Small molecule inhibitors of lytic transglycosylase to potentiate beta-lactam antibiotics

裂解性转糖基酶小分子抑制剂可增强 β-内酰胺抗生素的作用

基本信息

批准号：
10078254
负责人：
FOCCO VAN DEN AKKER
金额：
$ 21.01万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-01-01 至 2022-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10078254
关键词：
Acinetobacter baumannii Active Sites Anabolism Animals Anti-Bacterial Agents Antibiotics Antimicrobial Resistance Bacteria Bacterial Antibiotic Resistance Bacterial Proteins Binding Biological Biological Assay Campylobacter jejuni Carbohydrates Cell Death Cell Wall Cells Chemicals Chemosensitization Crystallography Cytolysis Cytoplasm Disaccharides Docking Drug Targeting Enterobacter cloacae Enzyme Inhibitor Drugs Enzymes Escherichia coli Glucosamine Helicobacter pylori Klebsiella aerogenes Knock-out Label Libraries Link Lytic Mechanics Microbiology Monobactams Multiple Bacterial Drug Resistance Natural Products Neisseria Neisseria gonorrhoeae Penicillin-Binding Proteins Peptidoglycan Pharmaceutical Chemistry Proteins Pseudomonas aeruginosa Pyrrolidines Resistance Shapes Structure Testing bacterial resistance base beta-Lactamase beta-Lactams biophysical techniques crosslink design global health glycosyltransferase improved in silico inhibitor/antagonist novel novel therapeutics pathogen pathogenic bacteria protein crosslink resistance mechanism scaffold screening small molecule inhibitor tool

项目摘要

The rapid emergence of antibiotic-resistant bacteria is a major global health threat. This spurs the need to revisit key antibacterial drug targets such as the peptidoglycan (PG) layer. The PG synthesis machinery is targeted by β-lactam antibiotics that inhibit penicillin-binding proteins (PBP) which crosslink PG strands. A main resistance mechanism is the expression of β-lactamases that can degrade β-lactams. There is thus a critical need for new antibiotics or for avenues to re-sensitize bacteria to β-lactam antibiotics. For the latter, one approach is developing β-lactamase inhibitors; unfortunately, the 5 current inhibitors do not inhibit certain key β-lactamases, and there are resistance mechanisms via inhibitor-resistant β-lactamases. A second approach is to inhibit PG degrading lytic transglycosylases (LT), the focus of this application. Inhibition of LTs or knocking out LTs genetically has been shown to restore the efficacy of β-lactam antibiotics in many serious pathogens including Escherichia coli, Neisseria meningitides, Pseudomonas aeruginosa, Enterobacter aerogenes, Acinetobacter baumannii, Helicobacter pylori, and Campylobacter jejuni. This β-lactam potentiation involves two possible mechanisms of which, depending on the pathogen, either or both contribute. In the first mechanism, the inhibition of both PBP and LT leads to long non-cross-linked PG strands that cause cell wall bulges, weakening the cell wall. In the second mechanism, LT activity generates disaccharide PG product that, when recycled to the cytoplasm, increases β-lactamase expression in certain pathogens. Despite these compelling observations, there is only one promising LT inhibitor known, bulgecin A; however, this natural product carbohydrate-based inhibitor is very challenging for medicinal chemistry efforts. As a result, bulgecin A has not been very amenable to advancing inhibition studies towards animal studies and beyond. This application proposes to overcome this key roadblock by developing new LT inhibitors with scaffold(s) different from bulgecin A via biased (Aim 1) and non-biased fragment-based approaches (Aim 2). Aim 1: To identify new inhibitor fragments that retain bulgecin A's key N-acetyl group. N-acetyl containing compounds will be selected or designed aided by docking; their LT binding and inhibition will be probed by biophysical techniques, protein crystallography, and enzymatic assays. Compounds will be tested against multiple LTs known to bind bulgecin A, and which are amenable to crystallography (E. coli, P. aeruginosa, and C. jejuni) in order to identify at least one fragment binding to one LT as a novel starting point for optimization. Aim 2: To identify non-acetyl containing fragments that bind to the active site of LT, we will screen non- biased fragments against LTs for binding and inhibition as in Aim 1. Such fragments could bind to the N-acetyl binding pocket or to the adjacent pockets. Compounds will be obtained from an sp3 fragment library and from in silico screening of larger libraries. Hits from Aims 1 and 2 will be modified/grown/linked to improve binding and inhibition of one or more of the LTs and also tested microbiologically for the potentiation of β-lactam antibiotics.

