966) caused a similar suppression of PGI-M.85 An additional study using nimesulide, a partially selective COX-2 inhibitor, showed that both COX-1 and COX-2 can contribute to prostacyclin production in patients with atherosclerosis.87 These studies, therefore, indicated that COX-2 inhibitors could be harmful in the cardiovascular system by blocking prostacyclin biosynthesis without affecting thromboxane production, in effect yielding a more favorable prothrombotic state.


Loss of Vascular Prostacyclin Synthesis Without Alteration in Platelet Thromboxane Synthesis Enhances Thrombotic Risk, Hypertension, and Atherosclerosis

The most straightforward explanation for the mechanistic basis by which COX-2 inhibition advances cardiovascular hazard is through experiments and clinical studies that show a decrease in COX-2 vascular prostacyclin production without a concomitant change in thromboxane levels. This leads to a state that is predisposed to thrombosis, hypertension, and atherosclerosis. Celecoxib or rofecoxib administration to volunteers causes a 65% to 80% decrease in prostacyclin synthesis from COX-2 on the basis of urinary metabolite measures as mentioned above.84-86 When blood is removed from these volunteers at peak drug concentration, the ability of the platelets to aggregate ex vivo is not affected as it is with a traditional NSAID (ibuprofen and indomethacin) that blocks both COX-1 and COX-2; consistent with this observation was the finding of no effect on platelet thromboxane synthesis from coxibs. These human studies, therefore, indicate that a constraint on the cardiovascular actions of thromboxane would be removed during coxib therapy.47,48,76,84-86

Key to this proposed mechanism is that COX-2-synthesized prostacyclin is indeed derived from important arterial sources. What is the evidence? Supporting data for the importance of COX-2 in vascular tissue comes from several sources. First, the human gene, or more specifically complementary DNA (cDNA) for COX-2, was first cloned from a blood vessel source, namely endothelial cells derived from human umbilical veins (HUVEC).27,28 Second, the COX-2 gene is induced by laminar shear stress using HUVEC, an in vitro model system, for blood vessels.88-90 This stress on the vascular wall is evident throughout most regions of the vascular tree and can be modeled in cultured cell systems.91 Third, COX-2 mRNA and protein can be found in human and mouse blood vessels in the endothelium and smooth muscle cells to varying degrees in normal and atherosclerotic states using techniques of in situ hybridization and immunohistochemistry.76,87,92-97 Fourth, a florid example of the relevance of COX-2 to vascular function is that in mice deficient in the enzyme, the ductus arteriosus (DA), a vessel that shunts blood away from the high -resistance lungs to the aorta during fetal life, remains patent after birth, resulting in neonatal death in ~40% of newborn mice.98 COX-2 generates PGH2 that is converted not only to prostacyclin but also to PGE2, which activates the EP4 prostaglandin receptor. This activation promotes DA closure via hyaluronic acid-mediated intimal cushion formation, a process that involves enhanced smooth muscle migration and proliferation leading to vascular lumen occlusion.99

The vascular endothelium, especially in areas of disturbed laminar shear where susceptibility to atherogenesis is highest, is under a consistent state of chronic low-grade inflammation.100 This may lead to variable responses to enhance COX-2 expression both in the endothelium and within lesional macrophages. In humans, it has not been possible to document the precise in vivo quantitation of COX-1 versus COX-2 in vascular endothelium throughout the arterial network. Immunohistochemistry techniques indicate COX-1 is found predominantly in normal, healthy blood vessels ex vivo.87,92-95 However, since COX-2 is an immediate-early gene that is induced rapidly and the turnover of COX-2 mRNA and protein is rapid and regulated by posttranscriptional mechanisms,23,101,102 it is likely that ex vivo immunohistochemistry measurements from autopsy specimens have not predicted an accurate amount of COX-2 in the vasculature and perhaps also in animal studies, where tissues can be harvested more quickly. Moreover, it is important to highlight that detection of COX isoforms by immunohistochemistry does not predict the amount of vascular prostacyclin being synthesized by each isoform, especially since COX-2 is much more active than COX-1 at low substrate and hydroperoxide concentrations.103 Even if COX-1 were in relative abundance as assessed by immunohistochemistry, along with the caveat of COX-2 stability vis-a vis COX-1,104 a preponderance of prostacyclin synthesis can result from an apparently low level of COX-2 in the vasculature, which has been mimicked in laminar shear stress-treated endothelial cells in vitro.90 Furthermore, in a novel mouse model in which the COX-1 gene was placed under the control of COX-2 gene regulatory elements, significantly less prostacyclin could be generated than by COX-2 as assessed by urinary PGI-M measures, likely due to the differential hydroperoxide and substrate requirements of the 2 COX isoforms.105

COX-2 inhibitors could be harmful in the cardiovascular system by blocking prostacyclin biosynthesis without affecting thromboxane production, in effect yielding a more favorable prothrombotic state.

