Abstract
Ex vivo depletion of alloreactive CD25+ T cells from a stem cell transplant (SCT) can reduce the incidence of graft-versus-host disease (GVHD) while preserving antimicrobial and perhaps antileukemia activity. However, the most effective methods for allodepleting T cells prior to transplant have not been determined. In this study, we have compared three agents that deplete CD25+ activated, alloreactive T cells. These included Ontak (Denileukin Diftitox), an IL-2 fusion toxin, anti-CD25 microbeads (MACS), an anti-CD25 immunotoxin (IT) and a combination of the IT and MACS. Peripheral blood mononuclear cells (PBMCs) activated in a primary mixed lymphocyte reaction (MLR) were allodepleted using optimal amounts of each agent, and the cells were then analyzed by flow cytometry. The treated cells were examined both for remaining alloreactivity and for the preservation of third party reactivity by testing them in a secondary MLR. Our data demonstrate that both the anti-CD25 IT and the anti-CD25 MACS were equally effective in depleting CD4+CD25+ cells and in sparing T cells that were reactive with third party cells. The anti-CD25 IT was, however, superior in depleting alloreactive CD8+CD25+ cells. In contrast, Ontak did not eliminate alloreactive cells and the Ontak-treated cells retained significant reactivity against the original stimulator cells.
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References
Ferrara JLM, Deeg HJ . Graft-versus-host disease. N Engl J Med 1991; 324: 667–674.
Murphy WJ, Blazar B . New strategies for preventing graft-versus-host disease. Curr Opin Immunol 1999; 11: 509–515.
Przepiorka D, Ippoliti C, Koberda J, Chan KW, Khouri IF, Fisher HE et al. Interleukin-2 for prevention of graft-versus-host disease after haploidentical marrow transplantation. Transplantation 1994; 58: 858–860.
Ferrara JL, Cooke KR, Teshima T . The pathophysiology of acute graft-versus-host disease. Int J Hematol 2003; 78: 181–187.
Ho VT, Soiffer RJ . The history and future of T cell depletion as graft-versus-host disease prophylaxis for allogeneic hematopoietic stem cell transplantation. Blood 2001; 98: 3192–3204.
O'Reilly RJ, Collins N, Dinsmore R, Kernan N, Siena S, Brochstein J et al. Transplantation of HLA-mismatched marrow depleted of T cells by lectin agglutination and E-rosette depletion. Tokai J Exp Clin Med 1985; 10: 99–107.
Michalek J, Collins RH, Vitetta ES . Clinical-scale selective depletion of alloreactive T cells using an anti-CD25 immunotoxin. Neoplasia 2003; 50: 296–299.
Solomon SR, Tran T, Carter CS, Donelly S, Hensel N, Schindler J et al. Optimized clinical-scale culture conditions for ex vivo selective depletion of host-reactive donor lymphocytes: a strategy for GVHD prophylaxis in allogeneic peripheral blood stem cell transplantation. Cytotherapy 2002; 4: 395–406.
Solomon SR, Mielke S, Savani BN, Montero A, Wisch L, Childs R et al. Selective depletion of alloreactive donor lymphocytes – a novel method to reduce the severity of graft-versus-host disease in older patients undergoing matched sibling donor stem cell transplantation. Blood 2005; 106: 1123–1129.
Andre-Schmutz I, Le Deist F, Hacein-Bey-Abina S, Vitetta E, Schindler J, Chedeville G et al. Immune reconstitution without graft-versus-host disease after haemopoietic stem-cell transplantation: a phase 1/2 study. Lancet 2002; 360: 130–137.
Amrolia P, Mucioli-Casadei G, Yvon E, Huls H, Sili U, Wieder D et al. Selective depletion of alloreactive T cells without loss of anti-viral or anti-leukemic responses. Blood 2004; 104: 1605 (Erratum).
Amrolia PJ, Mucioli-Casadei G, Huls H, Gotsolva T, Durett A, Weiss H et al. Adaptive immunotherapy with allodepleted donor T cells improves immune reconstruction after haploidentical stem cell transplant. 2005, submitted.
Eklund JW, Kuzel TM . Denileukin diftitox: a concise clinical review. Expert Rev Anticancer Ther 2005; 5: 33–38.
Jansen J, Hanks S, Akard LP, Thompson JM, Burns S, Chang Q et al. Immunomagnetic CD4+ and CD8+ cell depletion for patients at high risk for severe acute GVHD. Bone Marrow Transplant 1996; 17: 377–382.
Herrera CS, Torres A, Garcia-Castellano JM, Roman J, Martin C, Serrano J et al. Prevention of graft-versus-host disease in high risk patients by depletion of CD4+ and reduction of CD8+ lymphocytes in the marrow graft. Bone Marrow Transplant 1999; 5: 443–450.
Garderet L, Snell V, Przepiorka D, Schenk T, Lu JG, Marini F et al. Effective depletion of alloreactive lymphocytes from peripheral blood mononuclear cell preparations. Transplantation 1999; 61: 124–130.
Fehse B, Goldmann M, Frerk O, Bulduk M, Zander AR . Depletion of alloreactive donor T cells using immunomagnetic cell selection. Bone Marrow Transplant Suppl 2000; 2: S39–S42.
