Ezh2-mediated Epigenetic Effects and Alloimmunity

Ezh2介导的表观遗传效应和同种免疫

• 批准号: 9028856
• 负责人: YI ZHANG
• 金额: $ 62.46万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9028856
• 关键词: Accounting; Acute Graft Versus Host Disease; Affect; Alloantigen; Allogenic; Antigen-Presenting Cells; Apoptosis; Biological Markers; Calcineurin inhibitor; Cell Survival; Cell physiology; Cells; Cessation of life; Clinical; Complex; Complication; Data; Development; Diagnosis; Disease; EZH2 gene; Effector Cell; Epigenetic Process; Experimental Models; Frequencies; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Goals; Hematological Disease; Hematopoietic Stem Cell Transplantation; Histocompatibility; Histone H3; Human; Immune response; Immunity; Injury; Interferon Type II; Lead; Life; Link; Lysine; Maintenance; Measures; Mediating; Memory; Messenger RNA; Methods; Molecular; Molecular Chaperones; Morbidity - disease rate; Mus; Onset of illness; Pathway interactions; Patients; Proteins; Regulation; Resistance; Role; SET Domain; Severities; Signal Transduction; Staging; Stream; Symptoms; T cell differentiation; T cell regulation; T cell response; T-Cell Receptor; T-Lymphocyte; T-bet protein; Testing; Time; Tissues; Transplantation; base; clinically relevant; design; gene repression; graft versus host disease induction; graft vs host disease; histone methyltransferase; improved; improved functioning; inhibitor/antagonist; isoimmunity; leukemia; mortality; mouse model; multicatalytic endopeptidase complex; mutant; novel; novel strategies; outcome forecast; overexpression; predictive marker; prevent; protein expression; protein function; public health relevance; response; targeted agent; therapeutic target; transcription factor

 DESCRIPTION (provided by applicant): Graft-versus-host disease (GVHD) remains a major cause of morbidity and mortality after hematopoietic stem cell transplantation (allo-HSCT), occurring after 30-70% of transplants and accounting for up to 15% of deaths. Acute GVHD is characterized by host tissue injury, mediated by donor T cells following interaction with either donor- or host-derived antigen presenting cells (APCs). The interaction between APCs and alloantigen-responsive donor T-cells leads to changes in abundance and function of transcription factors, which in turn stimulate or repress expression of genes crucial for alloreactive T cell responses and GVHD. Epigenetic changes are thought to be important in regulation of gene expression in alloreactive T-cells, but it remains largely unknown how epigenetic processes help regulate alloreactive T cell responses by affecting gene transcription. Our long- term goal is to identify key epigenetic regulator(s) that can be targeted to modulate GVHD. The objective of this application is to investigate how Ezh2, a histone methyltransferase that acts primarily as a gene silencer, orchestrates the expression of transcription factors criticl for alloreactive T cell responses and identify an optimal pharmacological approach to modulate Ezh2 in GVHD. We previously discovered that genetic inactivation of Ezh2 function reduced GVHD but preserved graft- versus-leukemia (GVL) responses, leading to improved survival of mice with leukemia treated with allo-HSCT. These findings form a central hypothesis that: A) Ezh2 is a master regulator of alloreactive T-cell responses, and B) targeting Ezh2 may lead to novel and clinically relevant strategies to modulate GVHD. Unexpectedly, our preliminary studies showed that newly discovered Ezh2 inhibitors that can selectively reduce H3K27me3, failed to control GVHD in mice. Thus, optimal methods to target Ezh2 remain an unmet need for controlling GVHD. Most recently, we identified that: 1) Ezh2 promoted the expression of transcription factors T-bet (which is essential for development of T helper (Th)1 cells) and Id3 (which is important for effector cell survival and memory cell formation) in alloreactive T cells. Loss of Ezh2 led to selectively impaired expansion and survival of alloreactive T cells producing IFN-γ during late stages of GVHD induction. This activation function of Ezh2 is in contrast to its previously described role as a gene silencer; 2) the Ezh2 SET domain, which contains an enzymatic unit, positively regulated Ezh2 protein stability and function in T cells. Mutant Ezh2 protein that lacked the SET domain was rapidly degraded by proteasomes, and failed to form a stable complex with Hsp90, a chaperone protein that helps stabilize protein expression and function. Pharmacological inhibition of Hsp90 resulted in rapid degradation of Ezh2 protein in T cell receptor-activated T cells and marked decrease of allogeneic T cell responses in GVHD mice. Notably, although Hsp90 has several key signaling intermediates, overexpression of Ezh2 rescued activated T cells from Hsp90 inhibition-induced apoptosis. This indicates that depleting Ezh2 protein by Hsp90 inhibition may represent a novel approach to target Ezh2 in allogeneic T cells; and 3) in humans, EZH2 activity, assessed by levels of its targets, was higher in T cells from allo-HSCT recipients at the time of GVHD onset compared to T cells from patients without GVHD. Our preliminary findings point to the importance of EZH2 in mediating calcineurin inhibitor-resistance in human T cells from aGVHD patients, and this effect can be quantified using flow cytometric analysis. This provides substantial evidence for the relevance of this epigenetic mechanism in humans and strengthens the translational potential of Ezh2 modulation. To further test our central hypothesis in detail, we will determine the critical roles f Id3 and T-bet in mediating Ezh2 regulation of allogeneic T cell responses in mice after allo-HSCT; determine the effects of targeting the Ezh2-Hsp90 complex on GVH responses; and characterize EZH2-mediated T-cell function in human GVHD and establish its utility as a GVHD biomarker. These studies would potentially lead to: A) novel and clinically relevant strategies to target Ezh2 activation function for improving the efficacy of allo-HSCT; and B) improved understanding of the epigenetic effects in alloreactive T cells and its regulation of T cell immunity.

