normal volunteer non-mobilized peripheral blood CFU-C were 
transduced. Preliminary data indicates that marrow 
harvested from patients treated with 5FU 10 days prior to 
harvest had an increased transduction efficiency. 
3 . 5 Feasibility of gene transfer to tumor cells : 
Retroviral vectors have been successfully used to mark a 
number of fresh human malignant cells as well as cell lines. 
Our laboratory has shown 10% transduction of human myeloma 
cell lines ( IM9 and HS-Sultan). However, we have not beerT 
able to obtain in vitro evidence for transduction of primary 
myeloma tumor cells because of the difficulty of growing 
these cells in culture as colonies. We have isolated and 
sequenced tumor-specific immunoglobulin gene rearrangements 
in individual myeloma patients, and created allele-specific 
primers that can be used in a PCR assay to sensitively and 
specifically detect tumor cells in any cell population. 51 
This technique may allow us to assay concurrently for the 
presence of a tumor-specific genetic sequence and the 
transduced Neo gene in FACS-sorted plasma cells, B-cell 
progenitors, and other cell populations after 
transplantation with transduced cells. In the murine system, 
our laboratory has shown that marked plasma cells are 
detectable in animals transplanted with bone marrow 
transduced with a Neo-IL6 retroviral vector. 52 
Transduction efficiency of the K562 chronic myelogenous 
leukemia cell line has been shown by many laboratories to be 
greater than 10%. Transduction efficiency of bcr/abl 
positive CML CFU-C in our preliminary experiments is 5-30% 
(Appendix D) We can detect the presence of both the bcr/abl 
translocation and a transduced Neo gene in individual 
colonies. Transduction efficiency of primary breast 
carcinoma cells has not been assessed, again due to the 
difficulty of growing these primary tumor cells in culture. 
Breast cancer cell lines can be transduced with efficiencies 
from 1-10% .( Personal communication. Dr. Ken Cowan) 
3 . 6 Safety of the transduction procedure : 
The LNL6 and GIN. 40 Neo* marking vectors: The retroviral 
vectors to be used in this protocol are safety-modified 
versions of the retroviral vector N2. This vector was 
constructed by replacing the viral gag, pol and env genes 
from the Moloney murine leukemia virus (MoMLV) with the 
bacterial neomycin phosphotransferase gene. Since these 
vectors contain no viral genes, after integration into the 
target cell chromosome they are incapable of producing the 
[14] 
Recombinant DNA Research, Volume 16 
