autologous cells infused after TBI, VP-16, and cyclophosphamide. 
On the other hand, the shorter remission duration in other patients 
suggests that residual systemic disease may be responsible for the 
relapse. The use of the NEO vector marking of autologous cells 
will permit the identification of the source of relapse and thus 
provide direction to the implementation of changes (more intensive 
preparative regimens or in vitro purging programs of autologous 
cells) to reduce the relapse rate. 
The use of the PCR assay for the bcr-abl mRNA of CML in this 
study permits the characterization of the NEO marked colonies with 
respect to their normal or leukemic origin, both at the time of 
autologous cell collection and at the time of relapse. 
The following issues relevant to therapy may also be 
ultimately clarified by such marking protocols: 
a) the clonality of relapse 
b) the efficiency of in vitro purging procedures 
c) the efficiency of systemic preparative therapy 
d) and in later protocols, the efficiency of methods to 
separate autologous early progenitor cells from normal or 
leukemic marrow. 
1. USE OF RECOMBINANT DNA FOR THERAPEUTIC PURPOSES: Not applicable 
2. TRANSFER OF DNA FOR OTHER PURPOSES: 
a. INTO WHAT CELLS WILL THE RECOMBINANT DNA BE TRANSFERRED? 
2 X 10^° nucleated cells (2 x 10® cells/kg) will be harvested 
from marrow following recovery from chemotherapy and 6 x 10^® 
nucleated cells from peripheral blood during the very early stages 
of recovery (0.2 - 0.8 wbc/mm^) . The latter collection has been 
shown to generate predominantly diploid cells and to accelerate 
hematopoietic recovery over marrow cells alone. These autologous 
cells (marrow and peripheral blood) will be separated by Ficoll 
hypaque gradients. Thirty percent of this population will be used 
for the vector marking experiments. The total population of 
peripheral blood and marrow nucleated cells has the capability of 
reconstituting marrow function in patients following TBI, VP-16, 
cyclophosphamide therapy in a median of 24 days. 
b. WHY IS RECOMBINANT DNA NECESSARY FOR THIS PURPOSE? 
Recombinant DNA marking is the only method available for 
marking cells used for transplantation which is safe and effective. 
The recombinant DNA marker is stable over the life of the cell. 
All daughter cells contain the same marker signal, and it is not 
diluted by cell division. If a cell dies, its marker signal also 
disappears. Each integration event of the marker gene serves as a 
lineage marker, identifying all progeny of that cell. 
Recombinant DNA Research, Volume 14 
[743] 
