Recombinant DNA Advisory Committee - 06/1-2/92 
integrated proviral DNA? Dr. Smith responded that the patients will be monitored by 
PCR analysis. 
Dr. Parkman asked if the investigators will be looking at the DNA rather than the RNA? 
The examination of DNA would be making the assumption that all of the RNA is going 
to be transcribed. Dr. Smith stated that he would address this issue later. 
Dr. Haselkom asked Dr. Smith to explain the fact that almost complete protection from 
HIV infection is observed if only 10-20% of the cell population has been transduced? 
Dr. Smith stated that one explanation would be that TAR decoy transduced cells are 
growing out and expanding during this time period and diluting out immunofluorescence 
assay (IFA) + and P24 producing cells. 
Dr. Smith said that one of the questions that he had attempted to address was whether 
or not the production of TAR decoy RNA adversely affects the immunologic function of 
CD4( + ) cells. A CD4( + ) cytotoxic T lymphocyte (CTL) clone was transduced with the 
DCTAR encoding provirus. G418 selection of neomycin resistance expressing subclones 
was performed. Subcloning was performed to obtain CTLs that express high levels of 
the DCTAR. These cells were tested in a chromium release assay and compared to 
control cells. No significant difference was observed in CTL function. 
Dr. Smith stated that transduced cells were assayed for malignant transformation. 
Following 21-25 days in culture with IL-2, the cells were concentrated, divided into two 
aliquots and recultured with or without IL-2. No evidence of IL-2 independent growth 
was observed. 
Dr. Smith said that assays were performed for the detection of replication competent 
murine retrovirus using the NIH 3T3 amplification assay. Culture supernatants from 
CD4( + ) DCTAR transduced cells were examined after several weeks in culture. No 
evidence of replication competent retrovirus was observed. 
In response to the reviewer's concerns about the autologous protocol, Dr. Smith stated 
that the apheresis procedure may adversely affect the HIV infected individuals because 
their CD4 counts are marginal. In addition, there is concern that the mitogen stimulus 
in these cells may actually increase the HIV burden when reintroduced into the patient. 
Therefore, the current focus should be on the syngeneic protocol. The syngeneic 
protocol is the only protocol he wants the RAC to consider today. 
Dr. Smith stated that he does not expect to observe a therapeutic effect from this 
protocol because of the small number of cells that will be infused. 
Dr. Smith addressed the RAC's concerns regarding the status of the preclinical data for 
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