Elucidating mechanisms of fibrosis associated with the Crohns disease-associated pathogenic variant in the metal transporter ZIP8

阐明与克罗恩病相关的金属转运蛋白 ZIP8 致病性变异相关的纤维化机制

• 批准号: 10571146
• 负责人: Joanna Miller Peloquin Melia
• 金额: $ 12.28万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10571146
• 关键词: Affect; American; Anti-Tumor Necrosis Factor Therapy; Biological Assay; Bone Marrow; CRISPR/Cas technology; Cells; Clinical; Colitis; Collagen; Colon; Crohn' s disease; Data; Disease; Exhibits; Family; Fibroblasts; Fibrosis; Future; Genetic; Genotype; Goals; Homeostasis; Human; Human Genome; Hydroxyproline; Immune; Impairment; Inflammatory; Inflammatory Bowel Diseases; Innate Immune Response; Interleukin-11; Interleukin-6; Intestinal Fibrosis; Knock-in; Knock-in Mouse; Link; Macrophage; Macrophage Activation; Manganese; Measures; Mediating; Metals; Methods; Micronutrients; Modeling; Mucous Membrane; Mus; Operative Surgical Procedures; Orthologous Gene; PTPRC gene; Pathogenesis; Pathogenicity; Pathology; Patients; Penetration; Phenotype; Population; Positioning Attribute; Predisposing Factor; Predisposition; Prevention; Process; Production; Recovery; Refractory; Refractory Disease; Regulation; Reporting; Risk; Role; STAT3 gene; Signal Pathway; Signal Transduction; Sodium Dextran Sulfate; Stimulator of Interferon Genes; Stromal Cells; TNF gene; Tissues; Ulcerative Colitis; Variant; Western Blotting; Work; Zinc; chemokine; clinical practice; cytokine; dextran sulfate sodium induced colitis; differential expression; experience; human data; human disease; insight; member; mouse model; novel; overexpression; pre-clinical; profibrotic cytokine; programs; response; risk variant; single-cell RNA sequencing; therapeutic target; transcriptome sequencing; translational model; treatment response

PROJECT ABSTRACT Inflammatory bowel diseases (IBD), including Crohn’s disease (CD) and ulcerative colitis, affect 1 in 200 Americans. Anti-tumor necrosis factor (TNF)-alpha therapies remain the cornerstone of therapy for patients with IBD, but approximately 40% of patients will experience primary or secondary loss of response, and “anti- TNF-experienced” patients are less likely to respond to all subsequent therapies in current clinical practice. Single-cell RNA sequencing has uncovered key cellular signatures associated with anti-TNF refractoriness that strongly implicate signaling between inflammatory macrophages and activated fibroblasts. There remains a major gap in our understanding of the patient-specific factors that predispose to this aberrant macrophage- fibroblast activation, how it links to anti-TNF unresponsiveness, and the potential to modulate it to change to disease course and treatment response. Our work has focused on the functional implications of a pathogenic variant in a metal transporter, ZIP8 A391T, that dysregulates manganese homeostasis and is associated with complicated (stricturing and penetrating) Crohn’s disease. In studying ZIP8 391-Thr in a mouse model (Zip8 393T-knock-in (KI)), we have shown increased susceptibility to colitis, but even more interestingly, we have observed enhanced fibrosis. These mice also exhibit marked induction of Il-11, a pro-inflammatory and pro- fibrotic cytokine that is a hallmark of aberrant signaling between inflammatory macrophages and activated fibroblasts in patients with Crohn’s disease – particularly anti-TNF refractory disease. We therefore hypothesize that study of aberrant macrophage-fibroblast signaling is a key feature of disease pathogenesis in the Zip8 393T-KI mice. Establishing the underlying disease mechanisms are important because (1) up to 25% of patients with Crohn’s disease carry ZIP8 391-Thr in some populations and (2) the Zip8 393T-KI could serve as a novel translational model for mechanistic studies, particularly related to anti-TNF non-response. We will study this hypothesis in two aims: In Aim 1, we will determine if Zip8 393T-KI perturbs macrophage innate immune responses; in Aim 2, we will compare fibroblast activation induced by Zip8 393T-KI vs. WT Zip8 macrophages. The long-term goal is to establish if ZIP8 391-Thr genotype has clinical implications for the prevention of and treatment of patients with Crohn’s disease and use study of ZIP8 391-Thr-related pathology as key opportunity to elucidate the role of Mn homeostasis in human disease. This R03 application builds from a K08 that established the altered Mn homeostasis and fibroinflammatory phenotype in colitis in Zip8 393T-KI mice. This application with strong translational relevance will provide critical mechanistic insight of the ZIP8 genotypic effect on macrophage-fibroblast signaling and the interaction with Mn homeostasis to prioritize therapeutic targets, including STAT3 or IL-11 inhibition, and patient studies as part of an R01 application.

