Impact of mTOR Inhibition on FoxP3+ Regulatory T Cells as Compared to Conventional T Cells after Allogeneic Bone Marrow Transplantation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 448-448 ◽  
Author(s):  
Robert Zeiser ◽  
Dennis B. Leveson-Gower ◽  
Elizabeth A. Zambricki ◽  
Jing-Zhou Hou ◽  
Robert Negrin
Keyword(s):  
Clinical Trials ◽  
T Cells ◽  
Regulatory T Cells ◽  
Allogeneic Bone Marrow Transplantation ◽  
Inhibitory Effect ◽  
Allogeneic Bone ◽  
Mtor Inhibition ◽  
The Impact

Abstract FoxP3+CD4+CD25+ regulatory T-cells (Treg) have been shown to effectively reduce the severity of experimental acute graft-versus-host disease (aGvHD) while sparing graft-versus-leukemia activity. These findings, in concert with the observation that human and murine Treg share functional characteristics, have fueled interest in clinical trials to control aGvHD. Recent data indicates that the immunosuppressant rapamycin (RAPA) in contrast to cyclosporine A does not interfere with in vivo function of Treg and could enhance Treg expansion in vitro by a yet unknown mechanism. To investigate the impact of mTOR inhibition on proliferating Treg and Tconv, both cell types were exposed to CD3/CD28 Mabs in the presence of different RAPA concentrations in vitro. Phosphorylation of mTOR downstream products p70S6K1 and 4E-BP1 were assessed by western blot and flow cytometry. Inhibition of the phosphorylation of p70S6K1 and 4E-BP1 was observed in both populations in the presence of RAPA. Interestingly, Treg were more resistant to mTOR inhibition as compared to Tconv and displayed significantly higher phosphorylated products in the presence of RAPA at 10 nM (MFI Treg vs Tconv, p<0.001) and at 100nM (MFI Treg vs Tconv, p<0.001). To investigate whether Treg and RAPA protect from aGvHD in a synergistic manner, BALB/c recipients were transplanted with H-2 disparate BM and 1.6x10e6 T-cells (FVB/N) after lethal irradiation (8 Gy). aGvHD lethality was only slightly reduced when suboptimal Tconv:Treg ratios were employed (4:1, 8:1), or when recipients were treated with a non-protective RAPA dose (0.5 mg/kg bodyweight). Combining a suboptimal Tconv:Treg ratio with a non-protective RAPA dose reduced expansion of luciferase expressing (luc+) Tconv and pro-inflamatory cytokines and improved survival indicative for an additive in vivo effect of RAPA and Treg. To evaluate the impact of RAPA on in vivo T cell expansion, either luc+ Tconv or luc+ Treg were adoptively transferred. In vivo bioluminescence imaging demonstrated that RAPA had a more potent inhibitory effect on proliferation of Tconv as compared to Treg (p<0.05 vs. NS). We did not observe RAPA to increase FoxP3+ Treg numbers in vivo, or to enhance GITR or CTLA-4 expression. Thus, increased Treg numbers observed in RAPA containing expansion cultures are likely due to a lower susceptibility of this cell population to mTOR inhibition. This could explain the observed synergistic effect of RAPA and Treg in aGvHD protection which has relevance for clinical trials utilizing Treg to prevent aGvHD.

scholarly journals Identification and expansion of highly suppressive CD8+FoxP3+ regulatory T cells after experimental allogeneic bone marrow transplantation

Blood ◽  
2012 ◽  
Vol 119 (24) ◽  
pp. 5898-5908 ◽  
Author(s):  
Renee J. Robb ◽  
Katie E. Lineburg ◽  
Rachel D. Kuns ◽  
Yana A. Wilson ◽  
Neil C. Raffelt ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Bone Marrow Transplantation ◽  
Regulatory T Cells ◽  
Marrow Transplantation ◽  
Allogeneic Bone Marrow Transplantation ◽  
Class I ◽  
Allogeneic Bone ◽  
Mesenteric Lymph Nodes

