Pre-IDE Submission: Clinical Protocol 
Neuroblastoma Bone Marrow Purging System 
BAXTER HEALTHCARE CORPORATION, HYLAND DIVISION 
Children’s Research Hospital for transduction and purging. Conditions used for 
transportation of harvested and frozen purged marrows have been described previously 
(Janssen et al, 1991). Alternatively, the patient may be admitted to St. Jude for 
harvesting as well as transduction and purging. 
Patients will have marrow harvested under general anesthesia using aseptic technique. 
Multiple aspirates will be obtained from the posterior and anterior iliac crests into 
heparinized syringes, and pooled in heparinized tissue culture medium. A volume of 
bone marrow and blood will be aspirated sufficient to provide >2.5x10* nucleated cells 
per kilogram of body weight. The specimen will be filtered through different size 
meshes to remove bony particles and aliquots will be obtained for in vitro laboratory 
studies. A marrow mononuclear cell fraction will be obtained using an automated cell 
separator as described in Appendix C. One third of the pooled bone marrow cells will 
be cryopreserved without further manipulation, and stored for rescue of the patient, 
should the remaining marrow fail to engraft. The remaining two thirds of the marrow 
cells will be used for transduction. A flow Chart illustrating the gene marking and 
purging procedures is shown in Figure 6. 1.5.4. 
Vector Transduction 
Two thirds (67 %) of the mononuclear cells will be used for cell-free vector transduction. 
One half of this will be transduced with LNL6 and one half with GINa. In each patient, 
one of the marked aliquots will be selected for subsequent immunomagnetic purging. 
The transduced aliquots will then be processed and cryopreserved using standard 
techniques. The patients will be treated with chemo-radiotherapy, following the protocol 
at the participating institution, and reinfused with the pooled transduced marrow. Should 
engraftment (ANC > 50C//xL) not occur within 43 days post-transplant, the patient will 
receive the backup marrow aliquot (unmarked) at the discretion of the attending physician 
in consultation with the principal investigator. 
Cell-free vector transduction will be performed as described in Appendix C. 
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Recombinant DNA Research, Volume 16 
