Regulatory T Cells Preserve Anti-Viral Immunity While Preventing Lethal Graft-Versus-Host Disease

Abstract Graft-versus-host disease (GvHD) is a frequent and life-threatening complication of allogeneic hematopoietic stem cell transplantation (HSCT) and is therefore one of the main factors that limits the broad application of HSCT. Over the last decades, several studies have reported a clinical association between GvHD and reactivation of cytomegalovirus (CMV). Using a lethal murine GvHD model with major MHC mismatch (C57BL/6 -> Balb/c), we were able to demonstrate that recipients latently infected with murine CMV (MCMV) before transplantation showed recurrence of CMV infection concomitant with the manifestation of GvHD. Moreover, these preinfected recipients showed an accelerated mortality compared to recipients that were not preinfected. The therapeutic co-infusion of CD4+CD25+ regulatory T cells (Tregs) with conventional T cells (Tcons) prevented lethal GvHD in preinfected mice and, markedly reduced the recurrence of MCMV infection. Remarkably, these mice showed clearance of MCMV 5 weeks post transplantation in contrast to mice receiving only Tcons in which massive virus infection persisted. Enhanced reconstitution of T lymphocytes and establishment of an anti-MCMV antibody titer from donor B cells in these animals suggest that CD4+CD25+ Tregs do not interfere with an anti-viral response while suppressing Tcon-mediated GvHD. Therefore, our study revealed that the suppressive function of CD4+CD25+ Tregs is not affected by CMV reactivation and more importantly, that Tregs do not adversely affect the anti-viral immunity in the recipient. In sum, these results provide important information on the correlation of GvHD and CMV reactivation and underline the possible clinical benefit of Treg application in GvHD patients. Disclosures: No relevant conflicts of interest to declare.

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Role of CD8 Regulatory T Cells versus Tc1 and Tc17 Cells in the Development of Human Graft-versus-Host Disease

Journal of Immunology Research ◽

10.1155/2017/1236219 ◽

2017 ◽

Vol 2017 ◽

pp. 1-11 ◽

Cited By ~ 4

Author(s):

Adriana Gutiérrez-Hoya ◽

Rubén López-Santiago ◽

Jorge Vela-Ojeda ◽

Laura Montiel-Cervantes ◽

Octavio Rodríguez-Cortés ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Graft Versus Host Disease ◽

Treg Cells ◽

Protective Role ◽

Hematopoietic Stem ◽

Host Disease ◽

Graft Versus Host ◽

Tc17 Cells

CD8+ T cells that secrete proinflammatory cytokines play a central role in exacerbation of inflammation; however, a new subpopulation of CD8 regulatory T cells has recently been characterized. This study analyzes the prominent role of these different subpopulations in the development of graft-versus-host disease (GVHD). Samples from 8 healthy donors mobilized with Filgrastim® (G-CSF) and 18 patients who underwent allogeneic hematopoietic stem cell transplantation (HSCT) were evaluated by flow cytometry. Mobilization induced an increase in Tc1 (p<0.01), Th1 (p<0.001), Tc17 (p<0.05), and CD8+IL-10+ cells (p<0.05), showing that G-CSF induces both pro- and anti-inflammatory profiles. Donor-patient correlation revealed a trend (p=0.06) toward the development of GVHD in patients who receive a high percentage of Tc1 cells. Patients with acute GVHD (aGVHD), either active or controlled, and patients without GVHD were evaluated; patients with active aGVHD had a higher percentage of Tc1 (p<0.01) and Tc17 (p<0.05) cells, as opposed to patients without GVHD in whom a higher percentage of CD8 Treg cells (p<0.01) was found. These findings indicate that the increase in Tc1 and Tc17 cells is associated with GVHD development, while regulatory CD8 T cells might have a protective role in this disease. These tests can be used to monitor and control GVHD.

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Nanoparticle-Encapsulated Allogeneic T Cells Mitigate Graft-Versus-Host Disease but Retain Graft-Versus-Leukemia Activity

Blood ◽

10.1182/blood.v124.21.3820.3820 ◽

2014 ◽

Vol 124 (21) ◽

pp. 3820-3820

Author(s):

Lingling Zhang ◽

Shuting Zhao ◽

Steven M. Devine ◽

Xiaoming He ◽

Jianhua Yu

Keyword(s):

