Synthetic rescue of antigen-driven T cells and alloimmunity

抗原驱动T细胞和同种免疫的综合拯救

• 批准号: 10543445
• 负责人: Jonathan A Soboloff
• 金额: $ 63.76万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-12 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10543445
• 关键词: Allogenic; Antigens; Artemisinins; Autoimmune Diseases; Automobile Driving; CD8B1 gene; Ca(2+)-Transporting ATPase; Calcineurin inhibitor; Cell Death; Cell Differentiation process; Cell Survival; Cell membrane; Cellular Assay; Cellular biology; Clinical; Cyclosporine; Cytosol; DNA Synthesis Inhibitors; Development; Differentiation Antigens; Disease; Endoplasmic Reticulum; Epigenetic Process; Gatekeeping; Generations; Goals; Homeostasis; Human; ITPR1 gene; Immunity; Immunotherapy; Impairment; Inbred BALB C Mice; Inflammatory; Investigation; Libraries; Ligation; Malaria; Malignant Neoplasms; Mediating; Methods; Mitochondria; Molecular; Mus; Na(+)-K(+)-Exchanging ATPase; Natural Compound; Outcome; Pathogenesis; Pathway interactions; Patients; Production; Proteins; Pump; Reaction; Regimen; Repression; Role; STIM1 gene; Signal Transduction; T cell response; T memory cell; T-Cell Activation; T-Cell Proliferation; T-Cell Receptor; T-Lymphocyte; Tacrolimus; Testing; Therapeutic; Toxic effect; Translating; Water; artesunate; calcium uniporter; chronic infection; disorder prevention; effector T cell; experimental study; graft vs host disease; graft vs leukemia effect; histone methyltransferase; improved; insight; interest; isoimmunity; mitochondrial metabolism; novel; novel strategies; nuclear factors of activated T-cells; pharmacologic; preservation; prophylactic; response; sensor; tissue injury; uptake

During graft-versus-host disease (GVHD), donor T cells require the histone methyltransferase Ezh2 for producing and sustaining effector T cells that mediate host tissue injury. We recently established that Ezh2 serves as a molecular gatekeeper for the generation of CD8 memory T cell precursors in GVHD, critical for the production of effector T cells in response to persistent antigen (Nat Commun 2017). However, our efforts to develop novel approaches to selectively target alloreactive effector T cells has been limited by the lack of understanding of why Ezh2 loss causes cell death of antigen-activated T cells. Stromal interaction molecule (Stim) proteins, Stim1 and Stim2, are crucial dynamic endoplasmic reticulum (ER) Ca2+ sensors and modulators of Ca2+ signals. Upon T cell receptor (TCR) ligation, Stim1 activation causes its translocation towards the plasma membrane, where it activates the Ca2+ channel Orai1, facilitating Ca2+ entry and driving T cell activation. Conditional Stim1 deletion inhibits GVHD in mice due to impaired effector differentiation. Remarkably, Stim1 deletion rescues antigen-activated Ezh2-null T cells, leading to restored production of alloreactive effector T cells in mice and severe GVHD. Therefore, we hypothesize that: A) Ezh2 and Stim1 operate coordinately to regulate the viability and function of antigen-driven T cells; and B) Ezh2/Stim1- regulated molecular pathway(s) are crucial for controlling alloreactive T cell-mediated GVHD. We further establish that the role of Stim1 in Ezh2-mediated cell death is to drive mitochondrial Ca2+ (mitoCa2+) overload since conditional deletion of the mitochondrial calcium uniporter (MCU), leads to rescue of antigen-activated Ezh2-null T cells. To establish the therapeutic potential of these findings, we performed a preliminary screen with an 800 compound library, finding 36 compounds that block T cell proliferation. Amongst them was artesunate (ART), a water-soluble derivative of artemisinin clinically approved for the treatment of malaria and known to target the Sarco/Endoplasmic Reticulum Ca2+ ATPase (SERCA), which pumps Ca2+ from the cytosol to the ER lumen. SERCA inhibition leads to Stim1/Orai1 activation and mitoCa2+ uptake. Preliminary investigations show that ART treatment reduces GVHD in BALB/C mice receiving allogeneic C57BL/6 T cells. Considered collectively, these findings suggest that Ezh2 regulates antigen-specific effector T cell survival through modulation cytosolic Ca2+ entry, thereby limiting mitochondrial Ca2+ loading and protecting against cell death. This hypothesis will be tested through three specific aims. In Aim-1, we will define the mechanisms that regulate the survival and differentiation of antigen-driven Ezh2/Stim1-null T cells. Aim-2 will determine the molecular mechanisms by which Ezh2 deficiency dysregulates cytosolic and mitochondria Ca2+ uptake in activated T cells. Finally, Aim-3 will examine the beneficial effect of enhancing T cell Ca2+ load to modulate GVHD and GVL activity in mice. Completion of these experiments will provide novel insights into T cell biology, T cell-mediated inflammatory disorders such as GVHD and autoimmune diseases, and lead to development of novel methods for improving the efficacy of immunotherapy for chronic infections and cancer.

