Abstract
Ataxia-telangiectasia (A-T) is an autosomal recessive disorder characterized by neurodegeneration, immunodeficiency, cancer predisposition, genome instability, and sensitivity to ionizing radiation (IR). We have previously shown that a herpes simplex virus type 1 (HSV-1) amplicon vector carrying the human ataxia-telangiectasia mutated (ATM) complementary DNA (cDNA) is able to correct aspects of the cellular phenotype of human A-T cells in culture, and is also able to transfer the ATM cDNA to the Atm-/- mouse cerebellum. In order to achieve stable gene replacement, we have generated an HSV/adeno-associated virus (AAV) hybrid amplicon vector carrying the expression cassettes for the ATM cDNA [(9.2 kilobases (kb)] and enhanced green fluorescent protein (EGFP), flanked by AAV inverted terminal repeats (ITRs). This hybrid vector, in the presence of AAV Rep proteins, mediates site-specific integration into the AAVS1 site on chromosome 19 in human cells and in Atm-/- mice carrying that human locus. The functional activity of the vector-derived ATM was confirmed in vitro and in vivo by ATM autophosphorylation at Ser-1981 after IR. This proof-of-principle study establishes the ability of HSV/AAV hybrid amplicon vectors to mediate functional targeted integration of the ATM cDNA into A-T cells in culture and in Atm-/- mice in vivo, thus laying a foundation for possible gene therapy approaches in the treatment of A-T patients.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 81-88 |
| Number of pages | 8 |
| Journal | Molecular Therapy |
| Volume | 16 |
| Issue number | 1 |
| DOIs | |
| State | Published - Jan 2008 |
| Externally published | Yes |
Bibliographical note
Funding Information:We thank Yosef Shiloh (Tel Aviv University, Israel) and Michael Kastan (St. Jude Children's Research Hospital, Memphis) for advice and for providing the ATM cDNA. We thank Joanna Bakowska (NIH/NINDS), John lacomini and Jessamyn Bagley (Children's Hospital, Boston, MA) for helpful discussions and reagents. From Massachusetts General Hospital (Boston), we acknowledge Miguel Sena-Esteves and Deborah Schuback for helpful discussions, Joanne Yetz-Aldape for the FACS analysis, Tammy Gillis for the sequencing analysis, Sowmya Srinivas for technical assistance, and Suzanne McDavitt for editorial assistance. This work was supported by the A-T Children's Project and NINDS R21 NS047503.
