Rho GTPase siRNA as a Therapeutic Strategy in Regulating Outflow Resistance

Rho GTPase siRNA 作为调节流出阻力的治疗策略

• 批准号: 8925285
• 负责人: Cynthia Lynn Pervan
• 金额: --
• 依托单位: EDWARD HINES JR VA HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-03-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8925285
• 关键词: Actins; Affect; Age; Age-Years; Anterior; Aqueous Humor; Attenuated; Blindness; Caring; Cells; Confocal Microscopy; Deposition; Development; Disease; Dose; Employee Strikes; Endothelin-1; Equilibrium; Extracellular Matrix; Eye; Family suidae; Glaucoma; Goals; Guanosine Triphosphate Phosphohydrolases; Healthcare; Human; Immunohistochemistry; Individual; Intervention; Laboratories; Lead; Mediating; Mediator of activation protein; Medical; Operative Surgical Procedures; Pathologic; Patients; Perfusion; Physiologic Intraocular Pressure; Prevalence; Primary Open Angle Glaucoma; Production; Publishing; Publishing Peer Reviews; Refractory; Rehabilitation therapy; Reporting; Resistance; Risk Factors; Signal Transduction; Small Interfering RNA; Source; Stress Fibers; Technology; Testing; Therapeutic; Time; Trabecular meshwork structure; Vasoconstrictor Agents; Veterans; cytokine; improved; novel; novel strategies; palliative; prevent; proendothelin 1; public health relevance; rho GTP-Binding Proteins; social; socioeconomics; therapeutic target; translational study

 DESCRIPTION (provided by applicant): Primary open-angle glaucoma (POAG) is a leading cause of blindness worldwide, affecting over 2 million individuals 45 years of age or older in the US. The personal, social, and medical burden of glaucoma within the VHA remains an extraordinarily significant concern. Despite its overwhelming prevalence and socioeconomic impact, the treatment of Veterans with POAG is currently restricted to non-specific interventions that lower elevated intraocular pressure (IOP). For many glaucomatous Veterans, pharmacological management of IOP remains clinically refractive. These Veterans often require more extensive interventions, including invasive surgical manipulation. The development of targeted therapeutic strategies directed at the cause of elevated IOP is critical for the management of our glaucomatous Veterans. In healthy eyes, IOP is maintained through balanced production and outflow of aqueous humor (AH). Increased resistance to AH outflow through the trabecular meshwork (TM) is considered to be a major contributor of aberrant elevation of IOP in Veterans with POAG. The mechanism by which this occurs remains poorly defined, but most likely involves a combination of enhanced TM cell contractility and altered ECM deposition. Independent studies have shown that TGF-ß2, a cytokine that facilitates Rho GTPase-mediated ECM deposition and actin stress fiber organization in TM cells, is markedly elevated in the AH of POAG patients. Endothelin-1 (ET-1) a potent vasoconstrictor, is also increased in the AH from POAG patients. The association between TGF-ß2, ET-1, and Rho GTPase signaling with POAG, however, remains unclear. Peer-reviewed and published studies from our laboratory demonstrate a striking association between TGF-ß2 and ET-1 production within human TM cells. Our preliminary findings included in this application further demonstrate for the first time that by selectively targeting and disrupting Rho GTPase signaling, we can successfully prevent harmful increases in TGF-ß2 and ET-1 expression in human TM cells. These findings strongly suggest that aberrant Rho GTPase signaling elevates IOP, in part, by facilitating pathologic increases in TGF-ß2 and ET-1 in the anterior segment of POAG Veterans. Novel translational studies are critically needed to develop therapeutic and rehabilitative strategies for the advanced care of our Veterans debilitated by glaucoma. siRNA technology is an exciting and promising new field that has realistic therapeutic/rehabilitative potential for the management of ocular disorders, including glaucoma. In this two-year SPiRE study, we will determine whether siRNA-mediated Rho GTPase knockdown therapeutically lowers experimentally-elevated IOP. Hypothesis: Therapeutically administered siRNA selectively targeting monomeric Rho GTPases will markedly lower pathologically elevated intraocular pressure by disrupting TGF-ß2 and ET-1 expression and signaling. Specific Aim 1 will determine whether administration of siRNAs targeting key monomeric Rho GTPases protects against experimentally elevated changes in outflow resistance as quantified by human and porcine anterior segment perfusion analyses. Specific Aim 2 will determine whether administration of siRNAs targeting key monomeric Rho GTPases attenuates expression of proET-1, ETA, ETB, TGF-ß2 or TGFßRI in human and porcine globes with experimentally elevated outflow resistance as quantified by immunohistochemistry and confocal microscopy. The immediate goal of this study is to establish siRNA-mediated Rho GTPase knockdown as a novel strategy by which to manage elevated IOP associated with POAG. Successful completion of this study will serve as a critical first step toward developing this technology for the management of Veterans with POAG.

