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Thromb Haemost 2007; 98(01): 192-200
DOI: 10.1160/TH07-01-0010
Platelets and Blood Cells
Schattauer GmbH

The active metabolite of prasugrel inhibits ADP-stimulated thrombo-inflammatory markers of platelet activation: Influence of other blood cells, calcium, and aspirin

Andrew L. Frelinger III
1   Center for Platelet Function Studies
3   Pediatrics and
,
Joseph A. Jakubowski
5   Lilly Research Laboratories, Indianapolis, Indiana, USA
,
You FuLi
1   Center for Platelet Function Studies
3   Pediatrics and
,
Marc R. Barnard
1   Center for Platelet Function Studies
3   Pediatrics and
,
Marsha L. Fox
1   Center for Platelet Function Studies
3   Pediatrics and
,
Matthew D. Linden
1   Center for Platelet Function Studies
3   Pediatrics and
,
Atsuhiro Sugidachi
6   Daiichi Sankyo Company Ltd., Tokyo, Japan
,
Kenneth J. Winters
5   Lilly Research Laboratories, Indianapolis, Indiana, USA
,
Mark I. Furman
1   Center for Platelet Function Studies
2   Division of Cardiovascular Medicine, and Departments of
4   Medicine, University of Massachusetts Medical School, Worcester, Massachussetts, USA
,
Alan D. Michelson
1   Center for Platelet Function Studies
3   Pediatrics and
4   Medicine, University of Massachusetts Medical School, Worcester, Massachussetts, USA
› Author AffiliationsFinancial support: This work was supported by a grant to Drs. Frelinger and Michelson from Eli Lilly & Co. and Daiichi Sankyo Co., Ltd. Dr. Furman receives research support from Eli Lilly & Co. and is a member of the Speakers’ Bureau of Sanofi-Aventis. Drs. Jakubowski and Winters are employees of Eli Lilly & Co. Dr. Sugidachi is an employee of Daiichi Sankyo Co., Ltd.
Further Information

Publication History

Received 05 January 2007

Accepted after resubmission 28 March 2007

Publication Date:
29 November 2017 (online)

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Summary

The novel thienopyridine prodrug prasugrel, a platelet P2Y12 ADP receptor antagonist, requires in vivo metabolism for activity. Although pharmacological data have been collected on the effects of prasugrel on platelet aggregation,there are few data on the direct effects of the prasugrel’s active metabolite, R-138727, on other aspects of platelet function. Here we examined the effects of R-138727 on thrombo-inflammatory markers of platelet activation, and the possible modulatory effects of other blood cells, calcium, and aspirin. Blood (PPACK or citrate anticoagulated) from healthy donors pre- and post-aspirin was incubated with R-138727 and the response to ADP assessed in whole blood or platelet-rich plasma (PRP) by aggregometry and flow cytometric analysis of leukocyte-platelet aggregates,platelet surface P-selectin, and GPIIb-IIIa activation. Low-micromolar concentrations of R-138727 resulted in a rapid and consistent in-hibition of these ADP-stimulated thrombo-inflammatory markers.These rapid kinetics required physiological calcium levels, but were largely unaffected by aspirin. Lower IC50 values in whole blood relative to PRP suggested that other blood cells affect ADP-induced platelet activation and hence the net inhibition by R-138727. R-138727 did not inhibit P2Y12-mediated ADP-induced shape change, even at concentrations that completely inhibited platelet aggregation, confirming the specificity of R-138727 for P2Y12. In conclusion, R-138727, the active metabolite of prasugrel, results in rapid, potent, consistent, and selective inhibition of P2Y12-mediated up-regulation of thromboinflammatory markers of platelet activation.This inhibition is enhanced in the presence other blood cells and calcium,but not aspirin.

