Broad spectrum antibacterials selectively targeting an un-drugged site on the ribosome

项目摘要 该项目的最终目标是开发一类新的广谱抗生素，重点是革兰氏阴性菌， 阴性细菌。受天然产物的启发，这类目标是细菌的未开发的结合位点。 选择性抑制细菌蛋白质合成的核糖体。随着越来越多的关于耐药性的报道， 尽管目前的一线治疗方法主要针对革兰氏阴性菌，但仍迫切需要新的治疗方法，但很少有新的治疗方法。 在开发管道中发现。因此，任何针对革兰氏阴性菌的新疗法都将 解决未满足的医疗需求。 天然产物和与T.嗜热核糖体已经被 用于指导药物化学工作。这些晶体结构揭示了一个高度保守的区域 尚未被任何现有抗生素利用的肽基转移酶中心（PTC）实际上是一种可行的 一种针对核糖体的新治疗方法。与PTC的该位点结合似乎 传递细菌选择性。使用这些配合物作为初步优化指导的分子建模 已经产生了新的线索，已经扩大了抗菌谱的活动，从单一的结核分枝杆菌，包括 对E. coli、克雷伯氏菌（K. pneumoniae和S.金黄色葡萄球菌（包括每一种的耐药菌株）。 这些类似物在nM浓度范围内抑制蛋白质合成，对蛋白质的选择性指数>400。 抑制细菌对真核生物蛋白质合成，对哺乳动物细胞的毒性非常有限。 这个为期五年的R 01项目最终将开发出一种新的强效、广谱抗菌药物候选药物， 专注于准备IND制备的革兰氏阴性病原体。本抗菌铅系列的开发 将通过以下目标来实现。目标1将优化铅系列，以扩大活动，以增加 革兰氏阴性病原体，特别是铜绿假单胞菌和A.鲍曼不动杆菌。目标2将定义作用机制， 生物化学/微生物学活性和体外ADME-Tox，通过评估抗细菌的活性， 和真核蛋白质合成（包括线粒体），一组代表革兰氏阴性和 革兰氏阳性菌（包括耐药菌）。将使用以下方法在体外评价初步ADME-Tox Vero细胞预测细胞毒性、微粒体稳定性、hERG和细胞色素P450抑制、艾姆斯试验 遗传毒性和44个主要分子靶点的体外受体结合试验。目标3将使用体内PK 在小鼠大腿感染模型中指导优化，然后确定MTD和体内疗效， 选择单一的先导化合物进行推进。目标4将通过确定 MIC 90，对其他抗生素的耐药性和潜在交叉耐药性的表征。Aim 5将在 对肺炎感染模型的体内功效和PK/PD。目标6将解决CMC问题，并提出初步的 用于目标7中GLP毒理学研究的cGMP批次。

Project Summary The ultimate goal of this project is developing a new class of broad spectrum antibiotics, focused on Gram- negative bacteria. Inspired by a natural product, this class targets an unexploited binding site of the bacterial ribosome that selectively inhibits bacterial protein synthesis. With ever increasing reports of resistance to frontline therapies addressing Gram-negatives, there is a critical need for new therapies, yet very little can be found in the development pipeline. As a result, any new therapeutic that targets Gram-negative bacteria will address an unmet medical need. Crystal structures of the natural product and Cūrza analogs bound to the T. thermophilus ribosome have been used to guide medicinal chemistry efforts. These crystal structures revealed that a highly conserved region of the peptidyl transferase center (PTC) that has yet to be exploited by any existing antibiotics is in fact a viable new therapeutic approach against a well-vetted target, the ribosome. Binding to this site of the PTC appears to convey bacterial selectivity. Molecular modeling using these complexes as a guide for preliminary optimization has produced new leads that have expanded the antibacterial spectrum of activity from solely Mtb to include potent activity against E. coli, K. pneumoniae and S. aureus (including drug-resistant strains for each of these). These analogs inhibit protein synthesis in the nM concentration range, exhibit selectivity indexes of >400 for inhibition of bacterial versus eukaryotic protein synthesis and have very limited toxicity to mammalian cells. This five-year R01 project will ultimately develop a new potent, broad spectrum antibacterial drug candidate, focusing on Gram-negative pathogens ready for IND preparation. Development of this antibacterial lead series will be accomplished by the following aims. Aim 1 will optimize the lead series to expand activity to additional Gram-negative pathogens, notably P. aeruginosa and A. baumannii. Aim 2 will define mechanism of action, biochemical/microbiological activity and in vitro ADME-Tox of analogs by evaluating activity against bacterial and eukaryotic protein synthesis (including mitochondrial), a panel of bacteria representing Gram-negative and Gram-positive organisms (including antibiotic-resistant). Preliminary ADME-Tox will be evaluated in vitro using Vero cells to predict cytotoxicity, microsomal stability, inhibition of hERG and cytochrome P450s, Ames test for genotoxicity and in an in vitro receptor binding assay of 44 primary molecular targets. Aim 3 will use in vivo PK to guide optimization followed by determining the MTD and in vivo efficacy in a mouse thigh infection model to select a single lead compound for advancement. Aim 4 will expand the microbiological profile by determining MIC90s, characterization of resistance and potential cross-resistance to other antibiotics. Aim 5 will expand in vivo efficacy and PK/PD to pneumonia infection models. Aim 6 will address CMC issues and produce an initial cGMP lot for use in GLP toxicology studies in Aim 7.