Low-Affinity Material Does not Contribute to the Antithrombotic Activity of Orgaran (Org 10172) in Human Plasma

Summary

Orgaran is a LMW heparinoid composed of heparan sulphate (83% wlw) of which 4-5% has high affinity for antithrombin, dermatan sulphate (12% w/w) and chondroitin sulphate (59 wlw). To examine the contribution of the low-affinity fraction to Orgaran’s antithrombotic activity we have quantitated the binding of plasma proteins to Orgaran and its component fractions in whole, hirudin-anticoagulated human plasma. Antithrombin, largely bound to the high-affinity fraction, and histidine-rich glycoprotein, interacting with low-affinity components, were the dominant proteins bound to Orgaran. Vitronectin, fibrinogen, fibronectin, heparin cofactor 11, and apolipoprotein B were also detected in small amounts. The ratio of bound antithrombin, histidine- rich glycoprotein and vitronectin to GAG was negatively correlated with the Orgaran concentration in plasma, implying that the efficacy of Orgaran may not be linearly related to dose. Binding of antithrombin to the high-affinity fraction was not decreased by other plasma proteins or affected by addition of low-affinity material. Moreover, the antithrombin and anti-factor Xa activities of the high-affinity material were unaltered by low-affinity GAGs. On the basis of our results we conclude that the low-affinity material does not contribute to the antithrombotic activity of Orgaran by binding non-anticoagulant plasma proteins and releasing the high-affinity chains to interact with antithrombin and its target proteinases.

• References

• 1 Dedem Gvan, de Leeuw den Bouter H. The nature of the glucosaminoglycan in Orgaran (Org 10172). Thromb Haemost 1993; 69: 652
• 2 Turpie ACG, Levine MN, Hirsh J, Carter CJ, Jay RM, Powers PJ, Andrew M, Magnani HN, Hull RD, Gent M. Double-blind randomized trial of ORG 10172 low-molecular-weight hepatinoid in prevention of deep-venous thrombosis in thrombotic stroke. Lancet 1987; 1: 523-6
• 3 Gerhart TN, Yett HS, Robertson LK, Lee MA, Smith M, Salzman EW. Low molecular wight hcparinoid comparcd with warfarin for prophylaxis of deep-vein thrombosis in patients who are operated on for fracture of the hip. A prospective. randomized trial. J Bone Joint Surg Am 1991; 73: 434-502
• 4 Bergqvist D. Orgaran in hip fracture surgery. Haemostasis 1992; 22: 104-8
• 5 Turpie AG. Orgaran in the prevention of deep-vein thrombosis in stroke patients. Haemostasis 1992; 22: 92-8
• 6 Meuleman DG. Orgaran (Org 10172): Its pharmacological profile in experimental models. Haemostasis 1992; 22: 58-65
• 7 van Dinther TG, Hol F, Meuleman DG. Effect of various heparin(oid)s on heparin cofactor I1 mediated anti-thrombin activity and inhibition of thrombin generation in vitro. Thromb Haemost 1987; 58: 423
• 8 Lijnen HR, Hojlaerts M, Collen D. Heparin binding properties of human histidine-rich glycoprotein. Mechanism and role in the neutralization of heparin in plasma. J Biol Chem 1983; 258: 3803-8
• 9 Lane DA, Flynn AM, Pejler G, Lindahl U, Choay J, Preissner K. Structural requirements for the neutralization of heparin-like saccharides by complement S proteinlvitronectin. J Biol Chem 1987; 262: 16343-8
• 10 Lane DA, Denton J, Flynn AM, Thunberg L, Lindahl U. Anticoagulant activities of heparin oligosaccharides and their neutralization by platelet factor 4. Biochem J 1984; 218: 725-32
• 11 Barrowcliffe TW, Merton RE, Havercroft SJ, Thunberg L, Lindahl U, Thomas DP. Low-affinity heparin potentiates the action of high-affinity heparin oligosaccharides. Thromb Res 1984; 34: 125-33
• 12 Young E, Hirsh J. Contribution of red blood cells to the saturable mechanism of heparin clearance. Thromb Haemost 1990; 64: 559-63
• 13 Young E, Prim M, Levine MN, Hirsh J. Heparin binding to plasma proteins, an important mechanism for heparin resistance. Thromb Haemost 1992; 67: 639-43
• 14 Fraker PJ, Speck JC. Protein and cell membrane iodinations with a sparingly soluble chloroamide. 1,3,4,6-tetrachloro-3a,6a-diphrenylglycoluril. Biochem Biophys Res Commun 1978; 80: 849-57
• 15 Zammit A, Pepper DS, Dawes J. Interaction of immobilised unfractionated and LMW heparins with proteins in whole human plasma. Thromb Haemost 1993; 70: 951-8
• 16 Daws J. Measurement of thc affinities of heparins. naturally occurring glycosaminoglycans, and other sullatcd polysaccharides for antithsombin III and thrombin. Anal Biochcrn 1988; 174: 177-86
• 17 Izumi M, Yamada KM, Hayajhi M. Vitroncctin exists in two structurally and fimctionallq distinct forms in human plasma. Biochim Biophys Acta 1989; 990: 101-8
• 18 Danhof M, de Boer A, Magnani HN, Stiebcma JCJ. Phasmacokinetic considerations on Orgaran (Org 10172) therapy. Flaemostasis 1992; 22: 73-84
• 19 Lane DA, Pejler G, Flynn AM, Thompson EA, Lindahl U. Ncutlialization of heparin-related saccharides by histidinc-rich glycoprotein and platelet factor 4. J Biol Chem 1986; 261: 3980-6
• 20 Sliimacia K, Kawarnoto A, Matsubayashi K, Orawa T. Histidinerich dvcoprokin docs not interfere with interactions betuccn antithrombin III and hcparin-like compounds on vascular endotheliai cells. Blood 1989; 73: 191-3
• 21 Lijncn HR, Van Hoef B, Collen U. Hislidine-rich glycoprotein modulates the anticoagulant activily of heparin in lrurlian plasma. Thromh Hacrnost 1981; 51: 266-8
• 22 Lijnen HR, Jacobs G, Collen D. Histirline-rich glycoprotein in a norinal and a clmical population. Thromb Res 1981; 22: 519-23
• 23 Engesscr L, Kluft C, Brict E, Brommcr JP. Familial elevation of plnsma histidine-rich glycoprotcin in a fanlily with rhrombopliiiin. Br J Haernatol 1987; 67: 355-8
• 24 Falkon L, Gari MI, Montsemt I, Borrcll M, Fontcuherta J. Familial elevation of plama histidine-rich glycoprotein. A case associated with recurrent venous thrombosis and high PAT-I levels. Thromb Res 1992; 66: 265-70
• 25 Castaman G, Ruggeri M, Burei F, Rodeghiero I. High levels of histidinerich glycoprotein and thrombotic diathesis. Report of two unrelated families. Thrornb Res 1993; 69: 297-305
• 26 Ehrcnforth S, Aygoren P, Fischbach P, Zlvinge B, PicadWillems B, Scharrer I. Elcvated histidinc-rich glycoprotei~l and idiopathic thrombosis. A study of 695 patients. Thromb Haemost 1993; 69: 1083