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Adeno-associated virus serotype 2 induces cell-mediated immune responses directed against multiple epitopes of the capsid protein VP1

Emma R. Cantwell^1,

Patricia A. Johnson^3,

Bernard P. Mahon^1,

¹ Cellular Immunology Laboratory, Institute of Immunology, National University of Ireland Maynooth, County Kildare, Ireland
² Regenerative Medicine Institute (REMEDI), National University of Ireland Galway, Galway, Ireland
³ Viral Immunology Laboratory, School of Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland

Correspondence
Bernard P. Mahon
bp.mahon{at}nuim.ie

Adeno-associated virus serotype 2 (AAV-2) has been developed as a gene therapy vector. Antibody and cell-mediated immune responses to AAV-2 or AAV-2-transfected cells may confound the therapeutic use of such vectors in clinical practice. In one of the most detailed examinations of AAV-2 immunity in humans to date, cell-mediated and humoral immune responses to AAV-2 were characterized from a panel of healthy blood donors. The extent of AAV-2-specific antibody in humans was determined by examination of circulating AAV-2-specific total IgG levels in plasma from 45 normal donors. Forty-one donors were seropositive and responses were dominated by IgG1 and IgG2 subclasses. Conversely, AAV-2-specific IgG3 levels were consistently low in all donors. Cell-mediated immune recall responses were detectable in nearly half the population studied. In vitro restimulation with AAV-2 of peripheral blood mononuclear cell cultures from 16 donors elicited gamma interferon (IFN-) (ten donors), interleukin-10 (IL-10) (eight donors) and interleukin-13 (IL-13) (four donors) responses. Using a series of overlapping peptides derived from the sequence of the VP1 viral capsid protein, a total of 59 candidate T-cell epitopes were identified. Human leukocyte antigen characterization of donors revealed that the population studied included diverse haplotypes, but that at least 17 epitopes were recognized by multiple donors and could be regarded as immunodominant. These data indicate that robust immunological memory to AAV-2 is established. The diversity of sequences recognized suggests that attempts to modify the AAV-2 capsid, as a strategy to avoid confounding immunity, will not be feasible.

These authors contributed equally to this work.

Present address: Department of Histopathology, Institute of Molecular Medicine, Trinity College Dublin, Dublin 2, Ireland.

A supplementary figure, showing verification of the capacity of the IgG subclass ELISA protocol to successfully detect antigen-bound IgG3, and two supplementary tables, listing sequences of the 20-mer peptides derived from the AAV-2 VP1 capsid protein and AAV-2 VP1 capsid sequences recognized by human PBMC, are available with the online version of this paper.