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) 是经过验证的 β-内酰胺药物靶标，但对 β-内酰胺具有耐药性 碳青霉烯类和头孢菌素等抗生素已导致全球健康问题。有一个 多种耐药机制，其中β-内酰胺降解β-内酰胺酶是罪魁祸首之一。 我们的目标是通过研究和克服通常与 β-内酰胺相关的耐药机制 开发一种不同类型的 PBP 抑制剂，吡唑烷酮。吡唑烷酮类 YU253434 和 YU253911 含有铁载体部分，以促进铁介导的吸收。我们发现这两个 吡唑烷酮类不能被 A、C 和 D 类 β-内酰胺酶水解，只能缓慢水解 通过（B 类）金属 β-内酰胺酶。 YU253434和YU253911也优于氨曲南， 微生物学上的头孢他啶、美罗培南、头孢他嗪/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、大肠杆菌和 鲍曼不动杆菌（所有四种均为 ESKAPE 病原体）。 YU253911 还显着降低了菌落- 在具有耐多药铜绿假单胞菌菌株的小鼠大腿感染模型中形成单位。吡唑烷酮类目标 并抑制 PBP3，我们已经通过晶体学描述了它们与铜绿假单胞菌 PBP3 的结合模式。 总的来说，我们发现这些吡唑烷酮具有一些有利的特性，但仍需进一步改进 在 PBP3 IC50、摄取和克服已知 PBP3 F533L 耐药性的能力方面需要 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. 通过针对 R2 的基于结构的修饰来提高吡唑烷酮亲和力 团体。根据晶体结构，我们假设疏水取代基添加到 铁载体连接子中的吡唑烷酮将与疏水桥残基 F533 和 V333 相互作用 PBP3 中，从而提高亲和力。此外，这些疏水相互作用旨在 抵消F533L耐药突变。我们将针对以下小组对这些化合物进行微生物学测试 经过充分研究的肺炎克雷伯菌、鲍曼不动杆菌、大肠杆菌和铜绿假单胞菌，测量 PBP 抑制 wt 和 F533L 铜绿假单胞菌 PBP3，并从晶体学、生物物理上探讨它们的结合模式， 并使用分子动力学模拟。 具体目标 2. 通过添加来改善吡唑烷酮类的非铁依赖性和铁介导的吸收 将含胺部分连接到铁载体连接子上（目标 2a），并通过将铁载体与 与羟基相邻的吸电子 –Cl 基团可提高铁亲和力（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 PBP 靶向药物 吡唑烷酮可能会导致一种新的治疗策略来对抗抗生素耐药性。