Novel antimicrobials in fighting carbapenem-resistant Klebsiella pneumoniae

项目概要/摘要： 克雷伯氏菌是引起社区获得性肺炎的常见革兰氏阴性病原菌，克雷伯氏菌是引起社区获得性肺炎的主要病原菌。 在美国，肺炎被认为是医院获得性肺炎的最常见原因。 States. K.肺炎是一种很难治疗的感染，因为这种细菌的厚包膜通常是最好的治疗方法。 使用最后一线抗生素治疗，如碳青霉烯类。然而，耐碳青霉烯类K. 肺炎克雷伯氏菌（CRKP）是碳青霉烯类耐药肠杆菌科（CRE）的一种， 耐药性医院感染与高发病率和死亡率相关。新疗法在 迫切需要控制CRKP诱导的感染。利用重组蛋白和转基因 在小鼠基因敲除模型中，我们已经证明了短腭、肺、鼻上皮克隆（PLP 1） （SPLUNC 1）参与肺宿主对K.肺炎引起的感染。一个新的SPLUNC 1- 从SPLUNC 1蛋白的抗微生物基序衍生的肽，α4-Short，表现出更有效的 与全长重组SPLUNC 1蛋白相比的抗菌特性和在鼠模型中的体内功效 呼吸道感染基于α4-Short的额外修改，我们最近开发了一系列 合理工程化的抗菌肽（AMP），其通过透化作用快速杀死其微生物靶标， 细菌膜上的细菌，而不管细菌的特定代谢状态。我们的一个主要AMP A4-153 已证明对多种难治性多药耐药（MDR） 病原体，包括CRKP。令人兴奋的是，A4-153对许多已产生耐药性的CRKP具有活性。 其他膜活性化合物，如天然AMP LL 37和粘菌素，最后的抗生素。在 此外，我们还发现，与A4-153相比， 标准抗生素和天然抗菌肽。重要的是，我们发现类似于天然AMP LL 37，A4-153 没有表现出可检测到的溶血，并且可以安全地施用于小鼠肺部，具有非常高的溶血率 浓度的我们建议在这项SBIR应用中探讨使用新开发的A4- 153来预防CRKP诱导的肺炎，通过杀死CRKP并根除非生物和非生物环境中的CRKP生物膜， 使用体外和体内模型的生物系统。顺利完成第一阶段的拟议目标 申请将为我们在随后的第二阶段申请中提出的IND使能研究做好准备， 包括多剂量MTD、啮齿类动物和大型动物中的GLP毒性以及GMP生产的启动。

Project Summary/Abstract: Klebsiella is a common Gram-negative pathogen causing community-acquired bacterial pneumonia, and K. pneumoniae pneumonia is considered the most common cause of hospital-acquired pneumonia in the United States. K. pneumoniae is a difficult infection to treat because of the organism's thick capsule that is usually best treated with the last line antibiotic therapy such as carbapenems. However, carbapenem-resistant K. pneumoniae (CRKP), one of the carbapenem-resistant Enterobacteriaceae (CRE), is an emerging cause of antibiotic-resistant nosocomial infections associated with high rates of morbidity and mortality. New therapies in controlling CRKP-induced infections are urgently needed. Using recombinant protein and genetically modified mouse knockout models, we have demonstrated that the short palate, lung, nasal epithelium clone (PLUNC) 1 (SPLUNC1) contributes to pulmonary host defense against K. pneumoniae induced infection. A novel SPLUNC1- derived peptide from the antimicrobial motif of the SPLUNC1 protein, α4-Short, demonstrated more potent antibacterial properties than the full-length recombinant SPLUNC1 protein and in vivo efficacy in a murine model of respiratory infection. Based on additional modifications of α4-Short, we have recently developed a series of rationally engineered antimicrobial peptides (AMPs) that rapidly kill their microbial targets by permeabilizing bacterial membrane regardless of the specific metabolic state of the bacteria. One of our lead AMPs, A4-153, has demonstrated potent bactericidal activity against diverse difficult-to-treat multidrug resistant (MDR) pathogens, including CRKP. Exciting, A4-153 is active against many CRKP that have developed resistance to other membrane-active compounds, such as the natural AMP LL37 and colistin, an antibiotic of last resort. In addition, we have found a substantially lower tendency for bacteria to develop resistance to A4-153 compared to standard antibiotic agents and natural AMPs. Importantly, we found that similar to natural AMP LL37, A4-153 displayed no detectable hemolysis and could be safely administered to mouse lungs with very high concentrations. We propose in this SBIR application to explore the feasibility of using the newly developed A4- 153 to prevent CRKP-induced pneumonia by killing CRKP and eradicating the CRKP biofilm in the abiotic and biotic system using in vitro and in vivo models. The successful completion of the proposed aims in this Phase I application will prepare us for IND-enabling studies to be presented in a subsequent Phase II application, including multidose MTD, GLP toxicity in rodents and large animals, and initiation of GMP manufacturing.