Recombinant CD95-Fc (APG101) prevents graft-versus-host disease in mice without disabling antitumor cytotoxicity and T-cell functions

Blood ◽  
2013 ◽  
Vol 121 (3) ◽  
pp. 556-565 ◽  
Author(s):  
Natalie Hartmann ◽  
Joanna J. Messmann ◽  
Frank Leithäuser ◽  
Maxi Weiswange ◽  
Michael Kluge ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Cell Function ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host ◽  
Cell Functions ◽  
T Cell Function

Abstract Graft-versus-host disease (GVHD) induced by transplant-derived T cells represents a major complication after allogeneic bone marrow transplantation (BMT). However, these T cells support engraftment, early T-cell immunity, and mediate the graft-versus-tumor (GVT) effect. Cytotoxic effector functions by transplanted T cells are predominantly mediated by the perforin/granzyme and the CD95/CD95L system. APG101, a novel recombinant human fusion protein consisting of the extracellular domain of CD95 and the Fc domain of an IgG1 antibody inhibited CD95L-induced apoptosis without interfering with T-cell function in vitro and was therefore tested for its ability to prevent GVHD in murine BMT models across minor or major histocompatibility barriers. Starting APG101 treatment either 1 day before or 6 days after transplantation effectively reduced clinical GVHD and rescued survival between 60% and 100% if GVHD was CD95L mediated. APG101 did not interfere with the GVT effect, because P815 mastocytoma and most importantly primary Bcr-Abl–transformed B-cell leukemias were completely eradicated by the alloantigen-specific T cells. Phenotype and homing of alloantigen-specific T cells or their perforin/granzyme-mediated cytotoxicity and proliferative capacity were not affected by APG101 treatment suggesting that APG101 therapy might be useful in GVHD prophylaxis without impairing T-cell function and most importantly preserving GVT activity.

scholarly journals Remodeling specific immunity by use of MHC tetramers: demonstration in a graft-versus-host disease model

Blood ◽  
2006 ◽  
Vol 107 (5) ◽  
pp. 2045-2051 ◽  
Author(s):  
Barry J. Kappel ◽  
Javier Pinilla-Ibarz ◽  
Adam A. Kochman ◽  
Jeffrey M. Eng ◽  
Vanessa M. Hubbard ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Allogeneic Bone Marrow Transplantation ◽  
Cell Populations ◽  
Solid Organ ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host ◽  
Mhc Molecules

Major histocompatibility complex (MHC) molecules carrying selected peptides will bind specifically to their cognate T-cell receptor on individual clones of reactive T cells. Fluorescently labeled, tetrameric MHC-peptide complexes have been widely used to detect and quantitate antigen-specific T-cell populations via flow cytometry. We hypothesized that such MHC-peptide tetramers could also be used to selectively deplete unique reactive T-cell populations, while leaving the remaining T-cell repertoire and immune response intact. In this report, we successfully demonstrate that a tetramer-based depletion of T cells can be achieved in a murine model of allogeneic bone marrow transplantation. Depletion of a specific alloreactive population of donor splenocytes (< 0.5% of CD8+ T cells) prior to transplantation significantly decreased morbidity and mortality from graft-versus-host disease. There was no early regrowth of the antigen-specific T cells in the recipient and in vivo T-cell proliferation was greatly reduced as well. Survival was increased more than 3-fold over controls, yet the inherent antitumor activity of the transplant was retained. This method also provides the proof-of-concept for similar strategies to selectively remove other unwanted T-cell clones, which could result in novel therapies for certain autoimmune disorders, T-cell malignancies, and solid organ graft rejection.

scholarly journals Critical role for CCR5 in the function of donor CD4+CD25+ regulatory T cells during acute graft-versus-host disease

Blood ◽  
2005 ◽  
Vol 106 (9) ◽  
pp. 3300-3307 ◽  
Author(s):  
Christian A. Wysocki ◽  
Qi Jiang ◽  
Angela Panoskaltsis-Mortari ◽  
Patricia A. Taylor ◽  
Karen P. McKinnon ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Graft Versus Host Disease ◽  
Critical Role ◽  
Allogeneic Bone Marrow Transplantation ◽  
Lymphoid Tissues ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host