在移植物抗宿主病(GVHD)期间，供者T细胞需要组蛋白 甲基转移酶Ezh2在产生和维持介导宿主组织的效应T细胞中的作用 受伤。我们最近证实，Ezh2作为一种分子守门人，用于产生 移植物抗宿主病中CD8记忆T细胞前体对产生效应T细胞至关重要 到持久性抗原(NAT Commun 2017)。然而，我们努力开发新的方法来 选择性靶向同种异体反应性T细胞因缺乏 了解Ezh2缺失导致抗原激活T细胞死亡的原因。基质相互作用 STIM分子蛋白STIM1和STIM2是至关重要的动态内质网(ER) 钙离子传感器和钙信号调制器。在T细胞受体(TCR)结扎后， STIM1的激活导致它向质膜移位，在那里它激活了 钙通道Orai1，促进钙离子进入并驱动T细胞活化。条件STIM1 缺失可抑制小鼠的移植物抗宿主病，原因是效应器分化受损。值得注意的是， STIM1缺失拯救抗原激活的Ezh2零T细胞，导致恢复 同种异体反应性T细胞在小鼠和严重移植物抗宿主病中的产生因此，我们假设 A)Ezh2和STIM1协调运作，以调节 B)Ezh2/STIM1调节的分子通路(S)在 控制同种异体反应性T细胞介导的GVHD。我们进一步确定STIM1的作用 在Ezh2介导的细胞死亡是驱动线粒体钙超载自条件性 线粒体钙单转运体(MCU)的缺失，导致抗原激活的拯救 EZH2-空T细胞。为了确定这些发现的治疗潜力，我们进行了一项 用800个化合物文库进行初步筛选，发现36种化合物可以阻断T细胞 扩散。其中包括青蒿琥酯(ART)，一种青蒿素的水溶性衍生物 临床上被批准用于治疗疟疾，并已知靶向Sarco/内质 网状Ca~(2+)-ATPase(SERCA)，将胞浆中的Ca~(2+)泵入内质网。SERCA 抑制导致STIM1/Orai1激活和线粒体钙摄取。初步 研究表明，ART治疗可降低接受同种异体C57BL/6 T的BALB/C小鼠的GVHD 细胞。综合考虑，这些发现表明Ezh2调节抗原特异性。 效应器T细胞通过调节胞浆内钙离子内流而存活，从而限制 线粒体Ca~(2+)负荷和保护细胞死亡。这一假设将得到检验。 通过三个具体目标。在AIM-1中，我们将定义调节生存的机制 抗原驱动的Ezh2/STIM1缺失T细胞的分化。AIM-2将决定 Ezh2缺乏症调节胞浆和线粒体的分子机制 活化T细胞的钙摄取。最后，AIM-3将检查增强 T细胞钙负荷调节小鼠移植物抗宿主病和移植物抗宿主病活性这些实验的完成将 提供对T细胞生物学、T细胞介导的炎症性疾病等方面的新见解 GVHD和自身免疫性疾病，并导致开发新的方法来改善 免疫治疗对慢性感染和癌症的疗效。