Abstract
We have previously shown that a single portal vein infusion of a recombinant adeno-associated viral vector (rAAV) expressing canine Factor IX (F.IX) resulted in long-term expression of therapeutic levels of F.IX in dogs with severe hemophilia B1. We carried out a phase 1/2 dose-escalation clinical study to extend this approach to humans with severe hemophilia B. rAAV-2 vector expressing human F.IX was infused through the hepatic artery into seven subjects. The data show that: (i) vector infusion at doses up to 2 × 1012 vg/kg was not associated with acute or long-lasting toxicity; (ii) therapeutic levels of F.IX were achieved at the highest dose tested; (iii) duration of expression at therapeutic levels was limited to a period of ∼8 weeks; (iv) a gradual decline in F.IX was accompanied by a transient asymptomatic elevation of liver transaminases that resolved without treatment. Further studies suggested that destruction of transduced hepatocytes by cell-mediated immunity targeting antigens of the AAV capsid caused both the decline in F.IX and the transient transaminitis. We conclude that rAAV-2 vectors can transduce human hepatocytes in vivo to result in therapeutically relevant levels of F.IX, but that future studies in humans may require immunomodulation to achieve long-term expression*.
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Notes
NOTE:In the version of this article originally published, Pradip Rustagi was inadvertently omitted from the author list; John Rasko should be listed as John E. J. Rasko; the affiliations of John E. J. Rasko and Katherine A. High were incorrect; and the author order was incorrect. The correct list of authors with their affiliations is:
Catherine S Manno1,2,15, Glenn F Pierce3,15, Valder R Arruda1,2,15, Bertil Glader4,15, Margaret Ragni5, John J E Rasko6, Margareth C Ozelo7, Keith Hoots8, Philip Blatt9, Barbara Konkle10, Michael Dake4, Robin Kaye1,2, Mahmood Razavi4, Albert Zajko10, James Zehnder4, Pradip K Rustagi11, Hiroyuki Nakai4, Amy Chew1,3, Debra Leonard2,12, J Fraser Wright3, Ruth R Lessard3, Jürg M Sommer3, Michael Tigges3, Denise Sabatino1, Alvin Luk3, Haiyan Jiang3, Federico Mingozzi1, Linda Couto3, Hildegund C Ertl1,13, Katherine A High1,2,14 & Mark A Kay4
1The Children’s Hospital of Philadelphia, Philadelphia and 2University of Pennsylvania School of Medicine, 3615 Civic Center Boulevard, Philadelphia, Pennsylvania, 19104, USA. 3Avigen Inc., 1301 Harbor Bay Parkway, Alameda, California 94502, USA. 4Stanford University School of Medicine, 300 Pasteur Drive, Room G305, Stanford, California 94305-5208, USA. 5University of Pittsburgh Medical Center, Hemophilia Center of Western Pennsylvania, 3636 Boulevard of the Allies, Pittsburgh, Pennsylvania 15213, USA. 6Royal Prince Alfred Hospital, Centenary Institute of Cancer Medicine & Cell Biology, Locked Bag No. 6, Newtown NSW 2042, Australia. 7University of Campinas, State University of Campinas, Caixa Postal 6198, CEP, 13083-970, Campinas-Sao Paolo, Brazil. 8University of Texas Houston Health Science Center, 1515 Holcombe Boulevard, Houston, Texas 77030, USA. 9Christiana Care, Christiana Hospital, 4755 Ogletown-Stanton Road, Newark, Delaware 19713, USA. 10University of Pennsylvania School of Medicine Presbyterian Hospital, MAB, Suite 103, 39th & Market Street, Philadelphia., Pennsylvania 19104, USA. 11Stanford University, School of Medicine, Department of Hematology, Jane Lathrop Building, 770 Welch Road, Suite 380, Palo Alto, California 94304, USA. 12Weill Medical College of Cornell University, 525 East 68th Street, New York, New York 10012, USA. 13Wistar Institute, 3601 Spruce Street, Philadelphia, Pennsylvania 19104, USA.. 14Howard Hughes Medical Institute, 3615 Civic Center Boulevard, Philadelphia, Pennsylvania 19104, USA. 15These authors contributed equally to this work. Correspondence should be addressed to K.A.H. (high@email.chop.edu).
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Acknowledgements
This work was supported by US National Institutes of Health grants U01 HL66948 (to M.A.K., K.A.H., C.S.M., B.G.), R01 HL71518 (to C.S.M.), P01 HL064190 (to K.A.H.), K01 DK60580 (to V.R.A.) and by the Howard Hughes Medical Institute. We thank each of the subjects for agreeing to participate in this study. We thank J. Bluestone and C. June for their scientific input. We thank J.W. Sun for assistance, critical reading and bioinformatics support.
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Glenn F. Pierce, Hiroyuki Nakai, Amy Chew, J. Fraser Wright, Ruth Lessard, Jürg M. Sommer, Michael Tigges, Alvin Luk, Haiyan Jiang, Linda Couto and Mark A. Kay are employed by or hold equity in Avigen, Inc., which manufactured the clinical-grade vector used in the study.
Supplementary information
Supplementary Fig. 1
T cell epitopes identified by ELISPOT assays (colored and underlined) are highly conserved in serotypes 1-8. (PDF 382 kb)
Supplementary Table 1
Inclusion/exclusion criteria. (PDF 45 kb)
Supplementary Table 2
Vector biodistribution by dose group and body fluid/tissue based on PCR assay for vector sequences. (PDF 67 kb)
Supplementary Table 3
AAV sequences in semen and semen fractions of subject A. (PDF 37 kb)
Supplementary Table 4
ELISA of human FIX antigen in plasma of subject E post infusion of AAV2-hFIX vector. (PDF 29 kb)
Supplementary Table 5
Neutralizing antibody (NAB) titers of AAV as a function of time after vector administration in subjects infused with AAV-F.IX. (PDF 37 kb)
Supplementary Table 6
Evaluation of elevated transaminases in subject E. (PDF 29 kb)
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Manno, C., Pierce, G., Arruda, V. et al. Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response. Nat Med 12, 342–347 (2006). https://doi.org/10.1038/nm1358
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DOI: https://doi.org/10.1038/nm1358
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