AbstractCD4+CD25+ regulatory T cells (Tregs) have been shown to inhibit graft-versus-host disease (GVHD) in murine models, and this suppression was mediated by Tregs expressing the lymphoid homing molecule l-selectin. Here, we demonstrate that Tregs lacking expression of the chemokine receptor CCR5 were far less effective in preventing lethality from GVHD. Survival of irradiated recipient animals given transplants supplemented with CCR5-/- Tregs was significantly decreased, and GVHD scores were enhanced compared with animals receiving wild-type (WT) Tregs. CCR5-/- Tregs were functional in suppressing T-cell proliferation in vitro and ex vivo. However, although the accumulation of Tregs within lymphoid tissues during the first week after transplantation was not dependent on CCR5, the lack of function of CCR5-/- Tregs correlated with impaired accumulation of these cells in the liver, lung, spleen, and mesenteric lymph node, more than one week after transplantation. These data are the first to definitively demonstrate a requirement for CCR5 in Treg function, and indicate that in addition to their previously defined role in inhibiting effector T-cell expansion in lymphoid tissues during GVHD, later recruitment of Tregs to both lymphoid tissues and GVHD target organs is important in their ability to prolong survival after allogeneic bone marrow transplantation.

scholarly journals Absence of donor T-cell–derived soluble TNF decreases graft-versus-host disease without impairing graft-versus-tumor activity

Blood ◽  
2007 ◽  
Vol 110 (2) ◽  
pp. 783-786 ◽  
Author(s):  
Chiara Borsotti ◽  
Anna R. K. Franklin ◽  
Sydney X. Lu ◽  
Theo D. Kim ◽  
Odette M. Smith ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host ◽  
Tnf Α ◽  
Donor T Cells ◽  
Membrane Bound

Abstract Tumor necrosis factor (TNF) plays an important role in graft-versus-host disease (GVHD) and graft-versus-tumor (GVT) activity after allogeneic bone marrow transplantation (allo-BMT). TNF can be expressed in a membrane-bound form (memTNF) and as a soluble (solTNF) molecule after being cleaved by the TNF-α converting enzyme (TACE). To study the contribution of donor T-cell–derived memTNF versus solTNF in GVHD and GVT, we used mice containing a noncleavable allele in place of endogenous TNF (memTNFΔ/Δ) as donors in murine BMT models. Recipients of memTNF T cells developed significantly less GVHD than recipients of wild-type (wt) T cells. In contrast, GVT activity mediated by memTNF T cells remained intact, and alloreactive memTNF T cells showed no defects in proliferation, activation, and cytotoxicity. These data suggest that suppressing the secretion of solTNF by donor T cells significantly decreases GVHD without impairing GVT activity.

scholarly journals Prevention of graft-versus-host disease by anti–IL-7Rα antibody

Blood ◽  
2007 ◽  
Vol 110 (8) ◽  
pp. 2803-2810 ◽  
Author(s):  
Brile Chung ◽  
Eric P. Dudl ◽  
Dullei Min ◽  
Lora Barsky ◽  
Nancy Smiley ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Murine Models ◽  
Antibody Treatment ◽  
Host Disease ◽  
Graft Versus Host

Abstract Graft-versus-host disease (GVHD) continues to be a serious complication that limits the success of allogeneic bone marrow transplantation (BMT). Using IL-7–deficient murine models, we have previously shown that IL-7 is necessary for the pathogenesis of GVHD. In the present study, we determined whether GVHD could be prevented by antibody-mediated blockade of IL-7 receptor α (IL-7Rα) signaling. C57/BL6 (H2Kb) recipient mice were lethally irradiated and underwent cotransplantation with T-cell–depleted (TCD) BM and lymph node (LN) cells from allogeneic BALB/c (H2Kd) donor mice. Following transplantation, the allogeneic BMT recipients were injected weekly with either anti–IL-7Rα antibody (100 μg per mouse per week) or PBS for 4 weeks. Anti–IL-7Rα antibody treatment significantly decreased GVHD-related morbidity and mortality compared with placebo (30% to 80%). IL-7Rα blockade resulted in the reduction of donor CD4+ or CD8+ T cells in the periphery by day 30 after transplantation. Paradoxically, the inhibition of GVHD by anti–IL-7Rα antibody treatment resulted in improved long-term thymic and immune function. Blockade of IL-7R by anti–IL-7Rα antibody resulted in elimination of alloreactive T cells, prevention of GVHD, and improvement of donor T-cell reconstitution.

