Mechanisms of cefiderocol resistance

头孢地罗耐药机制

• 批准号: 10115606
• 负责人: Yohei Doi
• 金额: $ 18.35万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10115606
• 关键词: Acinetobacter baumannii; Active Biological Transport; Active Sites; Address; Amino Acid Substitution; Amino Acids; Antibiotics; Antimicrobial Resistance; Binding; Binding Sites; Catechols; Ceftazidime; Cephalosporinase; Cephalosporins; Clinic; Clinical; Clinical Treatment; Collection; Complex; Cyclophosphamide; Drug resistance; Enterobacter; Enterobacteriaceae; Enterobacteriaceae Infections; Enzymes; Escherichia coli; Exposure to; Generations; Genes; Goals; Gram-Negative Bacteria; Hydrolysis; Infection; Kinetics; Klebsiella pneumoniae; Laboratories; Lactose; Membrane; Minimum Inhibitory Concentration measurement; Modeling; Mutation; Organism; Pharmaceutical Preparations; Play; Predisposition; Property; Pseudomonas aeruginosa; Public Health; Reporting; Resistance; Resolution; Risk; Role; Side; Siderophores; Stenotrophomonas maltophilia; Structure; Structure-Activity Relationship; Testing; Variant; X-Ray Crystallography; alpha helix; beta-Lactamase; beta-Lactams; carbapenem resistance; carbapenem-resistant Enterobacteriaceae; carbapenemase; clinical development; clinical encounter; clinically relevant; de novo mutation; design; inhibitor/antagonist; insight; mutant; next generation; novel; resistance mechanism; resistant strain; surveillance study

Abstract Antimicrobial resistance has become one of the greatest threats to public health, with rising resistance to carbapenems being a particular concern. Cefiderocol is a novel catechol-substituted siderophore cephalosporin which is currently undergoing late-stage clinical development. It is actively transported across the Gram-negative outer membrane and is stable against all classes of beta-lactamases, resulting in potent activity across Gram-negative bacterial species. While rare cefiderocol-resistant strains have been reported in surveillance studies, the mechanisms underlying cefiderocol resistance are largely unknown. When we tested susceptibility of our collection of carbapenem-resistant Enterobacteriaceae strains to cefiderocol, we encountered strains with MICs of 1 mg/L or higher, and sometimes greater than 4 mg/L, which is the susceptibility breakpoint for this agent. Among KPC-producing, carbapenem-resistant K. pneumoniae strains, those that produced KPC with certain amino acid substitutions associated with resistance to ceftazidime- avibactam, such as D179Y, V240G and D179Y/T243M, showed 2 to 8-fold higher cefiderocol MICs compared with strains producing wild-type KPC-3. These differences were reproduced in isogenic E. coli laboratory strains. In addition, several non-KPC-producing, carbapenem-resistant Enterobacter spp. clinical strains also showed resistance to cefiderocol. In one of these strains that showed cefiderocol MIC of >16 mg/L, we identified chromosomal AmpC beta-lactamase that contained a two amino acid deletion in the R2 loop structure. This variant AmpC conferred reduced cefiderocol susceptibility when expressed in E. coli, and was confirmed to hydrolyze cefiderocol. The structures of apo-enzyme and the enzyme–drug complex revealed the role of the deletion in extending the loop in the alpha-helix structure allowing for the accommodation of the bulky R2 side chain of cefiderocol. These preliminary findings have led us to hypothesize that specific structural changes in broad-spectrum beta-lactamases allow them to accommodate and hydrolyze cefiderocol thereby conferring reduced susceptibility or frank resistance to cefiderocol, and that some of these changes also impact hydrolysis of ceftazidime and/or binding of avibactam. To address these hypotheses, we propose the following Specific Aims: (i) To elucidate the kinetics and structure of cefiderocol-hydrolyzing beta- lactamases, and (ii) To characterize de novo variant beta-lactamases and non-enzymatic resistance mechanisms that emerge upon exposure to cefiderocol. With its unparalleled spectrum of activity across Gram- negative species, cefiderocol is likely to become a crucially important agent in the treatment of carbapenem- resistant Gram-negative infections, but information regarding the mechanisms of resistance and the risk of their emergence is non-existent. Our proposal will address these key questions to optimize use of cefiderocol in the clinic. Furthermore, insights into the structure-activity relationship of substrate-binding sites of cefiderocol-hydrolyzing beta-lactamases will inform designing of the next-generation cephalosporins.

摘要 抗生素耐药性已成为对公共卫生的最大威胁之一， 碳青霉烯类是一个特别关注的问题。Cefiderocol是一种新的儿茶酚取代的铁载体 头孢菌素，目前正在进行后期临床开发。它被积极地传送到 革兰氏阴性外膜，对所有类型的β-内酰胺酶都是稳定的， 在革兰氏阴性细菌物种中的活性。虽然在美国已经报道了罕见的头孢地罗克耐药菌株， 尽管在监测研究中，头孢地罗可耐药的潜在机制在很大程度上是未知的。当我们测试 我们收集耐碳青霉烯类肠杆菌科菌株对头孢地罗可的敏感性， 遇到MIC为1 mg/L或更高的菌株，有时大于4 mg/L，这是 敏感性断点。在KPC产生菌中，碳青霉烯类耐药K.肺炎菌株， 那些产生具有与头孢他啶抗性相关的某些氨基酸取代的KPC的人- 阿维巴坦（如D179 Y、V240 G和D179 Y/T243 M）的头孢地罗可MIC是头孢地罗可的2 - 8倍， 用产生野生型KPC-3的菌株。这些差异在等基因E.大肠杆菌实验室 菌株此外，几种不产KPC的碳青霉烯类耐药肠杆菌属（Enterobacter spp.）临床菌株还 对头孢地罗克耐药。在显示头孢地罗可MIC>16 mg/L的这些菌株之一中，我们 鉴定了一种在R2环中含有两个氨基酸缺失的染色体AmpC β-内酰胺酶 结构当在E.大肠杆菌， 证实水解头孢地罗可。脱辅基酶和酶-药物复合物的结构表明， 缺失在延伸α-螺旋结构中的环中的作用，从而允许调节 Cefiderocol的大R2侧链。这些初步的发现使我们假设， 广谱β-内酰胺酶的结构变化使其能够适应和水解头孢地罗醇 从而降低对头孢地罗可的敏感性或产生明显耐药性， 也影响头孢他啶的水解和/或阿维巴坦的结合。为了解决这些问题，我们建议 以下具体目的：（i）阐明头孢地罗醇水解β- 内酰胺酶，和（ii）表征新生变异β-内酰胺酶和非酶耐药性 暴露于头孢地罗可后出现的机制。凭借其无与伦比的活动范围在革兰氏- 阴性种属，头孢地罗可可能成为碳青霉烯治疗中至关重要的药物， 耐药革兰氏阴性菌感染，但有关耐药机制和风险的信息， 他们的出现是不存在的。我们的提案将解决这些关键问题，以优化头孢地罗可的使用 在诊所里此外，深入了解底物结合位点的结构-活性关系， 头孢地洛水解β-内酰胺酶将为下一代头孢菌素的设计提供信息。