Epinephrine and antihistamines will be available at the bedside on a 
resuscitation cart as a precaution. Patients will be treated with benadryl 
(25mg) and Tylenol (625mg) 30 minutes prior to PSC infusion. Temperature, 
pulse, respiratory rate and blood pressure will be monitored every 15 minutes. 
Symptoms and signs of infusional toxicity will be recorded. 
L. ESTIMATION OF ENGRAFTMENT 
The first two patients receiving genetically corrected CD34 + cells will not receive 
myeloabl ation prior to treatment. As mentioned earlier in this application, some data 
suggests that engraftment can occur using whole bone marrow in an unprepared host’ 1 . These 
data provide some support for the concept that a habitable environment (niche) exists in 
which hematopoietic stem cells can engraft. While this concept remains controversial, it 
deserves testing in man because of the potentially reduced risks afforded by this approach. 
It would be to the patient's advantage to be able to autologously transplant genetically 
corrected cells without myeloablation. Therefore, we feel obligated to evaluate this 
possibility. Furthermore, enrichment for stem cells by CD34 selection increases the chance 
of engraftment of cells with the potential of bone marrow reconstitution on a long term 
basis. Enrichment for CD34 + cells may provide a greater chance of success than earlier 
studies using whole bone marrow. This favorably increases the odds of establishing 
permanent fraction of bone marrow that is genetically corrected and able to produce 
enzymatically competent macrophages. A critical appraisal to be made is whether a 
fractional correction of the marrow with genetically and enzymatically repaired cells has 
a chance of being therapeutic. Animal and human studies suggest that this is possible. 
In studies of the gus/gus model of mucopolysaccharidosis VII (gl ucocerebrosidase 
deficiency), bone marrow transplantation with genetically corrected whole marrow, in which 
expression of the transgene was suboptimal, resulted in enzymatic correction of tissues to 
only 2-5% of control levels 1891 . Despite this, the visceral manifestations of the disease 
were dramatically reversed. Furthermore, in Gaucher disease and Pompe disease, orthotopic 
liver transplantation results in a small fractional chimerization of the recipient marrow 
by donor marrow. Chimerization is believed to be a contributing factor to the therapeutic 
response in these disorders 1901 . Furthermore, targeting of glucocerebrosidase to macrophages 
results in only a temporary and incomplete correction of enzymatically competent cells, yet 
is therapeutic. From available data on the genetic correction of the LLD receptor in the 
Watanabe rabbit, it would also appear that a partial correction of functional gene product 
is sufficient to result in a significant reversal of phenotype 1911 . These collective 
experiences suggest that a small correction of functional enzymatic activity in GD will 
result in a therapeutic response. 
The protocol is designed to evaluate the possibility that engraftment of genetically 
corrected CD34 + cells will result in a biochemical and clinical response without 
myeloablation before proceeding to a regimen of partial myleoabl ation. (See Assessment of 
Risk/Benefit for a detailed discussion). The assessment of engraftment in these steps of 
the protocol depends on the estimation of enzymatic activity and immunocytochemical staining 
in peripheral blood leukocytes and bone marrow and on the measurement of indicators of 
therapeutic response in two clinical laboratory values in patients with Gaucher disease who 
have had a therapeutic response to enzyme replacement (see below). 
In mice, we have routinely observed an increase of 4-5 times the endogenous activity 
of glucocerebrosidase in freshly transduced donor marrow, primary CFU-S, secondary CFU-S, 
hematopoietic tissues from long-term reconstituted mice, and macrophages cultured from long- 
term reconstituted mice. In CD34 + cells from normal individuals, we have observed an 
increase in specific activity of GC of 2-4 fold. The transduction efficiency in these 
targets was 10-20% or 0. 1-0.2 transgene/genome. In CD34 + cells from Gaucher patients, the 
increment of activity is approximately 10 times greater owing to the fact that the 
endogenous activity of these enzyme deficient cells is approximately 10% of the control. 
Immunocytochemical staining of the patient cells is negative. Transduced cells show 
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Recombinant DNA Research, Volume 17 
