Precise Combination Strategies Targeting Carbapenem-Resistant Klebsiella pneumoniae

项目总结/摘要：耐碳青霉烯类肺炎克雷伯菌（CR-Kp）导致生命- 威胁感染，与不可接受的高死亡率相关。CR-Kp特别是 治疗具有挑战性，因为它通常具有无数的分子抗性决定因素，使其能够生长 在大多数抗生素存在的情况下。新型β-内酰胺/β-内酰胺酶抑制剂（BL/BLI），如头孢他啶- 阿维巴坦作为单药治疗不是CR-Kp的可持续治疗解决方案，因为有可能 当单独使用时，耐药性的产生和临床失败率仍然非常高。高达95%的CR- Kp仍然对至少一种氨基糖苷类（AMG）敏感。然而，几乎所有的CR-Kp分离株 含有至少一种氨基糖苷修饰酶（AME），可使AMG的一个子集失活。 因此，选择针对AMEs和其他AMG抗性决定因素（菌株特异性的）定制的AMG， AMG）具有创新性，并为该应用提供了基础，该应用专注于开发 CR-Kp的基于AMG的分子精确组合。我们的核心假设是， 包括最佳剂量、菌株特异性AMG和BL/BLI的短期疗程的方案可以最大限度地杀灭 和CR-Kp的抗性抑制。我们有希望的初步数据高度支持我们的创新 机械的方法。我们开发了一个初步的模型来预测菌株特异性AMG的基础上， 分离物的AMG抗性基因。我们生成了菌株特异性AMG与 a BL/BLI（头孢他啶-阿维巴坦、美罗培南-戊硼巴坦和亚胺培南-阿贝巴坦）在两种药物中均抗CR-Kp 中空纤维感染模型（HSP70）和小鼠肺炎模型，其中组合高度 协同作用。我们还开发了一种新的测定方法来定量细胞内AMG浓度。利用我们 最新的测定，我们将阐明负责菌株之间的组合的协同作用的机制- CR-Kp中的特异性AMG和BL/BLI，以合理优化它们用于未来的临床试验。在目标1中， 每个AMG抗性决定子对AMG活性的影响将被定义，并且新的AMG- 阻力将被阐明。在目标2中，我们将设计新的短程AMG治疗方案， 灵验。我们将使用系统定义AMG耐受性和抗性出现的时间过程 药理学和蛋白质合成测定。在目标3中，我们将合理优化AMG和BL/BLI组合 通过鉴定协同作用机制和评估抗生素靶位点浓度。障碍者号将被用于 以确定组合的药效学并确定抗生素给药的最佳时间。在 目的4，我们将开发定量和系统药理学（QSP）模型，整合我们的机制， 数据，并根据AMG耐药决定因素合理优化AMG联合给药策略 在CR-Kp中。QSP优化的组合将在小鼠肺炎模型中进行前瞻性验证。这个项目 将开发新的，分子精确的AMG为基础的组合，以打击CR-Kp的紧迫威胁。

PROJECT SUMMARY/ABSTRACT: Carbapenem-resistant Klebsiella pneumoniae (CR-Kp) cause life- threatening infections that are associated with unacceptably high mortality rates. CR-Kp is particularly challenging to treat since it often possesses a myriad of molecular resistance determinants that enable it to grow in the presence of most antibiotics. New β-lactam/β-lactamase inhibitors (BL/BLI), such as ceftazidime- avibactam, are not a sustainable therapeutic solution for CR-Kp as monotherapy since there is potential for development of resistance and clinical failure rates remain remarkably high when used alone. Up to 95% of CR- Kp still remain susceptible to at least one of the aminoglycosides (AMG). However, nearly all CR-Kp isolates harbor at least one of the aminoglycoside-modifying enzymes (AME), which inactivate a subset of the AMGs. Thus, selecting an AMG that is tailored to the AMEs and other AMG-resistance determinants (strain-specific AMG) is innovative and provides the foundation for this application, which is focused on development of molecularly precise AMG-based combinations for CR-Kp. Our central hypothesis is that novel combination regimens including short-courses of an optimally dosed, strain-specific AMG and a BL/BLI can maximize killing and resistance suppression of CR-Kp. Our promising preliminary data are highly supportive of our innovative and mechanistic approach. We developed a preliminary model to predict the strain-specific AMG based on an isolate’s AMG-resistance genes. We generated the first data for the combination of the strain-specific AMG with a BL/BLI (ceftazidime-avibactam, meropenem-vaborbactam, and imipenem-relebactam) against CR-Kp in both the hollow fiber infection model (HFIM) and the mouse pneumonia model, where the combination was highly synergistic. We have also developed a novel assay to quantify intracellular AMG concentrations. Leveraging our latest assays, we will elucidate the mechanisms responsible for synergy of the combination between a strain- specific AMG and BL/BLI in CR-Kp to rationally optimize them for future clinical trials. In Aim 1, the precise influence of each AMG-resistance determinant on AMG activity will be defined and novel predictors of AMG- resistance will be elucidated. In Aim 2, we will design novel short-course AMG treatment regimens that are efficacious. We will define the time-course of AMG-tolerance and resistance emergence using systems pharmacology and protein synthesis assays. In Aim 3, we will rationally optimize AMG and BL/BLI combinations by identifying mechanisms of synergy and assessing antibiotic target site concentrations. The HFIM will be used to define the pharmacodynamics of combinations and determine optimal timing of antibiotic administration. In Aim 4, we will develop quantitative and systems pharmacology (QSP) models that integrate our mechanistic data and rationally optimize AMG-based combination dosing strategies based on AMG-resistance determinants in CR-Kp. QSP-optimized combinations will be prospectively validated in mouse pneumonia models. This project will develop novel, molecularly precise AMG-based combinations to combat the urgent threat of CR-Kp.