T Cells ◽

Graft Versus Host Disease ◽

Conflicts Of Interest ◽

Hematopoietic Stem ◽

Host Disease ◽

Graft Versus Host ◽

Electrostatic Deposition ◽

Graft Versus Leukemia ◽

Donor T Cells

Abstract Allogeneic hematopoietic stem cell transplantation (HSCT) has curative potential for hematological malignancies, but is often associated with life-threatening complications including graft-versus-host disease (GVHD). The graft-versus-leukemia (GVL) activity which accompanies HSCT is responsible for eradication of tumor cells and prevention of relapse. GVHD and GVL are usually associated with each other and the separation of the two activities occurs in limited circumstances. In this study, we aimed to mitigate GVHD but retain GVL through transplantation of allogeneic T cells encapsulated with bio-degradable nanoparticle materials. For the above purpose, donor T cells were encapsulated with chitosan and alginate through layer-by-layer coating using electrostatic deposition. Encapsulated donor T cells were characterized in vitro, and their ability to inhibit GVHD and retain GVL was determined in vivo after being transplanted together with non-encapsulated donor bone marrow (BM) cells in a C57BL/6 → BALB/c HSCT mouse model. We found 85.7% of donor T cells were successfully encapsulated by the above method (Fig 1A). In vitro studies showed that the encapsulation did not change the phenotype of T cells as defined through the following parameters: size, viability, proliferation, antibody binding, cytokine secretion, and cytotoxicity of T cells (Fig. 1B and data not shown). Mice transplanted with encapsulated allogeneic T cells exhibited less severe acute GVHD and prolonged survival (Fig. 1 C-E). The mice showed a lower GVHD score, less liver damage, a smaller CD8/CD4 T cell ratio, and a higher number of donor BM-derived cells following transplantation with encapsulated donor T cells (Fig. 1 C-E and data not shown). When this GVHD model was combined with implantation of A20 lymphoma cells, GVL of encapsulated T cells was not compromised, while GVHD was still suppressed and the mouse survival also prolonged (Figure 2). In summary, nanoencapsulation of T cells with bio-degradable materials attenuated the severity of GVHD but retained GVL, presenting a novel and potentially safer and effective approach of allogeneic HSCT for future clinical application. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

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LYG1 Deficiency Attenuates the Severity of Acute Graft-Versus-Host Disease Via Skewing T Cells Polarization Towards Treg Cells

Blood ◽

10.1182/blood-2020-141877 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 4-4

Author(s):

Huihui Liu ◽

Zhengyu Yu ◽

Bo Tang ◽

Shengchao Miao ◽

Chenchen Qin ◽

...

Keyword(s):

T Cells ◽

Graft Versus Host Disease ◽

Cd4 T Cells ◽

Bone Marrow Cells ◽

Conflicts Of Interest ◽

Treg Cells ◽

Hematopoietic Stem ◽

Host Disease ◽

Graft Versus Host ◽

Acute Graft

Acute graft-versus-host disease (aGVHD) is a lethal complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT). As a complex immunopathology, aGVHD depends on the recognition of host antigens by donor T cells and induces augmented response of alloreactive T cells. Despite considerable achievements in the treatment of aGVHD, it remains a major clinical problem for the patients undergoing allo-HSCT. Therefore, it is necessary to further illustrate new mechanisms and develop novel therapeutic strategies of aGVHD. Previously we reported LYG1 (Lysozyme G-like 1) as a novel classical secretory protein promoted antitumor function of T cell. In this study, the role of LYG1 in aGVHD was investigated. Firstly, we examined whether LYG1 affected the alloreactivity of CD4+ T cells in vitro by MLR assay and discovered that LYG1 deficiency reduced the activation of CD4+ T cells and Th1 ratio, but increased Treg ratio. Then we confirmed these observations using a major MHC mismatched aGVHD model by transferring T cells sorting from WT B6 or Lyg1-/- mice with bone marrow cells from WT B6 mice into lethally irradiated BALB/c mice. The alloreactive CD4+ T cells and the proportions of Th1 cells decreased whereas the proportions of Treg cells increased in spleens and livers in mice receiving Lyg1-/- T cells. LYG1-deficient T cells attenuated aGVHD severity, inhibited the expression of CXCL9 and CXCL10 and restrained CD4+ T cells infiltrating in livers. Furthermore, administration of recombinant LYG1 protein intraperitoneally aggravated aGVHD by promoting IFN-γ production. More importantly, LYG1 deficiency did not affect GVT (graft-versus-tumor) effects. In summary, we demonstrate LYG1 regulates aGVHD via modulating the alloreactivity of CD4+ T cells and differentiation of Th1/Treg cells. Our study indicates that LYG1 may be a novel target in aGVHD by mitigating aGVHD without impairing GVT function. The therapeutic effect of targeting LYG1 is required in future investigations. Funding This study was supported by grant from The National Natural Science Foundation of China (NSFC) (Grant Number 81600144) and grant from Research Foundation of Peking University First Hospital. Disclosures No relevant conflicts of interest to declare.

