4. Growth and selection of transduced lymphocytes Transduced lymphocytes will be 
expanded and selected by culture in CM with 400 ug/ml G418 for 7-8 days. Efficiency 
of selection will be monitored in parallel by viability of non-transduced cultures. After 
selection is complete, G418 will be removed and the cultures expanded up to 3 X 10 9 /kg 
for reinfusion. 
5. Infusion of transduced, expanded autologous Ivmohocvtes . The transduced cells will be 
harvested, washed, and resuspended in normal saline. The final cell preparation will be 
filtered through a platelet filter and transferred into a syringe or transfusion pack for 
infusion. Patients will be admitted to the Clinical Research Facility or the inpatient 
service of the V.A. Medical Center. Interim history and physical examination will be 
performed by a study physician. Intravenous catheterization with standard sterile 
technique will be performed and good venous return and absence of infiltration will be 
verified immediately before infusion. The infusion will be of not more than 5x1 0 8 
cells/kg of body weight. (The dose target per infusion in the ADA Clinical protocol, ref. 
22, was 1-3 x 10 9 per kg). Total volume of infused cells should not exceed 10 ml/kg of 
body weight. After an initial test infusion of 1-5% of the total volume, cells will be 
infused over the next 60-120 minutes. During infusion, the cell suspension will be 
mixed gently approximately every 5 minutes while the patient is being observed for 
acute and subacute toxicity. Vital signs will be monitored before infusion and every 15 
minutes during and 2 hours after infusion or until the patient is stable. The infusion 
will be terminated if systolic blood pressure falls below 80 mm Hg, oxygen saturation 
falls below 90%, or other evidence of major systemic toxicity occurs. Patients will have 
nursing observation for 24 hours after infusion; patients will be discharged after 24 
hours if there are no complications. A blood sample or samples will be drawn prior to 
discharge for initial studies detailed in the following section. 
D. Follow-up studies 
The patients in this study will have direct access to an investigating physician via 
telephone or pager. The following patient visits and data collection are. planned (on the 
schedule indicated in table 1 below): 
1. Interim history and physical examination by an investigating physician. 
2. Isolation of PBLs for: 
Recombinant DNA Research, Volume 18 
[393] 
