AbstractAllogeneic hematopoietic cell transplantation provides effective control of hematopoietic malignancies, but with an associated risk of graft-versus-host disease (GVHD) related morbidity and mortality. Several advances in hematopoietic cell transplantation including high resolution HLA typing, development of reduced intensity conditioning regimens, infectious prophylaxis and treatment, and novel immunosuppressive agents have resulted in improved outcomes and improved access to transplantation, but GVHD remains a major obstacle. This clinico-pathological syndrome, mediated by donor alloreactive T cells, occurs often despite prophylactic immunosuppressive therapy. Regulatory T cells, a suppressive subset of the T-cell repertoire, may offer promise as a novel cellular therapy for more effective prevention of GVHD. While advances have been made in pre-clinical experimental animals, several challenges remain in the translation of this work to human trials. Strategies to effectively produce ex vivo expanded alloantigen-specific regulatory T cells specific for ubiquitous alloantigens but sparing hematopoietic- or tumor-associated antigens hold promise to prevent GVHD while allowing a preserved graft versus malignancy effect.
Graft versus host disease
Allogeneic hematopoietic cell transplantation offers effective control and potential cure of hematopoietic malignancies. This benefit is realized in part by the reduction in tumor burden by cytotoxic therapy, but is also due to the ongoing immune surveillance termed graft versus malignancy effect. This is thought to be mediated by donor T-cell recognition of disparate major and minor histocompatibility antigens, as well as tumor-associated antigens. This disparity, however, is also responsible for the development of acute graft versus host disease (GVHD). This process is initiated when high-dose chemo- and/or radio-therapy disrupts tissues leading to activation of dendritic cells, which present antigen to alloreactive T cells. Inflammatory cytokines and cytotoxic effector T cells mediate the tissue injury that manifests as the clinical syndrome of acute GVHD, primarily affecting the skin, liver, and gastrointestinal tract.1–7
There has been a considerable amount of work performed to define the optimal GVHD prevention strategy. Tacrolimus (TAC) and methotrexate (MTX) used in combination constitute the current standard of care for GVHD prevention after allogeneic HCT. Two large randomized trials have shown that TAC/MTX is superior to cyclosporine (CSA)/MTX in the prevention of acute GVHD. Grade II–IV acute GVHD was significantly lower with TAC/MTX compared to CSA/MTX in both sibling donor (32% vs. 44%; P=0.01), and unrelated donor (56% vs. 74%; P=0.0002) transplant trials.8, 9
Despite these preventive measures, grade II–IV aGVHD remains a significant obstacle to successful transplantation. Importantly, complete response to front-line therapy with 1–2 mg/kg of glucocorticoids is achieved in only 30–40% of patients. Additionally, acute GVHD responsive to the frontline therapy of high-dose corticosteroids portends 50–60% survival, while those with steroid-refractory disease have a reported long-term survival of only 5–30%. Finally, up to 70% of recipients of allogeneic hematopoietic stem cell transplant will develop chronic GVHD.10–12 Acute and later chronic GVHD, as well as its associated immunosuppressive treatment and infectious complications therein, constitute a major source of transplant related morbidity and mortality.
Characterization of regulatory T cells
Recent insights into the biology of regulatory T cells (Tregs) have resulted in a surge of interest in their role in health and disease, most notably in the fields of autoimmunity and control of alloresponse after hematopoietic cell transplantation.13–15 A naturally occurring population comprising less than 5–10% of the human T-cell repertoire, Tregs are characterized by their constitutive expression of CD4 and CD25, the IL-2Rα chain receptor. They also express high levels of a nuclear transcription factor, FOXP3, which is critical for their development and suppressive function. Regulatory T cells express little or no CD127, the IL-7Rα chain receptor. Accordingly, the constitutive expression of CD25 and low expression of CD127 has been proposed as a means of identifying and purifying regulatory T cells. However, it is clear that there is heterogeneity among Treg populations. As reviewed by Feuerer et al., there appear to be several distinct sub-phenotypes among regulatory T cells which differ with respect to activation, tissue localization, transcriptional program, and function.16 Sakaguchi et al. have demonstrated several distinct subpopulations among FoxP3(+)CD4(+) T cells, including suppressive CD45RA(+)FoxP3(lo) resting Treg cells (rTreg cells) and CD45RA(−)FoxP3(hi) activated Treg cells (aTreg cells) and non-suppressive CD45RA(−)FoxP3(lo) T cells.17 Human data suggest that the tumor-necrosis factor receptor family member CD27, that is expressed on memory T cells, and high expression of the adhesion molecule CD44 distinguish highly suppressive T cells.18,19 Regulatory T cells require engagement of their T-cell receptor and co-stimulatory molecules for activation. While their suppressive function is lost upon stimulation and proliferation, this appears to be enhanced upon removal from these stimulatory signals; ex vivo expanded CD4CD25 cells have been shown to effectively suppress otherwise lethal GVHD in murine models.13,14,20,21
Regulatory T cells mediate suppression of immune responses, as supported by multiple lines of evidence. Early work illustrated the development of a severe autoimmune syndrome leading to multi-organ dysfunction in mice lacking regulatory T cells. Regulatory T cells can suppress alloreactive effector T cells in vitro in MLR assay. Additionally, regulatory T cells can abrogate GVHD in murine models of MHC mismatched allogeneic stem cell transplantation. Also, regulatory T cells protect from autoimmune-mediated development of diabetes in the NOD murine model. While the exact means of suppression in vivo is not known, there is an increasing understanding of potential mechanisms including suppression by cell to cell contact with antigen-presenting cells and effector T cells by way of the CTLA-4/CD80/CD86 signaling pathway. 14, 22–26
Plasticity in CD4 T-cell differentiation
As recently reviewed by Zhou et al., emerging evidence challenges a previously accepted paradigm of CD4 T-cell differentiation as a single, terminal event.27 Rather, it appears that naïve CD4 T cells differentiate under the influence of antigen presentation and specific cytokines to Th1, Th2, Th17, or Treg phenotypes. However, these CD4 subtypes are not fixed, but rather conversion is possible amongst these phenotypes. In particular, Tregs can, under the influence of IL-6 and IL-21, become IL-17 producing Th17 cells, and can conversely convert to Tfh cells under the influence of B-cell signaling and CD40-CD40L interaction. This process of conversion may be directed by epigenetic modification of genes coding for key cytokines and transcription factors. In total, this evidence highlights novel aspects of CD4 T-cell biology, but also raises concerns about the application of Tregs in human studies. In particular, the infusion of regulatory T cells in human allogeneic hematopoietic cell transplantation to prevent aGVHD could be compromised by conversion of Tregs to Th17 cells, which have been demonstrated to have an important role in autoimmunity,28,29 as well as contribute to the pathogenesis of aGVHD.30,31