scholarly journals Nrf2 regulates CD4+ T cell–induced acute graft-versus-host disease in mice

Blood ◽  
2018 ◽  
Vol 132 (26) ◽  
pp. 2763-2774 ◽  
Author(s):  
Jennifer J. Tsai ◽  
Enrico Velardi ◽  
Yusuke Shono ◽  
Kimon V. Argyropoulos ◽  
Amanda M. Holland ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Cd8 T Cells ◽  
Cell Function ◽  
Host Disease ◽  
Graft Versus Host ◽  
T Cell Function ◽  
Donor T Cells ◽  
Acute Graft

Abstract Nuclear factor erythroid-derived 2-like 2 (Nrf2) is a ubiquitously expressed transcription factor that is well known for its role in regulating the cellular redox pathway. Although there is mounting evidence suggesting a critical role for Nrf2 in hematopoietic stem cells and innate leukocytes, little is known about its involvement in T-cell biology. In this study, we identified a novel role for Nrf2 in regulating alloreactive T-cell function during allogeneic hematopoietic cell transplantation (allo-HCT). We observed increased expression and nuclear translocation of Nrf2 upon T-cell activation in vitro, especially in CD4+ donor T cells after allo-HCT. Allo-HCT recipients of Nrf2−/− donor T cells had significantly less acute graft-versus-host disease (GVHD)-induced mortality, morbidity, and pathology. This reduction in GVHD was associated with the persistence of Helios+ donor regulatory T cells in the allograft, as well as defective upregulation of the gut-homing receptor LPAM-1 on alloreactive CD8+ T cells. Additionally, Nrf2−/− donor CD8+ T cells demonstrated intact cytotoxicity against allogeneic target cells. Tumor-bearing allo-HCT recipients of Nrf2−/− donor T cells had overall improved survival as a result of preserved graft-versus-tumor activity and reduced GVHD activity. Our findings characterized a previously unrecognized role for Nrf2 in T-cell function, as well as revealed a novel therapeutic target to improve the outcomes of allo-HCT.

CEACAM-1 Is Involved in Graft-Versus-Host-Disease in Murine Allogeneic Bone Marrow Transplantation Models.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 67-67
Author(s):  
Sydney X. Lu ◽  
Lucy Willis ◽  
Marsinay Smith ◽  
David Suh ◽  
Christopher King ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Bone Marrow Transplantation ◽  
T Cell ◽  
Small Bowel ◽  
Graft Versus Host Disease ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host

Abstract Carcinoembryonic antigen associated cell adhesion molecule 1 (CEACAM-1) belongs to a family of carcinoembryonic antigen-associated glycoproteins. It is expressed on leukocytes, endothelium, and epithelium. Microarray analysis showed that CEACAM-1 mRNA is increased in the small bowel during gut graft-versus-host-disease (GVHD) after allogeneic bone marrow transplantation (allo-BMT). Using CEACAM-1−/− mice as recipients or sources of donor bone marrow or T cells caused significantly worse GVHD mortality (p<0.05) compared to wildtype (WT) controls. Histopathological analysis of GVHD target organs from CEACAM-1−/− recipients of WT T cells or WT recipients of CEACAM-1−/− T cells revealed increased GVHD of the large bowel (p<0.05) but not liver or small bowel compared to WT control. Alloreactive splenic CD8 CEACAM-1−/− T cells from recipients with GVHD had increased levels of α4β7 integrin compared to WT controls. We also found increased numbers of small bowel intraepithelial lymphocytes and mesenteric lymph node cellularity in CEACAM-1−/− recipients of WT T cells and WT recipients of CEACAM-1−/− T cells, with a corresponding decrease of cellularity in peripheral lymph nodes and the liver. Adoptive transfer of CFSEhi CEACAM-1−/− T cells into WT hosts, or of WT T cells into CEACAM 1−/− hosts revealed more profound activation of T cells in CEACAM-1 deficient settings, shown by increased early CD25 expression and CD62L down-regulation on splenic CFSEdim alloreactive T cells. We found no significant differences in serum levels of TNF or IFNγ, T cell proliferation kinetics upon adoptive transfer, percentages of alloactivated CD4 or CD8 cells, intracellular levels of T-bet or IFNγ, CD8 T cell cytolytic efficiency, percentages of splenic regulatory T cells, or levels of T cell apoptosis in WT recipients of CEACAM-1−/− T cells or CEACAM-1−/− recipients of WT T cells with GVHD as compared with controls. Finally, irradiation of non-transplanted CEACAM-1−/− mice revealed increased radiation sensitivity, shown by earlier and greater lethality and increased small bowel crypt apoptosis, suggesting a role for CEACAM-1 in conditioning-related toxicity and subsequent GVHD amplification. We conclude that CEACAM-1 deficiency on donor T cells or transplant recipients results in increased gut and systemic GVHD due to increased T cell activation and elevated expression of the gut homing integrin α4β7. This suggests that the use of CEACAM-1 agonists could be a novel theraputic strategy for ameliorating acute intestinal and systemic graft-versus-host-disease.

