Differentiation of pathogenic granzyme A-producing CD4+ T cells in graft-versus-host-disease

移植物抗宿主病中致病性颗粒酶 A 产生的 CD4 T 细胞的分化

• 批准号: 10664183
• 负责人: Matthew R Olson
• 金额: $ 34万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-10 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10664183
• 关键词: Acute Graft Versus Host Disease; Anti-Cytokine Therapy; Binding; CD4 Positive T Lymphocytes; CRISPR screen; Cell Differentiation process; Cell Lineage; Cell surface; Cells; Chromatin; Clinical; Cytokine Signaling; Data; Development; Disease; Disease model; Donor person; GZMA gene; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Goals; Granzyme; Helper-Inducer T-Lymphocyte; Hematopoietic Neoplasms; Human; Immune; Individual; Inflammation; Inflammatory; Interferon Type II; Interleukin-2; Interleukin-4; Interleukin-6; Intestinal Diseases; Intestines; Laboratories; Lead; Maps; Mediating; Molecular; Mus; Organ; Outcome; Pathogenicity; Pathology; Pathway interactions; Patients; Population; Pre-Clinical Model; Process; Regulation; Reporter; Role; STAT1 gene; STAT3 gene; STAT6 gene; Serine Protease; Serum; Severity of illness; Signal Pathway; Signal Transduction; Signaling Protein; Source; Stat5 protein; Syndrome; Technology; Testing; Tissues; Work; anti-cancer; blood treatment; clinical prognosis; combinatorial; cytokine; druggable target; experimental study; genetic signature; genome-wide; graft vs host disease; hematopoietic cell transplantation; improved; in vivo; inhibitor; innovation; insight; mortality; new therapeutic target; novel; novel therapeutics; prevent; programs; promoter; response; targeted treatment; therapeutic target; transcription factor; transcriptome sequencing; virtual

PROJECT SUMMARY/ABSTRACT. In graft-versus-host disease (GVHD), a detrimental allo-immune response that occurs after hematopoietic cell transplant (HCT), donor CD4+ T helper (Th) cells traffic to multiple organs and their presence in the intestines correlates with worsened clinical prognosis. Despite their known role in GVHD, how Th cells contribute to intestinal disease remains unclear. My laboratory has recently identified a novel population of intestinal Th cells that produced the serine protease granzyme A (GrA) which were critical for intestinal damage and GVHD-associated mortality. However, GrA+ Th cells were dispensable for the beneficial anti-cancer effects of HCT and therefore represent a promising therapeutic target in GVHD patients. As effective GrA inhibitors are currently unavailable, defining the molecular mechanisms that lead to GrA+ Th cell differentiation will uncover novel drug targets for GVHD and provide insight into how pathogenic Th cells differentiate within the intestines. Here we show that GrA+ Th cells differentiate independently from other Th lineages and require the integration of both STAT3-dependent and -independent cytokine signaling pathways for their optimal differentiation during disease. GrA+ Th cells expressed high levels of the STAT3- induced transcription factors (TFs) BATF and Aiolos that were also required for their development. However, the STAT-activating cytokines that initiate GrA+ Th cell differentiation and how STAT3-induced TFs guide this process during GVHD remain undefined. As STAT3 is required for optimal GrA+ Th cell differentiation, we hypothesize that the STAT3-activating cytokines IL-6 and IL-21 function in parallel with other cytokine/STAT pathways to initiate GrA+ Th cell differentiation and that a STAT3/BATF/Aiolos network cooperatively promotes the GrA+ Th cell lineage during aGVHD. To test this hypothesis we will 1) Define the cytokine/STAT signaling requirements that regulate pathogenic GrA+ Th cell differentiation during GVHD and 2) We will determine how STAT3/BATF/Aiolos TF network and its downstream effector genes regulate the GrA+ Th cell differentiation program using combined gene expression, chromatin occupancy analysis and a novel genome-wide CRISPR screen. Upon completion of these studies we will have defined the inflammatory and transcription factor requirements that are required for the development of GrA+ Th cells in GVHD and validated these factors as therapeutic targets in pre-clinical models of disease. This work is highly significant as GrA+ Th cells are a major driver of intestinal pathology and lethal disease. Therefore, the signaling pathways and genes involved in the GrA+ Th cell transcriptional signature that we identify may be potential druggable targets for GVHD patients. These experiments are also highly innovative as we will leverage a novel Gzma-GFP reporter mouse with cutting-edge sequencing technologies (RNA-Seq, CUT&RUN) to uncover how a STAT3-driven TF network drives pathogenic Th cell differentiation outside of the traditional Th17/T follicular helper cell paradigm.

项目摘要/摘要。在移植物抗宿主病（GVHD）中，有害的同种免疫 造血细胞移植 (HCT) 后发生的反应，供体 CD4+ T 辅助性 (Th) 细胞转运至 多个器官及其在肠道中的存在与临床预后恶化相关。尽管他们的 虽然 Th 细胞在 GVHD 中的作用已知，但 Th 细胞如何导致肠道疾病仍不清楚。我的实验室最近 鉴定出一种新的肠道 Th 细胞群，可产生丝氨酸蛋白酶颗粒酶 A (GrA)， 对于肠道损伤和 GVHD 相关死亡率至关重要。然而，GrA+ Th细胞可有可无 HCT 具有有益的抗癌作用，因此是 GVHD 的一个有前途的治疗靶点 患者。由于目前尚无有效的 GrA 抑制剂，因此需要确定导致 GrA+ Th 细胞分化将发现 GVHD 的新药物靶点，并深入了解 GVHD 的致病机制 Th 细胞在肠道内分化。在这里，我们表明 GrA+ Th 细胞独立分化 其他 Th 谱系，需要整合 STAT3 依赖性和非依赖性细胞因子信号传导 疾病期间它们最佳分化的途径。 GrA+ Th 细胞表达高水平的 STAT3- 诱导转录因子 (TF) BATF 和 Aiolos 也是其发育所必需的。然而， 启动 GrA+ Th 细胞分化的 STAT 激活细胞因子以及 STAT3 诱导的 TF 如何指导这一过程 GVHD 期间的过程仍未明确。由于 STAT3 是最佳 GrA+ Th 细胞分化所必需的，我们 假设 STAT3 激活细胞因子 IL-6 和 IL-21 与其他细胞因子/STAT 并行发挥作用 STAT3/BATF/Aiolos 网络协同促进启动 GrA+ Th 细胞分化的途径 aGVHD 期间的 GrA+ Th 细胞谱系。为了检验这个假设，我们将 1) 定义细胞因子/STAT 信号传导 GVHD 期间调节致病性 GrA+ Th 细胞分化的要求以及 2) 我们将确定如何 STAT3/BATF/Aiolos TF网络及其下游效应基因调控GrA+ Th细胞分化 使用组合基因表达、染色质占据分析和新型全基因组 CRISPR 的程序 屏幕。完成这些研究后，我们将定义炎症和转录因子 GVHD 中 GrA+ Th 细胞发育所需的要求，并验证了这些因素： 疾病临床前模型中的治疗靶点。这项工作非常重要，因为 GrA+ Th 细胞是主要的 肠道病理和致命疾病的驱动因素。因此，参与该过程的信号通路和基因 我们确定的 GrA+ Th 细胞转录特征可能是 GVHD 患者的潜在药物靶标。 这些实验也具有高度创新性，因为我们将利用新型 Gzma-GFP 报告小鼠 尖端测序技术（RNA-Seq、CUT&RUN）揭示 STAT3 驱动的 TF 网络如何 驱动致病性 Th 细胞在传统 Th17/T 滤泡辅助细胞范式之外分化。