Heparins from porcine and bovine intestinal mucosa: Are they similar drugs?

Summary

Increasing reports of bleeding and peri- or post-operative blood dyscrasias in Brazil were possibly associated with the use of heparin from bovine instead of porcine intestine. These two pharmaceutical grade heparins were analysed for potential differences. NMR analyses confirmed that porcine heparin is composed of mainly trisulfated disaccharides →4-α-IdoA2S-1→4-α-GlcNS6S-1→. Heparin from bovine intestine is also composed of highly 2-sulfated α-iduronic acid residues, but the sulfation of the α-glucosamine units vary significantly: ~50% are 6-and N-disulfated, as in porcine heparin, while ~36% are 6-desulfated and ~14% N-acetylated. These heparins differ significantly in their effects on coagulation, thrombosis and bleeding. Bovine heparin acts mostly through factor Xa. Compared to porcine heparin on a weight basis, bovine heparin exhibited approximately half of the anticoagulant and antithrombotic effects, but similar effect on bleeding. These two heparins also differ in their protamine neutralisation curves. The doses of heparin from bovine intestine required for effective antithrombotic protection and the production of adverse bleeding effects are closer than those for porcine heparin. This observation may explain the increasing bleeding observed among Brazilian patients. Our results suggest that these two types of heparin are not equivalent drugs.

Keywords

Venous thrombosis - glycosaminoglycans - heparins - serpins - thrombosis

^* These two authors contributed equally to the work.

• References

• 1 Spyropoulos AC. Brave new world: The current and future use of novel anticoagulants. Thomb Res 2008; 123: S29-S35.
• 2 Ryckelynck JP, Chouraqui D, Lobbedez T. Anticoagulant of the extracorporeal circuit in chronic hemodialysis. Nephrologie 1998; 19: 223-225.
• 3 Liu H, Zhang Z, Linhardt RJ. Lessons learned from the contamination of heparin. Nat Prod Rep 2009; 26: 313-321.
• 4 Mulloy B, Gray E, Barrowcliffe TW. Characterization of unfractionated heparin: comparison of materials from the last 50 years. Thromb Haemost 2000; 84: 1052-1056.
• 5 Guerrini M, Naggi A, Guglieri S. et al. Complex glycosaminoglycans: profiling substitution patterns by two-dimensional nuclear magnetic resonance spectroscopy. Anal Biochem 2005; 337: 35-47.
• 6 Melo EI, Pereira MS, Cunha RS. et al. Heparin quality control in the Brazilian market: implications in the cardiovascular surgery. Rev Bras Cir Cardiovasc 2008; 23: 169-174.
• 7 ANVISA, alert about the increasing reports of bleedin and peri- or post-operative blood dyscrasias in Brazil possibly associated with the use of heparin from bovine instead of porcine intestine (24 oct 2008). Available at: http://www.anvisa.gov.brfarmacovigilancia/alerta/federal/2008/federal_3_08.htm
• 8 Blossom DB, Kallen AJ, Patel PR. et al. Outbreak of adverse reactions associated with contaminated heparin. N Engl J Med 2008; 359: 2674-2684.
• 9 Castro MO, Pomin VH, Santos LL. et al. A unique 2-sulfated β-galactan from the egg jelly of the sea urchin Glyptocidaris crenularis: conformational flexibility versus induction of the sperm acrosome reaction. J Biol Chem 2009; 284: 18790-18800.
• 10 Farndale RW, Buttle DJ, Barret AJ. Improved quantitation and discrimination of sulfated glycosaminoglycans by use of dimethylmethylene blue. Biochem Biophys Acta 1986; 883: 173-177.
• 11 Mourão PA, Bastos IG. Highly acidic glycans from sea cucumbers. Isolation and fractionation of fucose-rich sulfated polysaccharides from the body wall of Ludwigothurea grisea . Eur J Biochem 1987; 166: 639-645.
• 12 Herbert JM, Bernat A, Maffrand JP. Importance of platelets in experimental venous thrombosis in rats. Blood 1992; 80: 2281-2286.
• 13 Zancan P, Mourão PAS. Venous and arterial thrombosis in rat models: dissociation of the antithrombotic effects of glycosaminoglycans. Blood Coagul Fibrinolysis 2004; 15: 45-54.
• 14 Li W, Johnson DJ, Esmon CT. et al. Structure of the antithrombin-thrombin-heparin ternary complex reveals the antithrombotic mechanism of heparin. Nat Struct Mol Biol 2004; 11: 857-862.
• 15 Zhang Z, Li B, Suwan J. et al. Analysis of pharmaceutical heparins and potential contaminations using 1H-NMR and PAGE. J Pharm Sci 2009; 99: 4017-4026.
• 16 Kusche M, Giangiacomo T, Casu B. et al. Biosynthesis of heparin. Availability of glucosaminyl 3-O-sulfation sites. J Biol Chem 1990; 265: 7292-7300.
• 17 US FARMACOPEIA. USP Heparin Information (June, 2009). Available at: http://www.usp.orgottopics/heparin.html
• 18 Cavalcante MCM, Allodi S, Valente AP. et al. Occurrence of heparin in the invertebrate Styela plicata (Tunicata) is restricted to cell layers facing the outside environment. An ancient role in defense? J Biol Chem 2000; 275: 36189-36196.
• 19 Yates EA, Santini F, Guerrini M. et al. ¹H and ¹³C NMR spectral assignments of the major sequences of twelve systematically modified heparin derivatives. Carb Res 1996; 194: 15-27.
• 20 Chuang W, Christ MD, Rabenstein DL. Determination of the primary structures of heparin- and heparan sulfate-derived oligosaccharides using band-selective homonuclear-decouped two-dimentional ¹H NMR experiments. Anal Chem 2001; 73: 2310-2316.
• 21 Guerrini M, Beccati D, Shriver Z. et al. Oversulfated chondroitin sulfate is a contaminant in heparin associated with adverse clinical events. Nat Biotechnol 2008; 26: 669-675.