Expansion of polyclonal regulatory T cells
Isolation of regulatory T cells by antibody-based affinity column selection has resulted in high purity separation, providing a substrate for ex vivo expansion. Given their low precursor frequency, several groups have employed diverse methods to expand polyclonal regulatory T cells ex vivo, including the following: Hoffman et al. produced large scale ex vivo expansion of human polyclonal regulatory T cells using IL-2, and anti-CD3/CD28 with a net increase of over 40,000-fold over four weeks.32 Cohen et al. generated a 20-fold expansion of regulatory T cells after 15 days of culture using splenocytes and IL-2.33 Godfrey et al. produced a 100-fold expansion of regulatory T cells over three weeks utilizing CD3/CD28 beads, IL-2, and CD4CD25 feeder cells.22 Alternately, transformation of primary T cells to regulatory T cells has been achieved by retroviral transduction with a Foxp3 containing vector, as well as TGF-beta mediated induction of Foxp3 expression during antigen stimulation.34 Importantly, Hoffman et al. have demonstrated that only the CD45RA and CD62L expressing cells within the in vitro expanded CD4CD25high T cells maintain the functional capacity of regulatory T cells in murine transplantation models.35,36 Ultimately, investigators have demonstrated that polyclonal expansion of isolated regulatory T cells can produce an expanded population that retains phenotypic and functional characteristics of regulatory T cells.22 Others have demonstrated enhanced suppressive activity of expanded regulatory T cells as compared to their native counterparts.20,37
Expansion of antigen-specific regulatory T cells
Both the unknown antigenic specificity of polyclonal regulatory T cells as well as their relatively low potency may limit the application of polyclonal regulatory T cells to human studies of aGVHD prevention. Conversely, several studies have demonstrated potential advantages of antigen-specific Tregs with this application in mind. Albert et al. demonstrated that antigen-specific regulatory T cells can abrogate effector T-cell response to allo-antigens in vitro and protect against GVHD in the setting of specific antigenic stimulus in a murine transplant model. The suppression mediated by these antigen-specific cells was 10-fold greater than that achieved with polyclonal Tregs.38 The increased potency of antigen-specific Tregs as compared to polyclonal Tregs is corroborated by experimental evidence from several other investigators.39–42 Tang et al. expanded a TCR transgenic Treg population over 200-fold utilizing anti-CD3/CD28.41 Bluestone et al. and Tarbell et al. have produced 20-fold expansion of regulatory T cells with peptide-pulsed antigen presenting cells and peptide-MHC tetramers.43,44 Luo et al. produced a 100-fold expansion of antigen-specific regulatory T cells using dendritic cells, antigen, and TGF-β.23 In these and other systems,45 antigenspecific regulatory T cells have been separated from nonspecific regulatory T cells through CFSE labeling and cell sorting based on low CFSE content after antigenic exposure and proliferation in cell culture. However, this methodology imposes restrictions on translation to human studies, given that this is not a GMP approved agent for human use. Expansion of purified regulatory T cells under selection pressure of rapamycin, that inhibits proliferation of effector T cells but is permissive for regulatory T cells, or cloning of antigen-specific regulatory T cells may provide an alternate strategy for human translational studies.
Pre-clinical and human clinical data
Several authors have demonstrated that ex vivo expanded regulatory T cells can suppress alloreactive CD4CD25 effector T cells. This has been examined both in vitro with MLR studies, as well as in vivo in several model systems including murine MHC-mismatch allogeneic stem cell transplantation models, solid organ transplant models, and autoimmune mediated diabetes/NOD murine models.13,22,23 This important pre-clinical work has greatly informed our understanding of the enormous potential of regulatory T cells in the prevention of untoward alloresponses. Hoffmann et al. demonstrated that CD4CD25 regulatory T cells effectively suppressed alloresponse (>90% inhibition) in the MLR reaction, and prevented otherwise lethal acute GVHD in a murine model of MHC-mismatched allogeneic stem cell transplantation.46 Several authors have described that transfer of both freshly isolated and ex vivo expanded regulatory T cells in a 1:1 ratio with effector T cells protects against GVHD mortality in murine models. 33,46
With regard to human studies, retrospective analyses have examined the relationship between peripheral blood regulatory T-cell numbers and GVHD by PCR for Foxp3 expression. Several groups have reported that decreased Foxp3 expression levels are correlated with acute and chronic GVHD.47,48 Importantly, several clinical trials are now underway to examine adoptive transfer of regulatory T cells as prevention against aGVHD in humans. In an ongoing phase I trial, Edinger et al. have infused freshly isolated donor Tregs alongside donor lymphocyte infusion. Similarly, Blaza et al. have isolated cord blood Treg cells and expanded them with anti-CD3 and –CD28 coated microbeads and IL-2 for use in double umbilical cord blood transplantation.15
Relative sparing of graft versus malignancy effect
While regulatory T cells have been shown in various in vitro and in vivomodels to abrogate GVHD by inhibiting the proliferation of alloreactive effector T cells, there is concern that their suppressive effect may also diminish the desirable graft versus malignancy effect. Several groups have examined this issue, and in these model systems, regulatory T cells abrogate graft versus host disease while allowing a preserved graft versus malignancy effect. The experimental conditions employed and the potential mechanisms responsible for preservation of graft versus malignancy response, in these studies differ however. Jones et al., utilizing an MHC-matched transplantation model, introduced the MMCBA6 leukemic cell line one day prior to irradiation and transplant. Regulatory T cells in a 1:3 ratio to effector CD8 T cells were given ten days following transplantation. Under these conditions, the administration of regulatory T cells did not compromise the CD8 mediated anti-leukemic response.37 Trenado et al. and Edinger et al. have employed different methods, using instead an MHCmismatched murine transplant model, administering Tregs at a 1:1 ratio with effector T cells (Tcon), and challenging with leukemic A20 cells at time of transplant, as well as in Edinger et al. with a BCL-1 lymphoma model administered nine days prior to HCT. Under these conditions, it appears that Tregs do not block Tcon activation, but rather decrease the early proliferation responsible for aGVHD. The Tcon remaining can still mediate graft versus malignancy response. This is thought to be mediated by cytotoxic effectors, as the GVL effect in this model system was compromised with FAS ligand or Perforin deficient Tcons.49,50 Trenado et al. have reported conflicting evidence regarding this effect, however, in that anti-tumor response to the P815 mastocytoma cell line was compromised by the addition of Tregs.50 These studies in total provide a compelling rationale for translation of this pre-clinical work to the clinical setting.
