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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的、典型的进行性炎症，最常影响末端回肠。的在超过200个相关的遗传基因座中，三个最重要的克罗恩病相关性包括NOD 2、IL 23 R和 PTGER 4.我们呈现了来自回肠组织和血液的单细胞RNASeq（scRNASeq）数据，涉及超过100，000例转录组，并确定了一个子集的治疗难治性患者表达的炎性单核细胞吞噬细胞（inf. MNP）模块。该模块包括炎性巨噬细胞，活化的成纤维细胞，成熟的树突细胞，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研究完善炎性单核吞噬细胞（inf. MNP）模块和外周血。我们将改进目前的细胞簇分类，并提供更精细的蛋白质通过CITE-seq进行验证，其中将对相同的表面蛋白进行定量测量。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）针对相对不常见的患者结果定义，细胞（成纤维细胞、树突状细胞），c）改善适应性（T、B和浆细胞）的血液至组织映射免疫，和d）细化PTGER 4和IL 23 R基因表达，假设其在治疗非免疫性疾病中起主要作用。响应（目标3）。在目标2中，我们将定义巨噬细胞-成纤维细胞串扰的早期扰动， NOD 2缺陷和建立抗TNF应答机制。NOD 2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 NOD 2与表达CD 14（血液单核细胞标志物）的细胞和单个细胞中高水平的胶原蛋白的关系; 这一关键发现提示了一个新的假说，即NOD 2致病性功能丧失部分是由于分化成居住的巨噬细胞，有利于更多的多能基质型细胞。目标3：通过利用IL 23 R的细胞特异性基因表达加速精确克罗恩病的进展， PTGER 4优先考虑抗TNF难治性患者的细胞和分子挽救机制。尽管尽管存在明显的抗IL 12/23补救，但仍存在大量无应答。利用这种未满足的医疗需求，我们将通过以下分析测试表达IL 23 R的免疫细胞与抗IL 12/23临床应答的相关性：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IL 23 R的细胞串扰介导对抗IL 12/23阻断的应答，但通过通过PTGER 4的异常原位成熟树突状细胞分化。这些抗IL 12/23的比较分析反应者与无反应者，将突出显示靶向治疗优先考虑的难治细胞和途径优先化。该提案结合了三个主要的关联信号，尖端的单细胞方法，目前尚未满足的医疗需求领域，以促进对人类克罗恩病机制基础的理解，疾病