treatment and many of the related clinical problems have resolved 127 281 . Both of these 
approaches are novel and are important because they provide some means of therapy where none 
previously existed. However, both approaches have significant limitations. Allogeneic bone 
marrow transplantation carries morbidity and mortality risks that are unacceptable for many 
patients. Furthermore, matched donors do not exist for the majority of patients. 
Macrophage-targeted enzyme replacement is a life-long treatment and extremely expensive 
therapy. Current costs are between $150,000 and $450,000 per year per patient. Clearly, 
some alternative treatment is required for this disease. 
Despite their limitations, the success of these strategies has demonstrated that 
enzymatic correction of only one cell type - the macrophage - results in effective therapy. 
This is important from the point of view of developing gene therapy for Gaucher disease 
because these approaches demonstrate the potential efficacy of gene therapy. The fact that 
marrow transplantation is effective allows one to consider hematopoietic stem cells (HSC) 
as an appropriate target cell for gene transfer. Successful transduction of a sufficient 
number of HSC could provide a life-long cure of the disease. Furthermore, because 
expression of the GC gene in the macrophage lineage alone may be sufficient for therapy, 
one could consider, as less permanent alternatives, gene transfer to committed macrophage 
precursors, peripheral blood monocytes, or to cultures of bone marrow capable of producing 
enzyme competent macrophage precursors. 
C. GENE TRANSFER/GENE THERAPY 
To be an effective permanent treatment, gene therapy requires the transfer and 
sustained expression of genes in cells important to the pathogenesis of the disease. 
Sufficient expression of a transduced glucocerebrosidase gene in the progeny of pluripotent 
bone marrow stem cells would correct the deficiency of the enzyme in all ceil lineages j 
including monocytes/macrophages. Experience from allogeneic marrow transplantation and 
macrophage-targeted enzyme replacement supports the idea that gene therapy would be curative 
provided adequate expression of the GC gene were accomplished in a sufficient number of 
macrophages. Much experience has been gained with systems to evaluate the efficiency of . 
gene transfer and expression in hematopoietic cells using retroviral vectors' 29,301 . Most of 
these studies have been conducted in mice where conditions to accomplish efficient gene j 
transfer to hematopoietic stem cells have been defined' 31 ' 411 . Recent data suggest that a high | 
percentage of stem cells can be transduced and survive to repopulate marrow' 401 . For the ! 
mouse, critical parameters for efficient retroviral gene transfer and repopulation of bone 
marrow stem cells include high titer virus producer cell lines (VPL), pretreatment of mice 
with 5-fluorouracil (5-FU) to initiate stem cell cycling' 38 ' 401 , pre-culture of bone marrow 
with growth factors including IL-3 and IL-6' 431 , and extended infection by coculture of the 
cells with the VPL for 2-6 days with growth factors' 401 . Several studies have shown that the ; 
GC gene can be transferred to mouse bone marrow cells, but only two studies have 
demonstrated sustained expression in animals' 41 ' 471 . 
One of the early problems encountered in the transfer and expression of genes in bone 
marrow has been the low transduction efficiency of cells in the bone marrow capable of 
hematopoietic reconstitution of the animal 148 ' 581 . Improvements in gene transfer vectors and 
advances in the knowledge of the biology of the bone marrow, particularly in the use of 
cytokines to maintain cycling stem cells, has improved the ability to efficiently transfer 
genes to hematopoietic cells' 59 ' 641 . While much of this information has been derived from and 
may be applicable only in murine systems, the factors important to the transfer of genes 
to cells capable of hematopoietic reconstitution in larger animal species including man are 
being defined' 65 711 . Expression of transferred genes in vivo for long periods of time has 
been more problematic. The reasons for inconsistent expression of the transferred gene are 
not well understood but hypotheses include promoter suppression in gene transfer constructs 
with multiple promoters' 721 , down regulation of heterologous viral promoters' 73 ' 741 , and clonal 
loss of transduced long term repopulating cells' 751 . Despite these apparent obstacles, some 
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Recombinant DNA Research, Volume 17 