 描述（由申请人提供）：移植物抗宿主病（GVHD）仍然是造血干细胞移植（allo-HSCT）后发病和死亡的主要原因，30-70%的移植后发生，占比高达15%急性 GVHD 的特点是宿主组织损伤，由供体 T 细胞与供体或宿主来源的抗原呈递细胞 (APC) 相互作用介导。 APC 和同种异体抗原反应性供体 T 细胞会导致转录因子的丰度和功能发生变化，进而刺激或抑制对同种异体反应性 T 细胞反应至关重要的基因表达，而表观遗传变化被认为在基因表达的调节中很重要。在同种反应性 T 细胞中，但表观遗传过程如何通过影响基因转录来帮助调节同种反应性 T 细胞反应仍然很大程度上未知。本申请的目的是研究 Ezh2（一种主要充当基因沉默子的组蛋白甲基转移酶）如何协调对同种异体反应性 T 细胞反应至关重要的转录因子的表达，并确定在 GVHD 中调节 Ezh2 的最佳药理学方法。先前发现，Ezh2 功能的基因失活可降低 GVHD，但保留移植物抗白血病 (GVL) 反应，从而提高接受治疗的白血病小鼠的生存率这些发现形成了一个中心假设：A) Ezh2 是同种异体反应性 T 细胞反应的主要调节因子，B) 靶向 Ezh2 可能会导致新的临床相关策略来调节 GVHD。新发现的可选择性降低 H3K27me3 的 Ezh2 抑制剂未能控制小鼠 GVHD，因此，针对 Ezh2 的最佳控制方法仍然是一个未得到满足的需求。最近，我们发现：1) Ezh2 促进转录因子 T-bet（对于 T 辅助细胞 (Th)1 细胞的发育至关重要）和 Id3（对于效应细胞存活和记忆细胞形成很重要）的表达。在同种反应性 T 细胞中，Ezh2 的缺失会导致在 GVHD 诱导的后期产生 IFN-γ 的同种反应性 T 细胞的增殖和存活选择性受损。 先前描述的作为基因沉默子的作用；2) Ezh2 SET结构域包含一个酶单元，在T细胞中正向调节Ezh2蛋白的稳定性和功能，缺乏SET结构域的突变Ezh2蛋白被蛋白酶体快速降解，并且无法形成。与 Hsp90 的稳定复合物，Hsp90 是一种伴侣蛋白，有助于稳定蛋白表达和功能。Hsp90 的药理学抑制导致 T 细胞中 Ezh2 蛋白快速降解。 GVHD 小鼠中受体激活的 T 细胞和同种异体 T 细胞反应显着降低 值得注意的是，尽管 Hsp90 具有几个关键的信号传导中间体，但 Ezh2 的过度表达可将激活的 T 细胞从 Hsp90 抑制诱导的细胞凋亡中拯救出来。可能代表了一种在同种异体 T 细胞中靶向 Ezh2 的新方法；3) 在人类中，根据其靶标水平评估，EZH2 活性更高与来自未发生 GVHD 的患者的 T 细胞相比，EZH2 在介导来自 aGVHD 患者的人类 T 细胞中钙调神经磷酸酶抑制剂耐药性方面发挥着重要作用。使用流式细胞术分析进行量化，这为这种表观遗传机制在人类中的相关性提供了实质性证据，并增强了 Ezh2 调节的转化潜力。我们将确定 f Id3 和 T-bet 在介导 Ezh2 对异体 HSCT 后小鼠同种异体 T 细胞反应的调节中的关键作用；确定靶向 Ezh2-Hsp90 复合物对 GVH 反应的影响；并表征 EZH2 介导的 T 细胞；这些研究可能会导致：A）针对 Ezh2 激活功能的新颖且临床相关的策略，以提高 Ezh2 的功效。同种异体造血干细胞移植；B) 提高了对同种异体反应性 T 细胞的表观遗传效应及其对 T 细胞免疫的调节的了解。