Abstract

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an effective therapy for many hematological malignancies, however, it remains to be limited by morbidity and mortality related to graft-versus-host disease (GVHD). During GVHD, donor T cells are activated by host antigen-presenting cells (APCs) and differentiate into effector cells that mediate host tissue damage. Development of alloreactive effector T cells requires orchestrated expression of myriad genes initiated by host APCs. However, little is known about the key epigenetic factor that controls this process. Histone methylation, which is catalyzed by histone methyltransferanses (HMT), has been correlated with genes encoding effector cytokines and transcription factors critical for effector differentiation. We recently demonstrated that inhibition of histone methylation using 3-Deazaneplanocin A (DZNep) arrested ongoing GVHD through induction of apoptosis in alloreactive T cells. However, since DZNep reduced multiple histone methylation marks, including trimethylation of histone H3 at lysine 4 (H3K4me3), H3K27me3, H3K36me3 and H4K20me3, the key HMT that regulates the transcription program important for allogeneic T cell responses remains unclear. Using genetic approaches and mouse GVHD models, we identify that the HMT Ezh2, which specifically catalyzes H3K27me3 and acts primarily as a gene silencer, plays a central role in regulating allogeneic T cell proliferation, differentiation, and function during the GVHD process. Transfer of donor T cells derived from Ezh2 conditional knockout C57BL/6 (B6) mice did not induce GVHD in lethally irradiated allogeneic BALB/C mice, with all recipients surviving free of disease. In contrast, wild-type (WT) donor B6 T cells caused uniformly lethal GVHD in these recipients. We found that loss of Ezh2 selectively impaired alloantigen-specific T cell responses. Three days after allo-HSCT, both Ezh2-deficient and WT T cells were similarly activated to proliferate in vivo. However, 7 days after transplantation, there was a significant reduction in the number of highly proliferative Ezh2-deficient T cells compared to WT T cells, leading to a dramatic reduction of infiltrating Ezh2-deficient T cells in GVHD target organs. Interestingly, conditionally deleting Ezh2 did not affect proliferation and survival of B6 T cells that were transferred into lethally irradiated syngeneic B6/SJL mice by 7 days after transplantation. Thus, Ezh2 is critical for the continual proliferation of alloantigen-responding T cells during later stages of GVHD induction. In contrast, Ezh2 is dispensable for both the initial activation and proliferation of T cells upon alloantigen-priming and homeostatic expansion of T cells in response to lymphopenia. Previous studies suggest that IFN-g-producing alloreactive effector cells cause damage to the liver and gastrointestinal tract. We found that loss of Ezh2 led to selective reduction in frequency of IFN-γ-producing effector cells 7 days after allo-HSCT. This effect was accompanied with decreased expression of transcription factors TBX21 and STAT4, both of which are crucial for effector differentiation. These data suggest that in addition to regulating the transcription program critical for producing sufficient numbers of alloreactive effector T cells needed to mediate GVHD, Ezh2 is also important for orchestrating genes required to promote IFN-γ production in effector cells. Importantly, we found that Ezh2 ablation retained anti-leukemia activity in alloreactive T cells, leading to improved overall survival of transplant recipients. Collectively, these findings identify Ezh2 as a key epigenetic regulator that controls allogeneic T cell responses and GVHD. We propose that pharmacological modulation of Ezh2 using specific inhibitors should be investigated as a novel therapeutic strategy after allo-HSCT.

Disclosures:

No relevant conflicts of interest to declare.

Author notes

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