scholarly journals Donor-derived thymic-dependent T cells cause chronic graft-versus-host disease

Blood ◽  
2006 ◽  
Vol 109 (4) ◽  
pp. 1756-1764 ◽  
Author(s):  
Yukimi Sakoda ◽  
Daigo Hashimoto ◽  
Shoji Asakura ◽  
Kengo Takeuchi ◽  
Mine Harada ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Mhc Class Ii ◽  
Graft Versus Host Disease ◽  
Cd4 T Cells ◽  
Chronic Gvhd ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host

Abstract Chronic graft-versus-host disease (GVHD) is the most common cause of poor long-term outcomes after allogeneic bone marrow transplantation (BMT), but the pathophysiology of chronic GVHD still remains poorly understood. We tested the hypothesis that the impaired thymic negative selection of the recipients will permit the emergence of pathogenic T cells that cause chronic GVHD. Lethally irradiated C3H/HeN (H-2k) recipients were reconstituted with T-cell–depleted bone marrow cells from major histocompatibility complex [MHC] class II–deficient (H2-Ab1−/−) B6 (H-2b) mice. These mice developed diseases that showed all of the clinical and histopathological features of human chronic GVHD. Thymectomy prevented chronic GVHD, thus confirming the causal association of the thymus. CD4+ T cells isolated from chronic GVHD mice were primarily donor reactive, and adoptive transfer of CD4+ T cells generated in these mice caused chronic GVHD in C3H/HeN mice in the presence of B6-derived antigen-presenting cells. Our results demonstrate for the first time that T cells that escape from negative thymic selection could cause chronic GVHD after allogeneic BMT. These results also suggest that self-reactivity of donor T cells plays a role in this chronic GVHD, and improvement in the thymic function may have a potential to decrease chronic GVHD.

scholarly journals Transient antibody targeting of CD45RC inhibits the development of graft-versus-host disease

2020 ◽  
Vol 4 (11) ◽  
pp. 2501-2515 ◽  
Author(s):  
Laetitia Boucault ◽  
Maria-Dolores Lopez Robles ◽  
Allan Thiolat ◽  
Séverine Bézie ◽  
Michael Schmueck-Henneresse ◽  
...  
Keyword(s):  
T Cells ◽  
Graft Versus Host Disease ◽  
Rat Model ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Host Disease ◽  
Graft Versus Host ◽  
Nsg Mice ◽  
Donor Specific Tolerance ◽  
Specific Tolerance

