The role of Wnt signaling in treating glucocorticoid-induced glaucoma

Wnt信号在治疗糖皮质激素诱发的青光眼中的作用

• 批准号: 10838949
• 负责人: Weiming Mao
• 金额: $ 3.05万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10838949
• 关键词: Anti-Inflammatory Agents; Antiinflammatory Effect; Aqueous Humor; Biological Assay; Biological Process; Blindness; Cell Line; Cell Survival; Cells; ChIP-seq; Chromatin; Collagen; Complex; Cornea; DNA; DNA purification; DNA sequencing; Data; Disease; Distal; Dose; Endotoxins; Enzymes; Epigenetic Process; Erinaceidae; Ethanol; Extracellular Matrix; Eye; Eye diseases; Fibronectins; Future; G-Quartets; General Population; Genes; Glaucoma; Glucocorticoid Receptor; Glucocorticoids; Goals; Guanine; HDAC1 gene; Human; Hydrogen Bonding; Immunoprecipitation; Knowledge; Libraries; Luciferases; Matrix Metalloproteinases; Medicine; Metabolism; Methyl-CpG-Binding Protein 2; Mission; Modification; Molecular; Mus; Nuclear; Ocular Hypertension; Parents; Pathologic; Pathway interactions; Patients; Perfusion; Physiologic Intraocular Pressure; Prevention; Primary Open Angle Glaucoma; Production; Promega; Public Health; Publishing; Receptor Signaling; Reporter; Research; Role; Scientist; Shapes; Signal Pathway; Structure; Testing; Therapeutic; Tissues; Trabecular meshwork structure; Training; Underrepresented Minority; Uveitis; Validation; WNT Signaling Pathway; beta catenin; career; experience; graduate school; in vivo; innovation; medical schools; notch protein; novel; prevent; recruit; responders and non-responders; response; small molecule; therapeutic target; training opportunity; translational applications

PROJECT SUMMARY/ABSTRACT Glucocorticoid (GC)-induced ocular hypertension (OHT) and glaucoma (GIG) occur in ~40% of the general population and ~90% primary open angle glaucoma (POAG) patients. Our published studies and preliminary data show that canonical Wnt signaling activation inhibits glucocorticoid receptor signaling and GC-induced OHT. We hypothesize that activation of canonical Wnt signaling inhibits GIG, and this inhibition requires nucle- ar β-catenin, glucocorticoid receptor (GR), and epigenetic modification enzymes including HDAC1 and MeCP2. In this supplemental application, our objective is to identify new epigenetic mechanisms of and therapeutic compounds for glucocorticoid-induced OHT/glaucoma. Our rationale is that Wnt activation is a potential ap- proach to prevent and treat OHT/GIG, and studying GIG will help us to better understand primary open angle glaucoma. We propose two specific aims. SA 1: Determine whether G-quadruplex is regulated by the glucocor- ticoid receptor (GR) signaling pathway in the trabecular meshwork (TM). Recently, emerging evidence showed the importance of a DNA secondary structure, G-quadruplex (G4) in many biological processes. We hypothe- size that GCs enhance G4 formation in the TM. We will treat primary human trabecular meshwork (pHTM) cells from glucocorticoid responders or non-responders with or without 100nM DEX or 0.1% ethanol for 1 day. The cells will be used for immunoprecipitation against G4 chromatin, and purified DNA containing G4 will be used for DNA sequencing. At least 3 glucocorticoid responder and non-responder TM cell strains will be tested. We expect to find that glucocorticoid-responder TM cells have more G4 structures and these G4s are associated with key genes such as cell survival and ECM metabolism. We also expect to find that glucocorticoids enhance G4 formation and this enhancement is greater in responder TM cells. SA 2: Identify novel Wnt signaling path- way activation compounds with high potency in inhibiting GR signaling in the TM. We have found that the acti- vation of the Wnt signaling pathway using a small molecule compound CHIR is effective in the inhibition of GR signaling. Our preliminary data from the parent R01 also showed that CHIR did not compromise DEX’s anti- inflammatory effects in mouse eyes with endotoxin-induced uveitis. However, CHIR needs to be applied at high concentrations. Therefore, it is important to find more potent small molecule Wnt signaling activators. The pro- posed study is significant because it provides new molecular mechanisms as well as therapeutic targets for the prevention and treatment of glucocorticoid-induced OHT/glaucoma. The findings, especially from SA2, will have translational applications. It is also innovative because the role of G4 in the TM, glaucoma, and glucocor- ticoid receptor signaling, to our best knowledge, is completely unknown. Our future studies will focus on 1) whether G4 can be inhibited by Wnt signaling; 2) validation of the downstream target of glucocorticoid-induced G4 in the TM; and 3) testing the compound hits in vivo.

项目摘要/摘要 糖皮质激素（GC）诱导的眼高血压（OHT）和青光眼（GIG）发生在〜40％的一般性中 人口和〜90％的初级开角素（POAG）患者。我们发表的研究和初步 数据表明，规范Wnt信号传导激活抑制了糖皮质激素受体信号传导和GC诱导的 OHT。我们假设典型的Wnt信号传导的激活抑制了GIG，并且这种抑制需要核 - ARβ-catenin，糖皮质激素受体（GR）和包括HDAC1和MECP2在内的表观遗传修饰酶。 在此补充应用中，我们的目标是确定新的表观遗传机制和治疗 糖皮质激素诱导的OHT/青光眼的化合物。我们的理由是，Wnt激活是一种潜在的AP- 预防和治疗OHT/演出以及学习演出将有助于我们更好地理解主要的开放角度 青光眼。我们提出了两个具体目标。 SA 1：确定G-四链体是否受到葡萄糖的调节 小梁网（TM）中的Ticoid受体（GR）信号通路。最近，显示了新的证据 在许多生物过程中，DNA二级结构，G四链体（G4）的重要性。我们假设 GC在TM中增强G4形成的大小。我们将处理原代人小梁网（PHTM）细胞 从有或没有100nm dex或0.1％乙醇的糖皮质激素反应者或1天的非反应器中。这 细胞将用于针对G4染色质的免疫沉淀，并将使用含有G4的纯化DNA 用于DNA测序。将测试至少3个糖皮质激素响应者和非反应器TM细胞菌株。我们 希望发现糖皮质激素反应器TM细胞具有更多的G4结构，并且这些G4是相关的 具有关键基因，例如细胞存活和ECM代谢。我们还希望发现糖皮质激素会增强 G4形成和这种增强在响应者TM细胞中更大。 SA 2：确定新型Wnt信号通路 - 在抑制TM中GR信号传导方面具有高效力的激活化合物。我们发现Acti- 使用小分子化合物Chir的Wnt信号通路的外观有效地抑制GR 信号。我们的父母R01的初步数据还表明，Chir并未损害DEX的反抗 内毒素诱导的葡萄膜炎的小鼠眼中的炎症作用。但是，需要在高处应用 浓度。因此，重要的是要找到更多潜在的小分子Wnt信号传导激活剂。专业人士 提出的研究很重要，因为它为新的分子机制以及治疗靶标的 糖皮质激素诱导的OHT/青光眼的预防和治疗。这些发现，尤其是来自SA2的发现 已经翻译了应用程序。这也具有创新性，因为G4在TM，青光眼和葡萄糖糖中的作用 据我们所知，Ticoid受体信号传导是完全未知的。我们未来的研究将重点关注1） wnt信号传导； Wnt信号传导是否可以抑制G4； 2）验证糖皮质激素诱导的下游目标 TM中的G4； 3）在体内测试化合物命中。