EGR-1 Mediated Revascularization and Arteriogenic Bypass

描述（由申请人提供）：动脉发生描述了侧支血管生长的过程，被认为是血管闭塞的关键生物学反应。虽然临床试验采用血管生成生长因子的“治疗性”管理通常产生令人失望的结果，但现在看来，动脉生成可能比血管生成更相关，可以创造一个强大的、大容量的、稳定的新血管系统，可以提供血管阻塞的天然“旁路”。在这方面，虽然缺血/缺氧可能是血管生成的主要生理触发因素，但响应血管剪切和其他应激的单核细胞募集已被认为是动脉生成的关键早期介质。然而，这种激活途径的分子基础尚不清楚，尽管生长因子的任意施用可能有害地绕过“上游”信号通路，但“动脉生成”转录因子尚未确定。EGR-1转录因子对剪切应力的变化作出反应，独立于缺氧的存在，并且可以激活一系列潜在的动脉生成生长因子和分子。在此背景下，我们对血管闭塞后EGR-1表达的快速上调，EGR-1缺失小鼠血管结扎后单核细胞募集和再灌注的几乎缺失，以及EGR-1给药后灌流的接近正常化的证明产生了兴趣。因此，我们推测EGR-1是将血管闭塞的生理后遗症转化为（动脉源性）新生血管反应的“缺失”信号。因此，目前的目标是验证以下假设：1)通过上调关键的动脉生成介质诱导动脉生成，作为对血管闭塞的关键反应；2)诱导单核细胞募集，作为该反应的催化剂（并在治疗上增强这一过程）；3)可以“治疗上”诱导稳定的血管再生，比“下游”血管生成介质诱导的更大。为了实现这些目标，我们将在野生型与EGR-1缺陷（egr敲除或LacZ敲入）小鼠、大鼠和高胆固醇血症猪心肌缺血模型中，通过急性或慢性（可调节和组织特异性）基因转移载体，使用血管结扎后的血管化、灌注和表达分析。这些研究应该阐明EGR-1介导的血管重建调控的微观解剖部位和分子信号通路中的位点，并提供关于通过动脉生成药物和血管生成药物，以及通过“主开关”转录因子和下游介质进行血管重建策略的益处的临床前数据。尽管在冠状动脉和周围血管疾病的预防和治疗方面不断取得进展，但据估计，仅在美国，就有大约5万至8万人因晚期疾病而限制了传统疗法的有效性或适用性，这些人预计每年将死于冠状动脉疾病，约有10万人将因周围血管疾病而丧失肢体。干预可以改变血管生物学，并通过生长侧支血管（动脉生成）增强人体自身绕过自身的能力，因此是传统治疗的有吸引力的替代方案。基于越来越多的证据，我们相信一种叫做Egr-1的基因可能是一种“缺失”的信号因子，它可以将血管阻塞的生理（机械）压力转化为稳定和持久的新血管系统的生长。本研究旨在阐明Egr-1在天然血管生长中的作用及其作为一种有效的“动脉源性”血运重建剂的潜在用途。

DESCRIPTION (provided by applicant): Arteriogenesis describes the process of collateral vessel growth that is thought to be a critical biologic response to vascular occlusion. While clinical trials employing the "therapeutic" administration of angiogenic growth factors have generally yielded disappointing results, it now appears that arteriogenesis may be more relevant than angiogenesis in creating a robust, large capacitance, stable neovasculature that can literally provide native "bypasses" of vascular obstructions. In this regard, while ischemia/hypoxia are likely the primary physiologic triggers of angiogenesis, monocyte recruitment in response to vascular shear and other stresses has been implicated as a key early mediator of arteriogenesis. The molecular basis of this activation pathway is unclear, however, and while the arbitrary administration of growth factors may deleteriously bypass "upstream" signaling pathways, an "arteriogenic" transcription factor has yet to be identified. The EGR-1 transcription factor is responsive to changes in shear stress independent of the presence of hypoxia, and can activate a portfolio of potentially arteriogenic growth factors and molecules. In this context, we became intrigued by our demonstration of the rapid upregulation of EGR-1 expression following vascular occlusion, the near- absence of monocyte recruitment and reperfusion following vascular ligation in EGR-1 null mice, and the near-normalization of perfusion following EGR-1 administration to ligated wild type animals. We consequently speculated that EGR-1 is the "missing" signal that transponds the physiologic sequelae of vascular occlusion into an (arteriogenic) neovascularization response. The current aims are therefore to test the hypotheses that EGR-1: 1) induces arteriogenesis as a critical response to vascular occlusion, by upregulation of key arteriogenic mediators, 2) induces monocyte recruitment as the catalyst for this response (and augment this process therapeutically), and 3) can "therapeutically" induce stable revascularization greater than that induced by "downstream" angiogenic mediators. To pursue these aims, we will use vascularization, perfusion and expression assays following vascular ligation in wild type vs. EGR-1 deficient (EGR-knockout or LacZ knock-in) mouse, rat and in a hypercholesterolemic swine myocardial ischemia models with or without VEGF, MCP-1 or EGR-1 administration, via acute or chronic (regulatable and tissue specific) gene transfer vectors. These studies should elucidate the microanatomic site and the locus in molecular signaling pathways of EGR-1 mediated regulation of revascularization, and provide pre-clinical data regarding the benefits of revascularization strategies via arteriogenic vs. angiogenic agents, and via a "master switch" transcription factor vs. downstream mediators. PUBLIC HEALTH RELEVANCE Despite ongoing advances in the prevention and treatment of coronary and peripheral vascular disease, it is estimated that advanced disease limits the effectiveness or applicability of conventional therapies in approximately 50,000 - 80,000 individuals in the U.S. alone, who can be expected to die annually from coronary disease, and approximately 100,000 individuals will suffer limb loss from peripheral vascular disease. Interventions that can alter vascular biology and enhance the body's own ability to bypass itself by growing collateral vessels (arteriogenesis) therefore represent attractive alternatives to conventional therapies. Based upon a growing number of lines of evidence, we believe that a gene called Egr-1 may be a "missing" signaling factor that can translate the physiologic (mechanical) stresses of vascular blockages into the growth of a stable and persistent new vasculature. This study seeks to elucidate the role of Egr-1 in native blood vessel growth and its potential use as a potent "arteriogenic" revascularization agent.