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951 An Ezh2 gain of function mutation gives regulatory T cells a gain of function
  1. Janneke GC Peeters1,
  2. Stephanie Silveria1,
  3. Merve Ozdemir1,
  4. Grigorios Georgolopoulos2 and
  5. Michel DuPage1
  1. 1University of California, Berkeley, Berkeley, CA, USA
  2. 2Genevia Technology, Tampere, Pirkanmaa, Finland
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background Regulatory T cells (Tregs) are key players in maintaining immune homeostasis due to their immunosuppressive function. By downregulating the immune response, Tregs are essential in preventing autoimmune diseases, such as multiple sclerosis, in which Treg dysfunction has been reported. In contrast, high Treg frequencies are reported in many cancers where, due to inhibition of the anti-tumor immune response, their presence is associated with poor prognosis. This crucial role of Tregs makes them an attractive target for therapies against the two extreme ends of a flawed immune system; autoimmunity and cancer. Understanding the epigenetic mechanisms that regulate Treg function, will allow for the development of therapies that can modulate Treg function and thereby can enhance or dampen the immune response.

Methods Ezh2, a histone H3 lysine 27 (H3K27) methyltransferase, is essential for Treg suppression, since loss of Ezh2 leads to Treg instability and improved cancer control. To assess how increasing H3K27me3 modifications affect Treg cell behavior, we generated mice expressing a Treg-specific Ezh2 gain of function mutation, Ezh2Y641F/+. Ezh2Y641F/+ Treg function was assessed by phenotypic analysis and by monitoring disease development in mice expressing Ezh2Y641F/+ Tregs using preclinical models of autoimmunity and cancer. We used Cut & Run for H3K27me3 in wild type (WT) and Ezh2Y641F/+ Tregs to assess which genes are being regulated by H3K27me3 in Tregs.

Results As anticipated for hyperactivated Ezh2, Ezh2Y641F/+ Tregs demonstrated increased global H3K27me3 levels. Functional analysis of Ezh2Y641F/+ Tregs in vivo demonstrated that these Tregs displayed an activated Treg effector phenotype with increased expression of ICOS, CD103, TIGIT, and PD-1. In addition, increased expression of the chemokine receptors CXCR3, CCR8 and CCR6, but decreased expression of CD25, suggested that Ezh2Y641F/+ Treg are primed for homing to organ tissues. In a competitive setting of WT and Ezh2Y641F/+ Tregs, Ezh2Y641F/+ Tregs outcompeted WT Tregs for localization to the site of inflammation in a mouse model of MS (Experimental Autoimmune Encephalomyelitis, EAE). In addition, expression of Ezh2Y641F/+ in Tregs led to more rapid remission from EAE compared to mice with WT Tregs.

Conclusions H3K27me3 regulates Treg function and a gain of function mutation of Ezh2 improves Treg effector function, reducing autoimmunity but potentially promoting cancer. Further exploration of the genes differentially trimethylated in Ezh2Y641F/+ Tregs will increase our understanding of Treg function, allowing for the development of immunotherapies aimed at manipulating the Treg epigenome to treat cancer or autoimmune disease.

Ethics Approval The study was approved by University of California, Berkeley, Ethics Board, approval number AUP-2017–05-9915–2.

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