CD137+CD154- Expression As a Regulatory T Cell (Treg)-Specific Activation Signature for Identification and Sorting of Stable Human Tregs from In Vitro Expansion Cultures

Frontiers in Immunology
Anna NowakAlexander Scheffold

Abstract

Regulatory T cells (Tregs) are an attractive therapeutic tool for several different immune pathologies. Therapeutic Treg application often requires prolonged in vitro culture to generate sufficient Treg numbers or to optimize their functionality, e.g., via genetic engineering of their antigen receptors. However, purity of clinical Treg expansion cultures is highly variable, and currently, it is impossible to identify and separate stable Tregs from contaminating effector T cells, either ex vivo or after prior expansion. This represents a major obstacle for quality assurance of expanded Tregs and raises significant safety concerns. Here, we describe a Treg activation signature that allows identification and sorting of epigenetically imprinted Tregs even after prolonged in vitro culture. We show that short-term reactivation resulted in expression of CD137 but not CD154 on stable FoxP3+ Tregs that displayed a demethylated Treg-specific demethylated region, high suppressive potential, and lack of inflammatory cytokine expression. We also applied this Treg activation signature for rapid testing of chimeric antigen receptor functionality in human Tregs and identified major differences in the signaling requirements regarding CD137 vers...Continue Reading

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Citations

Nov 30, 2018·International Journal of Cancer. Journal International Du Cancer·Takuma MatobaSayuri Yamazaki
Sep 22, 2019·Nature Reviews. Drug Discovery·Leonardo M R FerreiraQizhi Tang
Jul 23, 2020·Frontiers in Immunology·Valentina BianchiGeorge Coukos
Sep 17, 2020·Oncoimmunology·Qun Zeng, Herbert Schwarz
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Dec 10, 2020·European Journal of Immunology·Patrick MaschmeyerMir-Farzin Mashreghi
Oct 29, 2020·Cellular & Molecular Immunology·Andrea Kristina HorstGisa Tiegs
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Jan 27, 2020·Trends in Biotechnology·Enrico FritscheMohamed Abou-El-Enein
May 1, 2021·Cancers·Justine CinierChristine Ménétrier-Caux

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Methods Mentioned

BETA
transgenic
Leukapheresis
FCS
transfection
PMA
FACS
PCR
Amplicon Sequencing

Software Mentioned

BiQ Analyzer HT
FlowJo
GraphPad PRISM

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