Discovery of metabolic pathways using phenotypic screening and chemical proteomics

项目摘要 肥胖症是一个主要的公共卫生问题，其特征在于脂肪组织的大量扩张。下 在慢性过量能量摄入的情况下，脂肪细胞膨胀以储存多余的营养物质。在肥胖症中， 增大的脂肪细胞不能再对胰岛素作出反应，不能正常分泌胰岛素增敏激素， 过量的脂质，然后存款在组织，如肝脏和肌肉，在那里他们阻碍胰岛素的作用。肥胖症- 相关的脂肪细胞功能障碍驱动代谢紊乱如2型糖尿病（T2 D）的发病机制， 非酒精性脂肪肝（NALFD）。可以逆转肥胖导致的脂肪细胞缺陷并恢复 组织间的正常脂质分配是人体中有用的胰岛素增敏剂。 Saez和Cravatt实验室先前开发了一种创新策略， 利用化学蛋白质组学来简化生物活性小分子的蛋白质靶点的鉴定， 在蛋白质组范围内快速发现新的代谢靶点。通过筛选独特的小分子库， 脂肪细胞中所需表型的分子（例如，增加脂质潴留），他们发现在串联 生理学相关的蛋白质和化学工具来扰乱这些蛋白质的功能，以促进它们的功能。 T2 D的功能表征和治疗验证。在过去的工作中，他们应用这种策略来研究 脂肪细胞生理学，以及鉴定和治疗验证的T2 D新靶点（例如，Ces3，PGRMC2）。 在这个项目中，我将通过筛选理想的效果来推进这种配体-蛋白质靶点发现策略， 脂肪细胞（例如，增加的脂质保留）立体化学限定的化学探针库。的 立体化学的引入允许一组化学探针具有相似的生理化学性质， 但在结合蛋白质的能力上不同，因此有助于鉴定 生物活性探针我的目标是应用这种整合的表型筛选和化学蛋白质组学策略 利用这个新的立体选择性探针库， 调节以恢复肥胖诱导的脂肪细胞缺陷。这是我的期望，一些新的途径，我 discover可能为肥胖相关疾病的新治疗策略的开发提供基础。 提案中包括的大量初步数据确定了这一方法的可行性。我已经 鉴定了几种立体选择性促进脂肪细胞分化和脂质储存的探针。在一个 例如，我还鉴定了与前脂肪形成化合物相互作用的蛋白质。因此，我准备 确定新的代谢途径，可以有针对性地治疗肥胖驱动的疾病。

PROJECT SUMMARY Obesity is a major public health problem that is characterized by a vast expansion of adipose tissue. Under conditions of chronic excess energy intake, adipocytes expand to store surplus nutrients. In obesity, chronically enlarged adipocytes can no longer respond to insulin, properly secrete insulin-sensitizing hormones, or retain excess lipids, which then deposit in tissues such as liver and muscle where they hinder insulin action. Obesity- linked adipocyte dysfunction drives the pathogenesis of metabolic disorders such as type 2 diabetes (T2D) and nonalcoholic fatty liver disease (NALFD). Agents that can revert obesity-driven adipocyte defects and restore normal lipid partitioning amongst tissues are useful insulin sensitizers in humans. The Saez and Cravatt labs previously developed an innovative strategy that integrates phenotypic screening with chemical proteomics to streamline the identification of protein targets of bioactive small molecules and rapidly uncover new metabolic targets on a proteome-wide scale. By screening unique libraries of small molecules for desirable phenotypes in adipocytes (e.g., increased lipid retention), they identified in tandem physiologically relevant proteins and chemical tools to perturb the function of these proteins to expedite their functional characterization and therapeutic validation in T2D. In past work, they applied this strategy to study adipocyte physiology, and identified and therapeutically validated new targets for T2D (e.g., Ces3, PGRMC2). In this project, I will advance this ligand-protein target discovery strategy by screening for desirable effects in adipocytes (e.g., increased lipid retention) a library of chemical probes that are stereochemically defined. The introduction of stereochemistry allows for a set of chemical probes to have similar physiochemical properties, yet differ in the ability to engage proteins, thus facilitating the identification of the relevant protein targets of bioactive probes. My goal is to apply this integrated phenotypic screening and chemical proteomics strategy using this new library of stereoselective probes to discover metabolic pathways that can be pharmacologically modulated to revert obesity-induced adipocyte defects. It is my expectation that some of the novel pathways I discover may provide the basis for the development of new therapeutic strategies for obesity-linked conditions. Extensive preliminary data included in the proposal establishes the feasibility of this approach. I have already identified several probes that stereoselectively promote adipocyte differentiation and lipid storage. In one instance, I have also identified the proteins that interact with the proadipogenic compound. Thus, I am poised to identify novel metabolic pathways that can be targeted to treat obesity-driven maladies.