Blood, Vol. 93 No. 8 (April 15), 1999:
pp. 2748-2749
CORRESPONDENCE
Chimerism Following Donor Lymphocyte Infusion for Chronic Myeloid
Leukemia
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LETTER |
To the Editor:
We read with interest the work reported recently by Baurmann et
al1 on the importance of defining the kinetics of the
graft-versus-leukemia (GVL) response after donor leukocyte infusions
(DLI) for relapsed chronic myeloid leukemia (CML) after allogeneic bone
marrow transplantation (BMT). Baurmann et al investigated the chimeric
profile in different hematopoietic lineages in 4 patients at various
timepoints who received DLI and/or stem cell infusions for relapsed
CML. Lineage-specific analysis was performed using chromosome-specific
fluorescent in situ hybridization (FISH) with simultaneous
immunophenotyping of interphase cells (FICTION) to define
chimeric status in the various lineages studied. Their results are
similar to previous work that we have reported assessing
lineage-specific chimerism by short tandem repeat polymerase
chain reaction (STR-PCR) of immunomagnetically separated fractions
after DLI alone for relapsed CML.2 At the time, we had
performed longitudinal chimerism studies on 4 patients who relapsed 1, 3, 6.5, and 7 years post-BMT and who were treated with DLI from the
original BMT donor. We observed that the conversion of the peripheral
blood (PB) T-cell subset from a mixed chimeric profile (~50%
recipient cells) to a predominantly donor profile (<5% recipient
cells) was the earliest indicator of a response to DLI. Molecular
monitoring defined subtle changes in the degree of mixed chimerism
before a response was seen in the other PB and bone marrow (BM)
fractions, ie, during the critical response time. The granulocytes were
the last lineage to convert to complete donor chimerism. We have now
performed longitudinal STR-PCR in 6 patients and have seen similar
response patterns.
The FICTION assay has the advantage of allowing dual staining of the
individual cells for CD and FISH markers; however, FISH analysis has
low sensitivity and the analysis of at least 200 cells is not always
possible in the post-DLI situation due to low cell numbers or
pancytopenia. Baurmann et al state that chimerism studies to date
have been hampered by the use of different tools to assess chimerism
post-BMT. Although we agree with this assertion in general, we
feel that our experience with the use of STR-PCR (which we first
reported in BLOOD in 19913) in more than 300 patients and
the use of lineage-specific STR-PCR allow us to provide reliable
results on chimeric profiles post-BMT. The separation of BM and PB into
lineage-specific fractions using immunomagnetic bead separation with a
high degree of purity allows analysis of a larger number of cells by
our radioactive STR-PCR (which has a sensitivity of detection of 1 in
105 of the minor cell population), therefore enabling
subtle changes in lymphopoietic chimerism to be detected. As indicated
above, the subsequent conversion of other lineages to donor chimerism can be reliably detected using this technique, and the observation of
the slower conversion of the granulocyte fraction has been reported by
ourselves and in other studies.4
In one of our patients, the CD34+ fraction was
predominantly donor (>90% donor) at the time of relapse when 40%
donor cells were detected in the total BM fraction, a finding similar
to that observed in patient no. 3 described by Baurmann et al. The
CD34+ fraction became predominantly recipient, during which
time the patient failed to show a clinical response to an initial DLI
dose of 0.9 × 107 CD3 cells/kg and disease progression
was seen. After a second DLI dose (1 × 108 CD3
cells/kg), the patient showed conversion of the CD34 fraction to a
fully donor profile. It would appear that the degree of mixed chimerism
and the kinetics of the GVL response are certainly different in those
patients with molecular relapse compared with patients with full
hematological relapse. The monitoring of the degree of chimerism
at the time of relapse is vital to predict the possibility of aplasia
after DLI; however, more detailed lineage-specific chimeric analysis
should give more insight into the possible outcome after DLI and
explain the failure of response in other patients. A larger scale
study with more patients receiving a single treatment regimen and
detailed monitoring of specific lineages is required to further
elucidate the kinetics of the GVL response after DLI and is
currently in progress with the help of the Chronic Leukaemia working
party of the European Blood and Marrow Transplant (EBMT) group.
Nicola Gardiner
Shaun R. McCann
Joan O'Riordan
Mark Lawler
Department of Haematology
St James
Hospital
Dublin, Ireland
| |
REFERENCES |
1.
Baurmann H, Nagel S, Binder T, Neubauer A, Siegert W, Huhn D:
Kinetics of the graft versus leukemia response after donor leukocyte infusions for relapsed chronic myeloid leukemia after bone marrow transplantation.
Blood
92:10, 1998
2.
Gardiner N, Lawler M, O'Riordan JM, DeArce M, McCann SR:
Monitoring of lineage specific chimaerism allows early prediction of response following donor lymphocyte infusions for relapsed chronic myeloid leukaemia.
Bone Marrow Transplantation
21:711, 1998[Medline]
[Order article via Infotrieve]
3.
