HEPATOCELLULAR TRANSPLANTATION AND TARGETING GENETIC MARKERS TO HEPATIC CELLS 
sensitivity of this method is only limited by the completeness of the restriction digest 
and probability of getting a false positive PCR product. The applicability of this 
method is also limited by the ability to identify such fortuitous polymorphism between 
host and donor. A final method would be to restrict these studies to the use of male 
donor cells and female recipient cells which would enable detection of transplanted 
cells with sex specific markers. 
We propose to mark transplanted hepatocytes with a recombinant gene which can be 
used to identify the transplanted cells. The marker gene we propose to use is the gene 
for neomycin phosphoribosyl- transferase (NEO-R) derived from the bacteriophage TN5 
(Southern and Berg, 1982). This gene has been recombined into a "defective" retroviral 
vector designated LNL6 (Bender et al, 1987; Miller and Rosman, 1989). The LNL6 vector 
is an amphotrophic retroviral vector which is capable of infecting human cells, 
integrating the NEO-R gene stably into the genome of the transduced cell, and directing 
expression of the NEO-R gene. The principle of using marker genes for htiman experiments 
has been explicitly approved or human use by the Recombinant DNA Advisory Committee 
(RAC) which reported: 
"The sense of the Subcommittee and outside consultants is supportive of the 
general concept of the use of recombinant vectors in gene transfer procedures 
for marking somatic cells in humans as an aid to the development of important 
new advances in clinical research. Because such procedures are not done 
primarily to benefit the subject, and may in fact be of no benefit to the 
individuals involved, proposals to carry out these experiments must be 
supported by a clear data base demonstrating if a specific procedure planned 
is safe and likely to yield knowledge of value." (Recombinant DNA Advisory 
Committee, 1988) 
We believe that there is sufficient existing data from laboratory, animal, and human 
studies to indicate that the LNL6 virus is "safe". We also believe that the published 
literature on animal experiments with HCT demonstrates the importance of having 
unequivocal genetic markers in order to "yield knowledge of value". While the LNL6 
vector may not be the optimal genetic marker for these experiments, it is the best 
marker available for human studies at this time and we believe there is a significant 
probability that the use of this vector will "yield knowledge of value" . As described 
below, the standards for clinical investigation which may be of no benefit to patients 
(children) require that the risks be commensurate with those of the underlying disease 
process or conventional therapy. We believe these criteria are satisfied for the use 
of marker genes in the proposed protocol. 
F. Retroviral mediated gene transfer and somatic gene therapy. 
The ability to mark hepatocytes with a vector such as LNL6 is an outgrowth of 
research directed at developing methods for somatic gene therapy. In several recent 
reviews, we have summarized various aspects of somatic gene therapy. A didactic review 
described the principles underlying gene therapy and their application to pediatric 
disease for pediatricians (Ledley, 1987a; 1987b). Another review surveys the 
Recombinant DNA Research, Volume 14 
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