Effect of immunosuppressive agents
A potential obstacle to successful utilization of regulatory T cells in human transplantation is the adverse impact of immunosuppressive agents on regulatory T cells. A common component of immunosuppressive regimens in clinical transplantation, calcineurin inhibitors including cyclosporine and tacrolimus have a deleterious effect on regulatory T cells. These calcineurin inhibitors interfere with T-cell signaling involved in activation by binding intracellular cyclophilin/immunophilin, which inhibits the function of calcium/calmodulin dependent calcineurin. This in turn blocks further downstream transcription factors that support IL-2 transcription. Pre-clinical evidence suggests that these agents interfere with the development of regulatory T cells, as these cells are dependent on IL-2. Conversely, other immunosuppressive agents like rapamycin and mycophenolate mofetil, which act through alternate mechanisms, are permissive for the expansion of regulatory T cells in pre-clinical animal models. In this setting, rapamycin has been shown to support expansion of regulatory T cells, preserve the potent CD27 subset of CD4CD25 regulatory T cells, not impair Foxp3 expression, and allow for a 16-fold greater suppressor function as compared to regulatory T cells treated with cyclosporine.18,51 Several investigators have demonstrated that rapamycin selectively expands CD4CD25FoxP3 regulatory T cells in vitro, and that these cells exert a potent suppressive effect in vivo murine systems.52,53 Rapamycin has also been shown to selectively expand regulatory T cells in human samples from those with type I diabetes, as well as healthy controls,54 and also in the setting of human pancreatic islet transplantation. 55 Evidence suggests that rapamycin may induce and expand Tregs due to differential cell signaling in Tregs as compared to effector T cells: Tregs in murine and human systems have been shown to not activate the phosphatidyl inositol 3-kinase (PI3-K)/AKT pathway after activation through the T-cell receptor, while effector T cells do. As rapamycin inhibits mTOR in this pathway, it may therefore selectively inhibit effector T cells.56–58 In the clinical setting, the net effect on regulatory T-cell reconstitution and function after transplantation when calcineurin inhibitors are used alongside agents such as rapamycin or mycophenolate mofetil remains unknown, as this has not yet been prospectively examined. Alternative approaches would include either the use of a calcineurin inhibitor-free immunosuppressive platform59 or the use of exogenous IL-2 to promote regulatory T-cell survival and function.60
While several pre-clinical models suggest that regulatory T cells do not compromise the graft versus malignancy response in exerting their inhibitory effect on alloreactive T cells responsible for GVHD, there is also evidence that their suppressive effect is not entirely specific.21 In addition, there is burgeoning evidence that suppression of regulatory T cells leads to enhanced anti-tumor effects.61–67 Taken together, successful clinical application of this immunotherapy will likely require additional safeguards promoting specific suppression of those alloreactive cells responsible for GVHD, while allowing for preserved graft versus malignancy response. This may be made possible by the development of antigen-specific regulatory T cells.
Several groups have produced alloantigen-specific regulatory T cells and demonstrated their specific suppression of alloresponse directed against such antigen without effect on third-party alloantigen. These results have been achieved in several different transplant model systems, and with attention to varied antigenic targets including allopeptide, allogeneic MHC class II molecules, HLA-A2 peptide, dendritic cells pulsed with allopeptide, and by vaccination with anti-donor specific T cells.68–74 In a murine allogeneic hematopoietic cell transplantation model, Albert and colleagues transduced an antigen-specific CD4 Th1 clone with foxp3 to generate antigen-specific regulatory T cells; in vitro and in vivo, these cells suppressed alloreactive T cells in an antigen-dependent manner and with greater suppressive potency than polyclonal regulatory T cells.38
Alternatively, tissue restriction of minor histocompatibility antigen expression may provide an avenue for the selection and expansion of antigen-specific regulatory T cells. Minor histocompatibility antigens, polymorphic proteins presented by HLA molecules, differ between donor and recipient; this disparity is responsible for activation and expansion of alloreactive donor T cells. An increasing body of knowledge supports their role in transplantation outcomes: H-Y antigens have been associated with increased GVHD in the context of male recipients of female donors. Mismatches at other minor histocompatibility antigens have also been associated with increased incidence of GVHD. Additionally, minor histocompatibility antigens manifest different tissue distribution (Table 1); while some have ubiquitous tissue expression, others have hematopoietic tissue restriction.75–79
Accordingly, we propose a model utilizing this concept for the production of antigen-specific regulatory T cells for GVHD prevention. This would take advantage of tissue distribution of minor histocompatibility antigens in the following way: antigen-specific Tregs with specificity for minor antigens of broad tissue distribution would be selected. If these antigen-specific Tregs could only or more selectively suppress donor alloreactive T cells with specificity for broad distribution minor antigens, this would selectively abrogate these cells responsible for the development of aGVHD. Conversely, the Ag-specific Tregs would not suppress those alloreactive T cells with specificity for minor antigens restricted to hematopoietic cells; in this way, the putative drivers of the graft versus malignancy reaction would not be affected. In so doing, this would allow for the application to human transplantation with potential for more specific GVHD prevention with an additional safeguard against primary disease relapse, thereby improving clinical outcomes. However, enthusiasm for the development of antigen-specific regulatory T cells must be tempered by the potential for ‘infectious’ or spreading tolerance, whereby regulatory T cells can enhance their suppressive ability by enlisting additional CD4 suppressor subsets.80,81 Therefore, the feasibility of this proposed approach allows further exploration.
