Evidence for gene transfer and expression of factor IX in haemophilia B patients treated with an AAV vector (original) (raw)

Nature Genetics volume 24, pages 257–261 (2000)Cite this article

Abstract

Pre-clinical studies in mice and haemophilic dogs have shown that introduction of an adeno-associated viral (AAV) vector encoding blood coagulation factor IX (F.IX) into skeletal muscle results in sustained expression of F.IX at levels sufficient to correct the haemophilic phenotype1,2. On the basis of these data and additional pre-clinical studies demonstrating an absence of vector-related toxicity, we initiated a clinical study of intramuscular injection of an AAV vector expressing human F.IX in adults with severe haemophilia B. The study has a dose-escalation design, and all patients have now been enrolled in the initial dose cohort (2×1011 vg/kg). Assessment in the first three patients of safety and gene transfer and expression show no evidence of germline transmission of vector sequences or formation of inhibitory antibodies against F.IX. We found that the vector sequences are present in muscle by PCR and Southern-blot analyses of muscle biopsies and we demonstrated expression of F.IX by immunohistochemistry. We observed modest changes in clinical endpoints including circulating levels of F.IX and frequency of F.IX protein infusion. The evidence of gene expression at low doses of vector suggests that dose calculations based on animal data may have overestimated the amount of vector required to achieve therapeutic levels in humans, and that the approach offers the possibility of converting severe haemophilia B to a milder form of the disease.

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References

  1. Herzog, R.W. et al. Stable gene transfer and expression of human blood coagulation factor IX after intramuscular injection of recombinant adeno-associated virus . Proc. Natl Acad. Sci. USA 94, 5804– 5809 (1997).
    Article CAS Google Scholar
  2. Herzog, R. et al. Long-term correction of canine hemophilia B by gene transfer of blood coagulation factor IX mediated by adeno-associated viral vector. Nature Med. 5, 56–63 ( 1999).
    Article CAS Google Scholar
  3. Ljung, R.C.R. Annotation. Can haemophilic arthropathy be prevented? Br. J. Haematol. 101, 215–219 ( 1998).
    Article CAS Google Scholar
  4. Lofqvist, T., Nilsson, I.M., Berntorp, E. & Pettersson, H. Haemophilia prophylaxis in young patients—a long term follow up. J. Intern. Med. 241, 395–400 (1997).
    Article CAS Google Scholar
  5. Mannucci, P.M. & Weitz, J.I. The hemophilias: progress and problems . Semin. Hematol. 36, 104– 117 (1999).
    CAS PubMed Google Scholar
  6. Parks, W.P., Boucher, D.W., Melnick, J.L., Taber, L.H. & Yow, M.D. Seroepidemiological and ecological studies of the adenovirus-associated satellite viruses. Infect. Immun. 2, 716–722 ( 1970).
    CAS PubMed PubMed Central Google Scholar
  7. Summerford, C. & Samulski, R.J. Membrane-associated heparan sulfate proteoglycan is a receptor for adeno-associated virus type 2 virions. J. Virol. 72, 1438–1445 (1998).
    CAS PubMed PubMed Central Google Scholar
  8. Qing, K. et al. Human fibroblast growth growth factor receptor 1 is a co-receptor for infection by adeno-associated virus 2. Nature Med. 5, 71–77 (1999).
    Article CAS Google Scholar
  9. Cheung, W.F. et al. Identification of the endothelial cell binding site for factor IX. Proc. Natl Acad. Sci. USA 93, 11068– 11073 (1996).
    Article CAS Google Scholar
  10. Matsushita, T. et al. Adeno-associated virus vectors can be efficiently produced without helper virus. Gene Ther. 5, 938– 945 (1998).
    Article CAS Google Scholar
  11. Burton, M. et al. Coexpression of factor VIII heavy and light chain adeno-associated viral vectors produces biologically active protein. Proc. Natl Acad. Sci. USA 96, 12725–12730 (1999).
    Article CAS Google Scholar
  12. Yoshitake, S., Schach, B.G., Foster, D.C., Davie, E.W. & Kuracki, K. Nucleotide sequence of the gene for human factor IX (antihemophilic factor B). Biochemistry 24, 3736–3750 (1985).
    Article CAS Google Scholar
  13. Kurachi, S., Hitomi, Y., Midori, F. & Kurachi, K. Role of intron I in expression of the human factor IX gene. J. Biol. Chem. 270, 5276–5281 (1995).
    Article CAS Google Scholar
  14. Manno, C.S., Larson, P.J., Cohen, A.R. & Flake, A.W. Guidelines for Research Involving Recombinant DNA Molecules (NIH Guidelines). Appendix M. Points to consider in the design and submission of protocols for the transfer of recombinant DNA molecules into one or more human subjects. (NIH Recombinant DNA Advisory Committee, Washington DC, 1999).
    Google Scholar
  15. Walter, J., You, Q., Hagstrom, J.N., Sands, M. & High, K.A. Successful expression of human factor IX following repeat administration of adenoviral vector in mice. Proc. Natl Acad. Sci. USA 93, 3056–3061 ( 1996).
    Article CAS Google Scholar
  16. Miller, J.H. Experiments in Molecular Genetics 352–355 (Cold Spring Harbor Press, Cold Spring Harbor, New York, 1972).
    Google Scholar

