autologous blood and marrow transplantation - Blog Science ...

516 Chapter 10: Graft Manipulation
performed on circulating peripheral blood mononuclear cells. The level of
detection was highest in marrow clonogenic assays, where it ranged from 1 to 15%.
Of interest, the level of transfer in clonogenic assays was around 1 log higher than
in peripheral blood, an observation also found in other gene transfer studies. 14
In
peripheral blood cells, expression was higher in granulocytes than in mononuclear
cells and higher in T lymphocyte lines than in B lymphocyte lines. These
observations show that the infused product contributes to both short- and long-term
hematopoietic and immune reconstitution after autologous transplantation.
Second-generation marking studies
Fifteen patients were treated in the second-generation study protocol between
October 1993 and October 1996. In five patients, aliquots of marrow were purged
with both 4HC and IL-2, and in three patients, aliquots were purged with 4HC and
CD15. Seven patients received marrow purged with 4HC alone, due to either
insufficient number of cells harvested or unavailability of one clinical grade
retroviral vector. Follow-up currently ranges from 25 to 58 months with a median
of 48 months. Two patients relapsed early at 2 and 3 months and were noninformative,
as marked malignant cells were not detected. A third patient relapsed at 20
months, and his malignant blasts were also negative.
The neo gene has been detected in normal hematopoietic and immune system
cells at a lower level than in the studies using unpurged marrow. PCR studies on
peripheral blood granulocytes and mononuclear cells have consistently shown a
stronger signal from the 4HC-purged fraction than that from the IL-2-purged
fraction, regardless of which vector was used for marking each aliquot. These
observations suggest that the 4HC-purged fraction is making a greater contribution
to hematopoietic reconstitution than the IL-2-purged fraction. We therefore discontinued
purging with IL-2 and substituted CD 15 antibodies.
Safety of gene marking
Patient follow-up is now >1100 patient months in this group, and no adverse
effects attributable to gene marking have occurred. Patient samples have been
analyzed for replication-competent retrovirus (RCR) at intervals requested by the
FDA, and all results have been negative. We have compared survival curves of
patients with AML in first remission enrolled in marking studies with patients
treated with the same chemotherapy and autoBMT protocol who received
unmarked marrow due to unavailability of clinical grade supernatant or refusal of
marking component. The survival curves shown in Fig. 1 show no significant
difference in leukemia-free survival between recipients of marked marrow (59% at
3 years) or unmarked marrow (39% at 3 years). The overall survival is 70% at 3