Vascular tissue is highly dynamic, acting much more than as a passive conduit of blood.80 The endothelial cells lining the vascular wall are dynamic sensors of wall shear stress and exhibit highly sensitive means of regulation in discrete, localized regions of the vasculature.106,107 This vascular heterogeneity at the level of individual endothelial cells has been examined by microarray analysis.107 COX-2 gene expression has been shown to be modulated by disturbed flow in this setting indicating that there are specific microdomains of the vasculature that express COX-2. In these regions, COX-2 is most likely acting as a cardioprotective mechanism that contributes to prostacyclin biosynthesis.

Assessing the cardiovascular risk with COX-2 inhibition is most easily studied in animal models. A genetic model of selective COX-2 inhibition has been established to investigate this matter by introducing a mutation into the COX-2 gene to disrupt COX activity.108,109 These mice and mice treated with 2 selective COX-2 inhibitors [celecoxib or 5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2(5H)-furanone (DFU)] show decreased urinary measures of PGI-M, just like in humans, with little if any effect on Tx-M urinary metabolites.108 These findings establish mice as a useful model for exploring the effects of COX-2 inhibition on mechanisms for increased cardiovascular risk in humans. In 3 different models of thrombosis, either genetic COX-2 deletion or pharmacological treatment (celecoxib or DFU) led to increased thrombosis.108 Moreover, there were also small but significant increases in both systolic and diastolic blood pressures associated with COX-2 inhibition in the same groups of mice.108

In another series of experiments in dogs, COX-2-derived prostacyclin was implicated in the protective response to induced thrombosis by electrolytic injury in the circumflex coronary artery.110 In these studies, oral aspirin administration combined with an endothelial recovery period after injury markedly prolonged the time to vascular occlusion. This prolongation was abrogated with celecoxib. In addition, the vasodilatory response to arachidonic acid was reduced significantly in celecoxib-treated animals.110

Salt-sensitive rats were shown to have an elevation of blood pressure with COX-2 inhibition using NS-398 and a COX-2 inhibitor (SC-58236, a lead compound in the development of celecoxib).111 Moreover, genetic COX-2 deficiency in mice reduced renal medullary blood flow, decreased urine flow, and enhanced the pressor effect of angiotensin II.112 Intravascular thrombosis was associated with COX-2 inhibition (SC-58236), including suppression of prostacyclin in a model of pulmonary hypertension in rats.113 Similarly, selective COX-2 inhibition with NS-398 suppressed prostacyclin (measured as 6-keto-PGF) and enhanced platelet-vessel wall interactions in vivo and platelet adhesion to hamster cheek pouch arterioles.114 Absence of the prostacyclin receptor, known as IP, which transduces the effects of prostacyclin, augmented the response to a thrombogenic stimulus in mice.115

In human coronary artery smooth muscle cells, prostacyclin stimulates thrombomodulin expression that constrains activation of the blood coagulant thrombin.116 Therefore, blockade of prostacyclin with COX-2 inhibition will lead to enhanced platelet activation and resultant thrombosis by promoting assembly of the prothrombinase complex.

Atherosclerosis is a chronic arterial wall inflammatory disease that develops over decades in humans, but the early stages of fatty streak formation can even be detected in the fetus of mothers with hypercholesterolemia.117 Long-term suppression of COX-2-derived prostacyclin by coxibs would be expected to predispose to a gradual elevation in cardiovascular risk over time.76 In atherosclerosis studies in mice, disruption of the prostacyclin receptor IP gave rise to enhanced atherosclerotic lesions; in mice lacking the thromboxane receptor TP, the mice were protected in their development of atherosclerotic lesions.118,119 COX-2 inhibition studied in mouse models of atherosclerosis, on the other hand, has yielded variable effects in terms of lesion development.120-126 The reason for this variation could be attributable to the disparate actions of the different prostaglandins in atherothrombosis.76 Thus, blocking COX-2 downstream at the level of PGE2 synthesis by disruption of the main gene that makes PGE2 (called microsomal PGE synthase-1) can reduce atherothrombosis,108,127 but blocking COX-2-derived prostacyclin is detrimental.118,119 Therefore, during coxib administration the relative contributions of COX-2-derived PGE2 versus PGI2 in the lesion and vicinity of the lesions would be important in determining the outcome of COX-2 inhibition on atherosclerosis. In the setting of acute plaque rupture within a coronary artery, a profound loss of prostacyclin synthesis by a coxib would be expected to result in a severely reduced capacity to dissolve a developing thrombus.