Challenges in translation to human studies
Regulatory T cells have emerged as a suppressive subset of the naturally occurring T cells, and may have an application in GVHD prevention. Motivated by their low precursor frequency, effective large scale ex vivo expansion of regulatory T cells has been achieved. Additionally, pre-clinical in vitro and murine data demonstrate the suppressive potential of these regulatory T cells, while graft versus malignancy effect appears preserved in some models. Finally, there are some early human clinical data that suggest a relationship between the development and severity of GVHD and numbers of circulating regulatory T cells. Accordingly, there is great interest in utilizing these expanded regulatory T cells for the prevention and treatment of GVHD.
Several important concerns regarding this application remain. First, the optimal timing of administration of regulatory T cells remains unknown. Given that the bulk of pre-clinical evidence supports their suppression of early activation of alloreactive T cells, a rationale exists for their administration prior to transplantation of the hematopoietic stem cell product, as this contains donor alloreactive T cells. Secondly, the optimal dose of regulatory T cells administered remains uncertain. While their suppressive effect is seen in pre-clinical models at low ratios of regulatory T cells to effector T cells, successful ex vivo expansion will need to take place given the low precursor frequency of naturally occurring regulatory T cells. Next, successful identification and separation of these regulatory T cells under clinical grade conditions will need to be performed in order to administer a product not contaminated by alloreactive T cells; administration of additional alloreactive T cells outside of those already contained in the hematopoietic stem cell product would invoke additional risk for GVHD, and thereby negate the intended benefit. Further efforts to refine current methods of isolation of Tregs are needed, as isolation based on cell surface marker expression may produce variable purity. The lack of a cell surface phenotype able to distinguish IL-10 producing Tr1 cells in particular limits the ability to isolate and utilize this regulatory T-cell subset for application in human studies.56 Finally, implementation of regulatory T-cell therapy in clinical investigation will require careful attention to any excess of primary disease relapse. Strategies for the creation of antigen-specific regulatory T cells may provide a means for selectively inhibiting those alloreactive T cells responsible for GVHD, while preserving the graft versus malignancy effect. Further work remains to be done to develop this approach.
- Funding: we acknowledge support from grants 3 P30-CA7692, 9 RO1-CA132197 and 5 K12-CA087989 from the NCI, and 5 K30-RR022270 from the National Center for Research Resources, National Institutes of Health.
- Authorship and Disclosures JP conducted the literature search and wrote the manuscript. CA provided mentorship and critically reviewed the manuscript.
- The authors have no financial conflicts or other conflicts of interest to report.
- Received August 18, 2009.
- Revision received September 29, 2009.
- Accepted October 15, 2009.
- Antin JH, Ferrara JL. Cytokine dysregulation and acute graft-versus-host disease. Blood. 1992; 80(12):2964-8. PubMedGoogle Scholar
- Chao NJ, Chen BJ. Prophylaxis and treatment of acute graft-versus-host disease. Semin Hematol. 2006; 43(1):32-41. PubMedhttps://doi.org/10.1053/j.seminhematol.2005.09.007Google Scholar
- Copelan EA. Hematopoietic stem-cell transplantation. N Engl J Med. 2006; 354(17):1813-26. PubMedhttps://doi.org/10.1056/NEJMra052638Google Scholar
- Hill GR, Ferrara JL. The primacy of the gastrointestinal tract as a target organ of acute graft-versus-host disease: rationale for the use of cytokine shields in allogeneic bone marrow transplantation. Blood. 2000; 95(9):2754-9. PubMedGoogle Scholar
- Horowitz MM, Gale RP, Sondel PM, Goldman JM, Kersey J, Kolb HJ. Graft-versus-leukemia reactions after bone marrow transplantation. Blood. 1990; 75(3):555-62. PubMedGoogle Scholar
- Thomas ED, Clift RA, Fefer A, Appelbaum FR, Beatty P, Bensinger WI. Marrow transplantation for the treatment of chronic myelogenous leukemia. Ann Intern Med. 1986; 104(2):155-63. PubMedGoogle Scholar
- Zittoun RA, Mandelli F, Willemze R, de Witte T, Labar B, Resegotti L. Autologous or allogeneic bone marrow transplantation compared with intensive chemotherapy in acute myelogenous leukemia. European Organization for Research and Treatment of Cancer (EORTC) and the Gruppo Italiano Malattie Ematologiche Maligne dell'Adulto (GIMEMA) Leukemia Cooperative Groups. N Engl J Med. 1995; 332(4):217-23. PubMedhttps://doi.org/10.1056/NEJM199501263320403Google Scholar
