Project 1

项目1

基本信息

批准号：
10625509
负责人：
Anuradha Ray
金额：
$ 53.43万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-06-01 至 2027-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10625509
关键词：
Address Adrenal Cortex Hormones Aftercare Algorithms Aspergillus Asthma Bioinformatics Biological Biological Markers Biological Products Bronchoalveolar Lavage CD8-Positive T-Lymphocytes CD8B1 gene CXCL10 gene Cell Aging Cells Cellular Indexing of Transcriptomes and Epitopes by Sequencing Chemotactic Factors Clinic Complex Cytometry Data Dinucleoside Phosphates Disease Enrollment Epithelial Cells Epithelium Event Experimental Models Flow Cytometry Funding Gene Expression Gene Expression Profile Genes Glucocorticoid Receptor Goals Grant Hematopoiesis Heterogeneity Human IL7 gene Immune Immune System Diseases Immune response Immunologics In Situ Innate Immune Response Interferon Type II Interleukin-10 Interleukin-4 Interleukin-5 Lymphocyte Memory Methods Modeling Molecular Molecular Abnormality Mus Outcome Measure Pathogenesis Pathway interactions Patient Selection Patients Peptide Hydrolases Periodicity Phenotype Population Production Qualifying Refractory Regulation Reporting Research Resolution Role STAT1 gene Signal Transduction Steroids T cell receptor repertoire sequencing T-Lymphocyte Time Tissues Work adaptive immune response airway epithelium airway hyperresponsiveness airway inflammation asthmatic airway asthmatic patient cell type cohort cytokine disease phenotype high dimensionality insight mouse model mucosal site novel participant enrollment profiles in patients programs response symptomatic improvement targeted treatment therapeutic development transcriptome transcriptome sequencing transcriptomics treatment response

项目摘要

Despite many advancements in the treatment of severe asthma (SA) with the advent of biologics, there still remain challenges because of incomplete understanding of the underlying immune and molecular aberrations. Towards this end, we analyzed the bronchoalveolar lavage (BAL) cells of patients enrolled in the severe asthma research program (SARP) and also of those in the P01-funded IMSA (Immune Mechanisms in Severe Asthma) program. In addition, we studied the transcriptome of these cells using bulk RNA-seq methods. Initial studies of the SARP cohort revealed that the BAL cells in 35-40% of SA patients harbor a high Type 1 (T1)/IFN-g immune signature and implicated IFN-g-activated STAT1 in steroid insensitivity due to co-operation with the glucocorticoid receptor (GR). Subjecting the BAL cells of 41 subjects in the IMSA cohort to mass cytometry/CyTOF and downstream bioinformatic analysis allowed clustering of the SA patients into two groups (PGs), PG2 and PG3, displaying distinct immune profiles. The PG2 group showed a heightened innate immune response dominated by FceRI+IL-4+ cells while PG3 was a lymphocyte-dominated group characterized by increased numbers of IFN-g+ CD4+ and CD8+ T cells, which included tissue-resident memory (TRM) cells. The immune cells were associated with gene modules by deconvolution of their transcriptomic data using our novel algorithm, ICLite. Module-associated gene expression suggests importance of FceRI- and IL-7-signaling in cytokine production by innate cells in PG2 and of co-stimulatory molecules in positive regulation of IFN-g production from TRMs in PG3. In the clinic, response to treatment of SA patients with the biologic, dupilumab, also highlights heterogeneity despite similar biomarker profiles of the treated patients. Thus, given the novel insight gained about distinct immune phenotypes of SA patients with specific transcriptomic signatures, we now have the opportunity to determine in fine resolution, immunologically and molecularly, how dupilumab differentially impacts the local immune (Proj. 1) and epithelial (Proj. 2) phenotypes in patients who show similar biomarker profiles and yet do not respond similarly to treatment. Collectively, these data allow us to hypothesize that two distinct immune mechanisms, one regulated by FceRI+ innate immune cells in conjunction with airway epithelial cells, and the second by T cells, with their impact on epithelial cells, collectively determines SA. To address this hypothesis, the specific aims of Project 1 are to: Aim 1. Characterize airway immune cells in high-resolution by CITE-seq and TCR-seq and determine impact of dupilumab on immune phenotype. Aim 2. Determine mechanisms that induce innate immune cell-driven SA phenotype using a novel fungal protease-driven model. Aim 3. Determine mechanisms that regulate a T1high adaptive immune response using an established HDM+cyclic dinucleotide-driven model. Collectively, the findings will qualify heterogeneity in immune response and response to therapy at the single cell level and establish key molecules/pathways that drive distinct immune phenotypes using novel experimental models.

尽管随着生物制剂的出现，严重哮喘 (SA) 的治疗取得了许多进展，但仍然存在一些问题由于对潜在的免疫和分子畸变的不完全了解，这仍然是挑战。为此，我们分析了重症患者的支气管肺泡灌洗（BAL）细胞。哮喘研究计划 (SARP) 以及 P01 资助的 IMSA（严重哮喘的免疫机制）哮喘）计划。此外，我们使用批量 RNA 测序方法研究了这些细胞的转录组。最初的 SARP 队列研究表明，35-40% SA 患者的 BAL 细胞具有较高的 1 型 (T1)/IFN-g 免疫特征和 IFN-g 激活的 STAT1 与合作导致的类固醇不敏感性有关与糖皮质激素受体（GR）。对 IMSA 队列中 41 名受试者的 BAL 细胞进行质量检查细胞计数/CyTOF 和下游生物信息分析将 SA 患者分为两组 (PGs)、PG2 和 PG3，显示出不同的免疫特征。 PG2组表现出更高的先天性免疫反应以FceRI+IL-4+细胞为主，PG3以淋巴细胞为主其特点是 IFN-g+ CD4+ 和 CD8+ T 细胞数量增加，其中包括组织驻留记忆（TRM）细胞。免疫细胞通过转录组的解卷积与基因模块相关联使用我们的新颖算法 ICLite 的数据。模块相关基因表达表明 FceRI- 和 PG2 中先天细胞和阳性共刺激分子产生细胞因子中的 IL-7 信号传导 PG3 中 TRM 产生 IFN-g 的调节。在临床上，SA 患者对治疗的反应尽管接受治疗的患者的生物标志物特征相似，但生物制剂 dupilumab 也凸显了异质性。因此，鉴于对具有特定特征的 SA 患者的不同免疫表型有了新的认识，转录组特征，我们现在有机会以高分辨率、免疫学和从分子角度看，dupilumab 如何对局部免疫（项目 1）和上皮细胞（项目 2）表型产生不同影响在表现出相似生物标志物特征但对治疗反应不相似的患者中。总的来说，这些数据使我们能够假设两种不同的免疫机制，一种由 FceRI+ 先天调节免疫细胞与气道上皮细胞结合，第二个由 T 细胞结合，其对上皮细胞共同决定SA。为了解决这一假设，项目 1 的具体目标是：目标 1. 通过 CITE-seq 和 TCR-seq 高分辨率表征气道免疫细胞并确定影响 dupilumab 对免疫表型的影响。目标 2. 确定诱导先天免疫细胞驱动的 SA 的机制使用新型真菌蛋白酶驱动模型进行表型分析。目标 3. 确定调节 T1high 的机制使用已建立的 HDM+环二核苷酸驱动模型进行适应性免疫反应。总的来说，研究结果将证明单细胞水平上免疫反应和治疗反应的异质性，使用新颖的实验模型建立驱动不同免疫表型的关键分子/途径。