Abstract FoxP3+ confers suppressive properties and is confined to regulatory T cells (Treg) that potently inhibit autoreactive immune responses. In the transplant setting, natural CD4+ Treg are critical in controlling alloreactivity and the establishment of tolerance. We now identify an important CD8+ population of FoxP3+ Treg that convert from CD8+ conventional donor T cells after allogeneic but not syngeneic bone marrow transplantation. These CD8+ Treg undergo conversion in the mesenteric lymph nodes under the influence of recipient dendritic cells and TGF-β. Importantly, this population is as important for protection from GVHD as the well-studied natural CD4+FoxP3+ population and is more potent in exerting class I–restricted and antigen-specific suppression in vitro and in vivo. Critically, CD8+FoxP3+ Treg are exquisitely sensitive to inhibition by cyclosporine but can be massively and specifically expanded in vivo to prevent GVHD by coadministering rapamycin and IL-2 antibody complexes. CD8+FoxP3+ Treg thus represent a new regulatory population with considerable potential to preferentially subvert MHC class I–restricted T-cell responses after bone marrow transplantation.

scholarly journals IL-2 administration increases CD4+CD25hi Foxp3+ regulatory T cells in cancer patients

Blood ◽  
2006 ◽  
Vol 107 (6) ◽  
pp. 2409-2414 ◽  
Author(s):  
Mojgan Ahmadzadeh ◽  
Steven A. Rosenberg
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Interleukin 2 ◽  
Selective Inhibition ◽  
High Dose ◽  
Protein Levels ◽  
And Function ◽  
The Impact

Abstract Interleukin-2 (IL-2) is historically known as a T-cell growth factor. Accumulating evidence from knockout mice suggests that IL-2 is crucial for the homeostasis and function of CD4+CD25+ regulatory T cells in vivo. However, the impact of administered IL-2 in an immune intact host has not been studied in rodents or humans. Here, we studied the impact of IL-2 administration on the frequency and function of human CD4+CD25hi T cells in immune intact patients with melanoma or renal cancer. We found that the frequency of CD4+CD25hi T cells was significantly increased after IL-2 treatment, and these cells expressed phenotypic markers associated with regulatory T cells. In addition, both transcript and protein levels of Foxp3, a transcription factor exclusively expressed on regulatory T cells, were consistently increased in CD4 T cells following IL-2 treatment. Functional analysis of the increased number of CD4+CD25hi T cells revealed that this population exhibited potent suppressive activity in vitro. Collectively, our results demonstrate that administration of high-dose IL-2 increased the frequency of circulating CD4+CD25hi Foxp3+ regulatory T cells. Our findings suggest that selective inhibition of IL-2-mediated enhancement of regulatory T cells may improve the therapeutic effectiveness of IL-2 administration. (Blood. 2006;107:2409-2414)

Critical and Opposing Effects of T Cell-Derived TNFα and Interferon-γ on IL-17 Induction Assessed in Vitro and in Vivo Following Allogeneic Bone Marrow Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3482-3482
Author(s):  
Minghui Li ◽  
Kai Sun ◽  
Mark Hubbard ◽  
Doug Redelman ◽  
Angela Panoskaltsis-Mortari ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Th17 Cells ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Allogeneic Bmt

Abstract IL-17-producing CD4 T cells (Th17) are a recently identified T helper subset that plays a role in mediating host defense to extracellular bacteria infections and is involved in the pathogenesis of many autoimmune diseases. In vitro induction of IL-17 in murine CD4+ T cells has been shown to be dependent on the presence of the proinflammatory cytokines TGF-β and IL-6 whereas IFNγ can suppress the development of Th17 cells. In the current study, we examined the roles of TNFα and IFNγ on IL-17 production by purified T cells in vitro and in vivo after allogeneic bone marrow transplantation (BMT). We present findings that expression of TNFα by the T cell itself is necessary for optimal development of Th17 under in vitro polarizing conditions. A novel role for T cell-derived TNFα in Th17 induction was observed when in vitro polarization of Tnf−/−CD4+ T cells resulted in marked reductions in IL-17+CD4+ T cells compared to Tnf+/+CD4+ T cells. In marked contrast, T cell-derived IFNγ markedly inhibited Th17 development as more IL-17+CD4+ T cells were found in Ifnγ−/−CD4+ T cells than in Ifnγ+/+CD4+ T cells, and of particular interest was the dramatic increase in IL-17+CD8+ cells from Ifnγ−/− mice. To determine if T cell-derived TNFα or IFNγ can regulate Th17 development in vivo we examined the differentiation of alloreactive donor T cells following allogeneic BMT. We have found that donor-derived Th17 cells can be found in lymphoid tissues and GVHD-affected organs after allogeneic BMT. However, transfer of Tnf−/− CD4+ T cells after allogeneic BMT resulted in marked reductions in Th17 cells in the spleen (18×103 vs 7×103, P<0.05). In agreement with the in vitro data and in contrast to what was observed with transfer of Tnf−/− CD4+ T cells, transfer of donor Ifnγ−/− T cells resulted in marked increases in not only IL-17+CD4+ but also IL-17+CD8+ T cells infiltrating the liver (7×103 vs 14×103, P<0.05; 4×104 vs 12.5×104, P<0.05). These results suggest that the donor T cell-derived TNFα and IFNγ opposingly regulate IL-17 induction of both CD4+ and CD8+ T cells in vitro and after allogeneic BMT which correlates with GVHD pathology.

