The molecular basis of IMiD induced neo-substrate recruitment to the CRL4CRBN ubiquitin E3 ligase

项目摘要 开发于20世纪50年代的沙利度胺用于治疗怀孕早期的晨吐， 导致数千名患有严重出生缺陷的儿童出生，随后退出市场。 今天，沙利度胺及其类似物来那度胺和泊马度胺(统称为IMiDS)是FDA 多发性骨髓瘤和DEL(5q)等恶性血液病的批准药物和有效治疗 MDS。但是，尽管经过60多年的研究，对这些分子如何导致 典型的出生缺陷才刚刚开始解体。由于IMIDS在临床上的广泛使用，以及 最近，靶向蛋白质降解疗法的发展呈爆炸性增长，这些疗法往往基于原始的 沙利度胺支架，比以往任何时候都更重要的是，我们必须全面了解IMIDS的潜在活动范围 挑衅。在这里，我们建议探索这些类似IMIDD的潜在活动的广泛范围 与E3连接酶CUL4-RBX1-DDB1-CRBN(CRL4CRBN)结合的分子并将其活性重定向为 不同的目标。具体地说，在目标1中，我们将设计一个体内Sall4降解的小鼠模型来解剖 沙利度胺介导的Sall4降解如何导致致畸。我们将通过以下组合来实现这一点 表达分析和发育测量以评估IMiD分子对 发育中的胚胎。这不仅有助于研究这些药物的致畸作用机制。 但也将提供一个关键的模型系统，用于测试当前和未来版本的这些 分子。这些分子不仅诱导功能增强以靶向新的蛋白质，而且它们还诱导 在目标2中，我们建议将新的基于质谱学的蛋白质组学技术用于 结合有效和选择性的工具化合物探索阻断CRBN的生物学后果 活动。我们之前已经证明，我们可以通过合理设计新的 能够降解与已知靶标相关的新蛋白质的IMID样分子。在目标3中，我们建议扩大 通过发展新的化学来靶向具有结构多样性的蛋白质，靶向范围进一步扩大 强大的治疗理论基础，从而为目标扩展提供概念验证，并提供化学先导 寻找新的治疗靶点。这笔赠款的首要目标是探索各种潜在的活动 广泛使用的CRL4CRBN连接酶通过识别和探索其降解的机制后果 生理和IMiD诱导的靶点，以及寻求将其靶点空间扩大到目前无法下药的 领地。

Project Summary Developed in the 1950s, Thalidomide was used to treat morning sickness in the early stages of pregnancy, which led to the birth of thousands of children with severe birth defects and subsequent withdrawal from the market. Today, thalidomide and its analogs lenalidomide and pomalidomide (collectively known as IMiDs) are FDA approved drugs and effective treatments for hematologic malignancies such as multiple myeloma and Del(5q) MDS. But despite over 60 years of research, the mechanistic understanding of how these molecules cause the characteristic birth defects is only just beginning to unravel. Due to the wide use of IMiDs in the clinic, and the recent explosion in the development of targeted protein degradation therapies based often on the original thalidomide scaffold, it is more critical than ever that we understand the full range of potential activities that IMiDs provoke. Here, we propose to explore the broad range of potential activities that result from these IMiD-like molecules binding to the E3 ligase CUL4-RBX1-DDB1-CRBN (CRL4CRBN) and redirecting its activity towards different targets. Specifically, in aim 1, we will engineer an in vivo mouse model of Sall4 degradation to dissect how thalidomide-mediated degradation of Sall4 leads to teratogenicity. We will do this through a combination of expression analyses and developmental measurements to assess the affect that IMiD molecules have on the developing embryo. This will not only enable the study of the mechanism of teratogenic activity of these drugs but will also provide a critical model system for testing the toxicity of current and future versions of these molecules. Not only do these molecules induce a gain-of-function to target new proteins, but they also induce a loss-of-function and in aim 2, we propose to use novel mass spectrometry-based proteomics technology in combination with potent and selective tool compounds to explore the biological consequences of blocking CRBN activity. We have previously shown that we can expand the scope of IMiD targets through rational design of new IMiD-like molecules capable of degrading new proteins related to known targets. In aim 3, we propose to expand the target scope even further by developing novel chemistry to target structurally diverse proteins that have a strong therapeutic rationale, thus providing proof-of-concept for target expansion and providing chemical leads for new therapeutic targets. The overarching goal of this grant is to explore the full range of potential activities of the widely utilized CRL4CRBN ligase by identifying and exploring the mechanistic consequences of degrading its physiological and IMiD-induced targets, as well as seeking to expand its target space into currently undruggable territories.