Intestinal single cell analyses and population differences in innate immunity in Crohn's disease drive treatment response and clinical heterogeneity: towards Precision IBD

肠道单细胞分析和克罗恩病先天免疫的群体差异驱动治疗反应和临床异质性：迈向精准 IBD

• 批准号: 9893616
• 负责人: JUDY H. CHO
• 金额: $ 73.6万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-05 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893616
• 关键词: Acetylmuramyl-Alanyl-Isoglutamine; Affect; Aftercare; Agonist; Antibodies; Area; Automobile Driving; B-Lymphocytes; Biological Markers; Biopsy; Blood; CD14 gene; Cell Differentiation process; Cells; Chronic; Classification; Clinical; Clinical Protocols; Collagen; Comparative Study; Crohn' s disease; Cytometry; Data; Data Set; Dendritic Cells; Development; Diagnosis; Disease; Distal part of ileum; Dose; Epithelial Cells; Failure; Fibroblasts; Foundations; Gene Expression; Genetic; Genetic study; Human; Immune; Immunity; Immunoglobulin G; In Situ; Individual; Inflammation; Inflammatory; Interleukin-12; Intestines; Libraries; Measurement; Mediating; Medical; Membrane Proteins; Molecular; Monoclonal Antibodies; Mononuclear; Natural Immunity; Pathogenicity; Pathway interactions; Patient-Focused Outcomes; Patients; Peptidoglycan; Phagocytes; Plasma Cells; Plasma Proteins; Plasmablast; Play; Population; Prostaglandin E Receptor; Proteins; RNA; Recurrence; Refractory; Refractory Disease; Role; Sampling; Signal Transduction; T-Lymphocyte; TNF gene; Testing; Time; Tissues; Validation; Zebrafish; base; biomedical referral center; cell type; chemokine; clinical heterogeneity; clinical practice; comparative; effective therapy; genomic locus; high risk; improved; in vivo; insight; loss of function; macrophage; monocyte; new therapeutic target; novel; peripheral blood; precision medicine; real world application; receptor; response; sensor; single cell analysis; single-cell RNA sequencing; sound; tool; transcriptome; transcriptome sequencing; treatment response

Crohn's disease is a chronic, typically progressive inflammation most commonly affecting the terminal ileum. Of the over 200 associated genetic loci, the three most significant Crohn's associations include NOD2, IL23R and PTGER4. We present single cell RNASeq (scRNASeq) data from ileal tissues and blood involving over 100,000 transcriptomes and define a subset of treatment refractory patients expressing an inflammatory mononuclear phagocyte (inf. MNP) module. This module includes inflammatory macrophages, activated fibroblasts, mature dendritic cells, as well as activated T cells and IgG producing plasmablasts. In Aim 1, we will protein validate and refine the inflammatory mononuclear phagocyte (inf. MNP) module through CITE-seq studies of ileal tissues and peripheral blood. We will improve on present cell cluster classifications and provide a more fine-scale protein validation through CITE-seq, where quantitative surface protein measurements will be performed on the same cells for which scRNASeq data are obtained. By so doing, these new CITE-seq studies will a) refine and validate transcriptome-based patient outcome definitions, b) refine key cell cluster definitions for relatively uncommon cells (fibroblasts, dendritic cells), c) improve blood to tissue mappings of adaptive (T, B, and plasma cells) immunity, and d) refine PTGER4 and IL23R gene expression, hypothesized to play major roles in treatment non- response (Aim 3). In Aim 2, we will define early perturbations in macrophage-fibroblast cross-talk driven by NOD2-deficiency and establish anti-TNF responsive mechanisms. NOD2 is an intracellular sensor of muramyl dipeptide (MDP), the minimal bioactive component of bacterial peptidoglycan. We have observed co-expression of NOD2 with cells expressing CD14 (blood monocyte marker) and high levels of collagens in individual cells; this key finding informs the novel hypothesis that loss-of-function NOD2 pathogenicity is partly driven by a failure to differentiate into residential macrophages, favoring more pluripotent stromal-type cells. In Aim 3, we seek to accelerate progress towards precision Crohn's disease by leveraging cell-specific gene expression of IL23R and PTGER4 to prioritize cellular and molecular salvage mechanisms in anti-TNF refractory patients. Despite appreciable anti-IL12/23 salvage, substantial non-response remains. Leveraging this unmet medical need, we will test for correlation of IL23R-expressing immune cells to anti-IL12/23 clinical response through analyses of multiple existing and newly-collected bulk RNA datasets. We hypothesize that inflammatory macrophage to IL23R-expressing cell cross talk mediates response to anti-IL12/23 blockade, but leaves pathogenicity via aberrant in situ mature dendritic cell differentiation via PTGER4. These comparative analysis of anti-IL12/23 responders vs. non-responders, will highlight refractory cells and pathways to be prioritized for target prioritization. This proposal combines the three major association signals, cutting-edge single cell approaches, with current areas of unmet medical needs to advance understanding of the mechanistic basis for human Crohn's disease.