Abstract Allogeneic bone marrow transplantation (BMT) is a widely spread treatment of many hematological diseases, but its most important side effect is graft-versus-host disease (GVHD). Despite the development of new therapies, acute GVHD (aGVHD) occurs in 30% to 50% of allogeneic BMT and is characterized by the generation of effector T (Teff) cells with production of inflammatory cytokines. We previously demonstrated that a short anti-CD45RC monoclonal antibody (mAb) treatment in a heart allograft rat model transiently decreased CD45RChigh Teff cells and increased regulatory T cell (Treg) number and function allowing long-term donor-specific tolerance. Here, we demonstrated in rat and mouse allogeneic GVHD, as well as in xenogeneic GVHD mediated by human T cells in NSG mice, that both ex vivo depletion of CD45RChigh T cells and in vivo treatment with short-course anti-CD45RC mAbs inhibited aGVHD. In the rat model, we demonstrated that long surviving animals treated with anti-CD45RC mAbs were fully engrafted with donor cells and developed a donor-specific tolerance. Finally, we validated the rejection of a human tumor in NSG mice infused with human cells and treated with anti-CD45RC mAbs. The anti-human CD45RC mAbs showed a favorable safety profile because it did not abolish human memory antiviral immune responses, nor trigger cytokine release in in vitro assays. Altogether, our results show the potential of a prophylactic treatment with anti-human CD45RC mAbs in combination with rapamycin as a new therapy to treat aGVHD without abolishing the antitumor effect.

Protection of the Intestinal Epithelium from Conditioning with R-spondin1 Ameliorates Graft-Versus-Host Disease

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 66-66 ◽  
Author(s):  
Shuichiro Takashima ◽  
Kazutoshi Aoyama ◽  
Motoko Koyama ◽  
Daigo Hashimoto ◽  
Takeshi Oshima ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Large Bowel ◽  
Immune Reconstitution ◽  
Allogeneic Bone ◽  
Gi Tract ◽  
Host Disease ◽  
Graft Versus Host ◽  
Tract Damage

Abstract Damage to the gastrointestinal (GI) tract by pretransplant conditioning regimen plays a critical role in amplifying graft-versus-host disease (GVHD). Thus protection of the GI tract from conditioning may represent a novel approach to prevent GVHD. R-Spondin1 (R-Spo1) is a novel class of soluble activator for Wnt/□-catenin signaling, and has potent and specific proliferative effects on the intestinal crypt cells; injection of R-Spo1 protects mice from chemotherapy-induced intestinal mucositis. We therefore hypothesized that administration of R-Spo1 could modulate GVHD by reducing the GI tract damage and improve outcome of allogeneic bone marrow transplantation (BMT). Lethally irradiated B6D2F1 (H-2b/d) mice were injected with 5 × 106 BM and 2 × 106 T cells from MHC-mismatched B6 (H-2b) donors on day 0. Mice were intravenously injected with 200 μg of R-Spo1 or diluent from days −3 to −1 and +1 to +3 after BMT. In vivo labeling assay of mitotic cells with BrdU demonstrated that the proliferative index, as determined by the percentages of BrdU-positive cells among crypt epithelial cells, was significantly greater in the small intestine of R-Spo1 treated mice than controls 4 days after BMT (57% ± 3% vs 48% ± 1%, P<0.05). Analysis of the mesenteric lymph nodes and spleens on day +7 demonstrated significantly reduced expansion of donor T cells in R-Spo1 treated recipients in association with reduced serum levels of IFN-□ and TNF-□ on day +7 when compared to controls (Table). GVHD was severe in allogeneic controls, with 12.5% survival by day +40, whereas 62.5% of R-Spo1-treated animals survived this period (Table). Histopathologic examination of the small and large bowel and liver showed significantly reduced GVHD pathology in R-Spo1 treated animals than in controls (Table). A flowcytometric analysis of the spleen and thymus after BMT showed that administration of R-Spo1 did not impair donor cell engraftment and T and B cell immune reconstitution. We next evaluated the impact of R-Spo1 on graft-versus-leukemia (GVL) effects. BMT was performed similarly as above with the addition of 5 × 104 host-type P815 leukemia cells (H-2d). All recipients of T cell-depleted BM died from leukemia by day +20 after BMT, while no leukemia death was observed in R-Spo1 treated allogeneic animals. Overall, R-Spo1 treatment improved outcome of allogeneic BMT by reducing GVHD, while maintaining immune reconstitution and GVL effects. Thus, administration of R-Spo1 reduces the GI tract damage and suppresses donor T cell activation and systemic GVHD, supporting a hypothesis that the GI tract plays a major role in the amplification of systemic GVHD. Brief treatment with R-Spo1 may serve as an effective adjunct to clinical regimens of GVHD prophylaxis. Pathology Scores Group Survivals on day+40 (%) Small bowel Large bowel Liver INF □ (ng/ml) TNF □ (pg/ml) TCD: T cell-depleted BMT, +T: T cell-repleted BMT, ND: not detected Data are expressed as mean ± SD. *P<0.01 vs control, **P<0.05 vs control TCD diluent 100 2.4± 0.9 3.5± 1.0 0.3± 0.3 ND ND +T diluent 12.5 8.3± 2.7 7.3± 1.9 2.0± 0.8 6.0 ± 2.4 103.7 ± 9.9 +T R-Spo1 62.5* 3.4±1.9** 3.9± 0.3** 0.8± 0.7** 2.3 ± 1.5** 55.4 ± 6.6**

