title: AAV8 induces tolerance in murine muscle as a result of poor APC transduction, T cell exhaustion, and minimal MHCI upregulation on target cells.
abstract: Following gene transfer of adeno-associated virus 2/8 (AAV2/8) to the muscle, C57BL/6 mice show long-term expression of a nuclear-targeted LacZ (nLacZ) transgene with minimal immune activation. Here, we show that pre-exposure to AAV2/8 can also induce tolerance to the more immunogenic AAV2/rh32.33 vector, preventing otherwise robust T-cell activation and allowing stable transgene expression. Depletion and adoptive transfer studies showed that a suppressive factor was not sufficient to account for AAV2/8-induced tolerance, whereas further characterization of the T-cell population showed upregulation of the exhaustion markers PD1, 2B4, and LAG3. Furthermore, systemic administration of Toll-like receptor (TLR) ligands at the time of AAV2/rh32.33-administration broke AAV2/8-induced tolerance, restoring T-cell activation and β-gal clearance. As such, AAV2/8 transduction appears to lack the inflammatory signals necessary to prime a functional cytotoxic T-cell response. Inadequate T-cell priming could be explained upstream by AAV2/8's poor transduction and activation of antigen-presenting cells (APCs). Immunohistochemical analysis indicates that AAV2/8 transduction also fails to upregulate major histocompatibility complex class I (MHCI) expression on the surface of myocytes, rendering transduced cells poor targets for T-cell-mediated destruction. Overall, AAV2/8-induced tolerance in the muscle is multifactorial, spanning from poor APC transduction and activation to the subsequent priming of functionally exhausted T-cells, while simultaneously avoiding upregulation of MHCI on potential targets.
authors.affiliation: Gene Therapy Program, Department of Pathology and Laboratory Medicine, Division of Transfusion Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Lauren E
authors.initials: LE
authors.lastname: Mays
authors.affiliation: Gene Therapy Program, Department of Pathology and Laboratory Medicine, Division of Transfusion Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.affiliation: Gene Therapy Program, Department of Pathology and Laboratory Medicine, Division of Transfusion Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Jianping
authors.initials: J
authors.lastname: Lin
authors.affiliation: Gene Therapy Program, Department of Pathology and Laboratory Medicine, Division of Transfusion Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Bell
authors.affiliation: Present address: Regeneron Pharmaceutical, Tarrytown, New York, USA.
authors.forename: Alison
authors.initials: A
authors.lastname: Crawford
authors.affiliation: Department of Microbiology and Institute for Immunology, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: E John
authors.initials: EJ
authors.lastname: Wherry
authors.affiliation: Gene Therapy Program, Department of Pathology and Laboratory Medicine, Division of Transfusion Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: James M
authors.initials: JM
authors.lastname: Wilson
mesh_terms: D000818:Animals; D000938:Antigen-Presenting Cells / Q000276:immunology* / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics / Q000276:immunology*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D006801:Humans; D007108:Immune Tolerance; D007249:Inflammation / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation / Q000276:immunology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018482:Muscle, Skeletal / Q000276:immunology* / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D050378:T-Lymphocytes, Regulatory / Q000276:immunology; D051193:Toll-Like Receptors / Q000378:metabolism; D014161:Transduction, Genetic; D019076:Transgenes / Q000235:genetics / Q000276:immunology
chemical_list: D015395:Histocompatibility Antigens Class I; D051193:Toll-Like Receptors