Stable integration of recombinant adeno-associated virus vector genomes after transduction of murine hematopoietic stem cells

Hum Gene Ther. 2008 Mar;19(3):267-78. doi: 10.1089/hum.2007.161.

Abstract

We previously reported that among single-stranded adeno-associated virus (ssAAV) vectors, serotypes 1 through 5, ssAAV1 is the most efficient in transducing murine hematopoietic stem cells (HSCs), but viral second-strand DNA synthesis remains a rate-limiting step. Subsequently, using double-stranded, self-complementary AAV (scAAV) vectors, serotypes 7 through 10, we observed that scAAV7 vectors also transduce murine HSCs efficiently. In the present study, we used scAAV1 and scAAV7 shuttle vectors to transduce HSCs in a murine bone marrow serial transplant model in vivo, which allowed examination of the AAV proviral integration pattern in the mouse genome, as well as recovery and nucleotide sequence analyses of AAV-HSC DNA junction fragments. The proviral genomes were stably integrated, and integration sites were localized to different mouse chromosomes. None of the integration sites was found to be in a transcribed gene, or near a cellular oncogene. None of the animals, monitored for up to 1 year, exhibited pathological abnormalities. Thus, AAV proviral integration-induced risk of oncogenesis was not found in our study, which provides functional confirmation of stable transduction of self-renewing multipotential HSCs by scAAV vectors as well as promise for the use of these vectors in the potential treatment of disorders of the hematopoietic system.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Bleomycin
  • DNA, Viral / genetics*
  • DNA, Viral / metabolism
  • Dependovirus / genetics*
  • Dependovirus / metabolism
  • Female
  • Genetic Therapy
  • Genetic Vectors*
  • Genome, Viral
  • Hematopoietic Stem Cell Transplantation
  • Hematopoietic Stem Cells / cytology
  • Hematopoietic Stem Cells / metabolism
  • Hematopoietic Stem Cells / virology*
  • Mice
  • Mice, Inbred C57BL
  • Proviruses / genetics
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transduction, Genetic*
  • Transgenes
  • Virus Integration*

Substances

  • DNA, Viral
  • Recombinant Fusion Proteins
  • Bleomycin
  • Zeocin