migrate to the tumor, thus theoretically producing high local TNF levels while 
potentially avoiding the systemic side effects. 
In addition, recently three human gene transfer protocols are being reviewed 
which serve to mark tumor cells in the bone marrow of patients undergoing autologous 
bone marrow transplantation (ABMT) (21). They will attempt to determine the role of 
these tumor cells on cancer relapse in these patients after ABMT. These protocols are 
the first to propose using retroviral stock on cells which will be immediately removed 
from the viral stock and injected into a patient, rather than first monitoring the cells in 
vitro for adverse side effects (i.e. , replication competent virus production or tumor 
formation due to an oncogenic event). However, assays to detect replication 
competent virus will allow this procedure to be performed safely since these assays 
can detect replication competent virus in the viral stock. 
The ability to transfer genes to non-transformed cells has become possible 
over the past decade. The most common method for gene transfer into non- 
transformed cells is by retroviral-mediated methods (22). Retroviral vectors, most 
commonly generated from the Moloney murine leukemia virus (M-MuLV), will 
efficiently transduce, stably insert into, and generate high levels of recombinant protein 
in host cells (23). The receptor for murine leukemia viruses is ubiquitous in 
mammalian species. A number of potential hazards exist when using the M-MuLV 
retroviral vector. First, the M-MuLV wild type retrovirus is a leukemia virus in some 
murine species. It can cause leukemia, in part, through protooncogene activation in 
the infected cell by the retroviral long terminal repeat (LTR). Second, though retroviral 
vectors are “crippled” retroviruses and thus are replication incompetent, they possess 
the ability to recombine with retroviruses within a packaging cell line to form a 
replication competent virus. 
Though the M-MuLV retroviral vector still possesses its LTR, the frequency with 
which oncogenesis occurs in the transduced cells appears to be very low. The 
overwhelming majority of reported experiments performed over the past decade with 
retroviral vectors demonstrate no association with transformed cells. These studies 
include experiments in murine, dog, monkey, and sheep models to name a few. 
However, we describe below an association of retroviral vectors with transformed cells, 
which appears to be low in frequency. 
The ability of retroviral vectors to undergo recombination events to produce 
replication competent virus can be detected by sensitive assays which have been 
established over the past decade. These assays are termed S + /L' and Helper Rescue 
Assay (24). If a recombination event occurs, it will almost always occur in the producer 
cell. The producer cell line contains both a helper virus and the retroviral vector. The 
producer cell line generates the viral particles used for transduction. Therefore, the 
producer cell line can be continually monitored for the appearance of replication 
competent virus. Cells transduced with replication incompetent viral stock, as 
[578] 
Recombinant DNA Research, Volume 15 