- Nash RA, Antin JH, Karanes C, Fay JW, Avalos BR, Yeager AM. Phase 3 study comparing methotrexate and tacrolimus with methotrexate and cyclosporine for prophylaxis of acute graft-versus-host disease after marrow transplantation from unrelated donors. Blood. 2000; 96(6):2062-8. PubMedGoogle Scholar
- Ratanatharathorn V, Nash RA, Przepiorka D, Devine SM, Klein JL, Weisdorf D. Phase III study comparing methotrexate and tacrolimus (prograf, FK506) with methotrexate and cyclosporine for graft-versus-host disease prophylaxis after HLA-identical sibling bone marrow transplantation. Blood. 1998; 92(7):2303-14. PubMedGoogle Scholar
- Martin PJ, Schoch G, Fisher L, Byers V, Anasetti C, Appelbaum FR. A retrospective analysis of therapy for acute graft-versus-host disease: initial treatment. Blood. 1990; 76(8):1464-72. PubMedGoogle Scholar
- Sierra J, Storer B, Hansen JA, Martin PJ, Petersdorf EW, Woolfrey A. Unrelated donor marrow transplantation for acute myeloid leukemia: an update of the Seattle experience. Bone Marrow Transplant. 2000; 26(4):397-404. PubMedhttps://doi.org/10.1038/sj.bmt.1702519Google Scholar
- Weisdorf D, Haake R, Blazar B, Miller W, McGlave P, Ramsay N. Treatment of moderate/severe acute graft-versus-host disease after allogeneic bone marrow transplantation: an analysis of clinical risk features and outcome. Blood. 1990; 75(4):1024-30. PubMedGoogle Scholar
- Albert MH, Anasetti C, Yu XZ. T regulatory cells as an immunotherapy for transplantation. Expert Opin Biol Ther. 2006; 6(4):315-24. PubMedhttps://doi.org/10.1517/147125188.8.131.525Google Scholar
- Nguyen VH, Zeiser R, Negrin RS. Role of naturally arising regulatory T cells in hematopoietic cell transplantation. Biol Blood Marrow Transplant. 2006; 12(10):995-1009. PubMedhttps://doi.org/10.1016/j.bbmt.2006.04.009Google Scholar
- Riley JL, June CH, Blazar BR. Human T regulatory cell therapy: take a billion or so and call me in the morning. Immunity. 2009; 30(5):656-65. PubMedhttps://doi.org/10.1016/j.immuni.2009.04.006Google Scholar
- Feuerer M, Hill JA, Mathis D, Benoist C. Foxp3+ regulatory T cells: differentiation, specification, subphenotypes. Nat Immunol. 2009; 10(7):689-95. PubMedhttps://doi.org/10.1038/ni.1760Google Scholar
- Miyara M, Yoshioka Y, Kitoh A, Shima T, Wing K, Niwa A. Functional delineation and differentiation dynamics of human CD4+ T cells expressing the FoxP3 transcription factor. Immunity. 2009; 30(6):899-911. PubMedhttps://doi.org/10.1016/j.immuni.2009.03.019Google Scholar
- Coenen JJ, Koenen HJ, van Rijssen E, Hilbrands LB, Joosten I. Rapamycin, and not cyclosporin A, preserves the highly suppressive CD27+ subset of human CD4+CD25+ regulatory T cells. Blood. 2006; 107(3):1018-23. PubMedhttps://doi.org/10.1182/blood-2005-07-3032Google Scholar
- Firan M, Dhillon S, Estess P, Siegelman MH. Suppressor activity and potency among regulatory T cells is discriminated by functionally active CD44. Blood. 2006; 107(2):619-27. PubMedhttps://doi.org/10.1182/blood-2005-06-2277Google Scholar
- Taylor PA, Lees CJ, Blazar BR. The infusion of ex vivo activated and expanded CD4(+)CD25(+) immune regulatory cells inhibits graft-versus-host disease lethality. Blood. 2002; 99(10):3493-9. PubMedhttps://doi.org/10.1182/blood.V99.10.3493Google Scholar
- Thornton AM, Shevach EM. Suppressor effector function of CD4+CD25+ immunoregulatory T cells is antigen nonspecific. J Immunol. 2000; 164(1):183-90. PubMedhttps://doi.org/10.4049/jimmunol.164.1.183Google Scholar
- Godfrey WR, Ge YG, Spoden DJ, Levine BL, June CH, Blazar BR. In vitro-expanded human CD4(+)CD25(+) T-regulatory cells can markedly inhibit allogeneic dendritic cell-stimulated MLR cultures. Blood. 2004; 104(2):453-61. PubMedhttps://doi.org/10.1182/blood-2004-01-0151Google Scholar
- Luo X, Tarbell KV, Yang H, Pothoven K, Bailey SL, Ding R. Dendritic cells with TGF-beta1 differentiate naive CD4+CD25− T cells into islet-protective Foxp3+ regulatory T cells. Proc Natl Acad Sci USA. 2007; 104(8):2821-6. PubMedhttps://doi.org/10.1073/pnas.0611646104Google Scholar
- Powrie F, Mason D. OX-22high CD4+ T cells induce wasting disease with multiple organ pathology: prevention by the OX-22low subset. J Exp Med. 1990; 172(6):1701-8. PubMedhttps://doi.org/10.1084/jem.172.6.1701Google Scholar
- Sakaguchi S, Fukuma K, Kuribayashi K, Masuda T. Organ-specific autoimmune diseases induced in mice by elimination of T cell subset. I. Evidence for the active participation of T cells in natural self-tolerance; deficit of a T cell subset as a possible cause of autoimmune disease. J Exp Med. 1985; 161(1):72-87. PubMedhttps://doi.org/10.1084/jem.161.1.72Google Scholar
- Sakaguchi S, Sakaguchi N, Asano M, Itoh M, Toda M. Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J Immunol. 1995; 155(3):1151-64. PubMedGoogle Scholar
