Appendix C 
SCIENTIFIC ABSTRACT 
A replication incompetent, recombination incompetent 
retroviral vector will be used to mark autologous peripheral blood 
and marrow cells stored at the time of cytogenetic remission or re- 
induction of chronic phase in Philadelphia chromosome positive CML 
patients who have developed blast crisis or accelerated phase and 
have been reinduced into chronic phase following Daunomycin, high- 
dose Ara-C, and GM-CSF therapy. We estimate that between 1.2 x 10^ 
and 1.2 x 10^ leukemic marrow blast cells and 4.2 x 10^ and 4.2 x 
10^ leukemic peripheral blood blastic CML cells will be marked with 
NEC in the autologous cells used for transplant. It is not known 
how many CML blastic leukemia cells are present in the systemic 
circulation following induction of the chronic phase or a 
cytogenetic remission by Daunomycin, high-dose Ara-C, and GM-CSF. 
The number must be low judging from the duration of remissions 
which may last greater than one year following autologous 
transplant. We will look for the number of NEO-marked cells using 
a methylcellulose late progenitor colony culture system and a PCR 
assay for the NEO gene used previously by Cornetta, et al (Human 
Gene Therapy 1:15-30, 1990), and Dr. Malcolm Brenner. In the CML 
cells under analysis, the percent of these NEO-marked cells which 
are leukemic can be determined by a PCR assay for the bcr-abl mRNA 
positive CML cells. We will also use PCR to determine the number 
of different vector integration sites and characterize the relapsed 
cells with respect to the presence of polyclonal or clonal vector 
integration sites. These studies will clarify if relapse arises 
from the leukemic CML blast cells present in the autologous cells 
infused after TBI, VP-16, and cytoxan (if polyclonal CML NEO-marked 
blastic cells appear at the time of relapse) , or if residual 
systemic disease contributes to relapse (if none of the CML 
leukemic blasts at the time of relapse contain the NEO gene) . 
Recombinant DNA Research, Volume 14 
[769] 
