Recombinant DNA Advisory Committee - 5/30-31/91 
gene vector and testing for G418 resistance. He said the neo gene could be detected in 
leukemic cells in 10-20% of the exposed cells and that PCR analysis confirmed that the neo 
gene was carried in 30% of these cells. Therefore he said the overall absolute frequency of 
cells shown to be neo positive in the chronic phase marrow was 4-5%. He said they went on 
to look at blast crisis marrow samples since the protocol is aimed at these, and they were able, 
using the same procedures, to show the same level of acquisition of resistance to G418 in the 
blast crisis marrow. Furthermore, using a specific messenger RNA marker for leukemic cells 
they were able by PCR to show that 75-100 percent of the blast colonies expressed the 
retroviral trans genome. He said the program could be considered feasible and the 
investigators are in a position to expect the blasts, if they are contributing to relapse from the 
marrow, would be marked with the retroviral trans genome. 
In terms of quantitative data. Dr. Deisseroth said they start out with 2 X 10^® cells from a 
marrow collection. They then concentrate the marrow, resulting in a 5-fold reduction to 4 X 
10^. Then only 30 percent of this will be used in the marking process, so that reduces the 
number to 1.2 X 10^. Then the marrow is subjected to further concentration to remove 
myeloid and many of the leukemia cells resulting in a 10-fold reduction down to 1.2 X 10® 
cells. At the time of cytogenetic remission the level of leukemic cells would be below 1 in 
100, and thus the range of total leukemic blasts that would be stored away would be anywhere 
between 10,000 and 1 million. Therefore, Dr. Deisseroth said he felt there was ample 
evidence to suggest that not only can the cells be marked, but they would be able to be 
detected at the time of relapse. He said the only remaining question was whether they would 
see marked cells in the marrow or not. 
Dr. Deisseroth then turned his attention to what is to be gained from this protocol. He noted 
that the patients themselves will not stand to gain from this protocol in an immediate sense 
because they are participating in a therapeutic program to control the blast crisis from which 
they have a 100% chance of dying and to reduce the level of these leukemic cells within their 
body so as to produce a state in which they may survive for several years. He underlined that 
the marking process itself has been proven not to pose any immediate risk to the patient and 
he pointed to long-term benefits to society in terms of the increasing use of intensive therapy 
requiring exogenous hematopoietic reconstitution in more common diseases such as breast 
cancer. He noted that this will not only help identify origins of relapse, but be helpful in 
understanding and cleaning up the fractionation process and refining purification of 
hematopoietic stem cells which is a very important technical obstacle to the application of 
genetic replacement strategies for inherited diseases. 
Dr. Leventhal said she was still unclear as to whether normal blasts would be labeled as well 
as leukemic blasts. Also she said she felt that the cause of the relapse may come partially 
from the autologous marrow and partially from residual systemic disease and that there may 
be a different time course involved for these two actions. She said she would like to see a 
plan of when tests will be done on the patients and what tests will be performed after the 
marrow is reinfused. She added that she was concerned about certain definitions in the early 
stopping rule and requested she be provided with a revised protocol so she could be assured 
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