Adeno-associated virus Rep-mediated targeting of integrase-defective retroviral vector DNA circles into human chromosome 19

• Published in: Biochemical and biophysical research communications Vol. 417; no. 1; pp. 78 - 83
• Main Authors: Huang, Shuohao, Kawabe, Yoshinori, Ito, Akira, Kamihira, Masamichi
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 06.01.2012
• Subjects: 60 APPLIED LIFE SCIENCES; AAVS1; Adeno-associated virus; Adeno-associated virus (AAV); ANIMAL CELLS; CATTLE; chromosome 19; Chromosomes, Human, Pair 19 - genetics; Circular DNA; Clinical trials; Dependovirus - genetics; DISEASE VECTORS; DNA; DNA, Circular - genetics; DNA-Binding Proteins - genetics; Expression vectors; FLUORESCENCE; Gene Targeting - methods; GENE THERAPY; Genetic Vectors - genetics; Genomes; HeLa Cells; Humans; insertional mutagenesis; Integrase-defective retroviral vectors; Integrases - genetics; Integration; MUTAGENESIS; Mutagenesis, Insertional; Packaging; Plasmids; PROTEINS; Recombination, Genetic; Rep; Rep protein; Targeted integration; Transgenes; Tropism; Viral Proteins - genetics; VIRUSES; Adeno-associated virus (AAV); RBS; Targeted integration; AAVS1; p5IEE; Integrase-defective retroviral vectors; LTR; EGFP; TRS; AAV; FBS; Rep; IDRV
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2011.11.059

► Adeno-associated virus (AAV) is capable of targeted integration in human cells. ► Integrase-defective retroviral vector (IDRV) enables a circular DNA delivery. ► A targeted integration system of IDRV DNA using the AAV integration mechanism. ► Targeted IDRV integration ameliorates the safety concerns for retroviral vectors. Retroviral vectors have been employed in clinical trials for gene therapy owing to their relative large packaging capacity, alterable cell tropism, and chromosomal integration for stable transgene expression. However, uncontrollable integrations of transgenes are likely to cause safety issues, such as insertional mutagenesis. A targeted transgene integration system for retroviral vectors, therefore, is a straightforward way to address the insertional mutagenesis issue. Adeno-associated virus (AAV) is the only known virus capable of targeted integration in human cells. In the presence of AAV Rep proteins, plasmids possessing the p5 integration efficiency element (p5IEE) can be integrated into the AAV integration site (AAVS1) in the human genome. In this report, we describe a system that can target the circular DNA derived from non-integrating retroviral vectors to the AAVS1 site by utilizing the Rep/p5IEE integration mechanism. Our results showed that after G418 selection 30% of collected clones had retroviral DNA targeted at the AAVS1 site.