Development of Selective Ribosomal P-site Inhibitors

项目摘要 该项目的长期目标是开发一类新的广谱抗生素，重点是 革兰氏阴性细菌和肺结核。此外，提议的项目还将教授如何优化抑制剂。 通过定义仅在细菌中可能的关键相互作用，对原核核糖体进行研究。灵感来自于一个自然的 产品，这个类别的目标是细菌核糖体的一个未被利用的结合部位。随着越来越多的关于 对一线抗菌药物的耐药性，迫切需要新的药物，但几乎找不到 在开发管道中。因此，任何针对革兰氏阴性细菌和/或 结核病将解决未得到满足的医疗需求。 发现新的治疗线索的一个可行的方法是重新评估现有的、但未被仔细审查的课程 天然产品。通过这种方法，一个天然的产品支架被确定为一个项目的领导者 针对结核病的抗生素。对其他细菌的活性初步评估表明， 抗菌谱仅限于分枝杆菌属。然而，根据化学结构和 对于相关化合物，我们希望能够生成具有更广泛光谱活性的类似物。 在与耶鲁大学汤姆·斯泰茨的实验室合作下，最初支架的结构和两个类似物 核糖体最近被解决了。结构研究表明，天然产物铅及其类似物 结合到肽基转移酶中心(PTC)高度保守的区域，似乎传递了 原核细胞的选择性，这一点在目前的治疗中还没有被利用。高度瞄准这一点 保守区域预计将导致缓慢的抗药性。 这种结构信息被用来产生两个更有效的类似物，具有良好的物理化学特性 属性。在一个由大约30种化合物组成的非常初步的搜救活动中，我们重新设计了我们领先的一部分 用于合成简单性和稳定性的支架，提供两个类似物，引入Mtb以外的活性，以 耐甲氧西林金黄色葡萄球菌、大肠埃希菌和肺炎克雷伯菌(包括一株耐碳青霉烯的菌株)，对 真核细胞(IC50和GT；100微米)。 该项目将探索并进一步定义抑制剂与细菌核糖体的关键结合作用。 这赋予了抑制细菌蛋白质合成和广谱抗菌活性的选择性，同时 维持对哺乳动物缺乏细胞毒性。目标1将阐述我们的假设，即化合物如何 在未用药的位置选择性地与原核糖体结合。如果我们的假设是正确的，这些抑制剂将 不主动对抗哺乳动物核糖体，从而减轻了其他核糖体靶向的局限性 抗生素。目标2和目标3将探索相关化合物家族的特征，根据我们的假设，这些家族是 预期有助于更有效地抑制细菌核糖体，同时保持选择性/安全性和 增强广谱抗菌活性。

Project Summary The long-term objective of this project is to develop a new class of broad spectrum antibiotics, focused on Gram-negative bacteria and tuberculosis. In addition, the proposed project will teach how to optimize inhibitors of the prokaryotic ribosome by defining critical interactions only possible in bacteria. Inspired by a natural product, this class targets an unexploited binding site of the bacterial ribosome. With increasing reports of resistance to frontline antibacterial therapies, there is a critical need for new agents, yet very little can be found in the development pipeline. As a result, any new therapeutic that targets Gram-negative bacteria and/or tuberculosis will address an unmet medical need. A viable approach to discover new therapeutic leads is re-evaluation of existing, but under-scrutinized classes of natural products. Through this approach a natural product scaffold was identified as a lead for a program directed toward antibiotics for tuberculosis. Initial evaluation of activity against other bacteria indicated that the antibacterial spectrum was limited to Mycobacterium spp. However, based on the chemical structure and related compounds, we expected to be able to generate analogs with more broad spectrum activity. In collaboration with Tom Steitz's lab at Yale, the structure of the initial scaffold and two analogues bound to the ribosome were recently solved. The structural studies revealed that the natural product lead and analogues bind to a highly conserved region of the peptidyl transferase center (PTC) in a manner that appears to convey prokaryotic selectivity and which has not been exploited in current therapeutics. Targeting this highly conserved region is expected to lead to slow rates of resistance. This structural information was used to generate two more potent analogues with favorable physiochemical properties. In a very preliminary SAR campaign of ~30 compounds, we re-engineered a portion of our lead scaffold for both synthetic simplicity and stability to provide two analogs that introduce activity beyond Mtb, to MRSA, E. coli and K. pneumoniae (including a carbapenem-resistant strain), and do not exhibit cytotoxicity to eukaryotic cells (IC50 >100 µM). This project will explore and further define critical binding interactions for inhibitors to the bacterial ribosome that impart selectivity for inhibiting bacterial protein synthesis and broad spectrum antibacterial activity, while maintaining the lack of mammalian cytotoxicity. Aim 1 will expound on our hypothesis for how compounds can selectively bind to prokaryotic ribosomes at an undrugged site. If our hypothesis is correct, these inhibitors will not be active against mammalian ribosomes, thereby mitigating limitations of other ribosome-targeting antibiotics. Aims 2 and 3 will explore features of related compound families that, based on our hypotheses, are expected to facilitate more potent inhibition of bacterial ribosomes while maintaining selectivity/safety and enhancing broad spectrum antibacterial activity.