Keywords

Anticoagulants - aspirin - blood cells - platelets - prasugrel - receptors
 

  • References

  • 1 Niitsu Y, Jakubowski JA, Sugidachi A. et al. Pharmacology of CS-747 (prasugrel, LY640315), a novel, potent antiplateletagent with in vivo P2Y12 receptor antagonist activity. Semin Thromb Hemost 2005; 31: 184-194.
    Thieme ConnectPubMedSearch in Google Scholar
  • 2 Cattaneo M. ADP receptor antagonists. In: Platelets. 2nd ed. San Diego: Elsevier/Academic Press; 2007
    Search in Google Scholar
  • 3 Hasegawa M, Sugidachi A, Ogawa T. et al. Stereoselective inhibition of human platelet aggregation by R-138727, the active metabolite of CS-747 (prasugrel, LY640315), a novel P2Y12 receptor inhibitor. Thromb Haemost 2005; 94: 593-598.
    Thieme ConnectPubMedSearch in Google Scholar
  • 4 Matsushima N, Jakubowski JA, Asai F. et al. Platelet inhibitory activity and pharmacokinetics of prasugrel (CS-747) a novel thienopyridine P2Y12 inhibitor: a multiple-dose study in healthy humans. Platelets 2006; 17: 218-226.
    CrossrefPubMedSearch in Google Scholar
  • 5 Jakubowski JA, Payne CD, Brandt JT. et al. The platelet inhibitory effects and pharmacokinetics of prasugrel after administration of loading and maintenance doses in healthy subjects. J Cardiovasc Pharmacol 2006; 47: 377-384.
    PubMedSearch in Google Scholar
  • 6 Asai F, Jakubowski JA, Naganuma H. et al. Platelet inhibitory activity and pharmacokinetics of prasugrel (CS-747) a novel thienopyridine P2Y12 inhibitor: a single ascending dose study in healthy humans. Platelets 2006; 17: 209-217.
    CrossrefPubMedSearch in Google Scholar
  • 7 Jernberg T, Payne CD, Winters KJ. et al. Prasugrel achieves greater inhibition of platelet aggregation and a lower rate of non-responders compared with clopidogrel in aspirin-treated patients with stable coronary artery disease. Eur Heart J 2006; 27: 1166-1173.
    CrossrefPubMedSearch in Google Scholar
  • 8 Wiviott SD, Antman EM, Winters Kj. etal. Randomized comparison of prasugrel (CS-747, LY640315), a novel thienopyridine P2Y12 antagonist, with clopidogrel in percutaneous coronary intervention: results of the Joint Utilization of Medications to Block Platelets Optimally(JUMBO)-TIMI 26 trial. Circulation 2005; 111: 3366-3373.
    CrossrefPubMedSearch in Google Scholar
  • 9 Sugidachi A, Asai F, Yoneda K. et al. Antiplatelet action of R-99224, an active metabolite of a novel thienopyridine-type G(i)-linked P2T antagonist, CS-747. Br J Pharmacol 2001; 132: 47-54.
    CrossrefPubMedSearch in Google Scholar
  • 10 Michelson AD, Linden MD, Barnard MR. et al. Flow cytometry. In: Platelets. 2nd ed. San Diego: Elsevier/Academic Press; 2007
    Search in Google Scholar
  • 11 Evangelista V, Manarini S, Dell’Elba G. et al. Clopidogrel inhibits platelet-leukocyte adhesion and platelet-dependent leukocyte activation. Thromb Haemost 2005; 94: 568-577.
    Thieme ConnectPubMedSearch in Google Scholar
  • 12 Storey RF, Judge HM, Wilcox RG. et al. Inhibition of ADP-induced P-selectin expression and platelet-leukocyte conjugate formation by clopidogrel and the P2Y12 receptor antagonist AR-C69931MX but not a spirin. Thromb Haemost 2002; 88: 488-494.
    Thieme ConnectPubMedSearch in Google Scholar
  • 13 Leon C, Ravanat C, Freund M. et al. Differential involvement of the P2Y1 and P2Y12 receptors in platelet procoagulant activity. Arterioscler Thromb Vasc Biol 2003; 23: 1941-1947.
    CrossrefPubMedSearch in Google Scholar
  • 14 Leon C, Alex M, Klocke A. et al. Platelet ADP receptors contribute to the initiation of intravascular coagulation. Blood 2004; 103: 594-600.
    CrossrefPubMedSearch in Google Scholar
  • 15 Klinkhardt U, Bauersachs R, Adams J. et al. Clopidogrel but not aspirin reduces P-selectin expression and formation of platelet-leukocyte aggregates in patients with atherosclerotic vascular disease. Clin Pharmacol Ther 2003; 73: 232-241.
    CrossrefPubMedSearch in Google Scholar
  • 16 Xiao Z, Theroux P. Clopidogrel inhibits platelet-leukocyte interactions and thrombinreceptor agonist peptide-induced platelet activation in patients with an acute coronary syndrome. J Am Coll Cardiol 2004; 43: 1982-1988.
    CrossrefPubMedSearch in Google Scholar
  • 17 Molero L, Lopez-Farre A, Mateos-Caceres PJ. et al. Effect of clopidogrel on the expression of inflammatory markers in rabbit ischemic coronary artery. Br J Pharmacol 2005; 146: 419-424.
    CrossrefPubMedSearch in Google Scholar