The In Vivo Impact of Human Regulatory T Cells on the Graft Versus Tumor Effect.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 349-349 ◽  
Author(s):  
Tuna Mutis ◽  
Henk Rozemuller ◽  
Maarten E. Emmelot ◽  
Tineke Aarts-Riemens ◽  
Vivienne Verweij ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Tumor Cells ◽  
Cell Reactivity ◽  
Human Pbmc ◽  
Ifn Γ ◽  
T Cell Reactivity ◽  
The Impact

Abstract The curative Graft-vs-Tumor effect (GvT) of allogeneic Stem cell transplantation (SCT) is frequently complicated with life threatening Graft-vs-Host Disease (GvHD). In mice, prevention of GvHD, without abrogation of GvT is possible by co-transplantation of naturally occurring regulatory T cells (Tregs) with SC grafts. Consistent with these murine studies, we recently demonstrated that also human Tregs possess potent GvHD-downregulatory capacities in a xenogeneic(x) model, where x-GvHD is induced by infusion of autologous human T cells in RAG2−/−γc−/− mice (Mutis et al. Clin. Cancer Res.2006, 12: 5520–5525). Towards clinical application of Tregs, we now explored the impact of human Treg-administration on GvT in a bioluminescence imaging (BLI) based human-GvT model in the RAG2−/−γc−/− mice. In this model, mice inoculated with luciferase (LUC)-transduced human myeloma (MM) cell lines developed BLI-detectable, progressive, MM-like multifocal tumors exclusively in the bone marrow (BM). Full blown tumors were effectively eliminated by infusion of allogeneic human PBMC. This treatment also caused lethal x-GvHD as expected. In this setting, co-infusion of human PBMC with autologous, in vitro cultured Tregs at a 1:1 Treg: T effector cell ratio had no adverse effects on the development of GvT while significantly reducing the lethality of x-GvHD. In vitro analyses of sacrificed mice at day 21 revealed that administered Tregs homed to BM and spleen, significantly downregulated the total numbers of IFN-γ-producing CD4+ and CD8+ T cells responding to CD3 mediated signals, but had no downregulatory effect on the frequencies of IFN-γ-producing T cells responding to tumor cells. There was also no downregulation of cytotoxic activity against tumor cells in Treg-treated mice. Conclusively, these results showed that Tregs, at doses which are inhibitory for x-GvHD-inducing T cells, could maintain the GvT effect by allowing T cell reactivity against tumor cells. Human Tregs thus still hold promise as attractive cellular tools for separating GvT from GvHD.

Donor CD4+CD25+ T cells promote engraftment and tolerance following MHC-mismatched hematopoietic cell transplantation

Blood ◽  
2005 ◽  
Vol 105 (4) ◽  
pp. 1828-1836 ◽  
Author(s):  
Alan M. Hanash ◽  
Robert B. Levy
Keyword(s):  
T Cells ◽  
Cell Transplantation ◽  
Hematopoietic Cell Transplantation ◽  
Allogeneic Bone Marrow Transplantation ◽  
Disease Recurrence ◽  
Hematopoietic Cell ◽  
Allogeneic Bone ◽  
Hematopoietic Stem

AbstractAllogeneic bone marrow transplantation (BMT) is a potentially curative treatment for both inherited and acquired diseases of the hematopoietic compartment; however, its wider use is limited by the frequent and severe outcome of graft-versus-host disease (GVHD). Unfortunately, efforts to reduce GVHD by removing donor T cells have resulted in poor engraftment and elevated disease recurrence. Alternative cell populations capable of supporting allogeneic hematopoietic stem/progenitor cell engraftment without inducing GVHD could increase numbers of potential recipients while broadening the pool of acceptable donors. Although unfractionated CD4+ T cells have not been shown to be an efficient facilitating population, CD4+CD25+ regulatory cells (T-reg's) were examined for their capacity to support allogeneic hematopoietic engraftment. In a murine fully major histocompatibility complex (MHC)-mismatched BMT model, cotransplantation of donor B6 T-reg's into sublethally conditioned BALB/c recipients supported significantly greater lineage-committed and multipotential donor progenitors in recipient spleens 1 week after transplantation and significantly increased long-term multilineage donor chimerism. Donor engraftment occurred without GVHD-related weight loss or lethality and was associated with tolerance to donor and host antigens by in vitro and in vivo analyses. Donor CD4+CD25+ T cells may therefore represent a potential alternative to unfractionated T cells for promotion of allogeneic engraftment in clinical hematopoietic cell transplantation. (Blood. 2005;105:1828-1836)

