PGE2 EP1 and EP3 receptors as therapeutic targets in intracerebral hemorrhage

描述(申请人提供)：PGE2、EP1和EP3受体作为脑出血的治疗靶点脑出血(ICH)是死亡率和发病率最高的卒中亚型；不幸的是，这一疾病受到的研究关注远远少于缺血性卒中。炎症机制，包括某些前列腺素(PGs)介导的炎症机制，已被认为有助于脑出血损伤的进展。特别是，在大脑中占主导地位的PGE2在血肿周围区域产生和积聚。PGE2通过四种G蛋白偶联受体亚型EP1-EP4发挥作用。在脑缺血的动物模型中，这些受体中的每一个都有不同的信号级联。在体内和体外，EP1和EP3受体的基因缺失或选择性抑制已被证明可以减少缺血性脑损伤。这样做的总体目标是 R01的目的是探讨抑制PGE2、EP1和EP3受体是否可用于脑出血的神经保护治疗，如果可以，则产生临床前疗效数据并阐明其潜在机制。我们一直在研究PGE2受体在脑出血损伤中的具体作用，并有初步数据表明，EP1或EP3受体的缺失或抑制可以减轻脑出血损伤并改善功能预后。我们的数据还表明，在脑出血后，Src激酶介导EP1毒性，而Rho Kinase(ROCK)介导EP3毒性。因此，我们将验证PGE2EP1受体阻断通过Src激酶信号通路减轻脑出血损伤的假说，而EP3受体阻断通过ROCK信号通路减轻脑出血损伤。我们利用自体血液脑出血模型和胶原酶诱导的脑出血模型设计了三个具体的靶点。考虑到PGE2的产生增加导致与年龄相关的炎症反应功能障碍，以及年龄和性别差异影响脑出血的预后，本研究将在年轻的雄性和雌性以及老年雄性小鼠中进行，以增强临床相关性。目的1将确定抑制EP1或EP3受体是否改善脑出血预后；目的2将确定抑制Src激酶信号是否有助于脑出血后EP1受体阻断所提供的神经保护，以及EP1受体抑制是否减少凝血酶诱导的脑损伤；以及目的3将确定抑制ROCK信号是否有助于脑出血后EP3受体阻断所提供的神经保护以及EP3受体抑制是否减轻凝血酶诱导的脑损伤。通过药理学、遗传学、影像学、组织学、分子生物学和细胞生物学的方法，这项工作将提供选择性PGE2EP1和EP3受体抑制对脑出血所致脑损伤的保护作用及其发生机制的新信息。这一概念验证信息是计划对选择性EP1和EP3受体拮抗剂在脑出血中进行更详细的临床前评估所必需的。

DESCRIPTION (provided by applicant): PGE2 EP1 and EP3 receptors as therapeutic targets in intracerebral hemorrhage Intracerebral hemorrhage (ICH) is the stroke subtype with the highest mortality and morbidity; unfortunately, this condition has received far less research attention than has ischemic stroke. Inflammatory mechanisms, including those mediated by certain prostaglandins (PGs), have been suggested to contribute to the progression of ICH injury. In particular, PGE2, which is predominant in the brain, is produced and accumulates in the perihematomal region. PGE2 acts through four G-protein-coupled receptor subtypes known as EP1- EP4. Each of these receptors has distinct signaling cascades in animal models of cerebral ischemia. Genetic deletion or selective inhibition of the EP1 and EP3 receptors has been shown to reduce ischemic brain injury both in vivo and in vitro. The overall objective of this R01 is to investigate whether inhibition of PGE2 EP1 and EP3 receptors can be used as neuroprotective therapy in ICH, and if so, to generate preclinical efficacy data and elucidate the underlying mechanisms. We have been investigating the specific roles of PGE2 receptors in ICH injury and have preliminary data showing that deletion or inhibition of the EP1 or EP3 receptor reduces ICH injury and improves functional outcomes. Our data also indicate that Src kinase mediates EP1 toxicity and Rho kinase (ROCK) mediates EP3 toxicity after ICH. Consequently, we will test the hypothesis that PGE2 EP1 receptor blockade reduces ICH injury through the Src kinase signaling pathway, whereas EP3 receptor blockade reduces ICH injury through the ROCK signaling pathway. We have designed three specific aims that utilize the autologous blood ICH model and collagenase-induced ICH model. Considering that an increase in PGE2 production contributes to age-related dysfunction of the inflammatory responses and that aging and sex differences affect ICH outcomes, this research will be carried out in young male and female and aged male mice to enhance the clinical relevance. Aim 1 will determine whether inhibition of EP1 or EP3 receptors improves ICH outcomes; Aim 2 will determine whether inhibition of Src kinase signaling contributes to the neuroprotection afforded by EP1 receptor blockade after ICH and whether EP1 receptor inhibition decreases thrombin-induced brain damage; and Aim 3 will determine whether inhibition of ROCK signaling contributes to the neuroprotection afforded by EP3 receptor blockade after ICH and whether EP3 receptor inhibition decreases thrombin-induced brain damage. Through pharmacologic, genetic, imaging, histologic, molecular, and cellular biologic approaches, this work will provide novel information about the efficacy of selective PGE2 EP1 and EP3 receptor inhibition to protect against ICH-induced brain injury and the mechanisms by which such protection occurs. This proof-of-concept information is required to plan more detailed preclinical assessment of selective EP1 and EP3 receptor antagonists in ICH.