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Acknowledgements

We thank A. Radu for assistance with immunohistochemical staining of muscle; M. Tanzer for assistance with coagulation assays; D. Leonard for assistance with PCR assays on human samples; the Nucleic Acid/Protein Research Core Facility at CHOP; R. Barth and M. King for assistance with ultrasound; and S.G. Madison for clinical support. This work was supported by National Institutes of Health Grants R01 HL53682 to M.A.K. and R01 HL53668, R01 HL61921, P50 HL54500 and a grant from the Hoxie Harrison-Smith Foundation to K.A.H. The work was also supported in part by NIH grant M01 RR00070 to the GCRC at Stanford University, by NIH grant M01 RR00240 to the GCRC at The Children's Hospital of Philadelphia and by Avigen.

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Authors and Affiliations

  1. Departments of Pediatrics, Stanford University School of Medicine, California, Palo Alto, USA
    Mark A. Kay & Bertil Glader
  2. Department of Genetics, Stanford University School of Medicine, California, Palo Alto, USA
    Mark A. Kay
  3. Department of Surgery, Stanford University School of Medicine, California, Palo Alto, USA
    Erik Skarsgard
  4. The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA
    Catherine S. Manno, Peter J. Larson, Amy J. Chew, Shing J Tai, Roland W. Herzog, Valder Arruda, Alan W. Flake & Katherine A. High
  5. Departments of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA
    Catherine S. Manno, Peter J. Larson & Katherine A. High
  6. Department of Surgery, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA
    Alan W. Flake
  7. Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
    Margaret V. Ragni
  8. Avigen, Inc, Alameda, California , USA
    Linda B. Couto, Alan McClelland, Fred Johnson & Ciaran Scallan

Authors

  1. Mark A. Kay
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  2. Catherine S. Manno
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  3. Margaret V. Ragni
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  4. Peter J. Larson
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  5. Linda B. Couto
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  6. Alan McClelland
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  7. Bertil Glader
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  8. Amy J. Chew
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  9. Shing J Tai
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  10. Roland W. Herzog
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  11. Valder Arruda
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  12. Fred Johnson
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  13. Ciaran Scallan
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  14. Erik Skarsgard
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  15. Alan W. Flake
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  16. Katherine A. High
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Corresponding author

Correspondence toKatherine A. High.

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Kay, M., Manno, C., Ragni, M. et al. Evidence for gene transfer and expression of factor IX in haemophilia B patients treated with an AAV vector.Nat Genet 24, 257–261 (2000). https://doi.org/10.1038/73464

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