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HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype.

Publication ,  Journal Article
Cortés, ML; Bakkenist, CJ; Di Maria, MV; Kastan, MB; Breakefield, XO
Published in: Gene Ther
August 2003

Ataxia-telangiectasia (A-T) is an autosomal recessive disorder characterized by neurodegeneration, immunodeficiency, cancer predisposition, genome instability, and radiation sensitivity. Previous research has shown that it is possible to correct the hereditary deficiency A-T by DNA transfection in cell culture, but the large size of the ATM cDNA (9 kb) limits the use of many vector types for gene replacement. HSV-1 amplicon vectors provide a means to deliver large genes to cells efficiently and without toxicity. In this study, the FLAG-tagged cDNA for human ATM was inserted into an HSV-1 amplicon under control of the CMV promoter (designated as HGC-ATM). FLAG-ATM expression was confirmed in 293T/17 cells and human A-T fibroblasts (GM9607) after transduction, by immunoprecipitation, Western analysis, and immunocytochemistry. Functional recovery was assessed by two independent assays. First, in vitro kinase assay showed that vector-derived ATM in GM9607 cells could successfully phosphorylate wt p53 using recombinant GST-p53(1-101). Second, in A-T cells infected with the HGC-ATM vector, the extent of accumulation in G2/M phase at 24 h postirradiation was similar to that observed in cells with wild-type endogenous ATM and lower than that observed in A-T cells infected with a control vector. Thus, these vectors provide a tool to test the feasibility of HSV-amplicons as gene therapy vectors for A-T.

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Published In

Gene Ther

DOI

ISSN

0969-7128

Publication Date

August 2003

Volume

10

Issue

16

Start / End Page

1321 / 1327

Location

England

Related Subject Headings

  • Tumor Suppressor Proteins
  • Transduction, Genetic
  • Protein Serine-Threonine Kinases
  • Humans
  • Herpesvirus 1, Human
  • Genetic Vectors
  • Genetic Therapy
  • Fluorescent Antibody Technique
  • Fibroblasts
  • DNA-Binding Proteins
 

Citation

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Cortés, M. L., Bakkenist, C. J., Di Maria, M. V., Kastan, M. B., & Breakefield, X. O. (2003). HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype. Gene Ther, 10(16), 1321–1327. https://doi.org/10.1038/sj.gt.3301996
Cortés, M. L., C. J. Bakkenist, M. V. Di Maria, M. B. Kastan, and X. O. Breakefield. “HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype.Gene Ther 10, no. 16 (August 2003): 1321–27. https://doi.org/10.1038/sj.gt.3301996.
Cortés ML, Bakkenist CJ, Di Maria MV, Kastan MB, Breakefield XO. HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype. Gene Ther. 2003 Aug;10(16):1321–7.
Cortés, M. L., et al. “HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype.Gene Ther, vol. 10, no. 16, Aug. 2003, pp. 1321–27. Pubmed, doi:10.1038/sj.gt.3301996.
Cortés ML, Bakkenist CJ, Di Maria MV, Kastan MB, Breakefield XO. HSV-1 amplicon vector-mediated expression of ATM cDNA and correction of the ataxia-telangiectasia cellular phenotype. Gene Ther. 2003 Aug;10(16):1321–1327.

Published In

Gene Ther

DOI

ISSN

0969-7128

Publication Date

August 2003

Volume

10

Issue

16

Start / End Page

1321 / 1327

Location

England

Related Subject Headings

  • Tumor Suppressor Proteins
  • Transduction, Genetic
  • Protein Serine-Threonine Kinases
  • Humans
  • Herpesvirus 1, Human
  • Genetic Vectors
  • Genetic Therapy
  • Fluorescent Antibody Technique
  • Fibroblasts
  • DNA-Binding Proteins