克罗恩病是一种慢性、典型进行性炎症，最常影响回肠末端。的 在 200 多个相关基因位点中，三个最重要的克罗恩病关联包括 NOD2、IL23R 和 PTGER4。我们提供来自回肠组织和血液的单细胞 RNASeq (scRNASeq) 数据，涉及超过 100,000 转录组并定义表达炎症单核细胞的难治性患者的子集 吞噬细胞（inf.MNP）模块。该模块包括炎症巨噬细胞、活化成纤维细胞、成熟细胞 树突状细胞，以及活化的 T 细胞和产生 IgG 的浆母细胞。在目标 1 中，我们将验证蛋白质 并通过回肠组织的 CITE-seq 研究完善炎症单核吞噬细胞 (inf. MNP) 模块 和外周血。我们将改进现有的细胞簇分类并提供更精细的蛋白质 通过 CITE-seq 进行验证，其中将在相同的表面蛋白上进行定量表面蛋白测量 获得 scRNASeq 数据的细胞。通过这样做，这些新的 CITE-seq 研究将 a) 完善和验证 基于转录组的患者结果定义，b) 完善相对不常见的关键细胞簇定义 细胞（成纤维细胞、树突状细胞），c) 改善血液与组织的适应性映射（T、B 和浆细胞） 免疫，d) 改善 PTGER4 和 IL23R 基因表达，假设它们在非免疫治疗中发挥重要作用。 响应（目标 3）。在目标 2 中，我们将定义由以下因素驱动的巨噬细胞-成纤维细胞串扰的早期扰动 NOD2 缺陷并建立抗 TNF 反应机制。 NOD2 是胞壁酰的细胞内传感器 二肽（MDP），细菌肽聚糖的最小生物活性成分。我们观察到共表达 NOD2 与表达 CD14（血液单核细胞标记物）的细胞和单个细胞中高水平的胶原蛋白； 这一关键发现为新的假设提供了依据，即 NOD2 功能丧失致病性部分是由失败引起的 分化为居住型巨噬细胞，有利于更多的多能基质型细胞。在目标 3 中，我们寻求 通过利用 IL23R 的细胞特异性基因表达加速精准克罗恩病的进展 PTGER4 优先考虑抗 TNF 难治性患者的细胞和分子挽救机制。尽管 抗 IL12/23 挽救明显，但仍存在大量无反应。利用这一未满足的医疗需求，我们 将通过分析来测试表达 IL23R 的免疫细胞与抗 IL12/23 临床反应的相关性 多个现有和新收集的批量 RNA 数据集。我们假设炎症巨噬细胞 表达 IL23R 的细胞串扰介导对抗 IL12/23 阻断的反应，但通过以下途径留下致病性： 通过 PTGER4 异常原位成熟树突状细胞分化。这些抗IL12/23的比较分析 响应者与非响应者，将突出显示难治性细胞和目标优先考虑的途径 优先顺序。该提案结合了三大关联信号、尖端的单细胞方法、 结合当前未满足的医疗需求领域，以增进对人类克罗恩病机制基础的了解 疾病。