A pilot study to define maintained alterations in mitochondrial DNA of eosinophilic esophagitis patients that may be exploited clinically

项目摘要 嗜酸性粒细胞性食管炎（EoE）是一种慢性、食物抗原介导的疾病，其特征是嗜酸性粒细胞增多， 食道炎症和症状，如腹痛、吞咽困难和食物嵌塞。最近的一 一项研究确定了EoE患者线粒体蛋白DHTKD 1编码基因的破坏性变异。 虽然这表明线粒体在EoE病理学中的潜在作用，但仍不清楚是否 线粒体DNA的改变存在于EoE中，并可用于改善EoE的临床护理 患者事实上，线粒体的房子活性氧（ROS）产生电子传递链 与核DNA相比，它们具有较不复杂的DNA修复机制， 线粒体DNA对ROS诱导的遗传改变高度敏感。这些信息与我们的 初步研究结果表明：（i）患者食管上皮中线粒体增加， 在非活动性EoE患者中未能正常化的活动性EoE;和（ii）在非活动性EoE患者中诱导线粒体功能障碍。 食管角化细胞暴露于EoE炎症环境后，导致我们假设， 线粒体DNA中的蛋白质可以在活动性EoE患者的食管粘膜中获得，并维持在 临床疾病缓解的非活动性患者亚组。为了验证这一假设，我们将进行以下研究： 具体目标：目标1：在来自横截面的活检标本中进行线粒体DNA测序 包括正常对照以及患有活动性和非活动性EoE的患者的人类受试者群组。 将对胃食管反流病患者进行评估，以确定是否鉴定出线粒体DNA 这些变化是EoE特有的。目的2：在来自一个组织的活检标本中进行线粒体DNA测序。 人类受试者的纵向队列，包括在两次活性EoE时具有可用组织的患者 和非活性EoE，并分类为治疗应答者或无应答者。本研究将（i）确定 线粒体DNA损伤是EoE的一个特征，包括活动性和非活动性;（ii）评估 EoE中线粒体DNA突变负荷和治疗反应;以及（iii）确定复发性 线粒体突变可能是治疗EoE的靶点。潜在的线粒体的例子- EoE的靶向治疗包括抗氧化剂化合物（例如辅酶Q10）或微量营养素的递送 (e.g.锌、生物素）改善线粒体功能，酶替代疗法恢复功能 由线粒体DNA编码的错误酶，以及腺病毒或CRISPR介导的对线粒体DNA的编辑。 线粒体基因组本研究产生的数据将为未来NIH R 01提案提供基础 这将描述线粒体DNA改变在EoE病理学中的功能意义， 使用临床前模型评估与线粒体功能障碍相关的治疗策略，包括原发性 上皮培养物、患者来源的食管类器官和鼠EoE模型。

Project Summary Eosinophilic Esophagitis (EoE) is a chronic, food antigen-mediated disease characterized by eosinophil-rich esophageal inflammation and symptoms such as abdominal pain, dysphagia and food impaction. A recent study identified damaging variants in the gene encoding the mitochondrial protein DHTKD1 in EoE patients. Although this indicates a potential role for mitochondria in EoE pathobiology, it remains unclear whether alterations in mitochondrial DNA are present in EoE and may be leveraged to improve clinical care of EoE patients. Indeed, mitochondria house reactive oxygen species (ROS)-producing electron transport chain components and have less sophisticated DNA repair mechanisms as compared to nuclear DNA, making mitochondria DNA highly susceptible to ROS-induced genetic alterations. This information in concert with our preliminary findings indicating (i) that there is an increase in mitochondria in esophageal epithelium of patients with active EoE that fails to normalize in inactive EoE patients; and (ii) mitochondrial dysfunction is induced in esophageal keratinocytes upon exposure to the EoE inflammatory milieu, led us to hypothesize that alterations in mitochondrial DNA may be acquired in esophageal mucosa of patients with Active EoE and maintained in a subset of Inactive patients in clinical disease remission. To test this hypothesis, we will pursue the following specific aims: Aim 1: Perform mitochondrial DNA sequencing in biopsy specimens from a cross-sectional cohort of human subjects comprising Normal controls as well as patients with Active and Inactive EoE. Gastroesophageal reflux disease patients will be evaluated to determine whether identified mitochondrial DNA alterations are specific to EoE. Aim 2: Perform mitochondrial DNA sequencing in biopsy specimens from a longitudinal cohort of human subjects comprising patients with tissues available at the time of both Active EoE and Inactive EoE and classified as therapeutic responders or non-responders. This study will (i) determine if mitochondrial DNA damage is a feature of EoE, both Active and Inactive; (ii) assess the relationship between mitochondrial DNA mutational burden and therapeutic response in EoE; and (iii) identify recurrent mitochondrial mutations that may be targeted therapeutically in EoE. Examples of potential mitochondria- targeted therapies for EoE include, delivery of antioxidant compounds (e.g. Coenzyme Q10) or micronutrients (e.g. Zinc, Biotin) to improve mitochondrial function, enzyme replacement therapy to restore the function of faulty enzymes encoded by mitochondrial DNA, and Adenoviral or CRISPR-mediated editing of the mitochondrial genome. Data generated in this study will provide the foundation for a future NIH R01 proposal that will delineate the functional significance of mitochondrial DNA alterations in EoE pathobiology and also evaluate therapeutic strategies related to mitochondrial dysfunction using preclinical models, including primary epithelial cultures, patient-derived esophageal organoids, and murine EoE models.