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- Title
Retroviral vector design studies toward hematopoietic stem cell gene therapy for mucopolysaccharidosis type I.
- Authors
Pan, D; Aronovich, E; McIvor, R S; Whitley, C B
- Abstract
To optimize a gene transfer system for hematopoietic stem ceil gene therapy of patients with mucopolysaccharidosis (MPS) type I, 10 retroviral vectors were constructed to express the human α-L-iduronidase (IDUA) cDNA. These vectors were designed to evaluate the potential effects of specific promoters, the addition of selectable markers, and the use of multiple promoters versus an internal ribosome entry site for expression of IDUA and selectable maker genes. The effect of vector design was investigated in primary patient fibroblasts (F[sub MPS]) or murine fibroblast cell lines; while overall comparison of transgene expression was determined in patients' peripheral blood lymphocytes (PBL[sub MPS]) and CD34[sup +] progenitors (PBPC[sub MPS]). We observed that the human PGK promoter introduced the highest IDUA activity per 1% relative transgene frequency in F[sub MPS]. Use of the same promoter to separately regulate both the therapeutic gene and a drug-resistance gene resulted in decreased expression of the unselected gene. Co-selection using bicistronic vectors not only increased the number of transductants, but also elevated transgene expression under selective pressure in transgene-positive progenitors. Bicistronic vector LP1CD overcame down-regulation and practically introduced the highest IDUA level in unselected PBL[sub MPS] and an intermediate level in PBPC[sub MPS]. These studies provide a better understanding of factors contributing to efficient gene expression in hematopoietic cells.
- Subjects
GENETIC transformation; MUCOPOLYSACCHARIDOSIS
- Publication
Gene Therapy, 2000, Vol 7, Issue 21, p1875
- ISSN
0969-7128
- Publication type
Article
- DOI
10.1038/sj.gt.3301298