Williams DP, Parker K, Bacha P, Bishai W, Borowski M, Genbauffe F et al. Diphtheria toxin receptor binding domain substitution with interleukin-2: genetic construction and properties of a diphtheria toxin-related interleukin-2 fusion protein. Protein Eng 1987; 1: 493–498.
Re GG, Waters C, Poisson L, Willingham MC, Sugamura K, Frankel AE . Interleukin 2 (IL-2) receptor expression and sensitivity to diptheria fusion toxin DAB389IL-2 in cultured hematopoietic cells. Cancer Res 1996; 56: 2590–2595.
Ghetie V, Thorpe P, Ghetie MA, Knowles P, Uhr JW, Vitetta ES . The GLP large scale preparation of immunotoxins containing deglycosylated ricin A chain and a hindered disulfide bond. J Immunol Methods 1991; 142: 223–230.
Jonuleit H, Schmitt E, Stassen M, Tuettenberg A, Knop J, Enk AH . Identification and functional characterization of human CD4+ CD25+ T cells with regulatory properties isolated from peripheral blood. J Exp Med 2001; 193: 1285–1294.
Bousvaros A, Stevens AC, Strom TB, Murphy J, Lamont JT . Interleukin-2 fusion protein (DAB389IL-2) selectively targets activated human peripheral blood and lamina propria lymphocytes. Dig Dis Sci 1997; 42: 1542–1548.
Shao RH, Tian X, Gorgun G, Urbano AG, Foss FM . Arginine butyrate increases the cytotoxicity of DAB(389)IL-2 in leukemia and lymphoma cells by upregulation of IL-2Rbeta gene. Leuk Res 2002; 26: 1077–1083.
Michalek J, Collins RH, Durrani HP, Vaclakova P, Ruff LE, Douek DC et al. Definitive separation of graft-versus-leukemia-and graft-versus-host-specific CD4+ T cells by virtue of their receptor β loci sequences. Proc Natl Acad Sci USA 2003; 100: 1180–1184.
Barrett AJ, Malkovaska V . Graft-versus-leukaemia: understanding and using the alloimmune response to treat haematological malignancies. Br J Haematol 1996; 93: 754–761.
Zeiser R, Marks R, Bertz H, Finke J . Immunopathogenesis of acute graft-versus-host disease: implications for novel preventive and therapeutic strategies. Ann Hematol 2004; 83: 551–565.
Barrett AJ, Mavroudis D, Tisdale J, Molldrem J, Clave E, Dunbar C et al. T cell-depleted bone marrow transplantation and delayed T cell add-back to control acute GVHD and conserve a graft-versus-leukemia effect. Bone Marrow Transplant 1998; 21: 543–551.
Cavazzana-Calvo M . Specific elimination of alloreactive T cells by an anti-interleukin-2 receptor B chain-specific immunotoxin. Transplantation 1990; 50: 1–7.
Ho VT, Zahrieh D, Hochberg E, Micale E, Levin J, Reynolds C et al. Safety and efficacy of denileukin diftitox in patients with steroid-refractory acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation. Blood 2004; 104: 1224–1226.
Shaughnessy PJ, Bachier C, Grimley M, Freytes CO, Callander NS, Essell JH et al. Denileukin diftitox for treatment of steroid-resistant acute graft-versus-host disease. Biol Blood Marrow Transplant 2005; 11: 188–193.
Oluwole SF, Oluwole OO, DePaz HA, Adeyeri AO, Witkowski P, Hardy MA . CD4+CD25+ regulatory T cells mediate acquired transplant tolerance. Transpl Immunol 2003; 11: 287–293.
BaecherAllen C, Brown JA, Freeman GJ, Hafler DA . CD4+CD25high regulatory cells in human peripheral blood. J Immunol 2001; 167: 1245–1253.
Sakaguchi S . Naturally arising CD4+ regulatory T cells for immunologic self-tolerance and negative control of immune responses. Annu Rev Immunol 2004; 22: 531–562.
Koenen HJ, Fasse E, Joosten I . CD27/CFSE-based ex vivo selection of highly suppressive alloantigen-specific human regulatory T cells. J Immunol 2005; 174: 7573–7583.
Mielke S, Rezvani K, Soloman SR, Kilical Y, Savani BN, Wisch L et al. Reconstitution of regulatory T cells after selective depletion of CD25+ host-reactive donor lymphocytes from allografts and association with acute graft-versus-host-disease (ASH abstract), 2005, in press.
Acknowledgements
We thank Ms Erica Garza and Ms Linda Berry for assistance in preparing the manuscript, Dr Victor Ghetie for preparing the immunotoxin, Ms Lien Le for providing us with the immunotoxin and antibodies, Drs Laurentiu Pop and Xiaoyun Liu for their technical advice; and Drs Nitin Karandikar, Radu Marches and Mr Paul Denton for reviewing of the manuscript.
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This work was supported by a grant from Lymphoma and Leukemia Society and by the Cancer Immunobiology Center.
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Vaclavkova, P., Cao, Y., Wu, L. et al. A comparison of an anti-CD25 immunotoxin, Ontak and anti-CD25 microbeads for their ability to deplete alloreactive T cells in vitro. Bone Marrow Transplant 37, 559–567 (2006). https://doi.org/10.1038/sj.bmt.1705286
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DOI: https://doi.org/10.1038/sj.bmt.1705286