Schema for Treatment Plan: 
I. Conventional dose chemotherapy. 
II. Collection of marrow to achieve a dose of 10,000 CFUGM/kg or a 
total nucleated count of 200 million nucleated cells/kg. The 
marrow will be concentrated using the 2991 COBE cell separator, 
and then separated using the CellPro Ceprate Concentrator to 
1 enrich the preparation of stem cells in CD34 positive cells. 
III. Exposure of the engrafting dose of hematopoietic cells to a 
safety-modified retrovirus which contains a MDR-1 cDNA will be 
undertaken following the protocol outlined in Appendix D. PCR 
assay for the MDR cDNA will be performed following the 
transduction using the assay outlined in Appendix*' D. The PCR 
assay will be used to identify if the transduction was 
successful. A transduction frequency of 1% will be considered 
successful. Functional assays (rhodamine dye exclusion and 
resistance to Taxol in vitro, will also be used to monitor the 
success of the transduction. The marrow will be frozen away 
using programmed freezing. The viability of the frozen marrow 
will be tested using the CFUGM assay. 
IV. Preparative therapy will be delivered and will involve 
cyclophosphamide and thiotepa, in doses outlined earlier in this 
, protocol. 
V. Infusion of autologous marrow and blood with PCR for MDR-1 of the 
thawed specimen before infusion. 
VI. PCR for MDR-1 and functional assays for MDR-1 on the 
hematopoietic cells following a recovery of an ANC of 2000/mm 3 is 
achieved. 
VII. Delivery of relatively intensive but not ablative levels of 
therapy, at a starting dose of 13 5 mg/m 2 every three to four 
weeks, and the dose will be increased by 50 mg/m 2 increments each 
course as is allowed by the rate of hematopoietic recovery. PCR 
assays will be conducted to assess the percentage of 
hematopoietic cells which are positive for the MDR-1 cDNA after 
each course of chemotherapy. 
VIII PCR for MDR-1 on colonies of peripheral blood and marrow 
hematopoietic cells every three to four weeks following each 
course of therapy, as long as that therapy is administered, or 
until relapse, at which time the therapy of Taxol maintenance 
will no longer be administered. In situ functional analysis of 
the number of cells in peripheral blood and marrow by dye 
exclusion will be performed at the same time as PCR of MDR-1 to 
enumerate the number of cells with increased MDR-1 in the marrow 
and peripheral blood. After discontinuation of Taxol therapy, 
the PCR assays will be conducted every three months for two 
years, and every six months until a 5 year period has elapsed 
following the initial transplantation of modified marrow. 
Recombinant DNA Research, Volume 17 
[671] 
