Structure-based inhibition of chemokine signaling in the inflamed pancreas

修改后的项目摘要/摘要部分 该项目的目标是确定CCL28在慢性胰腺炎中的作用，并使用基于结构的药物发现方法来确定这种分泌蛋白的小分子抑制物。CCL28是一种粘膜趋化因子，通过招募表达G蛋白偶联受体CCR10的调节性T细胞(Treg)来促进多种器官中的肿瘤生长。根据我们已发表的和初步的结果，我们推测胰腺导管上皮细胞分泌趋化因子CCL28也会导致慢性炎症进展为恶性疾病。我们假设抑制CCL28活性将改变胰腺粘膜微环境，从而减少癌前炎症并增强现有化疗和免疫治疗的活性。为了实现这一目标，我们提出了三个在概念上相互联系但在实验上独立的具体目标。首先，我们将在慢性胰腺炎的动物模型中验证我们的机制假说，以实验证明CCL28通过CCR10在体内的纤维炎症反应中发挥关键作用，并开发一个可用于筛选有前景的抑制剂的体外模型(目标1)。通过其G蛋白偶联受体CCR10识别CCL28的关键元素将使用核磁共振和分子建模进行详细绘制，我们将使用最先进的受体药理学分析平台定义这种趋化因子受体的完整细胞内信号图谱(AIM 2)。利用已解决的CCL28的核磁共振结构和其硫酪氨酸结合口袋的知识，我们将采用Volkman实验室开发的基于结构的策略，该策略能够发现与特定趋化因子靶标结合并抑制细胞迁移的小分子(目标3)。总而言之，拟议的研究将在以下方面提供基本的进展：(1)胰腺中Treg的功能和病理生理学；(2)配体-受体选择性的结构基础和在很大程度上未被研究的趋化因子信号轴上的GPCR药理学；(3)粘膜趋化因子在慢性炎症和肿瘤的促癌界面上的可药性。

Modified Project Summary/Abstract Section The goal of this project is to establish the role of CCL28 in chronic pancreatitis and use structure-based drug discovery methods to identify small molecule inhibitors of this secreted protein. CCL28 is a mucosal chemokine that promotes tumor growth in a variety of organs by recruiting regulatory T cells (Tregs) that express the G protein-coupled receptor CCR10. Based on our published and preliminary results, we postulate that secretion of the chemokine CCL28 by pancreatic ductal epithelial cells also drives chronic inflammation that progresses to malignant disease. We hypothesize that inhibition of CCL28 activity will alter the pancreatic mucosal microenvironment in a manner that reduces pre-malignant inflammation and enhances the activity of existing chemotherapeutics and immunotherapies. To achieve this objective, we propose three conceptually linked but experimentally independent specific aims. First, we will test our mechanistic hypothesis in animal models of chronic pancreatitis, to demonstrate experimentally that CCL28 activity through CCR10 plays a key role in the fibroinflammatory response in vivo and develop an in vitro model that can be used to screen promising inhibitors (aim 1). Key elements of CCL28 recognition by its G protein-coupled receptor CCR10 will be mapped in detail using NMR and molecular modeling, and we will define the complete intracellular signaling profile of this chemokine receptor using state-of-the-art assay platforms for receptor pharmacology (aim 2). Using the solved NMR structure of CCL28 and knowledge of its sulfotyrosine binding pocket, we will employ a structure-based strategy developed in the Volkman lab that enables the discovery of small molecules that bind a specific chemokine target and inhibit cell migration (aim 3). Collectively, the proposed studies will provide fundamental advances in our understanding of (1) Treg function and pathophysiology in the pancreas, (2) the structural basis for ligand-receptor selectivity and GPCR pharmacology in a largely unexamined chemokine signaling axis, and (3) the druggability of a mucosal chemokine at the tumor-promoting interface of chronic inflammation and neoplasia.