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Inhibition of prostaglandin E-2 synthesis by SC-560 is independent of cyclooxygenase 1 inhibition

Pharmazentrum Frankfurt, ZAFES, Klinikum der Johann Wolfgang Goethe-Universität Frankfurt, Theodor-Stern-Kai 7, Frankfurt 60590, Germany.
The FASEB Journal (Impact Factor: 5.48). 08/2006; 20(9):1352-60. DOI: 10.1096/fj.05-5346com
Source: PubMed

ABSTRACT Prostaglandin E2 (PGE2) produced by cyclooxygenase-2 (COX-2) and microsomal prostaglandin E2 synthase-1 (mPGES-1) plays an important role in the pathophysiology of inflammation, pain, and fever. We investigated the actions of TNFalpha toward stimulation of PGE2 synthesis in primary spinal cord neurons. TNFalpha induced COX-2 and mPGES-1 expression in neurons, followed by formation of PGE2, which was blocked by a selective COX-2 inhibitor. Surprisingly, the "selective COX-1" inhibitor SC-560 completely inhibited TNFalpha-induced PGE2 synthesis in neurons at nanomolar concentrations. Moreover, SC-560 inhibited PGE2 and thromboxane A2 synthesis in human monocytes and platelets with IC50 of 1.8 nM and 2.5 nM, respectively. SC-560 treatment neither altered TNFalpha-induced COX-2 or mPGES-1 expression nor did the addition of the calcium ionophore A23187 or arachidonic acid reverse the inhibition by SC-560. Moreover, no influence of SC-560 on PGE2 synthase activities or PGE2 transport was seen. Most importantly, SC-560 blocked TNFalpha-induced PGE2 synthesis in COX-1-deficient spinal cord neurons, demonstrating a COX-1-independent inhibition of PGE2 synthesis. Although SC-560 inhibited LPS-induced PGE2 synthesis in neurons and RAW264.7 macrophages in whole cell assays, no inhibition was observed in lysates of the same cells. Taken together our data demonstrate that SC-560 acts at least in some cell types as an unselective COX inhibitor despite its selectivity toward COX-1 under cell-free conditions.

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    • "Separate experiments have been performed to confirm, in addition to earlier studies (Yu et al., 2005), that platelet aggregation and TXB 2 formation using platelet rich plasma from healthy donors is purely dependent on COX-1. Here, platelet rich plasma prepared from 7 donors was pretreated with SC-560 (0.001–1 mmol/l) and 5,5- dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2(5H)- furan one (DFU) (0.1–100 mmol/l), which are highly selective inhibitors of COX-1 and COX-2, respectively (Brenneis et al., 2006; Riendeau et al., 1997). Five minutes later, platelets were stimulated with 1 mmol/l arachidonic acid. "
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    ABSTRACT: Nonsteroidal anti-inflammatory drugs (NSAIDs) may interfere with the anti-platelet activity of aspirin at the level of the platelet cyclooxygenase-1 (COX-1) enzyme. In order to examine the interference of common NSAIDs with the anti-platelet activity of aspirin the human platelet rich plasma from voluntary donors was used for arachidonic acid-induced aggreation and determination of thromboxane synthesis. Further, docking studies were used to explain the molecular basis of the NSAID/aspirin interaction. The experimental results showed that celecoxib, dipyrone (active metabolite), ibuprofen, flufenamic acid, naproxen, nimesulide, oxaprozin, and piroxicam significantly interfere with the anti-platelet activity of aspirin, while diclofenac, ketorolac and acetaminophen do not. Docking studies suggested that NSAIDs forming hydrogen bonds with Ser530, Arg120, Tyr385 and other amino acids of the COX-1 hydrophobic channel interfere with antiplatelet activity of aspirin while non interfering NSAIDs do not form relevant hydrogen bond interactions within the aspirin binding site. In conclusion, docking analysis of NSAID interactions at the COX-1 active site appears useful to predict their interference with the anti-platelet activity of aspirin. The results, demonstrate that some NSAIDs do not interfere with the antiplatelet action of aspirin while many others do and provide a basis for understanding the observed differences among individual non-aspirin NSAIDs.
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    • "However, direct evidence of a decrease in endothelial PGI 2 synthesis caused by COX-1 −/− has not yet been reported. Also, some COX-1 inhibitors may have effects independent of their intended target (Brenneis et al. 2006). Thus, it would also be of interest to determine how COX-1 −/− directly affects vascular PGI 2 synthesis in situ. "
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    ABSTRACT: This study was to determine whether the endothelium of mouse major arteries produces prostacyclin (PGI(2)) and, if so, to determine how PGI(2) affects vasomotor reactivity and whether cyclo-oxygenase-1 (COX-1) contributes to PGI(2) synthesis. Abdominal aortas, carotid and femoral arteries were isolated from wild-type mice and/or those with COX-1 or -2 deficiency (COX-1(-/-); COX-2(-/-)) for biochemical and/or functional analyses. The PGI(2) metabolite 6-keto-PGF(1α) was analysed with high-performance liquid chromatography-mass spectroscopy, while vasoreactivity was determined with isometric force measurement. Results showed that in the abdominal aorta, ACh evoked endothelium-dependent production of 6-keto-PGF(1α), which was abolished by COX-1(-/-), but not by COX-2(-/-). Interestingly, COX-1(-/-) enhanced the dilatation in response to ACh, while PGI(2), which evoked relaxation of the mesenteric artery, caused contraction that was abolished by antagonizing thromboxane prostanoid (TP) receptors in the abdominal aorta. However, the TP receptor agonist U46619 evoked similar contractions in the abdominal aorta and mesenteric artery. Also, antagonizing TP receptors enhanced the relaxation in response to PGI(2) in mesenteric arteries. Real-time PCR showed that the PGI(2) (IP) receptor mRNA level was lower in the abdominal aorta than in mesenteric arteries. In addition, COX-1(-/-) not only abolished the contraction in response to ACh following NO inhibition in abdominal aorta, but also those in the carotid and femoral arteries. These results demonstrate an explicit role for endothelial COX-1 in PGI(2) synthesis and suggest that in given mouse arteries, PGI(2) mediates not dilatation but rather vasoconstrictor activity, possibly due to a low expression or functional presence of IP receptors, which enables PGI(2) to act mainly on TP receptors.
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    • "Interestingly, Ivanov et al. (2002) report downregulation of COX-1 mRNA in hypothalamus and peripheral tissues during the later phases of LPSinduced fever, consistent with COX-1 influences being exerted over particular time domains. Caution is warranted in interpreting the results of such pharmacologic interventions, as off-target effects of isoform-selective COX inhibitors may occur with variations in dose or experimental setting (Brenneis et al., 2006). Supporting the validity of the present findings are the observations that a selective COX-2 inhibitor failed to affect LPS-induced HPA secretory responses or tissue PGE 2 levels at a time point at which COX-1 blockade was effective, and that COX-1-deficient mice displayed a compatible dampening of LPS-induced recruitment of HPA control circuitry. "
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