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Ability to transfer marrow progenitor cells into non-ablated recipients 
Using mouse models, investigators have been able to demonstrate the successful transfer 
of marrow stem cells into recipient mice without first "making space" with ablative 
treatments (32-36). These studies also have demonstrated that these non-ablative 
transfers can be repeated with a cumulative effect. Based on these rodent experiments 
it is rational to expect engraftment of autologous cells removed from a patient and treated 
ex vivo with retroviral vectors if the patient’s marrow is not ablated. 
Some human diseases have been successfully treated with similar non-ablative marrow 
transplantation regimens. These diseases include the allogeneic transplantation of marrow 
into patients with T-cell or severe combined immune deficiencies (ADA negative SCID, 
ADA positive SCID, Short-limbed dwarf). In these patients the donor cells have 
engrafted successfully and corrected the underlying immune defect. The donor marrow 
can be shown to have repopulated the immune system (T-cells) preferentially. Donor 
engraftment in other compartments is more difficult to demonstrate but has been seen. 
The ability to serially engraft patients with repetitive transplants has been shown in some 
of these diseases as well. In one form of immune deficiency, a patient with Wiskott- 
Aldrich syndrome who received a transplant from his father after ablation remained 
chimeric and thrombocytopenic. This patient then received marrow from his father again 
without ablation. This transplant was shown to engraft by HLA and antigen studies of 
various marrow populations as well as platelets (Horowitz, personal communication). 
The patients thrombocytopenia was corrected with platelets of parental origin. These 
human experiences support the concept that "making space" is not absolutely necessary 
to obtain engraftment of marrow cells. 
Rational e/Justification 
As outlined in the previous sections, retroviral vectors can efficiently transfer and express 
the GC (glucocerebrosidase) cDNA into rodent, primate and human hematopoietic cells 
(including Gaucher disease patient cells). The level of expression is appropriate for 
therapeutic benefits and the gene is expressed in the appropriate target cells 
(monocyte/macrophages). Clinical experience with similar vectors also demonstrate the 
utility of RMGT (retroviral mediated gene transfer) to transduce and express in the 
appropriate target cells. Human and animal experience suggests that it should be possible 
to engraft hematopoietic stem cells without requiring ablation. All components necessary 
to execute the plan in this protocol have been demonstrated in both animal and human 
protocols. 
It is estimated that there is one stem cell in every 10 s murine mononuclear bone marrow 
cells. If this ratio of total bone marrow cells to stem cells applies to humans, then we 
will be transducing an average of 1-10 X 10 4 stem cells from each patient. Even with 
a transduction efficiency of 1 % or lower, it would be likely that we would be able to 
detect stem cell transduction in each patient as if it was actually occurring in vitro under 
these transduction conditions. 
Recombinant DNA Research, Volume 17 
