title: Tyrosine Mutation in AAV9 Capsid Improves Gene Transfer to the Mouse Lung.
abstract: OBJECTIVE
  Adeno-associated virus (AAV) vectors are being increasingly used as the vector of choice for in vivo gene delivery and gene therapy for many pulmonary diseases. Recently, it was shown that phosphorylation of surface-exposed tyrosine residues from AAV capsid targets the viral particles for ubiquitination and proteasome-mediated degradation, and mutations of these tyrosine residues lead to highly efficient vector transduction in vitro and in vivo in different organs. In this study, we evaluated the pulmonary transgene expression efficacy of AAV9 vectors containing point mutations in surface-exposed capsid tyrosine residues.
  
  
  METHODS
  Eighteen C57BL/6 mice were randomly assigned into three groups: (1) a control group (CTRL) animals underwent intratracheal (i.t.) instillation of saline, (2) the wild-type AAV9 group (WT-AAV9, 1010 vg), and (3) the tyrosine-mutant Y731F AAV9 group (M-AAV9, 1010 vg), which received (i.t.) self-complementary AAV9 vectors containing the DNA sequence of enhanced green fluorescence protein (eGFP). Four weeks after instillation, lung mechanics, morphometry, tissue cellularity, gene expression, inflammatory cytokines, and growth factor expression were analyzed.
  
  
  RESULTS
  No significant differences were observed in lung mechanics and morphometry among the experimental groups. However, the number of polymorphonuclear cells was higher in the WT-AAV9 group than in the CTRL and M-AAV9 groups, suggesting that the administration of tyrosine-mutant AAV9 vectors was better tolerated. Tyrosine-mutant AAV9 vectors significantly improved transgene delivery to the lung (30%) compared with their wild-type counterparts, without eliciting an inflammatory response.
  
  
  CONCLUSIONS
  Our results provide the impetus for further studies to exploit the use of AAV9 vectors as a tool for pulmonary gene therapy.
authors.affiliation: Laboratory of Cellular and Molecular Physiology, Carlos Chagas Filho Institute of Biophysics, Federal University of Rio de Janeiro, Rio de Janeiro, Ilha do Fundx00E3;o, Brazil.
authors.forename: Sabrina V
authors.initials: SV
authors.lastname: Martini
authors.forename: Adriana L
authors.initials: AL
authors.lastname: Silva
authors.forename: Debora
authors.initials: D
authors.lastname: Ferreira
authors.forename: Rafael
authors.initials: R
authors.lastname: Rabelo
authors.forename: Felipe M
authors.initials: FM
authors.lastname: Ornellas
authors.forename: Karina
authors.initials: K
authors.lastname: Gomes
authors.forename: Patricia R M
authors.initials: PR
authors.lastname: Rocco
authors.forename: Hilda
authors.initials: H
authors.lastname: Petrs-Silva
authors.forename: Marcelo M
authors.initials: MM
authors.lastname: Morales
mesh_terms: D000818:Animals; D036022:Capsid Proteins / Q000235:genetics*; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics*; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008168:Lung / Q000378:metabolism*; D008297:Male; D008810:Mice, Inbred C57BL; D017354:Point Mutation; D011897:Random Allocation; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection / Q000379:methods*; D019076:Transgenes / Q000235:genetics; D014443:Tyrosine / Q000235:genetics*
chemical_list: D036022:Capsid Proteins; D016207:Cytokines; D036341:Intercellular Signaling Peptides and Proteins; D049452:Green Fluorescent Proteins; D014443:Tyrosine