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.

项目总结/摘要 糖皮质激素（GC）诱导的高眼压（OHT）和青光眼（GIG）发生在约40%的一般人群中。 人群和约90%的原发性开角型青光眼（POAG）患者。我们发表的研究和初步的 数据显示典型的Wnt信号传导激活抑制糖皮质激素受体信号传导和GC诱导的 好的。我们假设经典Wnt信号的激活抑制了GIG，这种抑制需要核- ar β-连环蛋白、糖皮质激素受体（GR）和表观遗传修饰酶，包括HDAC 1和MeCP 2。 在这个补充申请中，我们的目标是确定新的表观遗传机制和治疗， 用于糖皮质激素诱导的OHT/青光眼的化合物。我们的理由是Wnt激活是一个潜在的AP- 探讨OHT/GIG的防治方法，研究GIG有助于我们更好地认识原发性开角型青光眼 青光眼我们提出两个具体目标。SA 1：确定G-四链体是否受葡萄糖皮质激素调节， 小梁网（TM）中的噻酚受体（GR）信号通路。最近，新的证据显示 DNA二级结构G-四链体（G4）在许多生物过程中的重要性。我们假造- 使GC大小增加TM中G4形成。我们将处理原代人小梁网（pHTM）细胞 来自糖皮质激素应答者或无应答者，用或不用100 nM DEX或0.1%乙醇1天。的 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 进行DNA测序将检测至少3种糖皮质激素应答者和非应答者TM细胞株。我们 我希望发现糖皮质激素应答的TM细胞具有更多的G4结构，这些G4结构与糖皮质激素应答的TM细胞的G4结构相关。 细胞存活和细胞外基质代谢等关键基因。我们还希望发现糖皮质激素能增强 G4形成，这种增强在响应TM细胞中更大。SA 2：识别新的Wnt信号传导路径- 在抑制TM中GR信号传导方面具有高效力的方式活化化合物。我们发现，行为-- 使用小分子化合物CHIR对Wnt信号通路的阻断在GR抑制中是有效的 信号我们对亲本R 01的初步数据也表明，CHIR没有损害DEX的抗- 在患有内毒素诱导的葡萄膜炎的小鼠眼睛中的炎症作用。然而，CHIR需要在高 浓度的因此，寻找更有效的小分子Wnt信号激活剂具有重要意义。亲- 这一研究具有重要的意义，因为它提供了新的分子机制以及治疗靶点。 预防和治疗糖皮质激素诱导的OHT/青光眼。这些发现，特别是来自SA 2的发现，将 具有翻译应用。它也是创新的，因为G4在TM，青光眼和葡萄糖皮质激素中的作用- 据我们所知，类可可脂受体信号传导是完全未知的。我们未来的研究将集中在1） G4是否可以被Wnt信号抑制; 2）验证糖皮质激素诱导的G4表达的下游靶点。 TM中的G4;和3）测试化合物的体内命中。