Comparison of insulators and promoters for expression of the Wiskott-Aldrich syndrome protein using lentiviral vectors

Hum Gene Ther Clin Dev. 2013 Jun;24(2):77-85. doi: 10.1089/humc.2012.244. Epub 2013 Jun 20.


Gene therapy for the treatment of Wiskott-Aldrich syndrome (WAS) presents an alternative to the current use of allogeneic bone marrow transplantation. We describe the development of a self-inactivating lentiviral vector containing chromatin insulators for treatment of WAS and compare a gammaretroviral (MND), human cellular (EF1α), and the human WASp gene promoter for expression patterns in vivo during murine hematopoiesis using the green fluorescent protein (GFP) marker. Compared with the EF1α and the WASp promoters, expression from the MND promoter in mouse transplant recipients was much higher in all lineages examined. Importantly, there was sustained expression in the platelets of secondary recipient animals, necessary to correct the thrombocytopenia defect in WAS patients. Analysis of WAS protein expression in transduced human EBV-immortalized B-cells and transduced patient peripheral blood mononuclear cells also demonstrated stronger expression per copy from the MND promoter compared with the other promoters. In addition, when analyzed in an LM02 activation assay, the addition of an insulator to MND-promoter-containing constructs reduced transactivation of the LM02 gene. We propose a clinical trial design in which cytokine-mobilized, autologous, transduced CD34(+) cells are administered after myelosuppression.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • B-Lymphocytes / cytology
  • B-Lymphocytes / metabolism
  • Blood Platelets / cytology
  • CCAAT-Enhancer-Binding Protein-alpha / genetics
  • CCAAT-Enhancer-Binding Protein-alpha / metabolism
  • Cells, Cultured
  • Chromatin / metabolism
  • Gammaretrovirus / genetics
  • Genetic Therapy
  • Genetic Vectors / genetics
  • Genetic Vectors / metabolism*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Humans
  • Lentivirus / genetics*
  • Leukocytes, Mononuclear / cytology
  • Leukocytes, Mononuclear / metabolism
  • Mice
  • Platelet Count
  • Promoter Regions, Genetic
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics
  • T-Lymphocytes / cytology
  • T-Lymphocytes / metabolism
  • Transduction, Genetic
  • Wiskott-Aldrich Syndrome / therapy*
  • Wiskott-Aldrich Syndrome Protein / genetics
  • Wiskott-Aldrich Syndrome Protein / metabolism*


  • CCAAT-Enhancer-Binding Protein-alpha
  • Chromatin
  • Recombinant Fusion Proteins
  • Wiskott-Aldrich Syndrome Protein
  • Green Fluorescent Proteins