Lawler M, Humphries P, McCann SR:
Evaluation of mixed chimerism by in vitro amplification of dinucleotide repeat sequences using the polymerase chain reaction.
Blood
77:2504, 1991[Abstract/Free Full Text]
4.
Garichochea B, Van Rhee F, Spencer A, Chase A, Lin F, Cross NCP, Goldman JM:
Aplasia after donor lymphocyte infusion (DLI) for CML in relapse after sex-mismatched BMT: Recovery of donor type haemopoiesis by non-isotopic in situ hybridization (ISH).
Br J Haematol
88:400, 1994[Medline]
[Order article via Infotrieve]
Response
In the April 1998 issue of Bone Marrow
Transplantation, Gardiner et al1 published their
results on monitoring lineage-specific chimerism in 4 cases of relapsed
CML after DLI. Because their article appeared after our work was
submitted for publication to BLOOD,2 both
groups reached their conclusions independently, and we are
pleased to state that the data on the kinetics of the GVL response are
quite similar. Our statement about the existence of a critical
switch period during which hematopoiesis suddenly reverts to complete
donor type is especially supported by the analysis of their
cases. Interestingly, Gardiner et al report a sequential switch
of T cells and granulocytes in 3 of their 4 cases, a feature not
described in our report. However, in this context, it is important to
note that, in contrast to all 8 cases of our series, the initial BMT
was T-cell depleted in their 3 patients. T-cell chimerism before DLI,
therefore, was mixed and not already predominantly of donor type, as in
our cases.
Gardiner et al state that their technique using lineage-specific
STR-PCR provides reliable results on the chimeric profile post-BMT and
may be more useful than the FICTION assay in situations with low
cell numbers. Although the important aspect of morphology and
individual cell by cell analysis is lost with the PCR approach, we
agree that STR-PCR is a most valuable tool in this context and
therefore complementary to our FICTION analysis. However, we doubt that
subtle changes in the degree of mixed chimerism can be seen with
a semiquantitative technique according to which mixed chimerism is
defined as a percentage of recipient cells between less than 90%
and greater than 10%. Indeed, in the work of Gardiner et al,
significant changes in every case were detected only concomitantly to
the onset of clinical graft-versus-host disease (GVHD), a result
similar to that obtained in our cases when using the AmpliType
Polymarker PCR kit to document non-lineage-specific DNA chimerism. In
contrast, when using our quantitative techniques, ie, FICTION and
competitive differential bcr-abl RT-PCR as an additional
disease-specific marker, the beginning of the critical switch period
could be detected several weeks before the onset of clinical GVHD.
Although we therefore very much support a larger scale study to better
understand the kinetics of the GVL response after DLI in
T-cell-depleted and non
T-cell-depleted BMT recipients, we strongly
suggest the use of true quantitative measures of lineage-specific chimerism. The FICTION method is especially helpful in small centers, because it can be performed without larger equipment or in situations in which morphological control of the selected population is of additional value. Another attractive tool is the combination of FACS
sorting and quantitative multiplex STR-PCR with fluorescent primers.3 This technique has been successfully applied by
members of our group to study subtle changes of subset chimerism
in patients undergoing nonmyeloablative stem cell
transplantation and was shown to be predictive of response, GVHD, and
disease recurrence.4
Herrad Baurmann
BMT
Center
Deutsche Klinik für Diagnostik
Wiesbaden,
Germany
Christian Thiede
Stefan Nagel
Thomas Binder
Andreas Neubauer
Wolfgang Siegert
Dieter Huhn
| |
REFERENCES |
1.
Gardiner N, Lawler M, O'Riordan JM, DeArce M, McCann SR:
Monitoring of lineage specific chimaerism allows early prediction of response following donor lymphocyte infusions for relapsed chronic myeloid leukemia.
Bone Marrow Transplant
21:711, 1998
2.
Baurmann H, Nagel S, Binder T, Neubauer A, Siegert W, Huhn D:
Kinetics of the graft versus leukemia response after donor leukocyte infusions for relapsed chronic myeloid leukemia after allogeneic bone marrow transplantation.
Blood
92:3582, 1998[Abstract/Free Full Text]
3. Thiede C, Florek M, Bornhäuser M, Ritter M, Mohr B, Brendel
C, Ehninger G, Neubauer A: Rapid quantification of mixed chimerism
using multiplex amplification of short tandem repeat markers and
fluorescence detection. Bone Marrow Transplant (in press)
4.
Thiede C, Brendel C, Mohr B, Florek M, Oelschlagel U, Ritter M, Naumann R, Geissler G, Ehninger G, Neubauer A, Bornhäuser M:
Comparative analysis of chimerism in the early post-transplantation period in cellular subsets of patients undergoing myeloablative and non-myeloablative allogeneic blood stem cell transplantation.
Blood
92:132a, 1998 (abstr, suppl 1)
