AAPS PharmSci

, 3:15 | Cite as

Low molecular weight protamine (LMWP) as nontoxic heparin/low molecular weight heparin antidote (II): In vitro evaluation of efficacy and toxicity

  • Li-Chien Chang
  • Jun Feng Liang
  • Hsiao-Feng Lee
  • Lai Ming Lee
  • Victor C Yang


Patients undergoing anticoagulation with heparin or low molecular weight heparin (LMWH) require a superior antidote that possesses more selective biological actions and a better safety profile than protamine. We had previously developed 2 low molecular weight protamine (LMWP) fractions (TDSP4 and TDSP5) from thermolysin-digested protamine as potential nontoxie, heparin-neutralizing agents. In this, the second article in this series, studies focused on in vitro evaluation of heparin/LMWH-neutralizing efficacy and putative toxicity. These LMWP fractions, particularly TDSP5, were effective and fully capable of neutralizing a broad spectrum of heparin-induced anticoagulant activities (ie, aPTT, anti-Xa, and anti-IIa activities). Additionally, these LMWP fractions could neutralize the activities of commercial LMWH. As assessed by the anti-Xa assay, TDSP5 was as effective as, although less potent than, protamine in reversing the activity of Mono-Embolex (molecular weight 5000–7000) and 2 other different sizes (molecular weight of 3000 and 5000 d) of LMWH preparations. Furthermore, compared with protamine, TDSP5 exhibited a much-reduced toxicity and thus an improved safety profile, as reflected by its reduced ability to activate the complement system and cross-react with the antiprotamine antibodies, which are 2 primary indices of protamine toxicity.

Key Words

Heparin/LMWH neutralization protamine toxicity aPTT clotting assay anti-Xa assay complement Activation immunogenicity cross-reactivity 


