Human Gene Therapy Subcommittee - 4/5/91 
to improve treatment for human disease. Ms. Meyers maintained that cancer was being 
over-emphasized in the protocols that had been presented to the subcommittee, and 
there had been a lack of emphasis on hereditary diseases. 
Dr. Walters called for a brief coffee break and asked the subcommittee to move to Dr. 
Brenner upon return. After the recess, Dr. Walters reconvened the subcommittee and 
asked Dr. Brenner to respond to the questions and comments of the subcommittee 
members. 
Dr. Brenner stated that the goal of the protocol was not merely to determine whether 
the relapse was coming from the reinfused marrow or from the patient's residual tumor. 
If the relapse could be proven to be coming from the marrow, it would be possible to 
use this technique of cell marking to monitor the efficacy of purging and determine if 
any one of the several purging techniques for neuroblastoma was superior to another. 
He presented data that showed that primary clonigenic tumor cells can be transduced, 
but that the efficiency of transduction is low, between .3% and 3%, when colonies are 
grown in G418. He addressed the issue of cycling and non-cycling cells, stating that 
although non-cycling cells cannot be detected they could still be transduced. It is a 
reasonable estimate to assume that 1% of clonigenic neuroblastoma cells can be marked. 
Dr. Parkman argued this point, stating that there were techniques, such as in situ 
hybridization, to determine whether non-cycling neuroblastoma cells were being 
transduced. Dr. Brenner said they could only detect neuroblastoma cells with the 
neomycin gene if it is expressed in a colony-forming assay and that individual cells 
expressing the neomycin resistance gene cannot be detected looking at clonigenic 
precursors in vitro. Dr. Brenner said that he understood Dr. Parkman's point and added 
that he would revise his estimate for transduction frequency and consider it to be 0.1%. 
Dr. Brenner then presented data showing that neuroblastoma colonies can be identified 
in vitro and the presence of the neomycin gene can be verified using PCR. He further 
showed that the efficiency of transduction was low but was detectable using PCR. He 
also presented data showing that, despite the low level of transduction, it was possible to 
detect marked relapses with sufficient frequency to be able to compare the efficacy of 
different purging techniques. This would provide up to 1,000 times greater sensitivity 
than any other technique of minimal residual disease detection and would provide the 
only available method for assessing whether marrow purging actually works. 
Dr. Leventhal questioned whether performing the experiment in vitro would yield the 
same results. Dr. Brenner said there would be no way to assess the survival of the cells 
in vivo, and since only a proportion of the cells are clonigenic, it would not answer some 
of Dr. Parkman's concerns about cycling and non-cycling cells being able to contribute to 
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