HDAC11 Inhibits T-Cell Response to Alloantigens and Reduces Gvhd In Mice.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1466-1466
Author(s):  
Dapeng Wang ◽  
Fengdong Cheng ◽  
Yu Yu ◽  
Kenrick Semple ◽  
Lirong Peng ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Allogeneic Bone Marrow Transplantation ◽  
T Cell Response ◽  
Allogeneic Bone ◽  
Allogeneic Bmt ◽  
Significant Difference

Abstract Abstract 1466 Background: Histone acetyltransferases and histone deacetylases (HDAC) regulate gene expression through acetylation-deacetylation of histones. HDACs are the target of a family of compounds known as HDAC inhibitors, which have been shown to suppress pro-inflammatory cytokines and reduce acute graft-versus-host disease (GVHD) while preserving the graft-versus-leukemia (GVL) effect after allogeneic bone marrow transplantation (BMT) in mice. However, the role of individual HDAC members in the development of GVHD is not clear. Recently, HDAC11, the newest member of the HDAC family has emerged as an important transcriptional regulator of inflammatory responses in antigen-presenting cells (APCs)1. Here, we evaluated the role of HDAC11 on APCs and T cells in the allogeneic BMT setting in mice with genetic disruption of HDAC11. Method: Proliferation of wild-type (WT) and HDAC11 knock-out (KO) T cells in response to allogeneic antigens was compared by [H3] thymidine incorporation assay. Using the same method, we also tested the antigen presentation ability of WT and HDAC11 KO APCs. For in vivo studies, we used a clinical relevant mouse model of BMT: C57BL/6 (B6) ® BALB/c. To evaluate the role of HDAC11 in the function of T cells and APCs, WT and KO mice on B6 background were used as donors and recipients, respectively. Recipient survival was monitored daily and GVHD symptom was evaluated at least twice a week. HDAC11 KO mice were supplied by Merck and Co., Inc. Results: In vitro, HDAC11 KO T cells proliferated stronger than WT T cells under the stimulation of allogeneic APCs. Recipients of HDAC11 KO T cells lost significantly more body weight (p < 0.05), and died significantly sooner than those of WT T cells (p < 0.01). The pathologic score of KO recipients was higher than that of WT recipients in each of GVHD target organs including lung, liver, small intestine and colon. Mechanistically, we found that there were significantly more total and IFNγ-producing donor T cells in the recipients of KO cells than those of WT cells (p < 0.05). Collectively, HDAC11 KO T cells have higher activity in response to alloantigens in vitro and induced more severe GVHD in vivo compared to WT T cells. In contrast, KO and WT APCs had a similar ability to stimulate allogeneic T cells in vitro, and no significant difference in GVHD development was observed in WT or KO recipients after allogeneic BMT. Conclusion: HDAC11 negatively regulates T-cell function, but has no significant effect on APC function. This finding provides a rationale to promote T-cell immunity or tolerance by inhibiting or enhancing HDAC11, respectively. 1 Villagra et al. Nature Immunology, 10:92-100, 2009. Disclosures: No relevant conflicts of interest to declare.

Only the CD62L+ subpopulation of CD4+CD25+ regulatory T cells protects from lethal acute GVHD

Blood ◽  
2005 ◽  
Vol 105 (5) ◽  
pp. 2220-2226 ◽  
Author(s):  
Joerg Ermann ◽  
Petra Hoffmann ◽  
Matthias Edinger ◽  
Suparna Dutt ◽  
Francis G. Blankenberg ◽  
...  
Keyword(s):  
T Cells ◽  
Allogeneic Bone Marrow Transplantation ◽  
Treg Cells ◽  
Allogeneic Bone ◽  
Protective Function ◽  
Graft Versus Host ◽  
Characteristic Features ◽  
Related Mortality

