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

基本信息

批准号：
10580608
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
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-05 至 2025-01-31
项目状态：
未结题

项目摘要

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，通常是渐进的炎症，最常见地影响末端回肠。的超过200个相关的遗传基因座，三个最重要的克罗恩的关联包括NOD2，IL23R和 PTGER4。我们提供了来自回肠组织的单细胞RNASEQ（SCRNASEQ）数据，涉及超过100,000的血液转录组并定义表达炎症单核的治疗难治性患者的一部分吞噬细胞（Inf。MNP）模块。该模块包括炎症巨噬细胞，活化的成纤维细胞，成熟树突状细胞，以及激活的T细胞和产生浆膜的IgG。在AIM 1中，我们将蛋白质验证并通过对回肠组织的Cite-Seq研究来完善炎性单核吞噬细胞（Inf。MNP）模块和外周血。我们将改进现有细胞簇分类，并提供更精细的蛋白质通过Cite-Seq验证，其中定量表面蛋白测量将在相同获得SCRNASEQ数据的单元。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局定义，b）完善相对罕见的关键细胞簇定义细胞（成纤维细胞，树突状细胞），c）改善自适应组织映射的血液（T，B和浆细胞）免疫力，d）精炼PTGER4和IL23R基因表达，假设在治疗非 - 响应（目标3）。在AIM 2中，我们将定义由巨噬细胞薄纤维细胞串扰的早期扰动 NOD2缺陷并建立抗TNF响应机制。 NOD2是Muramyl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的细胞具有表达CD14（血液单核细胞标记）和单个细胞中高水平的胶原蛋白的细胞；这一关键发现证明了新的假设，即功能丧失nod2致病性部分是由失败驱动的分化为住宅巨噬细胞，有利于多能基质型细胞。在AIM 3中，我们试图通过利用IL23R和 PTGER4优先考虑抗TNF难治性患者的细胞和分子打捞机制。尽管明显的抗IL12/23打捞，大量无响应仍然存在。利用这种未满足的医疗需求，我们通过分析，将测试IL23R的免疫细胞与抗IL12/23的临床反应的相关性多个现有和新收集的批量RNA数据集。我们假设炎症性巨噬细胞表达IL23R的细胞交谈介导了对抗IL12/23封锁的反应，但通过致病性通过通过PTGER4异常原位成熟的树突状细胞分化。这些抗IL12/23的比较分析响应者与非反应者，将重点介绍难治性的细胞和途径，以优先考虑目标优先级。该提案结合了三个主要关联信号，尖端的单细胞方法，有了当前未经掌握的医疗领域，需要提高人们对人类克罗恩的机械基础的理解疾病。