CCR1 Is Required for Treg-Mediated Protection From Lethal Acute Graft-Versus-Host Disease.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1337-1337
Author(s):  
Michael J Carlson ◽  
James M. Coghill ◽  
Michelle L. West ◽  
Angela Panoskaltsis-Mortari ◽  
Bruce R. Blazar ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Graft Versus Host Disease ◽  
Chemokine Receptor ◽  
Lymphoid Tissues ◽  
Allogeneic Bone ◽  
Widespread Application ◽  
Host Disease ◽  
Graft Versus Host ◽  
Target Organs

Abstract Abstract 1337 Poster Board I-359 INTRODUCTION Graft-versus-host disease (GVHD) is a major complication following allogeneic bone marrow transplantation (BMT). Despite advances in understanding the etiology of GVHD it remains a formidable obstacle to the widespread application of BMT. A number of studies have demonstrated that T regulatory (Treg) cells represent a potential therapy for GVHD as Tregs have been shown to inhibit GVHD while preserving the beneficial graft-versus-leukemia (GVL) effect. Numerous groups, including our own, have demonstrated the importance of T cell migration in the pathology of GVHD. Following conditioning, donor T cells migrate to secondary lymphoid tissues. Once activated in the lymphatics, T cells migrate to GVHD target organs including; the skin, liver, lung and the gastrointestinal (GI) tract in response to the local production of chemokines. Disruption of chemokine-chemokine receptor interactions has been demonstrated to affect the pathology of GVHD. Previously, we have shown that Tregs lacking the chemokine receptor CCR5, which binds CCL3, CCL4, and CCL5, do not protect animals from lethal GVHD as well as WT Tregs, due to their impaired migration to the liver and lung. Thus, a greater understanding of the function of chemokine receptors on Tregs is important in deciphering how Tregs function and whether targeting these cells to lymphoid tissue or GVHD target organs would be preferable for treating patients in clinical trials. METHODS We utilized a parent into F1 haploidentical model to assess the role of CCR1 in Treg-mediated protection from GVHD. Here we demonstrate Tregs lacking CCR1, another receptor for CCL3 and CCL5, were unable to protect animals against lethal acute GVHD. While 67% of B6D2 recipients given 1×106 WT Tregs supplemented with 5×106 WT T cells and 3×106 B6 T cell-depleted BM cells survived, only 15% of the recipients given CCR1−/− Tregs survived (p < 0.03; Fisher's exact test). B6D2 recipient mice given WT Tregs had significantly reduced clinical scores for GVHD compared to B6D2 recipients of CCR1−/− Tregs (p <0.05) with elevated GVHD scores starting on day 28 post-transplant. Histopathology revealed significantly worse pathology in the liver (p < 0.03) and colon (p < 0.05) of CCR1−/− Treg recipients vs. WT Treg recipients. In vitro analysis demonstrated that CCR1−/− Tregs were capable of suppressing T cell responses to allo-antigen equally as well as WT Tregs, and CCR1−/− Tregs attained a normal activation phenotype. Interestingly, preliminary experiments suggested that CCR1−/− Tregs migrated to and/or expanded in GVHD target organs to a similar extent as WT Tregs. CONCLUSIONS Treg expression of CCR1 is required for the inhibition of GVHD. Tregs lacking CCR1 led to significantly more tissue destruction in the liver and colon, two predominant sites of GVHD pathology. Of interest, the migration of CCR1−/− Tregs to GVHD target organs and secondary lymphoid tissues did not appear to be compromised suggesting that CCR1 may be required for the function of Tregsin vivo. Disclosures No relevant conflicts of interest to declare.

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