Chemical, Structural and Cell-Signaling Interrogation of 15-Prostanglandin Dehydrogenase in Tissue Repair and Regeneration

抽象的。前列腺素E2(PGE2)调节多个器官的组织生长和修复。一个保守的人 合成和降解机制调节PGE2水平以应对创伤、炎症和 疾病。特别是，15-前列腺素脱氢酶(15-PGDH)是PGE2的主要降解酶 酶，因此是组织修复和再生的关键调节器。15-前列腺素脱氢酶是有吸引力的药物靶点 用于以组织损伤为特征的疾病。我们团队成功地研制出了第一个小分子 具有体内活性的15-前列腺素脱氢酶抑制剂。在啮齿动物身上，我们的抑制剂1)加速了骨骼的恢复 骨髓移植，2)加速溃疡性结肠炎的康复，或预防，3)加速再生 肝部分切除后的肝组织，4)改善博莱霉素性疾病的肺纤维化 5)提高成年小鼠新生海马神经元的存活率，6)保护认知功能和 减少小鼠创伤性脑损伤后的神经元损伤。独立报告描述了 15-前列腺素脱氢酶抑制在肾脏疾病和肺纤维化模型中的有益作用。 我们现在提出一个协作性的化学，结构和细胞信号询问的作用和活性的 15-前列腺素脱氢酶。我们的专长包括药物化学、生物化学、神经科学、药理学和 结构生物学。在目标1中，我们将定义和开发小分子抑制15-PGDH的结构基础 分子。这一目标建立在15-PGDH的第一个低温EM结构(2.3？分辨率)和两个不相关的 15-前列腺素脱氢酶的低NM抑制剂支架。拟议的研究旨在解决15-PGDH的结构 与新的小分子抑制剂或底物形成复合体。将采用计算方法来 询问底物/抑制物结合和酶机制。在目标2中，我们将定义细胞， 由15-前列腺素脱氢酶调控并由15-前列腺素脱氢酶参与的蛋白质和细胞因子信号网络 促进组织再生和修复的抑制剂。这一目标的基础包括第一个 15-前列腺素脱氢酶在脑内的活性证明，巨噬细胞和小胶质细胞是主要的 外周组织和脑组织中15-PGDH表达的储存库，以及细胞和 细胞因子网络对抑制15-前列腺素脱氢酶有反应。我们现在建议使用单细胞RNA测序来 确定表达15-前列腺素脱氢酶的细胞类型。类似的方法将确定细胞信令网络 诱导的细胞因子和被激活的细胞类型来表达它们。这些研究将在小鼠身上进行。 受伤后接受15-前列腺素脱氢酶抑制剂治疗的患者，以及适当的对照组。最后， 我们将设计巨噬细胞和小胶质细胞靶向的15-前列腺素脱氢酶基因敲除，以确定15-前列腺素脱氢酶的作用 巨噬细胞和小胶质细胞表达介导对PGE2和15-P保守的跨组织反应 前列腺素脱氢酶抑制剂。该数据集将为以15-15为目标的治疗学的未来发展提供基础 和其他调节组织再生的药物靶点。

Abstract. Prostaglandin E2 (PGE2) regulates tissue growth and repair in multiple organs. A conserved mechanism of synthesis and degradation modulates PGE2 levels in response to trauma, inflammation and disease. In particular, the enzyme 15-prostaglandin dehydrogenase (15-PGDH) is the main PGE2-degrading enzyme and therefore a key regulator of tissue repair and regeneration. 15-PGDH is an attractive drug target for diseases characterized by tissue damage. Our team successfully developed the first small molecule inhibitors of 15-PGDH with in vivo activities. In rodents, our inhibitors 1) accelerate recovery following bone marrow transplantation, 2) accelerate recovery from, or prevent, ulcerative colitis, 3) accelerate regrowth of liver tissue following partial hepatectomy, 4) ameliorate pulmonary fibrosis in a bleomycin-induced disease model, 5) enhance survival of new hippocampal neurons in adult mice, and 6) preserve cognitive function and minimize neuronal damage in mice following traumatic brain injury. Independent reports have described beneficial effects of 15-PGDH inhibition in models of renal disease and pulmonary fibrosis. We now propose a collaborative chemical, structural and cell-signaling interrogation of the role and activity of 15-PGDH. Our expertise includes medicinal chemistry, biochemistry, neuroscience, pharmacology, and structural biology. In Aim 1, we will define and exploit the structural basis for inhibition of 15-PGDH by small molecules. This aims builds on the first cryoEM structure (2.3 Å resolution) of 15-PGDH and two unrelated scaffolds of low-nM inhibitors of 15-PGDH. Proposed research aims to solve the structure of 15-PGDH in complex with new small molecule inhibitors or substrate. Computational approaches will be employed to interrogate substrate/inhibitor binding and the enzymatic mechanism. In Aim 2, we will define the cellular, protein and cytokine signaling networks that are regulated by 15-PGDH and that are engaged by 15-PGDH inhibitors to potentiate tissue regeneration and repair. The foundation of this aim includes the first demonstration of 15-PGDH activity in the brain, the identification of macrophages and microglia as major reservoirs of 15-PGDH expression in peripheral tissues and brain, respectively, and the discovery of cell and cytokine networks that respond to inhibiting 15-PGDH. We now propose to use single-cell RNA sequencing to determine the cell types that express 15-PGDH. Similar approaches will identify the cell-signaling network of induced cytokines and the cell types activated to express them. These studies will be performed in mice recovering from injury that have been treated with 15-PGDH inhibitors, along with appropriate controls. Finally, we will engineer macrophage- and microglia-targeted 15-PGDH knockouts to define the role of 15-PGDH expression in macrophages and microglia in mediating a conserved, cross-tissue response to PGE2 and 15- PGDH inhibitors. This data set will provide a foundation for future advancement of therapeutics targeting 15- PGDH and additional drug targets that modulate tissue regeneration.