Generation of allo-antigen-specific induced Treg stabilized by vitamin C treatment and its application for prevention of acute graft versus host disease model

Kasahara, Hidenori; Kondo, Taisuke; Nakatsukasa, Hiroko; Chikuma, Shunsuke; Ito, Minako; Ando, Makoto; Kurebayashi, Yutaka; Sekiya, Takashi; Yamada, Taketo; Okamoto, Shinichiro; Yoshimura, Akihiko

doi:10.1093/intimm/dxx060

Cited by 49 publications

(48 citation statements)

References 46 publications

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“…It is interesting to point out that some clones acquired total demethylation TSDR, so the refinement in their characterization could improve the isolation of stable iTregs with an established TSDR demethylation pattern over the course of the culture. In this context, recent reports have shown that is possible to increase the demethylation status of the TSDR region by adding vitamin C to iTreg cultures (56,57), which was reported to be an activator of TET enzymes, which, in turn, control the stability of FOXP3 (47).…”

Section: Discussionmentioning

confidence: 99%

Large-Scale Generation of Human Allospecific Induced Tregs With Functional Stability for Use in Immunotherapy in Transplantation

et al. 2020

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Regulatory T cells play an important role in the control of autoimmune diseases and maintenance of tolerance. In the context of transplantation, regulatory T cells (Tregs) have been proposed as new therapeutic tools that may induce allospecific tolerance toward the graft, avoiding the side effects induced by generalized immunosuppressors. Although most clinical trials are based on the use of thymic Tregs in adoptive therapy, some reports suggest the potential use of in vitro induced Tregs (iTregs), based on their functional stability under inflammatory conditions, indicating an advantage in a setting of allograft rejection. The aim of this work was to generate and expand large numbers of allospecific Tregs that maintain stable suppressive function in the presence of pro-inflammatory cytokines. Dendritic cells were derived from monocytes isolated from healthy donors and were co-cultured with CTV-labeled naïve T cells from unrelated individuals, in the presence of TGF-β1, IL-2, and retinoic acid. After 7 days of co-culture, proliferating CD4 + CD25 ++ CTV − cells (allospecific iTregs) were sorted and polyclonally expanded for 6 weeks in the presence of TGF-β1, IL-2, and rapamycin. After 6 weeks of polyclonal activation, iTregs were expanded 230,000 times, giving rise to 4,600 million allospecific iTregs. Allospecific iTregs were able to specifically suppress the proliferation of autologous CD4 + and CD8 + T cells in response to the allo-MoDCs used for iTreg generation, but not to third-party allo-MoDCs. Importantly, 88.5% of the expanded cells were CD4 + CD25 + FOXP3 + , expressed high levels of CCR4 and CXCR3, and maintained their phenotype and suppressive function in the presence of TNF-α and IL-6. Finally, analysis of the methylation status of the FOXP3 TSDR locus demonstrated a 40% demethylation in the purified allospecific iTreg, prior to the polyclonal expansion. Interestingly, the phenotype and suppressive activity of expanded allospecific iTregs were maintained after 6 weeks of expansion, despite an increase in the methylation status of the FOXP3 TSDR. In conclusion, this is the first report that demonstrates a large-scale generation of allospecific iTregs that preserve a stable phenotype and suppressor function in the presence of pro-inflammatory cytokines and pave the way for adoptive cell therapy with iTregs in transplanted patients.

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Section: Discussionmentioning

confidence: 99%

Large-Scale Generation of Human Allospecific Induced Tregs With Functional Stability for Use in Immunotherapy in Transplantation

et al. 2020

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“…37 Indeed, in a GVHD murine model, vitamin C stabilizes alloreactive T regulatory cells by demethylation of the FoxP3 promoter, the intronic cis-regulatory elements on DNA molecules. 38 Ascorbic acid is also a cofactor for the enzyme ten-eleven translocation (TET) which mediates promoter hypomethylation of the FOX P3 locus amongst other loci involved in hematopoiesis. 39,40 Huijskens et al showed that AA enhances ex vivo expansion of NK cells in cell culture systems as well as their differentiation from hematopoietic progenitors.…”

