Enhanced transduction of mouse bone marrow-derived dendritic cells by repetitive infection with self-complementary adeno-associated virus 6 combined with immunostimulatory ligands.

Academic Article

Abstract

  • The potential of adeno-associated virus (AAV)-based vectors in human gene therapy is being explored for several diseases. Although sustained transgene expression and low vector-associated cellular immunity are attractive features of recombinant (r) AAV, the wider application of rAAV vectors encapsidated in serotype 2 capsid is hampered by poor transduction efficiency in many target tissues. These include ex vivo-generated dendritic cells (DC), which have demonstrated promising immunotherapeutic activity. We report here that efficient transduction of mouse bone marrow-derived DC can be achieved with self-complementary (sc) rAAV encapsidated in serotype 6 capsid. Sequential exposure of DC precursor cultures to IL-4 and GM-CSF with sc rAAV6 encoding the human tumor antigen, carcinoembryonic antigen (CEA), for 7 days followed by activation with CpG oligodeoxynucleotides (ODN) and anti-mouse CD40 antibody resulted in highly efficient transduction of DC. DC surface markers as determined by flow cytometry analysis of sc rAAV6-transduced DC were comparable to nontransduced DC. Efficiency of vector transduction and transgene expression were confirmed by immunostaining and real-time PCR. Microarray analysis of RNA from CpG ODN and CD40 antibody stimulated sc AAV6-transduced DC revealed upregulation of transcription factors and cytokines involved in immune activation and downregulation of inhibitory factors, suggesting a possible role of transcriptional activation in the observed effect. The adoptive transfer into syngeneic mice of the ex vivo-transduced and activated DC resulted in the development of CEA-specific antibody and T-helper 1-associated immune responses. Immunized mice also developed antibody to AAV6 capsid protein, which did not crossreact with AAV2 capsid protein. These studies demonstrate the potential utility of sc rAAV serotype 6-based vectors in transduction of DC for genetic vaccination approaches.
  • Published In

  • Gene Therapy  Journal
  • Keywords

  • Adoptive Transfer, Animals, Antibodies, Viral, Antibody Specificity, Antigen Presentation, Bone Marrow Cells, CD40 Antigens, Carcinoembryonic Antigen, Cell Line, CpG Islands, Dendritic Cells, Dependovirus, Enzyme-Linked Immunosorbent Assay, Flow Cytometry, Gene Expression, Genetic Engineering, Genetic Therapy, Genetic Vectors, Green Fluorescent Proteins, Humans, Immunohistochemistry, Interferon-gamma, Mice, Oligonucleotide Array Sequence Analysis, Reverse Transcriptase Polymerase Chain Reaction, Transduction, Genetic, Transgenes
  • Digital Object Identifier (doi)

    Author List

  • Aldrich WA; Ren C; White AF; Zhou S-Z; Kumar S; Jenkins CB; Shaw DR; Strong TV; Triozzi PL; Ponnazhagan S
  • Start Page

  • 29
  • End Page

  • 39
  • Volume

  • 13
  • Issue

  • 1