Modulation of retinoid reactivity and pathological signaling in retinal therapeutics

项目摘要 老年性黄斑变性是60岁以上人群视力丧失的主要原因 包括退伍军人。它的特点是在中央视野中失去视力，在那里锐利，多色 图像是在强光条件下产生的，现代人通常在 醒着的时间。丧失这种高敏锐度的色觉会导致严重的残疾。AMD的高患病率 每年在与AMD相关的疾病上花费超过980亿美元，给医疗系统带来了巨大的负担 美国的医疗成本。目前，还没有非常有效的方法治疗最常见的 这种疾病称为地理性萎缩，约占晚期AMD的90%。在过去的退伍军人管理局资助期间 期间，我们研究了抑制视觉周期作为治疗AMD的一种潜在方法，并表征了酶 建议对视觉周期通路的正常功能作出贡献。根据我们在以下期间所做的主要发现 在最初的资助期内，我们现在建议调查与食道癌发病机制有关的相关途径。 AMD是通过视觉周期产生的视黄醛(Ral)参与的。此外，我们还将 继续我们对一种名为Des1的酶的研究，该酶被认为是介导锥体再生的酶 视觉色素，视网膜中央的颜色感应分子。我们将通过 具体目标如下：1.使用新的RPE-阐明Des1在RPE中的生物学作用- 特定的Cre小鼠。此前，我们发现Müler细胞中的Des1蛋白不是11-cis的主要贡献者- 视锥细胞感光细胞视黄醇的合成。然而，单细胞RNA-Seq分析表明，RPE是 视网膜中Des1表达的主要部位引发了关于其在该组织中的生物学作用的问题。使用 我们将研究Des1功能丧失对RPE和RPE的影响。 将这些小鼠与RPE特异性Cre小鼠和 使用各种功能和成像技术对它们进行表征。Des1在从头开始中也发挥着关键作用 神经酰胺的产生，已知的RPE细胞死亡的中介。我们假设Des1在 RPE将调节组织对化学诱导的毒性的敏感性，这是RPE细胞的模型 发生在地理萎缩中的死亡。这些研究将测试DES1作为AMD潜在靶点的可行性 治疗学。2.先进的下一代视觉周期调制器(VCM)，具有选择性 药效学。视觉周期调节器最初被设计为抑制RPE65，以便抑制 病理性脂褐素积聚和视网膜疾病进展缓慢。我们发现了一种新的机制 这些化合物的作用：与激活的视觉视蛋白释放的al直接反应，以限制形成 病理性的前额内收物。我们已经产生了视觉周期调节器，它对血管紧张素转换酶具有优先活性 对视网膜病变具有保护作用的隔离，减少了对视觉周期活动的影响。 基于我们的初步研究，我们建议合成和表征一套设计合理的新视觉 我们假设的周期调节器将具有增强的治疗活性和减少的视觉周期 压制。3.开发磷酸二酯酶(PDE)抑制剂作为视网膜疾病的治疗方法。 先前的研究表明，异常的GPCR信号与丙二醛的毒性有关。磷酸二酯酶是主要的 GPCRs的效应者和调节者，并已成功地靶向于临床应用。我们假设 PDE的抑制剂将在不损害视觉功能的情况下提供对视网膜侮辱的保护作用。 我们的初步数据表明，PDE4抑制剂在低剂量的动物模型中特别有效 丙二醛毒性。我们将筛选这些化合物和相关的衍生物，以阐明它们在体内的作用部位和机制。 使用视网膜病变动物模型的保护作用。总之，这些研究可能会发现小分子 可以很容易地转化为退伍军人的视网膜疾病治疗。

Project Summary Age-related macular degeneration is a principal cause of vision loss in individuals over the age of 60 years including Veterans. It is characterized by a loss of sight in the central visual field where sharp, polychromatic images are generated under the bright light conditions that modern humans are typically exposed to during waking hours. Loss of this high acuity color vision leads to significant disability. The high prevalence of AMD places a large burden on the healthcare system with upwards of 98 billion dollars spent yearly on AMD-related healthcare costs in the US. Currently, there are no highly effective treatments for the most common form of the disease, known as geographic atrophy, which makes up ~90% of advanced AMD. During the past VA funding period, we studied inhibition of the visual cycle as a potential treatment for AMD and characterized enzymes proposed to contribute to the normal function of the visual cycle pathway. Based on key findings we made during the initial funding period, we now propose to investigate related pathways that are tied to the pathogenesis of AMD through the involvement of retinaldehyde (RAL) derived from the visual cycle. Additionally, we will continue our studies on an enzyme known as Des1 that has been proposed to mediate the regeneration of cone visual pigments, the color sensing molecules in the central retina. We will explore these pathways through the following Specific Aims: 1. Elucidate biological roles for Des1 within the RPE using novel RPE- specific Cre mice. Previously, we showed that Des1 protein in Müller cells is not a major contributor to 11-cis- retinol synthesis for cone photoreceptors. However, single cell RNA-Seq analysis revealed that the RPE is the principal site of Des1 expression in the retina raising questions about its biological role in this tissue. Using a validated floxed Des1 mouse model, we will investigate the impact of Des1 loss of function on RPE and photoreceptor health and visual cycle function by crossing these mice with RPE-specific Cre mice and characterizing them using a variety of functional and imaging techniques. Des1 also plays a key role in de novo ceramide production, a known mediator of apoptotic RPE cell death. We hypothesize that Des1 deletion in the RPE will modulate susceptibility of the tissue to chemical-induced toxicity, which serves as a model for RPE cell death that occurs in geographic atrophy. These studies will test the viability of Des1 as a potential target for AMD therapeutics. 2. Advance next-generation visual cycle modulators (VCMs) with selective pharmacodynamics. Visual cycle modulators were originally designed to inhibit RPE65 in order to suppress pathological lipofuscin accumulation and slow retinal disease progression. We discovered a novel mechanism of action for these compounds: direct reaction with RAL released from activated visual opsins to limit formation of pathological RAL adducts. We have generated visual cycle modulators with preferential activity towards RAL sequestration that possess protective effects against retinopathy with reduced effects on visual cycle activity. Based on our initial studies, we propose to synthesize and characterize a new set of rationally-designed visual cycle modulators that we hypothesize will possess augmented therapeutic activity and diminished visual cycle suppression. 3. Develop phosphodiesterase (PDE) inhibition as a treatment for retinal disease. Prior research has implicated aberrant GPCR signaling in RAL toxicity. Phosphodiesterase enzymes are major effectors and regulators of GPCRs and have been successfully targeted for clinical applications. We hypothesize that inhibitors of PDEs will confer protective effects against retinal insults without impairing visual function. Our preliminary data indicates that PDE4 inhibitors are particularly effective at low doses in animal models of RAL toxicity. We will screen these compounds and related derivatives to elucidate their site and mechanism of protective action using animal models of retinopathy. Together, these studies may uncover small molecules that could readily be translated into retinal disease treatments for veterans.