  1. 1.
    Jones GR, Hashim R, Power DM. A comparison of the strength of binding of antithrombin III, protamine and poly(1-lysine) to heparin samples of different anticoagulant activities. Biochim Biophys Acta. 1986;883:69–76.PubMedCrossRefGoogle Scholar
  2. 2.
    Metz S, Horrow JC. Protamine and newer heparin antagonists. In: Stoelting RK. ed. Pharmacology & Physiology in Anesthetic Practice. JB Lippincott Co. Philadelphia, PA. 1994:1–15.Google Scholar
  3. 3.
    Porsche R, Brenner ZR. Allergy to protamine sulfate. Heart Lung. 1999;28:418–428.PubMedCrossRefGoogle Scholar
  4. 4.
    Weiler JM, Gellhaus MA, Carter JG, et al. A prospective study of the risk of an immediate adverse reaction to protamine sulfate during cardiopulmonary bypass surgery. J Allergy Clin Immunol. 1990;85:713–719.PubMedCrossRefGoogle Scholar
  5. 5.
    Levy JH, Zaidan JR, Faraj B. Prospective evaluation of risk of protamine reactions in patients with NPH insulin-dependent diabetes. Anesth Analg. 1986;65:739–742.PubMedCrossRefGoogle Scholar
  6. 6.
    Morel DR, Lowenstein E, Nguyenduy T, et al. Acute pulmonary vasoconstriction and thromboxane release during protamine reversal of heparin anticoagulation in awake sheep: Evidence for the role of reactive oxygen metabolites following nonimmunological complement activation. Circ Res. 1988;62:905–915.PubMedCrossRefGoogle Scholar
  7. 7.
    Sela M. Antigenicity: Some molecular aspects. Science. 1969;166:1365–1374.PubMedCrossRefGoogle Scholar
  8. 8.
    Yang VC, Port FK, Kim JS, Teng CL, Till GO, Wakefield TW. The use of immobilized protamine in removing heparin and preventing protamine-induced complications during extracorporeal blood circulation. Anesthesiology 1991;75:288–297.PubMedCrossRefGoogle Scholar
  9. 9.
    Chang LC, Lee HF, Yang Z, Yang VC. Low molecular weight protamine as nontoxic heparin low molecular weight heparin antidote (I): Preparation and characterization. AAPS PhamSci. 2001: 3 (2) article 17 ( Scholar
  10. 10.
    Byun Y, Singh VK, Yang VC. Low molecular weight protamine: A potential nontoxic heparin antagonist. Thromb Res. 1999;94:53–61.PubMedCrossRefGoogle Scholar
  11. 11.
    Mayer MM, In: Kabat EA, Mayer MM, eds. Expermental immunochemistry (Thomas, Springfield, II., 1961:113.Google Scholar
  12. 12.
    Cooper HN, Paterson Y. Production of antibodies. In: Coligan JE, Kruisbbel AM, Margulies DH, eds. Current protocols in immunology (Green Publishing Associates and Wiley-Interscience. New York, NY. 1991;241–247.Google Scholar
  13. 13.
    Hirsh J, Dalen JE, Deykin D, Poller L. Heparin: Mechanism of action, pharmacokinetics, dosing considerations, monitoring, efficacy, and safety. Chest. 1992;102:337S-351S.PubMedCrossRefGoogle Scholar
  14. 14.
    Oosta GM, Gardner WT, Beeler DL, Rosenberg RD. Multiple functional domains of the heparin molecule. Proc Natl Acad Sci U S A. 1981;78:829–833.PubMedCentralPubMedCrossRefGoogle Scholar
  15. 15.
    Casu B. Structure and biological activity of heparin. Adv Carbohydr Chem Biochem. 1985;43:51–134.PubMedCrossRefGoogle Scholar
  16. 16.
    Rosenberg R. The heparin-antithrombin system: A natural anticoagulant mechanism. In: Colman RW, Marder VJ, Hirsh J, eds. Hemostasis and thrombosis: Basic principles and clinical practice. JB Lippincott, Philadelphia, PA: 1987:1373–1392.Google Scholar
  17. 17.
    Bernat A, Herbert JM. Protamine sulphate inhibits pentasaccharide (sr80027)-induced bleeding without affecting its antithrombotic and anti-factor Xa activity in the rat. Haemostasis. 1996;26:195–202.PubMedGoogle Scholar
  18. 18.
    Buchanan MR, Ofosu FA, Fernandez F, Van Ryn J. Lack of relationship between enhanced bleeding induced by heparin and other sulfated polysaccharides and enhanced catalysis of thrombin inhibition. Semin Thromb Hemost 1986;12:324–327.PubMedCrossRefGoogle Scholar
  19. 19.
    Coccheri S. Low molecular weight heparins: An introduction. Haemostasis. 1990;20(suppl 1):74–80.PubMedGoogle Scholar
  20. 20.
    AHFS Drug Information 1996. Protamine sulfate; 2504-2505.Google Scholar
  21. 21.
    Kirklin JW, Barratt-Boyes BG. Cardiac surgery: Morphology, diagnostic criteria, natural history, techiques, results, and indications. Wiley, New York, NY; 1986.Google Scholar
  22. 22.
    Kirklin JK, Chenoweth DE, Naftel DC, et al. Effects of protamine administration after cardiopulmonary bypass on complement, blood elements, and the hemodynamic state. Ann Thorac Surg. 1986;41:193–199.PubMedCrossRefGoogle Scholar
  23. 23.
    Rent R, Ertel N, Eisenstein R, Gewurz H. Complement activation by interaction of polyanions and polycations. I. Heparin-protamine induced consumption of complement. J Immunol. 1975;114:120–124.PubMedGoogle Scholar
  24. 24.
    Cavarocchi NC, Schaff HV, Orszulak TA, Homburger HA, Schnell WA Jr, Pluth JR. Evidence for complement activation by protamine-heparin interaction after cardiopulmonary bypass. Surgery. 1985;98:525–531.PubMedGoogle Scholar
  25. 25.
    Nell LJ, Thomas JW. Frequency and specificity of protamine antibodies in diabetic and control subjects. Diabetes. 1988;37:172–176.PubMedCrossRefGoogle Scholar

Copyright information

© American Association of Pharmaceutical Scientists 2001

Authors and Affiliations

  • Li-Chien Chang
    • 1
  • Jun Feng Liang
    • 2
  • Hsiao-Feng Lee
    • 2
  • Lai Ming Lee
    • 2
  • Victor C Yang
    • 2
  1. 1.School of PharmacyNational Defense Medical CenterTaipeiTaiwan
  2. 2.College of PharmacyThe University of MichiganAnn Arbor

Personalised recommendations