- Zhou L, Chong MM, Littman DR. Plasticity of CD4+ T cell lineage differentiation. Immunity. 2009; 30(5):646-55. PubMedhttps://doi.org/10.1016/j.immuni.2009.05.001Google Scholar
- Park H, Li Z, Yang XO, Chang SH, Nurieva R, Wang YH. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat Immunol. 2005; 6(11):1133-41. PubMedhttps://doi.org/10.1038/ni1261Google Scholar
- Jin D, Zhang L, Zheng J, Zhao Y. The inflammatory Th 17 subset in immunity against self and non-self antigens. Autoimmunity. 2008; 41(2):154-62. PubMedhttps://doi.org/10.1080/08916930701776605Google Scholar
- Carlson MJ, West ML, Coghill JM, Panoskaltsis-Mortari A, Blazar BR, Serody JS. In vitro-differentiated TH17 cells mediate lethal acute graft-versus-host disease with severe cutaneous and pulmonary pathologic manifestations. Blood. 2009; 113(6):1365-74. PubMedhttps://doi.org/10.1182/blood-2008-06-162420Google Scholar
- Kappel LW, Goldberg GL, King CG, Suh DY, Smith OM, Ligh C. IL-17 contributes to CD4-mediated graft-versus-host disease. Blood. 2009; 113(4):945-52. PubMedhttps://doi.org/10.1182/blood-2008-08-172155Google Scholar
- Hoffmann P, Eder R, Kunz-Schughart LA, Andreesen R, Edinger M. Large-scale in vitro expansion of polyclonal human CD4(+)CD25high regulatory T cells. Blood. 2004; 104(3):895-903. PubMedhttps://doi.org/10.1182/blood-2004-01-0086Google Scholar
- Cohen JL, Trenado A, Vasey D, Klatzmann D, Salomon BL. CD4(+)CD25(+) immunoregulatory T Cells: new therapeutics for graft-versus-host disease. J Exp Med. 2002; 196(3):401-6. PubMedhttps://doi.org/10.1084/jem.20020090Google Scholar
- Horwitz DA, Zheng SG, Gray JD, Wang JH, Ohtsuka K, Yamagiwa S. Regulatory T cells generated ex vivo as an approach for the therapy of autoimmune disease. Semin Immunol. 2004; 16(2):135-43. PubMedhttps://doi.org/10.1016/j.smim.2003.12.009Google Scholar
- Ermann J, Hoffmann P, Edinger M, Dutt S, Blankenberg FG, Higgins JP. Only the CD62L+ subpopulation of CD4+CD25+ regulatory T cells protects from lethal acute GVHD. Blood. 2005; 105(5):2220-6. PubMedhttps://doi.org/10.1182/blood-2004-05-2044Google Scholar
- Hoffmann P, Eder R, Boeld TJ, Doser K, Piseshka B, Andreesen R. Only the CD45RA+ subpopulation of CD4+CD25high T cells gives rise to homogeneous regulatory T-cell lines upon in vitro expansion. Blood. 2006; 108(13):4260-7. PubMedhttps://doi.org/10.1182/blood-2006-06-027409Google Scholar
- Jones SC, Murphy GF, Korngold R. Post-hematopoietic cell transplantation control of graft-versus-host disease by donor CD425 T cells to allow an effective graft-versus-leukemia response. Biol Blood Marrow Transplant. 2003; 9(4):243-56. PubMedhttps://doi.org/10.1053/bbmt.2003.50027Google Scholar
- Albert MH, Liu Y, Anasetti C, Yu XZ. Antigen-dependent suppression of alloresponses by Foxp3-induced regulatory T cells in transplantation. Eur J Immunol. 2005; 35(9):2598-607. PubMedhttps://doi.org/10.1002/eji.200526077Google Scholar
- Jaeckel E, von Boehmer H, Manns MP. Antigen-specific FoxP3-transduced T-cells can control established type 1 diabetes. Diabetes. 2005; 54(2):306-10. PubMedhttps://doi.org/10.2337/diabetes.54.2.306Google Scholar
- Masteller EL, Warner MR, Tang Q, Tarbell KV, McDevitt H, Bluestone JA. Expansion of functional endogenous antigen-specific CD4+CD25+ regulatory T cells from nonobese diabetic mice. J Immunol. 2005; 175(5):3053-9. PubMedhttps://doi.org/10.4049/jimmunol.175.5.3053Google Scholar
- Tang Q, Henriksen KJ, Bi M, Finger EB, Szot G, Ye J. In vitro-expanded antigen-specific regulatory T cells suppress autoimmune diabetes. J Exp Med. 2004; 199(11):1455-65. PubMedhttps://doi.org/10.1084/jem.20040139Google Scholar
- Tarbell KV, Petit L, Zuo X, Toy P, Luo X, Mqadmi A. Dendritic cell-expanded, islet-specific CD4+ CD25+ CD62L+ regulatory T cells restore normoglycemia in diabetic NOD mice. J Exp Med. 2007; 204(1):191-201. PubMedhttps://doi.org/10.1084/jem.20061631Google Scholar
- Tarbell KV, Yamazaki S, Olson K, Toy P, Steinman RM. CD25+ CD4+ T cells, expanded with dendritic cells presenting a single autoantigenic peptide, suppress autoimmune diabetes. J Exp Med. 2004; 199(11):1467-77. PubMedhttps://doi.org/10.1084/jem.20040180Google Scholar
- Bluestone JA, Abbas AK. Natural versus adaptive regulatory T cells. Nat Rev Immunol. 2003; 3(3):253-7. PubMedhttps://doi.org/10.1038/nri1032Google Scholar
- Koenen HJ, Fasse E, Joosten I. CD27/CFSEbased ex vivo selection of highly suppressive alloantigen-specific human regulatory T cells. J Immunol. 2005; 174(12):7573-83. PubMedhttps://doi.org/10.4049/jimmunol.174.12.7573Google Scholar
- Hoffmann P, Ermann J, Edinger M, Fathman CG, Strober S. Donor-type CD4(+)CD25(+) regulatory T cells suppress lethal acute graft-versus-host disease after allogeneic bone marrow transplantation. J Exp Med. 2002; 196(3):389-99. PubMedhttps://doi.org/10.1084/jem.20020399Google Scholar