In upsetting prostacyclin synthesis with a COX-2 inhibitor, an important restraint on endogenous mediators with an array of adverse cardiovascular effects is removed.

COX-2 is expressed within the kidney in the macula densa of the juxtaglomerular apparatus and other vascular regions of the kidney and is capable of generating prostacyclin.128-130 Prostacyclin within the kidney can regulate water and salt balance, glomerular filtration, and renal blood flow.130 Moreover, elegant studies in mice have shown that COX-2-derived prostacyclin is a potent renin secretagogue and regulator of renovascular hypertension.131 Loss of prostacyclin capability to signal through its receptor IP132,133 and inhibition of COX-2108 elevate blood pressure and augment the blood pressure response to dietary sodium. In addition, both disruption of the IP receptor and COX-2 inhibition modulate vascular remodeling induced by hemodynamic stress such as hypertension in vivo.76,96

One other important area should be mentioned relating to ischemic preconditioning. This phenomenon elicits marked COX-2 upregulation and activity in the heart that is protective in the latter phases against myocardial stunning and infarction.134 These beneficial actions appear to be mediated via either prostacyclin or PGE2 or perhaps both. This mechanism has been postulated to be integral to the process whereby the heart adapts to stress and an important cardioprotective function of COX-2.134

Chemical Properties of Coxibs

The coxibs, although generally belonging to the diaryl heterocycle class of drugs, each possess unique chemical, pharmacodynamic, and pharmacokinetic properties that influence their distribution in the body and locally within cell membranes within vascular tissue. Rofecoxib, possessing a methyl sulfone moiety, but not celecoxib/valdecoxib with a sulfonamide group, exhibited enhanced susceptibility to oxidative modification of low-density lipoproteins and lipids within membranes in vitro.135,136 This oxidative mechanism was independent of COX activity and could be detected by an elevation of isoprostanes, free radical-catalyzed oxidative stress markers. The prooxidant activity of rofecoxib could be inhibited by an antioxidant. A biophysical approach to this study using small-angle x-ray diffraction indicated that rofecoxib could disrupt the phospholipid headgroup to evoke a disordered conformation of the phospholipid acyl chains.135 This would result in altered packing of lipids within the membrane and a facilitation of lipid peroxidation by enhanced free radical diffusion. Celecoxib, on the other hand, bound the hydrocarbon core of the membrane, consistent with its greater lipophilicity, and did not disorder lipid structure. Thus, chemical properties of the coxibs could influence their relative cardiotoxicity. Whether these chemical property mechanisms are actually at play in vivo is subject to debate because measurement of urinary isoprostane levels at normal dosing with coxibs did not reveal elevation of these oxidative stress markers (FitzGerald GA, et al, unpublished observations).

A recent study found that rofecoxib but not celecoxib inhibited prostacyclin synthase activity.137 The inhibitory effect was not due to altered levels or changes in intracellular localization of the enzyme but was hypothesized to be a direct interaction of the drug with the enzyme by a noncompetitive mechanism perhaps involving the heme site.137

Enhanced Leukotriene Biosynthesis From COX-2 Inhibition Via Shunting of Arachidonic Acid

There have been suggestions 79,138 that COX-2 inhibition could shift arachidonic acid metabolism through the 5-lipoxygenase (5-LO)/leukotriene (LT) pathway by shunting of the common substrate. Recent findings implicate the 5-LO/LT pathway in cardiovascular disease, and this topic has been the subject of several recent reviews.139-141 Although the shunting hypothesis in relation to coxibs is intriguing, it has not yet been experimentally verified in any relevant animal or in vitro system to our knowledge. For this mechanism to work, one would have to envision that endothelial cell-released arachidonate would be taken up by inflammatory cells in intimate rolling/adherent contact with the endothelium to be converted to LTs. 5-LO is not usually expressed in endothelial cells, but it can be induced in some settings142; as a result, this particular mechanism remains speculative.

Modulation of Tissue Factor Expression

Tissue factor (TF), an important modulator of coagulation, influences the etiology of atherosclerosis and thrombosis.143 Prostacyclin can inhibit TF directly in monocytic cells and indirectly in endothelial cells.144 Celecoxib, but not rofecoxib or NS-398, reduced the induction of TF activity by the cytokine TNF-α in human aortic endothelial cells via a c-jun N-terminal kinase (JNK) pathway.145 This effect was observed with only 1 of 3 coxibs, so it was postulated to occur independently of its COX-2 activity via a direct effect on JNK phosphorylation. These results suggest, therefore, heterogeneity of response to coxibs. Rofecoxib would be predicted to have a more exaggerated response to plaque rupture (when TF is exposed) than for celecoxib.

