Blood, 1 May 2001, Vol. 97, No. 9, pp. 2915-2917
CORRESPONDENCE
To the editor:
Transient mixed hematopoietic chimerism in dogs given
thymic irradiation before and pharmacologic immunosuppression after
marrow transplantation
We used postgrafting immunosuppression with mycophenolate
mofetil (MMF) and cyclosporine (CSP) to control both graft-versus-host and host-versus-graft reactions in a canine model of dog leukocyte antigen (DLA)-identical littermate marrow transplants. This way, the
single pretransplantation dose of total body irradiation (TBI) otherwise needed for sustained allografts could be lowered from 920 cGy
to the sublethal level of 200 cGy.1 We next substituted 450 cGy irradiation targeted to the cervical, thoracic, and upper abdominal lymph node chain for 200 cGy TBI in this model.2 Prompt and sustained engraftment was seen in unirradiated marrow and
lymph nodes as early as week 6. This indicated that creation of marrow
space by cytotoxic agents was not required for homing of transplanted
stem cells. Rather, some degree of host immunosuppression was
sufficient to set the stage for successful allografts. The concept was
validated in a 30-year-old patient with common variable immunodeficiency disease who received no conditioning therapy and in
whom durable allogeneic marrow engraftment was achieved solely with
postgrafting MMF/CSP.3
The field of lymph node chain irradiation in these studies included the
thymus. Reports by others on nonmyeloablative regimens for major
histocompatibility complex (MHC)-matched and mismatched murine and
porcine transplantations have emphasized the pivotal importance of
thymus irradiation in the success of grafts.4,5 It is not
clear whether the engraftment of the transplanted cells was facilitated
through creation of thymic space or elimination of thymic
alloreactivity by thymic irradiation. Here, we evaluated in the canine
model whether the success of central lymph node chain radiation could
largely be attributed to inclusion of the thymus in the radiation
field. We asked whether sustained grafts of DLA-identical littermate
marrow could be achieved using 450 cGy thymic irradiation before and
MMF/CSP after transplantation.
Litters of beagles weighing from 7.5 to 11.4 kg (median, 9.0 kg) and 6 to 9 months old (median, 7 months) were used. Research was conducted
per the principles outlined in the Guide for Laboratory Animal
Facilities and Care (National Academy of Sciences, National Research Council). The Institutional Animal Care and Use
Committee of the Fred Hutchinson Cancer Research Center approved
the research protocol. Kennels are certified with the American
Association for Accreditation of Laboratory Animal Care. A high-energy
linear accelerator (Varian CLINAC 6, Palo Alto, CA) delivered
450 cGy thymic irradiation at 200 cGy/min in a single setting with a
tightly collimated 6 million electron volt beam using 2 isocentric
parallel-opposed anterior and posterior ports, 2.5 cm wide and 3.5 cm
long, to include the thymus with margins. A radiograph verified the
portal placement. Selection of DLA-matched pairs, transplants of 1.5 to
6.2 × 108 (median, 2.7 × 108) nucleated
marrow cells/kg (day 0), posttransplantation care, administration and
dose schedules of MMF and CSP, assessment of engraftment, and
histopathologic examinations were done as previously described.1,2
All 5 recipients showed initial evidence of mixed
donor-host hematopoietic chimerism, with the donor contributions
ranging from 1% to 5% (Table 1). Four
dogs lost their grafts between 9 and 14 weeks after
transplantation, and they survived with complete autologous
recovery. One dog died on day 23 with systemic canine herpes virus
infection, at a time when donor cell contribution in the marrow was
1%. We speculate that this dog would have rejected his graft over the
next 6 to 10 weeks, similarly to the other 4 dogs in the study. The
dogs' median platelet and granulocyte counts showed minimal transient
changes after transplantation, whereas median lymphocyte counts
transiently declined from 3500/µL to almost 600/µL between weeks 2 and 4, presumably due to the combined effects of thymic radiation and
postgrafting MMF (Figure 1). Analysis of
peripheral blood samples (dog E751) by microsatellite markers provides
an example of the transient nature of donor engraftment.
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Table 1.
Marrow grafts from DLA-identical littermates after
conditioning with 450 cGy thymus irradiation and with postgrafting
MMF/CSP
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| Figure 1.
Effects of thymic radiation and postgrafting
MMF/CSP.
(A) Platelet, granulocyte, and lymphocyte changes in 5 dogs given 450 cGy thymus irradiation, DLA-identical littermate marrow grafts, and
MMF/CSP after transplantation. (B) Results of microsatellite
marker studies of donor and recipient (dog E751) cells before
transplantation (lanes 1 and 2) and recipient cells after marrow
transplantation (lanes 3-10). Gran indicates granulocytes; MNC,
mononuclear cells.
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We conclude that thymic irradiation in itself provided
insufficient host immunosuppression for durable donor-cell engraftment. The duration of engraftment was comparable to that seen in dogs conditioned with 100 cGy of TBI.1 In comparison,
mixed hematopoietic chimerism lasting at most 3 weeks was seen
in dogs not given pretransplantation irradiation and only treated with
postgrafting MMF/CSP.1 The current data do not support a
pivotal role of thymus irradiation for durable allografts in this
MHC-matched model. Rather, it seems that irradiation of a larger
proportion of recipient lymphoid tissues is required, such as is
afforded by the previously described irradiation of the lymph node
chain.2
Rainer Storb and George Sale
Fred Hutchinson Cancer Research Center and University
of Washington Seattle, WA
Todd Barnett
Swedish Medical Center Tumor Institute Seattle, WA
Cong Yu, Eustacia Zellmer, and Marie-Térèse Little
Fred Hutchinson Cancer Research Center Seattle, WA
Acknowledgments
Supported in part by National Institutes of Health grants
CA78902, CA15704, DK42716, and HL36444. R.S. also received support from
the Laura Landro Salomon Endowment Fund and a prize from the Josef
Steiner Krebsstiftung, Bern, Switzerland; M.T.L. received additional
support from a Lady Tata Memorial Trust International research grant,
London, United Kingdom.
References
1.
Storb R, Yu C, Wagner JL, et al.
Stable mixed hematopoietic chimerism in DLA-identical littermate dogs given sublethal total body irradiation before and pharmacological immunosuppression after marrow transplantation.
Blood.
1997;89:3048-3054[Abstract/Free Full Text].
2.
Storb R, Yu C, Barnett T, et al.
Stable mixed hematopoietic chimerism in dog leukocyte antigen-identical littermate dogs given lymph node irradiation before and pharmacologic immunosuppression after marrow transplantation.
Blood.
1999;94:1131-1136[Abstract/Free Full Text].
3.
Woolfrey AE, Nash RA, Frangoul HA, et al.
Nonmyeloablative transplant regimen used for induction of multilineage allogeneic hematopoietic mixed donor-host chimerism in patients with T-cell immunodeficiency [abstract].
Blood.
1998;92(suppl 1):520a.
4.
Fuchimoto Y, Huang CA, Yamada K, et al.
Mixed chimerism and tolerance without whole body irradiation in a large animal model.
J Clin Invest.
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[Order article via Infotrieve].
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Sharabi Y, Sachs DH.
Mixed chimerism and permanent specific transplantation tolerance induced by a nonlethal preparative regimen.
J Exp Med.
1989;169:493-502[Abstract/Free Full Text].