项目摘要 炎症性肠病 (IBD)，包括克罗恩病 (CD) 和溃疡性结肠炎，影响每 200 人中就有 1 人 美国人，抗肿瘤坏死因子 (TNF)-α 疗法仍然是患者治疗的基石。 患有 IBD，但大约 40% 的患者会经历原发性或继发性反应丧失，并且“抗- 在当前的临床实践中，“经历过 TNF 治疗的”患者不太可能对所有后续治疗产生反应。 单细胞 RNA 测序揭示了与抗 TNF 耐药性相关的关键细胞特征， 强烈暗示炎症巨噬细胞和活化的成纤维细胞之间仍然存在信号传导。 我们对导致这种异常巨噬细胞的患者特异性因素的理解存在重大差距 成纤维细胞激活，它如何与抗 TNF 无反应相关，以及调节其改变的潜力 我们的工作重点是致病性的功能影响。 金属转运蛋白 ZIP8 A391T 中的变体，它会失调锰稳态并与 在小鼠模型中研究 ZIP8 391-Thr（Zip8）。 393T-敲入（KI）），我们已经显示出对结肠炎的易感性增加，但更有趣的是，我们 观察到这些小鼠的纤维化增强，IL-11 是一种促炎和促炎症因子。 纤维化细胞因子是炎症巨噬细胞和活化巨噬细胞之间信号异常的标志 因此，我们对克罗恩病（尤其是抗 TNF 难治性疾病）患者的成纤维细胞进行了研究。 研究表明，异常巨噬细胞-成纤维细胞信号传导的研究是疾病发病机制的一个关键特征 建立 Zip8 393T-KI 小鼠的潜在疾病机制很重要，因为 (1) 高达 25%。 克罗恩病患者在某些人群中携带 ZIP8 391-Thr，并且 (2) Zip8 393T-KI 可以发挥作用 作为机制研究的新型转化模型，特别是与抗 TNF 无反应相关的机制研究。 研究这一假设有两个目标：在目标 1 中，我们将确定 Zip8 393T-KI 是否干扰巨噬细胞先天 免疫反应；在目标 2 中，我们将比较 Zip8 393T-KI 与 WT Zip8 诱导的成纤维细胞活化 长期目标是确定 ZIP8 391-Thr 基因型是否对巨噬细胞具有临床意义。 克罗恩病患者的防治及ZIP8 391-Thr相关病理学的应用研究 作为阐明 Mn 稳态在人类疾病中的作用的关键机会，此 R03 应用程序基于此构建。 a K08 在 Zip8 393T-KI 中建立了结肠炎中改变的 Mn 稳态和纤维炎症表型 该应用具有很强的翻译相关性，将为 ZIP8 提供关键的机制见解。 基因型对巨噬细胞-成纤维细胞信号传导的影响以及与锰稳态的相互作用以优先考虑 治疗目标，包括 STAT3 或 IL-11 抑制，以及作为 R01 应用一部分的患者研究。