 描述（由申请人提供）： 原发性开角型青光眼（POAG）是全球范围内致盲的主要原因，在美国影响超过200万45岁或以上的个体。VHA内青光眼的个人、社会和医疗负担仍然是一个非常重要的问题。尽管其压倒性的患病率和社会经济影响，POAG退伍军人的治疗目前仅限于降低眼内压（IOP）升高的非特异性干预。对于许多患有青光眼的退伍军人来说，药物治疗IOP在临床上仍然是屈光性的。这些退伍军人往往需要更广泛的干预，包括侵入性手术操作。针对IOP升高原因的靶向治疗策略的开发对于我们的青光眼退伍军人的管理至关重要。在健康的眼睛中，通过平衡的产生和流出的房水（AH）来维持IOP。AH通过小梁网（TM）流出的阻力增加被认为是POAG退伍军人IOP异常升高的主要原因。发生这种情况的机制仍然不清楚，但最有可能涉及TM细胞收缩性增强和ECM沉积改变的组合。独立的研究表明，TGF-β 2，一种促进TM细胞中Rho GTP酶介导的ECM沉积和肌动蛋白应力纤维组织的细胞因子，在POAG患者的AH中显著升高。内皮素-1（ET-1）是一种强有力的血管收缩剂，在POAG患者的AH中也增加。然而，TGF-β 2、ET-1和Rho GT3信号传导与POAG之间的关联仍不清楚。来自我们实验室的同行评审和发表的研究表明，在人TM细胞内TGF-β 2和ET-1的产生之间存在显著的关联。本申请中包括的我们的初步发现首次进一步证明，通过选择性靶向和破坏Rho GT3信号传导，我们可以成功地防止人TM细胞中TGF-β 2和ET-1表达的有害增加。这些发现强烈提示异常Rho GT3信号传导升高IOP，部分是通过促进POAG退伍军人前段中TGF-β 2和ET-1的病理性增加。迫切需要新的转化研究来制定治疗和康复策略，为青光眼衰弱的退伍军人提供先进的护理。siRNA技术是一个令人兴奋和有前途的新领域，具有现实的治疗/康复潜力，用于管理眼部疾病，包括青光眼。在这项为期两年的SPiRE研究中，我们将确定siRNA介导的Rho GT3基因敲低是否能在治疗上降低实验性升高的IOP。假设：治疗性施用的选择性靶向单体Rho GTP酶的siRNA将通过破坏TGF-β 2和ET-1表达和信号传导而显著降低病理性升高的眼内压。具体目标1将确定靶向关键单体Rho GTP酶的siRNA的施用是否保护免于流出阻力的实验性升高的变化，如通过人和猪眼前段灌注分析所定量的。具体目标2将确定靶向关键单体Rho GTP酶的siRNA的施用是否减弱具有实验性升高的流出阻力的人和猪球体中proET-1、ETA、ETB、TGF-β 2或TGF β RI的表达，如通过免疫组织化学和共聚焦显微镜定量的。本研究的直接目标是建立siRNA介导的Rho GT3基因敲低作为一种新的策略，通过该策略来管理与POAG相关的IOP升高。这项研究的成功完成将成为开发这项技术用于POAG退伍军人管理的关键第一步。