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An in vitro system for efficiently evaluating gene therapy approaches to hemoglobinopathies

Gene Therapy volume 7, pages 215–223 (2000)Cite this article

Abstract

A variety of gene therapy strategies are under development for the treatment of sickle cell anemia and other hemoglobinopathies. A number of alternative vectors have been developed to transfer and express the β-globin gene and other therapeutic molecules, but none has resulted in efficient transduction and stable long-term expression in primary hematopoietic cells. One reason for this problem is that most vectors are initially evaluated in immortalized cell lines which may not faithfully recapitulate the biology of primary erythroid cells. In order to provide a more relevant system for efficiently evaluating alternative vector constructs for β-globin disorders, we have developed (1) a simple method for generating primary human red blood cell (RBC) precursors in liquid culture established with mononuclear cells obtained from normal donors as well as patients with Hb SC disease; (2) a high titer retroviral vector which can be easily modified to optimize gene transfer and transgene expression; and (3) methods for transducing the RBC precursors at high efficiency. The development of simple and efficient methods and reagents for generating and transducing primary human RBC precursors provides a facile and effective means for screening alternative gene therapy strategies.

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Acknowledgements

RH was supported by NIH Fellowship training grant 5T32CA09307 and NIH HL57606. We wish to thank M Telen for her generous gift of the E6 antibody, and the nurses in labor and delivery and the medical oncology treatment center at Duke University Medical Center for their assistance in collecting UCB and peripheral blood samples. In addition, we thank Immunex for providing Flt-3-ligand.

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Authors and Affiliations

  1. Division of Hematology/Oncology, Department of Pediatrics, Duke University Medical Center, Durham, NC, USA

    R P Howrey

  2. Center for Genetic and Cellular Therapies, Duke University Medical Center, Durham, NC, USA

    R P Howrey, M El-Alfondi, K L Phillips, L Wilson, B Rooney, N Lan, B Sullenger & C Smith

  3. Division of Oncology and Stem Cell Transplantation, Department of Medicine, Duke University Medical Center, Durham, NC, USA

    C Smith

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  1. R P Howrey
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  2. M El-Alfondi
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  3. K L Phillips
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  4. L Wilson
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  6. N Lan
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  7. B Sullenger
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Howrey, R., El-Alfondi, M., Phillips, K. et al. An in vitro system for efficiently evaluating gene therapy approaches to hemoglobinopathies. Gene Ther 7, 215–223 (2000). https://doi.org/10.1038/sj.gt.3301064

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