Novel pathways in eicosanoid biosynthesis and metabolism

摘要 环氧合酶、脂氧合酶和细胞色素P450氧化转化花生四烯酸 上升为内源性脂质调节剂，调节体内平衡和疾病的细胞过程。这些脂类 调解人形成了一个不断进化和扩大的家庭，称为二十烷类。此应用程序包含两个 这两个项目都与二十烷类生物化学中的新转化有关，并提出了重要的 使用非类固醇抗炎药(NSAIDs)抑制其生物合成的后果。这个 第一个项目以5-LOX/COX-2交叉生物合成途径为中心，而第二个项目是 关注二十烷类化合物的新的代谢转化，以及这些转化对血浆的使用意味着什么 以及尿前列腺素代谢产物作为药物反应的标志物。今天，二十烷类的新成员 在脂质组学方法中，家族经常被发现，因为它们与已知的二十烷类化合物相似。我们有 采用了一种基于了解生物合成途径、酶、 和底物，使我们能够预测新的转变。这导致了对 5-LOX/COX-2交叉生物合成途径形成半缩酮二十烷(HKE2和HKD2)和5-LOX-2 羟基前列腺素，阿司匹林乙酰化环氧合酶-2形成的15R-前列腺素的鉴定，以及 PGD2代谢为11-脱氢-β-内酰胺酶的Baeyer-Villiger氧化途径的鉴定 血栓烷。突显我们方法的相关性的是这样一个事实，即我们拥有的新二十烷类化合物 在体外和体内形成的化合物在脂类组学分析中尚未被鉴定，可能是因为它们的 不寻常的特性，使其难以在标准分析中检测到。我们设计了分析性的 允许在体外和体内检测和定量新型二十烷类化合物的程序。我们计划继续 鉴定新二十烷类化合物并建立它们的细胞靶点和在动态平衡和生物平衡中的生物学作用 在炎症性疾病的模型中。我们正在进行的关于杂交的生物学效应的调查 二十烷类化合物已经确定受体酪氨酸激酶(RTK)VEGFR2是促血管生成的靶点 HKE2的活性，以及其他RTK和一个未知的靶点，介导抑制血小板激活 还有待进一步分析。对于5-羟基前列腺素，我们也计划采用筛选方法 作为前列腺素受体的靶向测试，以确定其细胞靶点并确定生物 效果。我们的目标是鉴定可以用来操纵5-LOX/COX-2交叉的生物合成的化合物。 不依赖于前列腺素和白三烯的形成。我们将继续 描述前列腺素的新代谢途径，并建立这些途径在体内的相关性。 新二十烷类化合物的鉴定和表征、生物学作用和新的前列腺素代谢 途径将导致对非类固醇抗炎药可能导致的病理生理状况的精细理解 阿司匹林等非甾体抗炎药的使用及其对体内前列腺素生物合成的影响。

Abstract Oxidative transformation of arachidonic acid by cyclooxygenases, lipoxygenases, and cytochromes P450 gives rise to endogenous lipid mediators that regulate cellular processes in homeostasis and disease. These lipid mediators form an evolving and expanding family referred to as eicosanoids. This application comprises two projects that both are concerned with novel transformations in eicosanoid biochemistry and present important ramifications for the use of non-steroidal anti-inflammatory drugs (NSAIDs) that inhibit their biosynthesis. The first project is centered around the 5-LOX/COX-2 cross-over biosynthetic pathway while the second project is concerned with novel metabolic transformations of eicosanoids, and what these mean for the use of plasma and urinary prostanoid metabolites as markers of drug response. Today, novel members of the eicosanoid family are often discovered in lipidomics approaches by their similarity with known eicosanoids. We have employed an approach based on understanding the structure-function of the biosynthetic pathways, enzymes, and substrates that allowed us to make predictions of novel transformations. This has led to the discovery of the 5-LOX/COX-2 cross-over biosynthetic pathway forming hemiketal eicosanoids (HKE2 and HKD2) and 5- hydroxy-prostaglandins, the identification of 15R-prostaglandins formed by aspirin-acetylated COX-2, and the identification of the Baeyer-Villiger oxidative pathway underlying the metabolism of PGD2 to 11-dehydro- thromboxanes. Underscoring the relevance of our approach is the fact that the novel eicosanoids we have shown to be formed in vitro and in vivo have not been identified in lipidomics analyses, likely due to their unusual properties that make them difficult to detect in standard analyses. We have designed analytical procedures that allow to detect and quantify the novel eicosanoids in vitro and in vivo. We plan to continue the identification of novel eicosanoids and to establish their cellular targets and biological role in homeostasis and in models of inflammatory disease. Our ongoing investigation into the biological effects of the cross-over eicosanoids has identified the receptor tyrosine kinase (RTK) VEGFR2 as a target for the pro-angiogenic activity of HKE2, as well as other RTK and an unknown target that mediates inhibition of platelet activation that are to be further analyzed. For the 5-hydroxy-prostaglandins we plan to employ screening approaches as well as targeted testing of prostanoid receptors in order to identify their cellular targets and determine biological effects. We aim to identify compounds that can be used to manipulate biosynthesis of 5-LOX/COX-2 cross- over eicosanoids independent from the formation of prostaglandins and leukotrienes. We will continue to characterize novel metabolic pathways of prostanoids and establish the relevance of these pathways in vivo. Identification and characterization of novel eicosanoids, their biological effects, and novel prostanoid metabolic pathways will result in a refined understanding of the pathophysiologic conditions for which NSAIDs may be used and effect of aspirin and other NSAIDs on prostanoid biosynthesis in vivo.