  • 18 Graff J, Harder S, Wahl O. et al. Anti-inflammatory effects of clopidogrel intake in renal transplant patients: effects on platelet-leukocyte interactions, platelet CD40 ligand expression, and proinflammatory biomarkers. Clin Pharmacol Ther 2005; 78: 468-476.
    CrossrefPubMedSearch in Google Scholar
  • 19 Harding SA, Sarma J, Din JN. et al. Clopidogrel reduces platelet-leucocyte aggregation, monocyte activation and RANTES secretion in type 2 diabetes mellitus. Heart 2006; 92: 1335-1337.
    CrossrefPubMedSearch in Google Scholar
  • 20 Payne CD, Brandt JT, Weerakkody GJ. et al. Superior inhibition of platelet aggregation following a loading dose of CS-747 (prasugrel, LY640315) versus clopidogrel: correlation with the pharmacokinetics of active metabolite generation. J Thromb Haemost 2005; 3: M109.
    PubMedSearch in Google Scholar
  • 21 Hardisty RM, Hutton RA, Montgomery D. et al. Secondary platelet aggregation: aquantitative study. Br J Haematol 1970; 19: 307-319.
    CrossrefPubMedSearch in Google Scholar
  • 22 Zhou L, Schmaier AH. Platelet aggregationtesting in platelet-richplasma: description of procedures with the aim to develop standards in the field. Am J Clin Pathol 2005; 123: 172-183.
    CrossrefPubMedSearch in Google Scholar
  • 23 O’Donnell CJ, Larson MG, Feng D. etal. Genetic and environmental contributions to platelet aggregation: the Framingham heartstudy. Circulation 2001; 103: 3051-3056.
    CrossrefPubMedSearch in Google Scholar
  • 24 Michelson AD, Ellis PA, Barnard MR. et al. Down-regulation of the platelet surface glycoprote in Ib-IX complex in whole blood stimulated by thrombin, adenosine diphosphate, or an in vivo wound. Blood 1991; 77: 770-779.
    PubMedSearch in Google Scholar
  • 25 Heptinstall S, Johnson A, Glenn JR. et al. Adenine nucleotide metabolism in human blood--important roles for leukocytes and erythrocytes. J Thromb Haemost 2005; 3: 2331-2339.
    CrossrefPubMedSearch in Google Scholar
  • 26 Koziak K, Sevigny J, Robson SC. et al. Analysis of CD39/ATP diphosphohydrolase (ATPDase) expression in endothelial cells, platelets and leukocytes. Thromb Haemost 1999; 82: 1538-1544.
    Thieme ConnectPubMedSearch in Google Scholar
  • 27 Dombrowski KE, Trevillyan JM, Cone JC. et al. Identification and partial characterization of anec toATPase expressed by human natural killercells. Biochemistry 1993; 32: 6515-6522.
    CrossrefPubMedSearch in Google Scholar
  • 28 Glenn JR, White AE, Johnson A. et al. Leukocyte count and leukocyte ecto-nucleotidase are major determinants of the effects of adenosine triphosphate and adenosine diphosphate on platelet aggregationin human blood. Platelets 2005; 16: 159-170.
    CrossrefPubMedSearch in Google Scholar
  • 29 El-Omar MM, Islam N, Broekman M. et al. The ratio of ADP-to ATP-ectonucleotidase activity is reducedinpatientswith coronaryartery disease. Thromb Res 2005; 116: 199-206.
    CrossrefPubMedSearch in Google Scholar
  • 30 Leal DB, Streher CA, Neu TN. et al. Characterization of NTPDase(NTPDase 1;ecto-apyrase; ecto-diphosphohydrolase; CD39; EC 3.6.1.5) activity in human lymphocytes. Biochim Biophys Acta 2005; 1721: 9-15.
    CrossrefPubMedSearch in Google Scholar
  • 31 Jin RC, Voetsch B, Loscalzo J. Endogenous mechanisms of inhibition of plateletfunction. Microcirculation 2005; 12: 247-258.
    CrossrefPubMedSearch in Google Scholar
  • 32 Drosopoulos JH. Roles of Asp54 and Asp213 in Ca2+ utilizationbysoluble human CD39/ecto-nucleotidase. Arch Biochem Biophys 2002; 406: 85-95.
    CrossrefPubMedSearch in Google Scholar
  • 33 Abraham EH, Sterling KM, Kim RJ. et al. Erythrocyte membrane ATP binding cassette (ABC) proteins: MRP1 and CFTR as well as CD39 (ecto-apyrase) involved in RBCATP transport and elevated blood plasma ATP of cysticfibrosis. Blood Cells Mol Dis 2001; 27: 165-180.
    CrossrefPubMedSearch in Google Scholar
  • 34 Small DS, Payne CD, Ferguson-Sells LR. et al. Increasing concentrations of prasugrel’s active metabolite produce increased inhibition of platelet aggregation. J Am Coll Cardiol 2006; 47: 379A-380A.
    CrossrefPubMedSearch in Google Scholar
  • 35 McEver R. P-selectin/PSGL-1 and other interactions between platelets, leukocytes and endothelium. In: Platelets. 2nd edn. NewYork: Academic Press; 2007
    Search in Google Scholar
  • 36 Bergmeier W, Wagner DD. Inflammation. In: Platelets. 2nd edn. San Diego: Elsevier/Academic Press; 2007
    Search in Google Scholar