抗生素耐药细菌的迅速出现是全球健康的主要威胁，这促使我们重新审视这一问题。肽聚糖 (PG) 层等关键抗菌药物靶点是 PG 合成机制的目标。 β-内酰胺类抗生素可抑制青霉素结合蛋白 (PBP)，该蛋白可交联 PG 链，是一种主要耐药性。机制是可以降解β-内酰胺的β-内酰胺酶的表达，因此迫切需要新的。抗生素或使细菌对β-内酰胺抗生素重新敏感的途径。对于后者，一种方法是。正在开发 β-内酰胺酶抑制剂；不幸的是，目前的 5 种抑制剂不能抑制某些关键的 β-内酰胺酶，第二种方法是抑制 PG。降解裂解性转糖基酶 (LT)，是本应用的重点。抑制 LT 或从基因上敲除 LT 已被证明可以恢复 β-内酰胺的功效许多严重病原体的抗生素，包括大肠杆菌、脑膜炎奈瑟菌、假单胞菌铜绿假单胞菌、产气肠杆菌、鲍曼不动杆菌、幽门螺杆菌和空肠弯曲杆菌。这种 β-内酰胺增强作用涉及两种可能的机制，其中任一机制取决于病原体，或者在第一种机制中，PBP 和 LT 的抑制都会导致长的非交联 PG。导致细胞壁凸起、削弱细胞壁的链在第二种机制中，LT 活性产生。二糖 PG 产品，当再循环到细胞质时，会增加某些细胞中 β-内酰胺酶的表达尽管有这些令人信服的观察结果，但已知只有一种有前途的 LT 抑制剂：bulgecin A；然而，这种基于碳水化合物的天然产物抑制剂对于药物化学工作来说非常具有挑战性。因此，bulgecin A 不太适合将抑制研究推进到动物研究和该申请建议通过开发新的 LT 抑制剂来克服这一关键障碍。通过有偏向（目标 1）和无偏向片段方法（目标 2）构建与 bulgecin A 不同的支架。目标 1：鉴定保留了 bulgecin A 的关键 N-乙酰基的新抑制剂片段。将通过对接来选择或设计化合物；将通过以下方法探测它们的 LT 结合和抑制作用：将针对生物物理技术、蛋白质晶体学和酶分析进行测试。已知多种 LT 可以结合 bulgecin A，并且适合晶体学分析（大肠杆菌、铜绿假单胞菌和 C. jejuni），以确定至少一个与一个 LT 结合的片段作为优化的新起点。目标 2：为了识别与 LT 活性位点结合的不含乙酰基的片段，我们将筛选非乙酰基片段。如目标 1 所示，偏向针对 LT 的片段进行结合和抑制。此类片段可以与 N-乙酰基结合结合口袋或邻近口袋的化合物将从sp3片段文库中获得。来自目标 1 和 2 的命中结果将被修改/生长/链接以改善结合和抑制一种或多种 LT，并通过微生物学测试 β-内酰胺抗生素的增强作用。

项目成果

期刊论文数量（2）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Turnover Chemistry and Structural Characterization of the Cj0843c Lytic Transglycosylase of Campylobacter jejuni.

空肠弯曲杆菌 Cj0843c 裂解性转糖基酶的周转化学和结构表征。

DOI：
发表时间：
2021-04-13
期刊：
影响因子：
2.9
作者：
Kumar, Vijay;Mathure, Snigdha A;Lee, Mijoon;Boorman, Jacob;Zeng, Ximin;Lin, Jun;Hesek, Dusan;Lastochkin, Elena;Mobashery, Shahriar;van den Akker, Focco
通讯作者：
van den Akker, Focco

Exploring the inhibition of the soluble lytic transglycosylase Cj0843c of Campylobacter jejuni via targeting different sites with different scaffolds.

通过使用不同支架靶向不同位点，探索对空肠弯曲杆菌可溶性裂解转糖基酶 Cj0843c 的抑制。