Original Article

Gene Therapy (2009) 16, 1452–1464; doi:10.1038/gt.2009.96; published online 6 August 2009

Retroviral vector integration in post-transplant hematopoiesis in mice conditioned with either submyeloablative or ablative irradiation

M A Sadat1,9, S Dirscherl1,9, L Sastry2,9, J Dantzer2,3, N Pech1, S Griffin2, T Hawkins2, Y Zhao2,3, C N Barese1, S Cross2, A Orazi4, C An4, W S Goebel1, M C Yoder1,5, X Li3,8,10, M Grez6, K Cornetta2,7,8, S D Mooney2,3,11 and M C Dinauer1,2,7

  1. 1Department of Pediatrics, Herman B Wells Center for Pediatric Research, Riley Hospital for Children, Indiana University School of Medicine, Indianapolis, IN, USA
  2. 2Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN, USA
  3. 3Center for Computational Biology and Bioinformatics, Indiana University School of Medicine, Indianapolis, IN, USA
  4. 4Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN, USA
  5. 5Department of Biochemistry & Molecular Biology, Indiana University School of Medicine, Indianapolis, IN, USA
  6. 6Department of Molecular Virology, Georg-Speyer-Haus, Frankfurt, Germany
  7. 7Department of Microbiology/Immunology, Indiana University School of Medicine, Indianapolis, IN, USA
  8. 8Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA

Correspondence: Professor MC Dinauer, Cancer Research Institute R4, Indiana University School of Medicine, 1044 West Walnut Street, Room 402A, Indianapolis, IN 46202 USA. E-mail: mdinauer@iupui.edu

9These authors contributed equally to this work.

10Current address: Burnett School of Biomedical Science, University of Central Florida, Orlando, FL, USA

11Current address: Buck Institute for Age Research, Novato, CA, USA

Received 18 May 2009; Accepted 9 June 2009; Published online 6 August 2009.

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Abstract

X-linked chronic granulomatous disease (X-CGD) is an inherited immunodeficiency with absent phagocyte NADPH-oxidase activity caused by defects in the gene-encoding gp91phox. Here, we evaluated strategies for less intensive conditioning for gene therapy of genetic blood disorders without selective advantage for gene correction, such as might be used in a human X-CGD protocol. We compared submyeloablative with ablative irradiation as conditioning in murine X-CGD, examining engraftment, oxidase activity and vector integration in mice transplanted with marrow transduced with a γ-retroviral vector for gp91phox expression. The frequency of oxidase-positive neutrophils in the donor population was unexpectedly higher in many 300cGy-conditioned mice compared with lethally irradiated recipients, as was the fraction of vector-marked donor secondary CFU-S12. Vector integration sites in marrow, spleen and secondary CFU-S12 DNA from primary recipients were enriched for cancer-associated genes, including Evi1, and integrations in or near cancer-associated genes were more frequent in marrow and secondary CFU-S12 from 300cGy-conditioned mice compared with fully ablated mice. These findings support the concept that vector integration can confer a selection bias, and suggest that the intensity of the conditioning regimen may further influence the effects of vector integration on clonal selection in post-transplant engraftment and hematopoiesis.

Keywords:

chronic granulomatous disease, NADPH oxidase, retrovirus, insertional mutagenesis