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A Selective TNFR2 Agonist Expands Host Treg Cells in Vivo to Protect from Acute Graft-Versus-Host Disease

Blood ◽

10.1182/blood.v124.21.1099.1099 ◽

2014 ◽

Vol 124 (21) ◽

pp. 1099-1099 ◽

Cited By ~ 1

Author(s):

Andreas Beilhack ◽

Martin Chopra ◽

Marlene Biehl ◽

Martin Vaeth ◽

Andreas Brandl ◽

...

Keyword(s):

T Cells ◽

Graft Versus Host Disease ◽

Conflicts Of Interest ◽

Ex Vivo ◽

Hematopoietic Stem ◽

Peripheral Tissues ◽

Host Disease ◽

Graft Versus Host ◽

Pre Treatment

Abstract Donor CD4+Foxp3+ regulatory T cells (Tregs) suppress graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (allo-HCT) while maintaining the anti-tumoral effect of transplanted conventional T cells in preclinical mouse models. Current clinical study protocols with donor Tregs for treatment or prophylaxis of GVHD rely on their ex vivo expansion and infusion in high numbers. Here we present a fundamentally novel strategy for inhibiting GVHD that is based on the in vivo expansion of recipient Tregs prior to allo-HCT, exploiting the crucial role of tumor necrosis factor receptor 2 (TNFR2) in Treg biology. To this end we constructed a recombinant nonameric TNFR2-specific agonist, mimicking the activity of murine membrane-bound TNF on TNFR2 without TNFR1 stimulation, thereby avoiding the inflammatory side effects observed with conventional TNF. In vitro, this TNFR2-agonist expanded natural Tregs from wild type but not from TNFR2 KO mice. Accordingly, a human variant of this TNFR2-specific agonist expanded human Tregsin vitro. In vivo treatment of healthy mice with the murine TNFR2-agonist significantly increased Treg numbers in secondary lymphoid organs and peripheral tissues, particularly in the gastrointestinal tract, a prime target of acute GVHD. Next, we pre-treated recipient mice with this novel TNFR2-agonist to expand host-type radiation resistant Tregs prior to of allo-HCT in two models across MHC barriers (C57BL/6, H-2b->Balb/c, H-2d and FVB/N, H-2q->C57BL/6, H-2b). TNFR2-agonist pre-treatment resulted in significantly prolonged survival and reduced GVHD severity when compared to TNFR2-deficient recipients or untreated allo-HCT recipients. This was accompanied by reduced donor T cell proliferation and infiltration into GVHD target organs as assessed by in vivo and ex vivo bioluminescence imaging, flow cytometry and immunofluorescence microscopy. While in vivo TNFR2-agonist pre-treatment protected allo-HCT recipients from GVHD, anti-tumor effects of transplanted T cells remained unaffected in two different murine B cell leukemia models. In vivo depletion of host derived Tregs completely abrogated the protective effect of TNFR2-agonist pre-treatment. Our study shows that the expansion of host Tregs by selective in vivo TNFR2-activation significantly improves the outcome after allo-HCT and results in prolonged tumor-free survival. Disclosures No relevant conflicts of interest to declare.

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Preparation of Human Alloantigen-Specific Natural Regulatory T-Cells for the Control of Graft Versus Host Disease.

Blood ◽

10.1182/blood.v114.22.47.47 ◽

2009 ◽

Vol 114 (22) ◽

pp. 47-47

Author(s):

Yamina Hamel ◽

Claude Baillou ◽

Maude Guillot-Delost ◽

Soumia Touil ◽

Mustapha Cherai ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Graft Versus Host Disease ◽

