Ferrochelatase as a mediator of ocular angiogenesis

新生血管眼病早产儿视网膜病变、增殖性糖尿病视网膜病变和新生血管 老年性黄斑变性是导致终生失明的主要原因。并不是所有的患者都有反应 因此，迫切需要找到可以靶向治疗的新的细胞成分 阻断这些疾病特有的病理性血管生成。铁络合酶(FECH)是一种 这种成分是血管内皮细胞在体外和体内增殖所必需的。费赫负责 将亚铁离子插入原卟啉IX，这是血红素生物合成的最后一步。前一批款期 发现FECH在小鼠和人的脉络膜和视网膜新生血管中上调， FECH抑制会导致内皮细胞中的血红素蛋白耗尽，而FECH的丢失会阻止血红素- 内皮细胞中依赖的氧化磷酸化。FECH活动减少也出人意料地阻止了 糖酵解。此外，第一个类药物FECH抑制剂SH-17023被开发出来，并被证明是 体外和体内抗血管生成作用。在这项工作的基础上，目前的目标是定义如何 血红素通过FECH和糖酵解合成影响内皮细胞生物学和新生血管。这个 这项研究的基本原理是FECH是血管生成的重要介质，是一个潜在的治疗靶点， 以及糖酵解酶的表达和功能的以前未被认识到的调节剂。整体而言 假说认为，FECH通过控制血红素的可获得性，是一种整合的多种 促血管生成途径，包括糖酵解。在强劲的初步数据的指导下，这一假设将得到检验 通过两个特定的目的：1.阐明FECH影响糖酵解和血管生成的机制。这个 内皮细胞中血红素合成抑制引起的糖酵解酶失调及其对血红素的依赖 转录调控将在内皮细胞中进行评估。糖酵解功能及其相关代谢 这些途径将通过海马、靶向代谢组学和稳定同位素示踪进行评估。的影响 这种依赖于血红素的糖酵解调节对内皮细胞功能的调节将与细胞- 这种效应的类型特异性。2.眼部首个类药物小分子FECH抑制剂的评价 新生血管。SH-17023的药代动力学和毒性将被量化。然后，这部小说 将测试分子对氧诱导视网膜病变Vldlr-/-和JR5558视网膜、视网膜下、 脉络膜新生血管模型，以及与抗血管内皮生长因子治疗的协同作用。总的来说，这项工作是 创新，因为这是第一次对血红素合成、糖酵解和后遗症之间的联系进行机制研究 眼部血管生成，以及用类药物小分子直接抑制FECH的首次特征 视网膜和脉络膜新生血管。这项工作意义重大，因为它将全面 阐明血红素代谢和糖酵解过程之间的联系，以前没有联系，以及 将FECH阻断作为一种可行的治疗方法，从而开发出新的防盲方法。

The neovascular eye diseases retinopathy of prematurity, proliferative diabetic retinopathy, and neovascular age-related macular degeneration are major causes of blindness through the lifespan. Not all patients respond to existing therapies, so there is thus a critical need to find novel cellular components that could be targeted to block the pathological angiogenesis that is characteristic of these diseases. Ferrochelatase (FECH) is one such component, necessary for proliferation of endothelial cells in vitro and in vivo. FECH is responsible for inserting ferrous ion into protoporphyrin IX, the final step in heme biosynthesis. The previous grant period yielded findings that FECH is upregulated in murine and human choroidal and retinal neovascularization, that FECH inhibition leads to depletion of hemoproteins in endothelial cells, and that FECH loss blocks heme- dependent oxidative phosphorylation in endothelial cells. Reduced FECH activity also surprisingly blocks glycolysis. In addition, the first drug-like FECH inhibitor, SH-17023 was developed and shown to be antiangiogenic in vitro and in vivo. Building on this work, the current goal is to define the mechanism of how heme synthesis through FECH and glycolysis impacts endothelial cell biology and neovascularization. The rationale for this research is that FECH is a significant mediator of angiogenesis, a potential therapeutic target, and a previously unappreciated regulator of the expression and function of glycolytic enzymes. The overall hypothesis is that FECH, via controlling heme availability, is an integrated master regulator of multiple proangiogenic pathways, including glycolysis. Guided by strong preliminary data, the hypothesis will be tested via two specific aims: 1. Delineate the mechanism of FECH’s influence on glycolysis and angiogenesis. The glycolysis enzymes dysregulated by heme synthesis inhibition in endothelial cells and their heme-dependent transcriptional regulation will be assessed in endothelial cells. Glycolytic function and related metabolic pathways will be assessed by Seahorse, targeted metabolomics, and stable isotope tracing. The influence of this heme-dependent glycolysis regulation on endothelial cell function will be determined, along with the cell- type specificity of this effect. 2. Evaluate the first drug-like small molecule FECH inhibitor in ocular neovascularization. The pharmacokinetics and toxicity of SH-17023 will be quantified. Then, this novel molecule will be tested for efficacy in the oxygen-induced retinopathy, Vldlr-/-, and JR5558 retinal, subretinal, and choroidal neovascularization models, plus synergy with anti-VEGF therapy. Overall, this work is innovative, as it is the first mechanistic study of the links between heme synthesis, glycolysis, and posterior ocular angiogenesis, and the first characterization of direct FECH inhibition with a drug-like small molecule for retinal and choroidal neovascularization. The work is highly significant because it will comprehensively elucidate linkages between heme metabolism and the process of glycolysis, not previously connected, and establish FECH blockade as a viable therapy, leading to development of new ways to prevent blindness.