Methods to manufacture regulatory T cells for cell therapy

MacDonald, Katherine N.; Piret, James M.; Levings, Megan K.

doi:10.1111/cei.13297

Cited by 85 publications

(92 citation statements)

References 100 publications

(150 reference statements)

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“…Engineering Tregs specific for a given antigen by transduction of a TCR, scFv, or BAR can render polyclonal Tregs more specific by increasing the number of cells that can target a given immune response, 8 as well as reduce potential non-specific suppression by Tregs of irrelevant specificities (see Brunstein et al 23 ). Aside from our laboratory, Levings and colleagues 9,24 previously reported the development of an scFv recognizing a human class I antigen, as well as the role of signaling domains in their efficacy. Interestingly, alterations of the latter did not improve the regulatory function significantly.…”

Section: Discussionmentioning

confidence: 99%

Genetic Engineering of T Cells for Immune Tolerance

Scott

2020

Molecular Therapy - Methods & Clinical Development

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Regulatory T cells (Tregs) play a role in the induction and maintenance of tolerance, as well as in modulating aberrant immune responses. While expanded Tregs have been used in clinical trials, they are polyclonal and the frequency of specific Tregs is very low. To overcome this issue, we have endeavored to "specify" Tregs by engineering them to express receptors that can recognize a given antigen and applied this protocol in autoimmunity, hemophilia and allergy. Thus, we have used retroviral transduction of a specific T cell receptor, single-chain variable fragments (Fvs), or antigen domains in Tregs to achieve this goal. This review summarizes our steps to achieve the ultimate goal of modulating human diseases.CD4 + regulatory T cells (Tregs) are well characterized for their immunosuppressive functions and maintenance of immunological tolerance. The importance of Tregs in immune regulation and brokering tolerance has been robustly demonstrated, 1-3 and expanded polyclonal Tregs are being developed for clinical applications. 4 Tregs can be broadly grouped into two categories, either "natural" or induced (i.e., peripherally derived). Natural Tregs (nTregs) represent between approximately 4% and 8% of CD4 + T cells in healthy donor peripheral blood, whereas induced Tregs can be generated from total CD4 T cells by expansion with anti-CD3 in the presence of transforming growth factor b (TGF-b). There T cells are polyclonal and reflect the entire repertoire. Herein, we summarize studies in our laboratory designed to render polyclonal Tregs antigen-specific by transduction of specific receptors such as T cell receptors (TCRs) or single-chain variable fragments (scFvs). More recently, we engineered antigenic domains in Tregs that then can directly be recognized by, or recognize B, cell receptors (BCRs). Our laboratory has concentrated recently on efforts to expand and "specify" Tregs, as well as CD8 cytotoxic cells, 2,5-7 and apply them to modulate adverse immune responses in autoimmunity, allergy, and hemophilia. Parallel work in other laboratories is summarized separately. 8-11 (DR1) peptide in the C2 region of FVIII. 12 The specific Teffs

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Section: Discussionmentioning

confidence: 99%

Genetic Engineering of T Cells for Immune Tolerance

Scott

2020

Molecular Therapy - Methods & Clinical Development

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“…Tregs are relatively rare cells that are hypo proliferative, so most protocols to expand Tregs for adoptive immunotherapy typically involve at least two polyclonal stimulations with anti-CD3 and -CD28 mAbs to obtain clinically relevant numbers (MacDonald et al, 2019b). To ask if repetitive stimulation could be detrimental for Treg function, we polyclonally stimulated Tregs weekly for 4 weeks and analyzed changes in their phenotype and function over time.…”

Section: Repetitive Polyclonal Treg Stimulation Leads To Decreased Sumentioning

confidence: 99%

“…Regulatory T cells (Tregs) control immune homeostasis and their adoptive transfer is a promising therapeutic approach with multiple trials completed, on-going or planned to test their efficiency in autoimmunity, solid organ transplantation or hematopoietic stem cell transplantation (reviewed in (Ferreira et al, 2019)). As Tregs are relatively rare, the majority of cell therapy protocols involve in vitro stimulation and expansion, in some cases culturing cells for up to 36 days to achieve > 2,000 fold expansion (MacDonald et al, 2019b). Although these in vitro-expanded cells typically remain FOXP3 + and suppressive in vitro, whether or not repeated stimulation of Tregs affects their in vivo function is unknown.…”

Section: Introductionmentioning

confidence: 99%

Repeated stimulation or tonic-signaling chimeric antigen receptors drive regulatory T cell exhaustion

Lamarche

Novakovsky

et al. 2020

Preprint

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Regulatory T cell (Treg) therapy is a promising approach to improve outcomes in transplantation and autoimmunity. In conventional T cell therapy, chronic stimulation can result in poor in vivo function, a phenomenon termed exhaustion. Whether or not Tregs are also susceptible to exhaustion, and if so, if this would limit their therapeutic effect, was unknown. We studied how two methods which induce conventional T cell exhaustion -repetitive stimulation or expression of a tonic-signaling chimeric antigen receptor (CAR) -affect human Tregs. With each repetitive polyclonal stimulation Tregs progressively acquired an exhausted phenotype, and became less suppressive in vitro. Tregs expressing a tonic-signaling CAR rapidly acquired an exhausted phenotype and had major changes in their transcriptome and metabolism. Although tonicsignaling CAR-Tregs remained stable and suppressive in vitro, they lost in vivo function, as tested in a model of xenogeneic graft-versus-host disease. The finding that human Tregs are susceptible to exhaustion has important implications for the design of Treg adoptive immunotherapy strategies.

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“…Conventional methods such as flow cytometry and ELISspot are regarded as the gold standards for single-cell analysis (see Figure 5a). [221][222][223] However, these approaches typically involve complex functionalization, immobilization, incubation, and washing steps, with a long assay time. Hence, they are limited to static measurements and do not completely satisfy the increasing demand of adding dynamic information in singlecell analysis.…”

Section: Biosensors For Label-free Detection Of Single-cell Protein Smentioning

confidence: 99%

Recent Advances and Perspectives in Microfluidics‐Based Single‐Cell Biosensing Techniques

Brimmo

Qasaimeh

et al. 2017

Small Methods

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Single‐cell analyses of secretory proteins are essential to fully understand cellular functional heterogeneity and unravel the underlying mechanisms of intercellular signaling and interactions. Retrieving dynamic information of protein secretion at single‐cell resolution reflects the precise, real‐time functional states of individual cells in physiological processes. Such measurements remain very challenging in single‐cell analysis, which requires highly integrated systems capable of performing on‐chip single‐cell isolation and subsequent real‐time protein detection. Here, recent advances in microfluidics‐based single‐cell manipulation and emerging approaches for label‐free single‐cell biosensing are reviewed. The advantages and limitations of these technologies are summarized and challenges to establish the integrated microfluidic biosensing systems for real‐time single‐cell secretomics are discussed. Recent efforts on integrated platforms for on‐chip single‐cell protein assays are highlighted and some perspectives on future directions in this field are provided.

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Methods to manufacture regulatory T cells for cell therapy

Cited by 85 publications

References 100 publications

Genetic Engineering of T Cells for Immune Tolerance

Genetic Engineering of T Cells for Immune Tolerance

Repeated stimulation or tonic-signaling chimeric antigen receptors drive regulatory T cell exhaustion

Recent Advances and Perspectives in Microfluidics‐Based Single‐Cell Biosensing Techniques

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