Nothing Special   »   [go: up one dir, main page]
Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

RNA aptamers as reversible antagonists of coagulation factor IXa

Nature volume 419, pages 90–94 (2002)Cite this article

Abstract

Many therapeutic agents are associated with adverse effects in patients. Anticoagulants can engender acute complications such as significant bleeding that increases patient morbidity and mortality1. Antidote control provides the safest means to regulate drug action. For this reason, despite its known limitations and toxicities, heparin use remains high because it is the only anticoagulant that can be controlled by an antidote, the polypeptide protamine2,3,4. To date, no generalizable strategy for developing drug–antidote pairs has been described. We investigated whether drug–antidote pairs could be rationally designed by taking advantage of properties inherent to nucleic acids to make antidote-controlled anticoagulant agents. Here we show that protein-binding oligonucleotides (aptamers) against coagulation factor IXa are potent anticoagulants. We also show that oligonucleotides complementary to these aptamers can act as antidotes capable of efficiently reversing the activity of these new anticoagulants in plasma from healthy volunteers and from patients who cannot tolerate heparin5. This generalizable strategy for rationally designing a drug–antidote pair thus opens up the way for developing safer regulatable therapeutics.

Access through your institution
Buy or subscribe

This is a preview of subscription content, access via your institution

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Sequences and secondary structure of aptamers isolated against FIXa.
Figure 2: Inhibition of FIXa function by aptamers 9.3t and Peg-9.3t.
Figure 3: Antidote oligonucleotides to aptamer 9.3t.
Figure 4: Properties of antidote oligonucleotide 5-2.
Figure 5: Antidote-controlled anticoagulation of plasma from patients with HIT.

Similar content being viewed by others

References

  1. Levine, M. N., Raskob, G., Landefeld, S. & Kearon, C. Hemorrhagic complications of anticoagulant treatment. Chest 119, 108S–121S (2001)

    Article  CAS  Google Scholar 

  2. Hirsch, J., Anand, S. S., Halperin, J. L. & Fuster, V. Guide to anticoagulant therapy: Heparin: A statement for healthcare professionals from the American Heart Association. Circulation 103, 2994–3018 (2001)

    Article  Google Scholar 

  3. Carr, J. A. & Silverman, N. The heparin-protamine interaction. A review. J. Cardiovasc. Surg. 40, 659–666 (1999)

    CAS  Google Scholar 

  4. Pifarre, R., Walenga, J. M. & Fareed, J. in New Anticoagulants for the Cardiovascular Patient (ed. Pifarre, R.) 1–7 (Hanley and Belfus, Philadelphia, 1997)

    Google Scholar 

  5. Warkentin, T. E., Chong, B. H. & Greinacher, A. Heparin-induced thrombocytopenia: towards consensus. Thromb. Haemost. 79, 1–7 (1998)

    Article  CAS  Google Scholar 

  6. Johnson, K. & Hung, D. Novel anticoagulants based on inhibition of the factor VIIa/tissue factor pathway. Coron. Artery Dis. 9, 83–87 (1998)

    CAS  PubMed  Google Scholar 

  7. Rusconi, C. P., Yeh, A., Lyerly, H. K., Lawson, J. H. & Sullenger, B. A. Blocking the initiation of coagulation by RNA aptamers to factor VIIa. Thromb. Haemost. 84, 841–848 (2000)

    Article  CAS  Google Scholar 

  8. Bock, L. C., Griffin, L. C., Latham, J. A., Vermaas, E. H. & Toole, J. J. Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Nature 355, 564–566 (1992)

    Article  ADS  CAS  Google Scholar 

  9. Tasset, D. M., Kubik, M. F. & Steiner, W. Oligonucleotide inhibitors of human thrombin that bind distinct epitopes. J. Mol. Biol. 272, 688–698 (1997)

    Article  CAS  Google Scholar 

  10. White, R. et al. Generation of species cross-reactive aptamers using “toggle” SELEX. Mol. Ther. 4, 567–573 (2001)

    Article  ADS  CAS  Google Scholar 

  11. Tuerk, C. & Gold, L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science 249, 505–510 (1990)

    Article  ADS  CAS  Google Scholar 

  12. Ellington, A. D. & Szostak, J. W. In vitro selection of RNA molecules that bind specific ligands. Nature 346, 818–822 (1990)

    Article  ADS  CAS  Google Scholar 

  13. Pieken, W. A., Olsen, D. B., Benseler, F., Aurup, H. & Eckstein, F. Kinetic characterization of ribonuclease-resistant 2′-modified hammerhead ribozymes. Science 253, 314–317 (1991)

    Article  ADS  CAS  Google Scholar 

  14. Davis, J. P., Janjic, N., Javornik, B. E. & Zichi, D. A. Identifying consensus patterns and secondary structure in SELEX sequence sets. Methods Enzymol. 267, 302–314 (1996)

    Article  CAS  Google Scholar 

  15. Watson, S. R. et al. Anti-L-selectin aptamers: binding characteristics, pharmacokinetic parameters, and activity against an intravascular target in vivo. Antisense Nucleic Acid Drug Dev. 10, 63–75 (2000)

