Large-scale analysis of adeno-associated virus vector integration sites in normal human cells

J Virol. 2005 Sep;79(17):11434-42. doi: 10.1128/JVI.79.17.11434-11442.2005.

Abstract

The integration sites of viral vectors used in human gene therapy can have important consequences for safety and efficacy. However, an extensive evaluation of adeno-associated virus (AAV) vector integration sites has not been completed, despite the ongoing use of AAV vectors in clinical trials. Here we have used a shuttle vector system to isolate and analyze 977 unique AAV vector-chromosome integration junctions from normal human fibroblasts and describe their genomic distribution. We found a significant preference for integrating within CpG islands and the first 1 kb of genes, but only a slight overall preference for transcribed sequences. Integration sites were clustered throughout the genome, including a major preference for integration in ribosomal DNA repeats, and 13 other hotspots that contained three or more proviruses within a 500-kb window. Both junctions were localized from 323 proviruses, allowing us to characterize the chromosomal deletions, insertions, and translocations associated with vector integration. These studies establish a profile of insertional mutagenesis for AAV vectors and provide unique insight into the chromosomal distribution of DNA strand breaks that may facilitate integration.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Chromosome Mapping
  • Chromosomes, Human / genetics
  • Chromosomes, Human / virology
  • DNA-Binding Proteins / genetics
  • Dependovirus / physiology*
  • Fibroblasts / virology*
  • Genetic Vectors / physiology*
  • Humans
  • Trans-Activators
  • Virus Integration*

Substances

  • CXXC1 protein, human
  • DNA-Binding Proteins
  • Trans-Activators