AbstractCD4+CD25+ regulatory T (Treg) cells are potent modulators of alloimmune responses. In murine models of allogeneic bone marrow transplantation, adoptive transfer of donor CD4+CD25+ Treg cells protects recipient mice from lethal acute graft-versus-host disease (aGVHD) induced by donor CD4+CD25- T cells. Here we examined the differential effect of CD62L+ and CD62L- subsets of CD4+CD25+ Treg cells on aGVHD-related mortality. Both subpopulations showed the characteristic features of CD4+CD25+ Treg cells in vitro and did not induce aGVHD in vivo. However, in cotransfer with donor CD4+CD25- T cells, only the CD62L+ subset of CD4+CD25+ Treg cells prevented severe tissue damage to the colon and protected recipients from lethal aGVHD. Early after transplantation, a higher number of donor-type Treg cells accumulated in host mesenteric lymph node (LN) and spleen when CD4+CD25+CD62L+ Treg cells were transferred compared with the CD62L- subset. Subsequently, CD4+CD25+CD62L+ Treg cells showed a significantly higher capacity than their CD62L- counterpart to inhibit the expansion of donor CD4+CD25- T cells. The ability of Treg cells to efficiently enter the priming sites of pathogenic allo-reactive T cells appears to be a prerequisite for their protective function in aGVHD.

Delayed Onset of T Cell-Mediated Xenogeneic Disease in Rag2−/− γc−/− mice after Co- Transplantation of in Vitro Expanded Human CD4+CD25highCD45RA+ Regulatory T Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4609-4609
Author(s):  
Tina J Boeld ◽  
Ellen Obermann ◽  
Reinhard Andreesen ◽  
Petra Hoffmann ◽  
Matthias Edinger
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Allogeneic Bone Marrow Transplantation ◽  
Disease Onset ◽  
Clinical Signs ◽  
Allogeneic Bone

Abstract In murine disease models, the adoptive transfer of donor CD4+CD25+ regulatory T cells (Treg) prevents lethal graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation. We recently described methods for the isolation and in vitro expansion of human Treg from adult peripheral blood and found that only CD45RA+ but not CD45RA− CD4+CD25high Treg (RA+ and RA− Treg, respectively) maintain FOXP3 expression as well as suppressive activity after prolonged culture (Hoffmann et al. Blood108:4260; 2006). Here, we compared the immunoregulatory capacity of these two different Treg subpopulations in vivo in a xenogeneic GVHD (xGVHD) model. Transplantation of 20x106 human peripheral blood mononuclear cells (huPBMCs) into immunodeficient BALB/c Rag2−/−γc−/− mice led to engraftment and extensive proliferation of human CD45+ cells (huCD45+), which was accompanied by clinical signs of xGVHD. Histological and flow cytometric evaluation revealed that mainly CD8+ T cells within PBMC caused xGVHD and that main targets were liver, lung and spleen as well as mouse hematopoiesis. When expanded CD45RA+ Treg were co-transplanted at a 1:1 ratio with CD4+ and CD8+ T cells contained in huPBMC, the frequency of huCD45+ cells in peripheral blood (PB) at d14 was reduced to 12 ± 4.9 % (n=15), while animals that received expanded RA− Treg had 30 ± 7.5 % (n=15) and animals without co-transfer of Treg 40 ± 8.3 % (n=14) huCD45+ cells. The expansion of conventional T cells, however, was only delayed but not prevented, as by d21 the percentage of huCD45+ cells in PB increased to 35 ± 9.2 % (n=15) when RA+ Treg were co-transplanted as compared to 40 ± 9.3 % (n=15) with RA− Treg and 50 ± 6.4 % (n=14) without Treg administration. Subsequently, all mice developed clinical signs of xGVHD and 73.3 % of mice co-transplanted with RA+ Treg, 86.7 % co-transplanted with RA− Treg and 78.6 % of mice transplanted with huPBMCs alone died within 100 days. Since Treg mainly inhibited early expansion of huPBMC, we examined the splenic cellularity of xeno-transplanted animals at d10 after cell transfer. While spleens of mice that received only huPBMC contained 20 ± 3x106 huCD45+ cells (n=10), co-transfer of RA− Treg reduced huCD45+ cells to 14 ± 4.8×106 (n=9) whereas animals that received RA+ Treg contained only 5 ± 1.1×106 huCD45+ cells (n=11) at that time. Furthermore, IL-2 and IFN-γ production of conventional T cells re-isolated from the spleen on d10 was reduced 5fold in mice co-transplanted with RA+ Treg as compared to recipients of only huPBMCs or additional RA− Treg. Thus, in vitro expanded human RA+ Treg suppress the proliferation and cytokine production of conventional T cells in early phases of xGVHD, but ultimately do not prevent disease onset and mortality.

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