- Miura Y, Thoburn CJ, Bright EC, Phelps ML, Shin T, Matsui EC. Association of Foxp3 regulatory gene expression with graft-versus-host disease. Blood. 2004; 104(7):2187-93. PubMedhttps://doi.org/10.1182/blood-2004-03-1040Google Scholar
- Zorn E, Kim HT, Lee SJ, Floyd BH, Litsa D, Arumugarajah S. Reduced frequency of FOXP3+ CD4+CD25+ regulatory T cells in patients with chronic graft-versus-host disease. Blood. 2005; 106(8):2903-11. PubMedhttps://doi.org/10.1182/blood-2005-03-1257Google Scholar
- Edinger M, Hoffmann P, Ermann J, Drago K, Fathman CG, Strober S. CD4+CD25+ regulatory T cells preserve graft-versus-tumor activity while inhibiting graft-versus-host disease after bone marrow transplantation. Nat Med. 2003; 9(9):1144-50. PubMedhttps://doi.org/10.1038/nm915Google Scholar
- Trenado A, Charlotte F, Fisson S, Yagello M, Klatzmann D, Salomon BL. Recipient-type specific CD4+CD25+ regulatory T cells favor immune reconstitution and control graft-versus-host disease while maintaining graft-versus-leukemia. J Clin Invest. 2003; 112(11):1688-96. PubMedhttps://doi.org/10.1172/JCI200317702Google Scholar
- Zeiser R, Nguyen VH, Beilhack A, Buess M, Schulz S, Baker J. Inhibition of CD4+CD25+ regulatory T-cell function by calcineurin-dependent interleukin-2 production. Blood. 2006; 108(1):390-9. PubMedhttps://doi.org/10.1182/blood-2006-01-0329Google Scholar
- Battaglia M, Stabilini A, Roncarolo MG. Rapamycin selectively expands CD4+CD25+FoxP3+ regulatory T cells. Blood. 2005; 105(12):4743-8. PubMedhttps://doi.org/10.1182/blood-2004-10-3932Google Scholar
- Tian L, Lu L, Yuan Z, Lamb JR, Tam PK. Acceleration of apoptosis in CD4+CD8+ thymocytes by rapamycin accompanied by increased CD4+CD25+ T cells in the periphery. Transplantation. 2004; 77(2):183-9. PubMedhttps://doi.org/10.1097/01.TP.0000101005.44661.3EGoogle Scholar
- Battaglia M, Stabilini A, Migliavacca B, Horejs-Hoeck J, Kaupper T, Roncarolo MG. Rapamycin promotes expansion of functional CD4+CD25+FOXP3+ regulatory T cells of both healthy subjects and type 1 diabetic patients. J Immunol. 2006; 177(12):8338-47. PubMedhttps://doi.org/10.4049/jimmunol.177.12.8338Google Scholar
- Hering BJ, Kandaswamy R, Harmon JV, Ansite JD, Clemmings SM, Sakai T. Transplantation of cultured islets from two-layer preserved pancreases in type 1 diabetes with anti-CD3 antibody. Am J Transplant. 2004; 4(3):390-401. PubMedhttps://doi.org/10.1046/j.1600-6143.2003.00351.xGoogle Scholar
- Allan SE, Broady R, Gregori S, Himmel ME, Locke N, Roncarolo MG. CD4+ T-regulatory cells: toward therapy for human diseases. Immunol Rev. 2008; 223:391-421. PubMedhttps://doi.org/10.1111/j.1600-065X.2008.00634.xGoogle Scholar
- Bensinger SJ, Walsh PT, Zhang J, Carroll M, Parsons R, Rathmell JC. Distinct IL-2 receptor signaling pattern in CD4+CD25+ regulatory T cells. J Immunol. 2004; 172(9):5287-96. PubMedhttps://doi.org/10.4049/jimmunol.172.9.5287Google Scholar
- Crellin NK, Garcia RV, Levings MK. Altered activation of AKT is required for the suppressive function of human CD4+CD25+ T regulatory cells. Blood. 2007; 109(5):2014-22. PubMedhttps://doi.org/10.1182/blood-2006-07-035279Google Scholar
- Schleuning M, Judith D, Jedlickova Z, Stübig T, Heshmat M, Baurmann H, Schwerdtfeger R. Calcineurin inhibitor-free GVHD prophylaxis with sirolimus, mycophenolate mofetil and ATG in Allo-SCT for leukemia patients with high relapse risk: an observational cohort study. Bone Marrow Transplant. 2009; 43(9):717-23. PubMedhttps://doi.org/10.1038/bmt.2008.377Google Scholar
- Zorn E, Mohseni M, Kim H, Porcheray F, Lynch A, Bellucci R. Combined CD4+ donor lymphocyte infusion and low-dose recombinant IL-2 expand FOXP3+ regulatory T cells following allogeneic hematopoietic stem cell transplantation. Biol Blood Marrow Transplant. 2009; 15(3):382-8. PubMedhttps://doi.org/10.1016/j.bbmt.2008.12.494Google Scholar
- Comes A, Rosso O, Orengo AM, Di Carlo E, Sorrentino C, Meazza R. CD25+ regulatory T cell depletion augments immunotherapy of micrometastases by an IL-21-secreting cellular vaccine. J Immunol. 2006; 176(3):1750-8. PubMedhttps://doi.org/10.4049/jimmunol.176.3.1750Google Scholar
- Curtin JF, Candolfi M, Fakhouri TM, Liu C, Alden A, Edwards M. Treg depletion inhibits efficacy of cancer immunotherapy: implications for clinical trials. PLoS ONE. 2008; 3(4):e1983. PubMedhttps://doi.org/10.1371/journal.pone.0001983Google Scholar
- Heier I, Hofgaard PO, Brandtzaeg P, Jahnsen FL, Karlsson M. Depletion of CD4+ CD25+ regulatory T cells inhibits local tumour growth in a mouse model of B cell lymphoma. Clin Exp Immunol. 2008; 152(2):381-7. PubMedGoogle Scholar
- Imai H, Saio M, Nonaka K, Suwa T, Umemura N, Ouyang GF. Depletion of CD4+CD25+ regulatory T cells enhances interleukin-2-induced antitumor immunity in a mouse model of colon adenocarcinoma. Cancer Sci. 2007; 98(3):416-23. PubMedhttps://doi.org/10.1111/j.1349-7006.2006.00385.xGoogle Scholar