Section: Discussionmentioning

confidence: 99%

Reduced Plasma Ascorbic Acid Levels in Recipients of Myeloablative Conditioning & Hematopoietic Cell Transplantation

Rasheed

Simmons

Fisher

et al. 2019

Preprint

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Hematopoietic cell transplantation (HCT) conditioned using myeloablative conditioning (MAC) is complicated by end organ injury due to endothelial dysfunction and graft versus host disease. Mucositis and oxidant injury results in micronutrient deficiency. Ascorbic acid (AA) levels were measured in 15 patients undergoing HCT conditioned with MAC (11 allogeneic and 4 autologous HCT). Ascorbate levels declined post conditioning to 27.3 (±14.1) by day 0 (p <0.05 compared with baseline), reaching a nadir level of 21.5 (±13.8) on day 14 (p <0.05) posttransplant. Patients undergoing allogeneic HCT continued to have low AA levels to day 60 post transplant, whereas recipients of autologous HCT recovered plasma AA levels to normal. The role of AA in maintaining endothelial function and hematopoietic as well as T cell recovery is provided, developing the rationale for repletion of vitamin C following HCT.

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“…74 Recent studies have demonstrated that demethylation of CNS2 by Tet enzymes is promoted by vitamin C in iTregs, leading to stabilized Foxp3 expression. 17,65,72,74 Vitamin C was shown to potentiate Tet activity, 75,76 thereby facilitating demethylation of the Foxp3 CNS2 region and increasing the stability of Foxp3 expression in TGF-βinduced iTregs. 77 Compared with untreated iTregs, iTregs generated under low oxygen conditions in the presence of vitamin C retained more stable Foxp3 expression in vitro and in vivo and exhibited stronger suppression activity in a colitis model.…”

Section: Treg-specific Epigenetic Modifications and Stability Of Tregsmentioning

confidence: 99%

“…We observed stable, very high FOXP3 expression in human iTregs that underwent vitamin C-treatment. 17 Vitamin C treatment of iTregs also shows promise for innovative clinical applications of adoptive Treg immunotherapy. Further understanding of the mechanisms that facilitate the establishment of a Treg-specific transcriptional program will allow the production of sufficient numbers of antigen-specific Tregs with appropriate stability.…”

Section: Use Of Itregs To Prevent Gvhdmentioning

confidence: 99%

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Regulatory T Cells: Pathophysiological Roles and Clinical Applications

et al. 2019

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Inflammation and immune responses after tissue injury play pivotal roles in the resolution of inflammation, tissue recovery, fibrosis, and remodeling. Regulatory T cells (Tregs) are responsible for immune tolerance and are usually activated in secondary lymphatic tissues. Activated Tregs subsequently regulate effector T cell and dendritic cell activation. For clinical applications such as the suppression of both autoimmune diseases and the rejection of transplanted organs, methods to generate stabilized antigen-specific Tregs are required. For this purpose, transcriptional and epigenetic regulation of Foxp3 expression has been investigated. In addition to conventional Tregs, there are some Tregs that reside in tissues and are called tissue Tregs. Tissue Tregs exhibit tissue-specific functions that contribute to the maintenance of tissue homeostasis and repair. Such tissue Tregs could also be useful for Treg-based cell therapy. We recently discovered brain Tregs that accumulate in the brain during the chronic phase of ischemic brain injury. Brain Tregs resemble other tissue Tregs, but are unique in expressing neural cell-specific genes such as the serotonin receptor (Htr7); consequently, brain Tregs respond to serotonin. Here, we describe our experiences in the use of Tregs to suppress graft-versus-host disease and to promote neural recovery after stroke.

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Generation of allo-antigen-specific induced Treg stabilized by vitamin C treatment and its application for prevention of acute graft versus host disease model

Cited by 49 publications

References 46 publications

Large-Scale Generation of Human Allospecific Induced Tregs With Functional Stability for Use in Immunotherapy in Transplantation

Large-Scale Generation of Human Allospecific Induced Tregs With Functional Stability for Use in Immunotherapy in Transplantation

Reduced Plasma Ascorbic Acid Levels in Recipients of Myeloablative Conditioning & Hematopoietic Cell Transplantation

Regulatory T Cells: Pathophysiological Roles and Clinical Applications

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