title: Mapping the structural determinants responsible for enhanced T cell activation to the immunogenic adeno-associated virus capsid from isolate rhesus 32.33.
abstract: Avoiding activation of immunity to vector-encoded proteins is critical to the safe and effective use of adeno-associated viral (AAV) vectors for gene therapy. While commonly used serotypes, such as AAV serotypes 1, 2, 7, 8, and 9, are often associated with minimal and/or dysfunctional CD8(+) T cell responses in mice, the threshold for immune activation appears to be lower in higher-order species. We have modeled this discrepancy within the mouse by identifying two capsid variants with differential immune activation profiles: AAV serotype 8 (AAV8) and a hybrid between natural rhesus isolates AAVrh32 and AAVrh33 (AAVrh32.33). Here, we aimed to characterize the structural determinants of the AAVrh32.33 capsid that augment cellular immunity to vector-encoded proteins or those of AAV8 that may induce tolerance. We hypothesized that the structural domain responsible for differential immune activation could be mapped to surface-exposed regions of the capsid, such as hypervariable regions (HVRs) I to IX of VP3. To test this, a series of hybrid AAV capsids was constructed by swapping domains between AAV8 and AAVrh32.33. By comparing their ability to generate transgene-specific T cells in vivo versus the stability of transgene expression in the muscle, we confirmed that the functional domain lies within the VP3 portion of the capsid. Our studies were able to exclude the regions of VP3 which are not sufficient for augmenting the cellular immune response, notably, HVRs I, II, and V. We have also identified HVR IV as a region of interest in conferring the efficiency and stability of muscle transduction to AAVrh32.33.
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.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.forename: Rebeca
authors.initials: R
authors.lastname: Tenney
authors.forename: Peter
authors.initials: P
authors.lastname: Bell
authors.forename: Hyun-Joo
authors.initials: HJ
authors.lastname: Nam
authors.forename: Jianping
authors.initials: J
authors.lastname: Lin
authors.forename: Brittney
authors.initials: B
authors.lastname: Gurda
authors.forename: Kim
authors.initials: K
authors.lastname: Van Vliet
authors.forename: Kyle
authors.initials: K
authors.lastname: Mikals
authors.forename: Mavis
authors.initials: M
authors.lastname: Agbandje-McKenna
authors.forename: James M
authors.initials: JM
authors.lastname: Wilson
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002213:Capsid / Q000276:immunology; D036022:Capsid Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000229:Dependovirus / Q000145:classification / Q000235:genetics / Q000276:immunology*; D018604:Epitope Mapping; D006824:Hybridization, Genetic; D008213:Lymphocyte Activation; D008253:Macaca mulatta / Q000821:virology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D012703:Serotyping; D013601:T-Lymphocytes / Q000276:immunology* / Q000821:virology*
chemical_list: D036022:Capsid Proteins; C442558:VP3 protein, Dependovirus