Abstract
Hepatic adeno-associated virus (AAV)-serotype 2 mediatedgene transfer results in transgene product expression that is sustained in experimental animals but not in human subjects. We hypothesize that this is caused by rejection of transduced hepatocytes by AAV capsid–specific memory CD8+ T cells reactivated by AAV vectors. Here we show that healthy subjects carry AAV capsid–specific CD8+ T cells and that AAV-mediated gene transfer results in their expansion. No such expansion occurs in mice after AAV-mediated gene transfer. In addition, we show that AAV-2 induced human T cells proliferate upon exposure to alternate AAV serotypes, indicating that other serotypes are unlikely to evade capsid-specific immune responses.
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Acknowledgements
This work was supported by US National Institutes of Health (NIH) grants P01 HL078810, M01-RR00240 (NIH GCRC award to Children's Hospital of Philadelphia), M01-RR000056 (NIH GCRC award to University of Pittsburgh), a grant to the Penn Center for AIDS Research P30 AI045008 and the Howard Hughes Medical Institute. D.J.H. was supported by training grant NIH T32 HL07439, S.L.M. by training grant NIH T32 DK07748 and D.E.S. by NIH F32 HL69647. We thank F. Lemonnier (Institute Pasteur) for kindly providing HLA-B*0702 transgenic mice. We thank M. Lasaro and M. Tigges for scientific input, and J. Sun for assistance in manuscript preparation.
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F.M. and M.V.M. performed the in vitro expansion experiments on normal donors and expansions on subjects with hemophilia B. They performed intracellular cytokine assays, the cytotoxic T lymphocyte (CTL) assay, the pentamer staining, the immunophenotyping of memory CD8+ T cells specific to the AAV capsid and the cross-reactivity experiments. D.J.H. performed experiments on human splenocytes and the AAV epitope characterization. D.E.S. performed in vitro expansion experiments on subjects with hemophilia B enrolled in the gene transfer study and part of the ELISpot studies on healthy donors. S.L.M. performed the experiments on HLA-B*0702 transgenic mice. H.J. and J.S. performed part of the ELISpot studies and AAV antibody titer determination on normal donors. C.S.M., M.V.R. and J.E.J.R. directed and/or participated in the clinical gene transfer study, including the care of hemophilic subjects, and collection of PBMCs used in the study. They provided assistance in drafting the manuscript. H.C.J.E. and G.F.P. collaborated on experimental design and interpretation, and helped to draft the manuscript. K.A.H. directed experimental design, conduct, data analysis and interpretation, and drafted the manuscript.
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J.S., H.J. and G.F.P. were employees of Avigen, Inc. at the time the work was done. F.M., D.J.H. and K.A.H. hold pending patent applications that may be affected by publication of this manuscript.
Supplementary information
Supplementary Fig. 1
Immunophenotyping of capsid-specific CD8+ T cells in a healthy donor. (PDF 192 kb)
Supplementary Table 1
AAV capsid epitopes defined by IFN-γ ELISpot assays of normal donors and of AAV-infused subjects (PDF 56 kb)
Supplementary Table 2
Neutralizing antibody and cellular immune responses, sera and PBMCs from adult human subjects (PDF 48 kb)
Supplementary Table 3
IFN-γ ELISpot responses to AAV capsid in unexpanded and peptide-expanded human splenocytes (PDF 58 kb)
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Mingozzi, F., Maus, M., Hui, D. et al. CD8+ T-cell responses to adeno-associated virus capsid in humans. Nat Med 13, 419–422 (2007). https://doi.org/10.1038/nm1549
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DOI: https://doi.org/10.1038/nm1549
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