Another mechanism for coxib -dependent prothrombotic actions has recently been proposed.146 Endothelial cell COX-2 metabolism of endocannabinoids coupled to prostacyclin synthase activates the nuclear receptor peroxisomal proliferator-activated receptor (PPAR)-delta that can led to a downregulation of TF expression and activity. Coxibs suppress PPAR-delta activity, which induces TF in vascular endothelium and leads to an elevation in circulating TF activity in vivo.

Differential Sensitivity of Coxibs to Endothelial and Platelet COX Inhibition

A recent article by Mitchell et al147 using cultured human aortic endothelial cells has offered another explanation for the mechanism of coxibs to promote increased thrombotic risk: preferential inhibition of endothelial COX-1-derived prostacyclin formation versus platelet COX-1-derived thromboxane formation. Thus, in static cultures (not exposed to laminar shear stress), only COX-1 was expressed, and COX-2 was present only if induced by an inflammatory stimulus (eg, interleukin-1β). Incubations of the aortic endothelial cells carried out in the presence of human platelet-rich plasma, to mimic drug binding to plasma proteins in vivo, revealed that COX-1 in endothelial cells was much more sensitive to inhibition by coxibs than COX-1 in platelets (2-log units for celecoxib and 1-log unit for rofecoxib). This differential inhibition of COX-1 in the 2 cell types was lost when the cells were homogenized, apparently because the cellular hydroperoxide environment was disrupted. Although coxibs generally are poor inhibitors of purified recombinant COX-1, COX-2 selectivity is lost at high enough concentrations. Moreover, in vitro assay conditions with arachidonate substrate and hydroperoxide strongly influence the ability of coxibs to inhibit COX-1.148 Therefore, these studies propose that drugs like rofecoxib and celecoxib can promote a prothrombotic environment. This is not a result of the fact that they inhibit COX-2 in endothelial cells, but because the postulate of a partisan sensitivity on endothelial prostacyclin formation by COX-1. One concern with this study is the concentration of coxib required to achieve the desired effects on COX-1 inhibition would likely not be attained in individuals taking these drugs. Also, it is unknown also whether the conditions of oxidant stress modeled in vitro pertain under any conditions in vivo.


Taken together, the data support a detrimental effect of COX-2-derived prostacyclin inhibition that would be expected to lead to conditions predisposing to hypertension, increased susceptibility to thrombosis, and atherosclerosis. In upsetting prostacyclin synthesis with a COX-2 inhibitor, an important restraint on endogenous mediators with an array of adverse cardiovascular effects is removed. The consequences of this disruption are particularly heightened in people with compromised endothelial function (eg, atherosclerosis) or renal function (eg, salt retention, fluid imbalances), and administration of a COX-2 inhibitor would be expected to lower further the threshold for cardiovascular prothrombotic events in these individuals. Arteriolar vascular endothelium is an extremely rich source of prostacyclin synthase.52,53 The endothelial cells that line the intimal surface throughout the circulatory tree in an average-sized man are estimated to cover a surface area equivalent to a half dozen tennis courts.80 Taking into account (1) this huge surface area of the vascular endothelium and the great capacity of this tissue to produce prostacyclin, (2) the dynamic nature of endothelium and rapid regulation of COX-2, and (3) that COX-2 inhibitors reduce 65% to 80% of urinary prostacyclin metabolites, indices sensitive to local vascular stimulation and short-term systemic administration of prostacyclin,74,149 we are presented with compelling evidence that suppression of vascular prostacyclin synthesis by COX-2 inhibitors translates into the elevation of cardiovascular risk with these drugs.

Certain questions remain for future studies. Is there pharmacogenetic heterogeneity in the cardiovascular response of individuals to different members of the COX-2 inhibitor class? If so, can this be explained mechanistically? Can more direct approaches to assessment of the precise amounts of COX-1-derived versus COX-2-derived prostacyclin within the coronary vasculature be devised? Will individuals who are taking COX-2 -selective inhibitors long-term and who are at low cardiovascular risk undergo risk transformation, and can we understand at the biochemical, genetic, and pharmacological levels if or why this occurs and identify the patients at emerging risk? The answer to these and other questions will ultimately lead to the answer of another important question posed recently: is it possible to make a COX-2 inhibitor that's safe?150


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