Infection-homing nanosystems as antibacterial therapeutics-delivery platforms

项目摘要 金黄色葡萄球菌和铜绿假单胞菌是医院获得性感染的主要原因 并对发病率和死亡率有显著影响[3，4]。感染的标准治疗需要重复 高剂量的抗生素给药，但治疗往往是无效的，由于交付不良， 感染部位和阻止抗生素进入细胞内药物靶点的耐药性机制（例如， 革兰氏阴性铜绿假单胞菌中的药物不可渗透的细胞壁）[5，6]。皮肤感染已经侵入了 游离抗生素制剂也难以到达肌肉和纤维， 治疗[7]。我们在这个建议中解决的障碍是：（1）抗生素流失到未感染的组织;（2） 小分子抗生素通过肾脏和胃肠道清除的快速清除;（3） 药物能穿过细菌细胞壁我们假设将抗生素装载到更长循环的纳米载体中 其将归巢到感染部位并随后促进药物摄取到感兴趣的细胞/细菌中， 克服上述挑战。在这里，我们建议通过三个方面来开发这样的纳米平台。 主要目标。在目标1中，我们将使用体内噬菌体展示来鉴定将归巢于以下细菌的肽： 感兴趣和/或受感染组织。我们将特别关注S。金黄色葡萄球菌和铜绿假单胞菌感染的模型 小鼠的深层皮肤（肌肉和纤维中的侵入）感染和肺炎。如果直接细菌- 靶向被证明是困难的，我们还将研究选择性结合感染组织和宿主的肽 细菌菌落周围的细胞，以及巨噬细胞靶向肽。因为这些肽将被 结合到纳米颗粒表面，然后我们将研究单一肽的结合特性， 和多价形式。在目标2中，我们将设计两个纳米平台：（1）基于肽的药物， 选择性地穿透细菌膜（即肽渗透剂） （2）多孔硅纳米颗粒（pSiNP），以装载具有差吸收的药物， 由于不利的物理化学性质（疏水性、高离子性等）而递送至感染部位。 这些纳米平台将使用我们以前发现的肽靶向感染部位， 目的1中鉴定的其他肽。将装载体内抗菌活性较差的模型药物 以及基于药物负载、释放动力学和用于体内的细胞摄取选择的最佳平台。 药代动力学除了单独的基于pSi和肽的纳米平台，我们将开发一种组合的纳米平台。 系统，其中细菌穿透药物缀合物被加载到靶向pSi纳米颗粒中，目标是 增强功效。最后，目标3将重点关注领先的纳米平台候选物的治疗性能 in vivo.该目的旨在证明生物安全性和治疗有效性（即细菌负荷 清除，组织恢复，提高存活率）的pSiNP和细菌穿透纳米系统。这 该项目将产生主动靶向受感染组织的工具，以及一组强大的纳米平台， 解决了目前体内抗菌药物活性的许多障碍。

PROJECT SUMMARY Staphylococcus aureus and Pseudomonas aeruginosa are the leading causes of hospital-acquired infections and contribute significantly to morbidity and mortality [3, 4]. Standard treatment of infection entails repetitive high-dose administrations of antibiotics, but the treatment is often rendered ineffective due to poor delivery to sites of infection and drug resistance mechanisms preventing antibiotic access to intracellular drug targets (e.g. the drug-impermeable cell wall in gram-negative P. aeruginosa) [5, 6]. Skin infections that have invaded down to the muscles and fibers are also difficult-to-reach by free-antibiotic formulations and require surgical treatment [7]. The obstacles we tackle in this proposal are: (1) loss of antibiotics to non-infected tissues; (2) rapid clearance of small molecule antibiotics by renal and gastrointestinal clearance; (3) poor penetration of drugs past the bacterial cell wall. We hypothesize that loading antibiotics into longer-circulating nanovehicles that will home to sites of infection and subsequently facilitate drug uptake into cells/bacteria of interest can overcome the abovementioned challenges. Here, we propose to develop such nanoplatforms through three major aims. In Aim 1, we will use in vivo phage display to identify peptides that will home to the bacteria of interest and/or infected tissue. We will focus specifically on S. aureus and P. aeruginosa infections in models of deep skin (invasion in muscles and fibers) infection and pneumonia in mice. In the event that direct bacteria- targeting proves to be difficult, we will also look at peptides that bind selectively to infected tissues and host cells surrounding the bacterial colonies, as well as macrophage-targeting peptides. As these peptides are to be conjugated to nanoparticle surfaces, we will then investigate the binding properties of the peptides in singular and multivalent forms. In Aim 2, we will engineer two nanoplatforms: (1) peptide-based agents that can selectively penetrate the bacterial membrane (i.e. peptide permeation agents) to which small molecule drugs will be tethered for increased uptake and (2) porous silicon nanoparticles (pSiNP) to load drugs that have poor delivery to sites of infection due to unfavorable physicochemical properties (hydrophobic, highly ionic, etc). These nanoplatforms will be targeted to sites of infection using peptides we have previously discovered or additional peptides to be identified in Aim 1. Model drugs with poor in vivo antibacterial activity will be loaded and optimal platforms selected based on drug loading, release kinetics, and cellular uptake for in vivo pharmacokinetics. In addition to individual pSi- and peptide-based nanoplatforms, we will develop a combined system in which bacteria-penetrating drug conjugates are loaded into targeted pSi nanoparticles with the goal of enhanced efficacy. Finally, Aim 3 will focus on the therapeutic performance of lead nanoplatform candidates in vivo. The goal of this aim is to demonstrate the biosafety and therapeutic efficacy (i.e. bacterial burden clearance, tissue recovery, improved survival) of the pSiNP and bacteria-penetrating nanosystems. This project will yield tools to actively target infected tissues as well as a strong set of nanoplatforms that can address many of the current barriers to in vivo antibacterial drug activity.