- Kline J, Brown IE, Zha YY, Blank C, Strickler J, Wouters H. Homeostatic proliferation plus regulatory T-cell depletion promotes potent rejection of B16 melanoma. Clin Cancer Res. 2008; 14(10):3156-67. PubMedhttps://doi.org/10.1158/1078-0432.CCR-07-4696Google Scholar
- Morse MA, Hobeika AC, Osada T, Serra D, Niedzwiecki D, Lyerly HK. Depletion of human regulatory T cells specifically enhances antigen-specific immune responses to cancer vaccines. Blood. 2008; 112(3):610-8. PubMedhttps://doi.org/10.1182/blood-2008-01-135319Google Scholar
- Nagai H, Horikawa T, Hara I, Fukunaga A, Oniki S, Oka M. In vivo elimination of CD25+ regulatory T cells leads to tumor rejection of B16F10 melanoma, when combined with interleukin-12 gene transfer. Exp Dermatol. 2004; 13(10):613-20. PubMedhttps://doi.org/10.1111/j.0906-6705.2004.00198.xGoogle Scholar
- Golshayan D, Jiang S, Tsang J, Garin MI, Mottet C, Lechler RI. In vitro-expanded donor alloantigen-specific CD4+CD25+ regulatory T cells promote experimental transplantation tolerance. Blood. 2007; 109(2):827-35. PubMedhttps://doi.org/10.1182/blood-2006-05-025460Google Scholar
- Jiang S, Tsang J, Game DS, Stevenson S, Lombardi G, Lechler RI. Generation and expansion of human CD4+ CD25+ regulatory T cells with indirect allospecificity: Potential reagents to promote donor-specific transplantation tolerance. Transplantation. 2006; 82(12):1738-43. PubMedhttps://doi.org/10.1097/01.tp.0000244932.29542.9eGoogle Scholar
- Nishimura E, Sakihama T, Setoguchi R, Tanaka K, Sakaguchi S. Induction of antigen-specific immunologic tolerance by in vivo and in vitro antigen-specific expansion of naturally arising Foxp3+CD25+CD4+ regulatory T cells. Int Immunol. 2004; 16(8):1189-201. PubMedhttps://doi.org/10.1093/intimm/dxh122Google Scholar
- Sanchez-Fueyo A, Sandner S, Habicht A, Mariat C, Kenny J, Degauque N. Specificity of CD4+CD25+ regulatory T cell function in alloimmunity. J Immunol. 2006; 176(1):329-34. PubMedhttps://doi.org/10.4049/jimmunol.176.1.329Google Scholar
- Tsang JY, Tanriver Y, Jiang S, Xue SA, Ratnasothy K, Chen D. Conferring indirect allospecificity on CD4+CD25+ Tregs by TCR gene transfer favors transplantation tolerance in mice. J Clin Invest. 2008; 118(11):3619-28. PubMedhttps://doi.org/10.1172/JCI33185Google Scholar
- Turnquist HR, Raimondi G, Zahorchak AF, Fischer RT, Wang Z, Thomson AW. Rapamycin-conditioned dendritic cells are poor stimulators of allogeneic CD4+ T cells, but enrich for antigen-specific Foxp3+ T regulatory cells and promote organ transplant tolerance. J Immunol. 2007; 178(11):7018-31. PubMedhttps://doi.org/10.4049/jimmunol.178.11.7018Google Scholar
- Wang J, Zhang L, Tang J, Jiang S, Wang X. Adoptive transfer of transplantation tolerance mediated by CD4+CD25+ and CD8+CD28− regulatory T cells induced by anti-donor-specific T-cell vaccination. Transplant Proc. 2008; 40(5):1612-7. PubMedhttps://doi.org/10.1016/j.transproceed.2008.02.079Google Scholar
- de Bueger M, Bakker A, Van Rood JJ, Van der Woude F, Goulmy E. Tissue distribution of human minor histocompatibility antigens. Ubiquitous versus restricted tissue distribution indicates heterogeneity among human cytotoxic T lymphocyte-defined non-MHC antigens. J Immunol. 1992; 149(5):1788-94. PubMedGoogle Scholar
- Goulmy E. Human minor histocompatibility antigens. Curr Opin Immunol. 1996; 8(1):75-81. PubMedhttps://doi.org/10.1016/S0952-7915(96)80108-7Google Scholar
- Marijt WA, Heemskerk MH, Kloosterboer FM, Goulmy E, Kester MG, van der Hoorn MA. Hematopoiesis-restricted minor histocompatibility antigens HA-1- or HA-2- specific T cells can induce complete remissions of relapsed leukemia. Proc Natl Acad Sci USA. 2003; 100(5):2742-7. PubMedhttps://doi.org/10.1073/pnas.0530192100Google Scholar
- Mutis T, Verdijk R, Schrama E, Esendam B, Brand A, Goulmy E. Feasibility of immunotherapy of relapsed leukemia with ex vivo-generated cytotoxic T lymphocytes specific for hematopoietic system-restricted minor histocompatibility antigens. Blood. 1999; 93(7):2336-41. PubMedGoogle Scholar
- Wang W, Meadows LR, den Haan JM, Sherman NE, Chen Y, Blokland E. Human H-Y: a male-specific histocompatibility antigen derived from the SMCY protein. Science. 1995; 269(5230):1588-90. PubMedhttps://doi.org/10.1126/science.7667640Google Scholar
- Jonuleit H, Schmitt E, Kakirman H, Stassen M, Knop J, Enk AH. Infectious tolerance: human CD25(+) regulatory T cells convey suppressor activity to conventional CD4(+) T helper cells. J Exp Med. 2002; 196(2):255-60. PubMedhttps://doi.org/10.1084/jem.20020394Google Scholar
- Stassen M, Schmitt E, Jonuleit H. Human CD(4+)CD(25+) regulatory T cells and infectious tolerance. Transplantation. 2004; 77(1 Suppl):S23-5. PubMedhttps://doi.org/10.1097/00007890-200401151-00009Google Scholar