Conflicts Of Interest ◽

Therapeutic Potential ◽

Third Party ◽

Host Disease ◽

Cell Ratio ◽

Graft Versus Host ◽

Teff Cell

Abstract Abstract 47 Adoptive transfer of natural regulatory T-cells (Tregs) may have a great therapeutic potential for the induction of tolerance in transplantation patients. This concept was demonstrated in murine models of graft versus host disease (GVHD) where alloantigen-specific Tregs (sTregs) were always more efficient to control GVHD than polyclonal Tregs (pTregs). Recently, we reported a procedure for expanding human pTregs in the presence of anti-CD3/anti-CD28 coated beads, cells being cultured in RPMI medium containing 10% human serum, interleukin (IL)-2 and rapamycin (namely TCM for Tregs Culture Medium) during 3 weeks. Whether or not alloantigen sTregs could be generated for the control of GVHD was investigated. In a first set of experiments, we compared the activation of Tregs when stimulated by allogeneic monocyte-derived dendritic cells (DCs) under different conditions. To follow cell divisions, Tregs (CD4+CD25high T-cells) were purified by FACS, stained with CFSE and co-cultured in TCM ± IL-15 in the presence of either immature (i) or mature (m) allogeneic DCs. Data showed that: i) mDCs yielded higher Treg divisions than iDCs; ii) combination of IL-2 + IL-15 triggered better cell division than IL-2 or IL-15 alone; iii) Tregs divided from day-3 to day-10 when stimulated by mDCs and cultured in TCM + IL-15. Next, the alloantigen specificity of divided Tregs was evaluated: Tregs stimulated by allogeneic mDCs (mDC1) were FACS-sorted at day-4 on the basis of CFSE-staining. CFSElow, CFSEintermediate (int) and CFSEhigh cell fractions were separated and expanded in TCM medium IL-15 in the presence anti-CD3/anti-CD28 coated beads. At day-21, they were assayed for their ability to inhibit within 4 days the proliferation of autologous effector T-cells (Teff) stimulated by either mDC1 or a third party of allogeneic mDCs (mDC2), used as control of specificity. In these mixed leukocyte cultures, different Tregs/Teff cell ratio were tested. Results showed that the CFSElow fraction (highly dividing Tregs) was greatly enriched in sTregs, by contrast to the CFSEhigh and CFSEint fractions. From these data, further experiments that are more suitable for clinical application were performed. Tregs selected by FACS or by immunomagnetic selection (MACs), based on CD25 expression, were cultured in TCM + IL-15 and stimulated twice (day-0 and day-10) by allogeneic mDCs (mDC1). At day-21, the alloantigen-specific suppression of sTregs was studied and compared to the suppression activity of pTregs. Results showed that sTregs better suppressed the proliferation of mDC1-stimulated Teff than pTregs. In addition, sTregs better maintained their suppressive activity than pTregs at low Tregs/Teff cell ratio. Under these culture conditions, sTregs can be expanded ∼12 fold within 3-weeks. Our data showed that human sTregs can be generated under clinical grade conditions. Further experiments using a xenogeneic GVHD model are now envisaged to compare the respective capacity for sTregs and pTregs to control GVHD. Disclosures: No relevant conflicts of interest to declare.

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Efficient and Selective Prevention of Graft-Versus-Host Disease by Ag-Specific Induced Regulatory T Cells In Mice.

Blood ◽

10.1182/blood.v116.21.3745.3745 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3745-3745

Author(s):

Kenrick Semple ◽

Yu Yu ◽

Dapeng Wang ◽

Claudio Anasetti ◽

Xue-Zhong Yu

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Graft Versus Host Disease ◽

Hematopoietic Cell Transplantation ◽

Conflicts Of Interest ◽

Therapeutic Potential ◽

Target Antigen ◽

Host Disease ◽

T Effector Cells ◽

Graft Versus Host

Abstract Abstract 3745 Naturally occurring regulatory T cells (nTregs) suppress the development of graft-versus-host disease (GVHD). The non-selective suppression against tumor associated antigens in some models severely dampened our enthusiasm for the application of nTregs in the control of GVHD after allogeneic hematopoietic cell transplantation (HCT). In this study, we used an alternative strategy to generate antigen-specific, induced Tregs (iTregs), and tested their potential in the prevention of GVHD in murine model of myeloablative BMT. CD4+CD25+Foxp3+ iTregs generated from OT-II TCR transgenic mice specific for OVA target antigen efficiently prevented GVHD induced by polyclonal T effector cells (Teffs) in allogeneic recipients that express OVA protein but not in those that do not express OVA. The efficacy of these antigen-specific iTregs was significantly higher than polyclonal iTregs in preventing GVHD. As controls, OT-II CD4+Foxp3− cells had no effect on GVHD development in OVA− recipients and exacerbated GVHD in OVA+ recipients when transplanted together with polyclonal Teffs. Mechanistically, OT-II iTregs expanded extensively, and significantly suppressed expansion and infiltration of Teffs in OVA+ recipients. In sharp contrast, OT-II iTregs failed to expand and had no effect on Teffs in OVA− recipients. These results reveal the therapeutic potential of TGFβ-induced, antigen-specific iTregs to prevent GVHD efficiently and selectively. Disclosures: No relevant conflicts of interest to declare.

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