Developing novel pyrazolidinone antibiotics targeting PBP3 to overcome resistance mechanisms

开发针对 PBP3 的新型吡唑烷酮抗生素以克服耐药机制

• 批准号: 10590839
• 负责人: FOCCO VAN DEN AKKER
• 金额: $ 24.15万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-21 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10590839
• 关键词: Acceleration; Acinetobacter baumannii; Affinity; Amines; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Antimicrobial Resistance; Autolysis; Aztreonam; Bacteria; Bacterial Antibiotic Resistance; Bacterial Counts; Binding; Biological Testing; Biophysics; Bypass; Carbapenems; Ceftazidime; Cell Death; Cell Wall; Cell division; Cephalosporins; Chemicals; Colony-forming units; Crystallography; Development; Dose; Drug Kinetics; Drug Targeting; ESKAPE pathogens; Electrons; Enzymes; Escherichia coli; Goals; Gram-Negative Bacteria; Horizontal Gene Transfer; Hydrophobic Interactions; Hydrophobicity; Hydroxyl Radical; Individual; Infection; Iron; Iron Chelating Agents; Iron Chelation; Klebsiella pneumoniae; Lactams; Length; Measures; Mechanics; Mediating; Membrane; Meropenem; Modeling; Multi-Drug Resistance; Mus; Mutation; Penicillin-Binding Proteins; Peptidoglycan; Periodicity; Predisposition; Property; Proteins; Pseudomonas aeruginosa; Reporting; Resistance; Shapes; Side; Siderophores; Site-Directed Mutagenesis; Structure; Tazobactam; Testing; Thigh structure; VDAC1 gene; analog; beta-Lactam Resistance; beta-Lactamase; beta-Lactams; carbapenem resistance; combat; crosslink; design; efflux pump; flexibility; genome sequencing; global health; high reward; high risk; hydroxyl group; improved; inhibitor; molecular dynamics; novel; novel therapeutic intervention; resistance gene; resistance mechanism; resistance mutation; resistant Klebsiella pneumoniae; resistant strain; trend; uptake; whole genome

Penicillin-binding proteins (PBPs) are a proven β-lactam drug target, yet resistance to β-lactam antibiotics, such as carbapenems and cephalosporins, has resulted in a global health problem. There are a number of resistance mechanisms of which β-lactam degrading β-lactamases is one of the main culprits. Our goal is to overcome the resistance mechanisms often associated with β-lactams by studying and developing a different type of PBP inhibitor, the pyrazolidinone. The pyrazolidinones YU253434 and YU253911 contain a siderophore moiety to facilitate iron-mediated uptake. We have found that these two pyrazolidinones cannot be hydrolyzed by Classes A, C, and D β-lactamases and are only slowly hydrolyzed by (Class B) metallo β-lactamases. YU253434 and YU253911 also compared favorably to aztreonam, ceftazidime, meropenem, ceftolozane/tazobactam, and ceftazidime/avibactam when microbiologically tested against panels of Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, and Acinetobacter baumannii (all four are ESKAPE pathogens). YU253911 also significantly lowered colony- forming units in a mouse thigh-infection model with an MDR P. aeruginosa strain. The pyrazolidinones target and inhibit PBP3, and we have delineated their binding modes to P. aeruginosa PBP3 crystallographically. Overall, we found that these pyrazolidinones have several favorable attributes, yet further improvements are needed in terms of PBP3 IC50, uptake, and ability to overcome the known PBP3 F533L resistance mutation. We propose these improvements in the following two specific Aims. Specific Aim 1. To improve the pyrazolidinone affinity via structure-based modifications targeting the R2 group. Based on the crystal structure, we hypothesize that hydrophobic substituents added to pyrazolidinones in the siderophore-linker will interact with the hydrophobic bridge residues F533 and V333 in PBP3 and thereby improve affinity. Additionally, these hydrophobic interactions are designed to counteract the F533L resistance mutation. We will test the compounds microbiologically against panels of well-studied K. pneumoniae, A. baumannii, Escherichia coli, and P. aeruginosa, measure PBP inhibition both wt and F533L P. aeruginosa PBP3, and probe their binding mode crystallographically, biophysically, and using molecular dynamics simulations. Specific Aim 2. To improve the iron–independent and iron-mediated uptake of pyrazolidinones via adding an amine-containing moiety to the siderophore-linker (Aim 2a) and by incorporating a siderophore with an electron-withdrawing –Cl group adjacent to the hydroxyl groups to improve iron affinity (Aim 2b). This high-risk, high-reward proposal aims to develop a more potent non-β-lactam PBP-targeting pyrazolidinone that could lead to a novel therapeutic strategy to combat antibiotic resistance.

青霉素结合蛋白（PBP）是β-内酰胺类药物的靶点，但对β-内酰胺类药物耐药 抗生素如碳青霉烯类和头孢菌素类已经导致了全球性的健康问题。有 β-内酰胺降解酶是其主要的耐药机制之一。 我们的目标是通过研究， 开发一种不同类型的PBP抑制剂，吡唑烷酮。吡唑烷酮YU 253434和 YU 253911含有铁载体部分，以促进铁介导的摄取。我们发现这两个 吡唑烷酮不能被A、C和D类β-内酰胺酶水解，只能缓慢水解 （B类）金属β-内酰胺酶。YU 253434和YU 253911也优于氨曲南， 头孢他啶、美罗培南、头孢洛扎/他唑巴坦和头孢他啶/阿维巴坦， 针对铜绿假单胞菌、肺炎克雷伯菌、大肠埃希菌和 鲍曼不动杆菌（所有四种都是ESKAPE病原体）。YU 253911还显著降低了菌落- 在具有MDR铜绿假单胞菌菌株的小鼠大腿感染模型中形成单位。吡唑烷酮靶向 并抑制PBP 3，并且我们已经在晶体学上描绘了它们与铜绿假单胞菌PBP 3的结合模式。 总的来说，我们发现这些吡唑烷酮具有几个有利的属性，但进一步的改进 在PBP 3 IC 50、摄取和克服已知的PBP 3 F533 L抗性的能力方面， 突变我们在以下两个具体目标中提出这些改进。 具体目标1。通过靶向R2的基于结构的修饰来改善吡唑烷酮亲和力 组基于晶体结构，我们假设疏水取代基添加到 铁载体-接头中的吡唑烷酮将与疏水桥残基F533和V333相互作用 从而提高亲和力。此外，这些疏水相互作用被设计成 抵消F533 L抗性突变。我们将对这些化合物进行微生物测试， 研究K。pneumoniae、A.鲍曼不动杆菌、大肠杆菌和铜绿假单胞菌测量PBP抑制作用 wt和F533 L铜绿假单胞菌PBP 3，并在晶体学，生物病理学， 并使用分子动力学模拟。 具体目标2。为了改善吡唑烷酮的铁非依赖性和铁介导的摄取， 将含胺部分引入铁载体-接头（Aim 2a），并通过将铁载体与含胺部分结合， 吸电子-Cl基团与羟基相邻，以提高铁亲和力（目标2b）。 这项高风险、高回报的提案旨在开发一种更有效的非β-内酰胺类PBP靶向药物 吡唑烷酮，可能导致一种新的治疗策略，以打击抗生素耐药性。