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

Gene Therapy volume 16, pages 1452–1464 (2009)Cite this article

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 300 cGy-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 300 cGy-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.

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Acknowledgements

We thank Robert Getty for assistance with LM-PCR and Shari Upchurch, Maureena Lewis, Melody Warman, and Catherine Matthews for help with preparing the article. This work was supported by National Institutes of Health Grants P01 HL53586 (MCD, MY, KC), K22 LM009135 (SM), K08 HL075253 (SG), RO1 HG004359 (XL), T32 CA111198 Cancer Biology Training Program (s.d.), T32 HL007910 Basic Science Studies on Gene Therapy of Blood Disease (TH) and the Riley Children's Foundation (MCD, MY, WSG). MAS, SD, LS and MCD designed, performed and analyzed experiments and helped draft the paper and figures; JD and TH analyzed data and prepared figures; NP, SG and SC performed and analyzed experiments; YZ and XL analyzed data, CB helped with experimental design and developed a critical procedure; AO and CA helped with analysis of leukemic tissue; WSG and MCY helped with interpretation of data and paper preparation; MG provided critical reagents and helped with data interpretation and preparation of the paper; KC and SM helped design, analyze and interpret the experiments and preparation of the paper; MCD oversaw this entire project including experimental design, analysis, interpretation of the data and preparation of the paper.

Author information

Author notes

  1. X Li

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

  2. S D Mooney

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

  3. M A Sadat, S Dirscherl and L Sastry: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Pediatrics, Herman B Wells Center for Pediatric Research, Riley Hospital for Children, Indiana University School of Medicine, Indianapolis, IN, USA

    M A Sadat, S Dirscherl, N Pech, C N Barese, W S Goebel, M C Yoder & M C Dinauer

  2. Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN, USA

    L Sastry, J Dantzer, S Griffin, T Hawkins, Y Zhao, S Cross, K Cornetta, S D Mooney & M C Dinauer

  3. Center for Computational Biology and Bioinformatics, Indiana University School of Medicine, Indianapolis, IN, USA

    J Dantzer, Y Zhao, X Li & S D Mooney

  4. Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN, USA

    A Orazi & C An

  5. Department of Biochemistry & Molecular Biology, Indiana University School of Medicine, Indianapolis, IN, USA

    M C Yoder

  6. Department of Molecular Virology, Georg-Speyer-Haus, Frankfurt, Germany

    M Grez

  7. Department of Microbiology/Immunology, Indiana University School of Medicine, Indianapolis, IN, USA

    K Cornetta & M C Dinauer

  8. Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA

    X Li & K Cornetta

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  1. M A Sadat
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Correspondence to M C Dinauer.

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Supplementary Information accompanies the paper on Gene Therapy website (http://www.nature.com/gt)

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Sadat, M., Dirscherl, S., Sastry, L. et al. Retroviral vector integration in post-transplant hematopoiesis in mice conditioned with either submyeloablative or ablative irradiation. Gene Ther 16, 1452–1464 (2009). https://doi.org/10.1038/gt.2009.96

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