1 Dept. Oncology and Hematology, BMT-Unit,
2 Dept. of TransfusionMedicine.
3 Dept. Pediactric Hematology, University Hospital Eppendorf, Hamburg,
The hematopoietic growth factors G-CSF and GM-CSF are able to stimulate
granulopoiesis after autologous transplantation of bone marrow or
blood stem cells (PBSC) and can also mobilize peripheral blood progenitor
cells [1, 2, 3, 4]. Rapid recovery of platelets has been observed
following addition of G-CSF mobilized PBSC to autologous bone marrow
transplants . The kinetics of engraftment are mainly dependent
on the number and qualitiy of progenitor/stem cells in the graft
[5, 6, 7, 8]. We have studied the impact of higher doses of G-CSF
(Filgrastim) on the yield of CD34 positive cells, CFU-GM and mononucelar
cells in leukapheresis products in patients with Non Hodgkin Lymphomas,
Hodgkins disease, testicular cancer, AML and neuroblastoma as well
as in normal donors for allogeneic progenitor cell transplantation.
We report our experience with mobilization of GCSF in two different
doses: 10 µg/kg per day versus 24 (2 x 12) µg/kg per day for stem
cell collection, separation and transplantation.
Materials and methods
Mobilization: G-CSF (Filgrastim) was administered subcutaneously
for 5- 6 days at a daily dose of 10 µg/kg (group A) or 2x12 µg/kg
(group B) to 45 patients. Beyond this a total of 18 mobilizations
followed by 1 to 4 leukapheresis procedures, was carried out in
normal donors. 15 transplants, involving mobilized progenitor cells
either alone or in conjunction with bone marrow, were carried out.
The characteristics of the autologous patients are shown in table
Table 1. Patient characteristics
15 patients with the following diagnosis: AML: 4, ALL: 5, Multiple
Myeloma: 1, Severe Aplastic Anaemia: 3, CML blastic crisis: 1, Myeloproliferative
Syndromes 1 received an allogeneic stem cell transplant. The donor-recipient
relationships were the following: matched siblings: 8, one mismatch:
2, parent to child: 3, child to parent: 1, matched unrelated donor:
1. Conditioning regimens: without conditioning as a rescue graft
for rescue from insufficient bone marrow function: 5; TBICyclophosphamide-VP16:
5; TBI-Cyclophosphamide: 1, Busulfan, Cyclophosphamid +- Etoposid:
Colony forming assay: Colony forming units granulocyte-monocyte
(CFU-GM) were assayed in Iscove's methylcellulose. Colonies were
enumerated after 14 days of culture in 37° C, 5 % CO2 and 100 %
humidity by using an inverted phase microscope (9).
Flow cytometric analysis: Surface antigen expression was evaluated
using the monoclonal antibody HPCA-2, directly conjugated to FITC
(Becton-Dickinson). Analysis was performed on a FACScan (Becton-Dickinson)
using Lysis II research software [5,10,11].
Statistics: Data are shown as median (range), statistical significance
was determined by the Student's t-test.
Autologous: The percentage of CD34+ mononuclear cells (MNC) in
the first leukapheresis product increased significantly (p=0.024)
from 0.68 (0.03-1.96) (group A) to 1.23 (0.4-4.28) (group B) (Fig.
Fig.1 Leukapheresis products 10µg
vs. 2x12µg G-CSF
Statistically significant differences were also found with respect
to total CD34+ MNC per leukapheresis product (LP1): 10.08x10high
7 (0.4-44.0) (group A) versus 42.51 x 10 high 7 (3.46-86.3) (group
B) (p=0.012) (Fig. 2, Tabl. 2).
Fig. 2 Leukapheresis products 10µg vs.
Table 2. Graft Data
CFU-GM increased from 37.05 (1.6- 95.1) x 105 (group A) to 106.5
(8.7- 390.5) (group B) (p = 0.004). These results confirm, that
higher doses of G-CSF can significantly increase the number of CD34
positive cells and CFU-GM in leukapheresis products.
Allogeneic: An average of 3 leukapheresis (range 1 to 4) were carried
out, which yielded 13 x 10 high8 (range 2.9 -29) mononuclear cells/kg
BW of the recipient, 5 x 10 high 6 CD34 positive cells/kg BW of
recipient (range 1.2- 39), CFC 3 x 10 high 5 /kg BW (0,88 60,4).
The mobilization with G-CSF was generally well tolerated with the
exception of moderate bone pain in 2/3 of the donors.
Engraftment in 9 evaluable patients revealed neutrophil engraftment
of more than 0,5/nl by day 14 (range 9 -22), neutrophils more than
1.0/nl day 15 (range 10- 24), thrombocytes more than 20/nl day 16
(range 8 -32), more than 50/nl 24 (range 13 to more than 100 days).
GvH grade 0 -1: 3 patients, grade 2: 4 patients, grade 4: 2 patients
and moderate chronic GvH in 1 patient. Total of 12 of the 15 patients
are alive in continuous complete remission, 3 have died of GvH and
infection (aspergillus 1, candida crusei 1 ).
We could show that the mobilization with a higher than usual dosis
of G-CSF is well tolerated in normal donors as well as in patients
and yields grafts which allow engraftment after myeloablative therapy.
GvH occured in 2/3 of evaluable patients and led in conjunction
with infection to death in 3 patients. 1 patient received an allogeneic
graft from a matched unrelated donor, showed engraftment but unfortunately
died of a preexisting fungus disease.
Based on our experience in the autologous setting, where we found
statistically significant differences in the number of CD34+ cells
and CFU-GM in patients mobilized with 10 µg/kg G-CSF compared with
24 µg/kg and based on the observation that the side effects in both
dose levels are comparable, we would recommend 24 µg/kg BW G-CSF
per day, given in 2 divided doses or continuously intravenous infusion
for mobilization of normal donors.
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