    Article  CAS  Google Scholar 

  16. Tucker, C. E. et al. Detection and plasma pharmacokinetics of an anti-vascular endothelial growth factor oligonucleotide-aptamer (NX1838) in rhesus monkeys. J. Chromatogr. B Biomed. Sci. Appl. 732, 203–212 (1999)

    Article  CAS  Google Scholar 

  17. High, K. A. & Roberts, H. R. in Molecular Basis of Thrombosis and Hemostasis (eds High, K. A. & Roberts, H. R.) 215–237 (Marcel Dekker, New York, 1995)

    Google Scholar 

  18. Spanier, T. B. et al. Selective anticoagulation with active site-blocked factor IXA suggests separate roles for intrinsic and extrinsic coagulation pathways in cardiopulmonary bypass. J. Thorac. Cardiovasc. Surg. 116, 860–869 (1998)

    Article  CAS  Google Scholar 

  19. Feuerstein, G. Z. et al. An inhibitory anti-factor IX antibody effectively reduces thrombus formation in a rat model of venous thrombosis. Thromb. Haemost. 82, 1443–1445 (1999)

    Article  CAS  Google Scholar 

  20. Choudhri, T. F. et al. Targeted inhibition of intrinsic coagulation limits cerebral injury in stroke without increasing intracerebral hemorrhage. J. Exp. Med. 190, 91–99 (1999)

    Article  CAS  Google Scholar 

  21. White, R. R., Sullenger, B. A. & Rusconi, C. P. Developing aptamers into therapeutics. J. Clin. Invest. 106, 929–934 (2000)

    Article  CAS  Google Scholar 

  22. Campbell, K. R. et al. Bivalirudin in patients with heparin-induced thrombocytopenia undergoing percutaneous coronary intervention. J. Invasive Cardiol. 12, 14F–19F (2000)

    PubMed  Google Scholar 

  23. Greinacher, A. et al. Recombinant hirudin (lepirudin) provides safe and effective anticoagulation in patients with heparin-induced thrombocytopenia: a prospective study. Circulation 99, 73–80 (1999)

    Article  CAS  Google Scholar 

  24. Lewis, B. E. et al. Argatroban anticoagulant therapy in patients with heparin-induced thrombocytopenia. Circulation 103, 1838–1843 (2001)

    Article  CAS  Google Scholar 

  25. Hicke, B. J. & Stephens, A. W. Escort aptamers: a delivery service for diagnosis and therapy. J. Clin. Invest. 106, 923–928 (2000)

    Article  CAS  Google Scholar 

  26. van Dijk, M. A. & van de Winkel, J. G. Human antibodies as next generation therapeutics. Curr. Opin. Chem. Biol. 5, 368–374 (2001)

    Article  CAS  Google Scholar 

  27. Fitzwater, T. & Polisky, B. A SELEX primer. Methods Enzymol. 267, 275–301 (1996)

    Article  CAS  Google Scholar 

  28. Hope, M. J., Bally, M. B., Webb, G. & Cullis, P. R. Production of large unilamellar vesicles by a rapid extrusion procedure. Characterization of size distribution, trapped volume and ability to maintain a membrane potential. Biochim. Biophys. Acta 812, 55–65 (1985)

    Article  CAS  Google Scholar 

  29. Silverman, S. K. & Cech, T. R. Energetics and cooperativity of tertiary hydrogen bonds in RNA structure. Biochemistry 38, 8691–8702 (1999)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank R. Califf and R. Harrington for their insight into the need for antidote-controlled anticoagulant and antithrombotic agents. This work was supported by grants from the American Heart Association to C.P.R. and the National Institutes of Health to B.A.S, T.L.O. and D.M.

Author information

Authors and Affiliations

  1. Department of Surgery, Program in Combinatorial Therapeutics, Duke University Medical Center, Durham, North Carolina, 27710, USA

    Christopher P. Rusconi, Elizabeth Scardino, Juliana Layzer, George A. Pitoc & Bruce A. Sullenger

  2. Departments of Medicine and Pathology, Duke University Medical Center, Durham, North Carolina, 27710, USA

    Thomas L. Ortel

  3. Center for Thrombosis and Haemostasis, University of North Carolina School of Medicine, Chapel Hill, North Carolina, 27599, USA

    Dougald Monroe

Authors
  1. Christopher P. Rusconi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Elizabeth Scardino
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Juliana Layzer
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. George A. Pitoc
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Thomas L. Ortel
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Dougald Monroe
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Bruce A. Sullenger
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Christopher P. Rusconi or Bruce A. Sullenger.

Ethics declarations

Competing interests

C.P.R., E.S., J.L., T.L.O. and B.A.S. are employed by the Duke University Medical Center, which has filed two patent applications on work described in this Letter.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Rusconi, C., Scardino, E., Layzer, J. et al. RNA aptamers as reversible antagonists of coagulation factor IXa. Nature 419, 90–94 (2002). https://doi.org/10.1038/nature00963

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature00963

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing