Recombinant DNA Advisory Committee - 12/3-4/92 
to determine the level of MDR expression at steady state following ABM transplant. 
Patients who have received taxol or vinblastine after transplant will be monitored for 
MDR expression to determine if selection is occurring. If protection is being conferred 
by MDR expression, a decrease should be observed in the duration and depth of the 
neutropenia/thrombocytopenia. Dr. O'Shaughnessy noted that there is an ongoing gene 
transfer protocol in which patients receive high dose ICE therapy with neomycin resistant 
(neo R ) transduced ABM stem cells. Patients who relapse and are treated with taxol or 
vinblastine will be used as controls for the MDR study. 
Dr. D. Miller said that this protocol should be directed at patients with tumor types that 
would benefit from this therapy. Since breast cancer patients already demonstrate MDR 
resistance following drug therapy, this therapy will probably not provide a therapeutic 
effect. Dr. O'Shaughnessy explained that 20 to 30% of breast cancer patients should 
benefit from the subsequent taxol or vinblastine therapy. Dr. Leventhal asked if eligible 
patients will have limited bone disease. Dr. O'Shaughnessy stated that patients with <; 3 
metastases on a bone scan will be eligible for this protocol. Metastatic breast cancer 
refers to metastases in the liver, lung, and soft tissues. 
Dr. Carmen inquired whether mixing experiments have been performed with tumor cells 
and transduced ABM cells. Dr. O'Shaughnessy stated that these experiments have been 
performed by Dr. E. J. Shpall. Dr. Shpall's data demonstrates that transduced ABM 
cells are detectable at concentration between 1 x 10 5 and 1 x 10 6 cells. 
Presentation--Dr. Nienhuis 
Dr. Nienhuis addressed the issue of in vitro expression of transduced cells and evidence 
of gene transfer. The DNA from MDR-1 transduced cells has been analyzed by PCR 
and reverse transcriptase (RT) PCR has been employed to determine RNA levels. 
Rhodamine efflux assays have been performed. Rhodamine is a dye that is released by 
cells as a function of protein expression. Transduced cells have been grown in selective 
culture, and colonies are screened for drug resistance. 
What is the evidence for in vivo gene expression? Dr. Nienhuis presented fluorescence 
activated cell sorter (FACS) analysis data using an antibody that is specific for an 
external epitope of the MDR protein. In the murine model, approximately 10 to 12% of 
the bone marrow cells were positive by FACS analysis. This antibody detection method 
would be used to monitor patients. Dr. Parkman asked about the level of expression 
following long-term in vitro culture. Dr. Nienhuis responded that the long-term culture 
experiments have not been performed. Dr. Parkman asked if MDR expression has been 
observed in human MDR-negative cells. Dr. Nienhuis stated that these experiments 
have not been performed. 
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Recombinant DNA Research, Volume 16 
