Biodegradable Nanoparticles for Targeted Antibiotic Delivery

描述（由申请人提供）：人类感染细胞内细菌病原体沙眼衣原体和肺炎原体提出了一个巨大的卫生保健问题。这些病原体的感染与产生和/或加重几种慢性疾病有关，其中一些衣原体感染已被证明对抗生素治疗是难治的。缺乏治疗效果的原因是在某些情况下感染衣原体的代谢率降低，再加上这些药物正常递送到宿主细胞质中衣原体所在的包涵体中所能达到的适度细胞内浓度。这里概述的研究的主要治疗目标是开发一种方法，通过这种方法，抗生素或其他治疗剂可以在有效浓度下以靶向方式递送到细胞内衣原体包涵体，而对宿主细胞或感染组织没有毒性。我们的初步研究表明，衣原体感染引起宿主细胞叶酸受体（FAR）的表达增加，叶酸缀合纳米颗粒可能为衣原体感染细胞提供一种新的、高效的细胞内递送治疗剂的方法。所提出的研究的工作假设是，感染沙眼原体或肺炎原体的宿主细胞可以通过纳米颗粒介导的靶向递送有效浓度的抗生素来清除活跃或持续感染的生物体。已知抗生素可以对抗活动性衣原体感染。我们进一步假设，纳米颗粒促进的递送将需要减少治疗材料的数量，这可能会显著降低医疗成本。为实现这一目标，将讨论三个具体目标。在Aim 1中，我们将定义沙眼C感染相关人类宿主细胞培养和沙眼C感染小鼠模型中叶酸受体表达的时间过程，并定义叶酸受体在宿主细胞和细胞质包涵膜之间的定位。在目标二中，我们将确定在体外感染后的什么时间点递送这些纳米颗粒最有效，以及通过纳米颗粒向感染细胞递送抗生素是否能有效清除沙眼原体感染。此外，利用荧光显微镜，我们将研究纳米颗粒在感染和未感染细胞中的细胞内运输，以了解纳米颗粒靶向包涵体的机制。在第三项研究中，利用体内成像、分子遗传学和定量分析方法，我们确定了向感染衣原体的小鼠靶向递送纳米颗粒抗生素是否能清除体内滑膜感染。如果成功，这项研究的结果将提出一种新的基于纳米技术的治疗方案，用于有效治疗一个重要的卫生保健问题。

DESCRIPTION (provided by applicant): Human infections by the intracellular bacterial pathogens Chlamydia trachomatis and C pneumoniae present an enormous health care problem. Infections by these pathogens have been associated with engendering and/or exacerbating several chronic diseases, and some of these Chlamydial infections have proved to be refractory to antibiotic therapy. The lack of therapeutic efficacy results from the attenuated metabolic rate of infecting chlamydiae under some circumstances, in combination with the modest intracellular concentrations achievable by normal delivery of such drugs to the inclusions within which chlamydiae reside in the host cell cytoplasm. The major therapeutic goal of the studies outlined here is to develop a means by which antibiotics or other therapeutic agents can be delivered in a targeted manner to the intracellular Chlamydial inclusion at effective concentrations, without toxicity to the host cell or infected tissue. Our initial studies indicate that Chlamydial infection elicits increased expression of host cell folic acid receptors (FAR), and that folic acid-conjugated nanoparticles may provide a novel and highly effective means of intracellular delivery of therapeutic agents to Chlamydia- infected cells. The working hypothesis underlying the studies proposed is that host cells infected with either C trachomatis or C pneumoniae can be cleared of actively- or persistently-infecting organisms via nanoparticle-mediated targeted delivery of effective concentrations of antibiotics known to work against active Chlamydial infections. We further hypothesize that nanoparticle-facilitated delivery will require reduced amounts of therapeutic materials, which could engender significant health care cost reductions. To pursue that end, three Specific Aims will be addressed. In Aim One, we will define the time-course of expression of folate receptors following C trachomatis infection of a relevant panel of human host cells in culture and in a mouse model of C trachomatis infection, and define the localization of folic acid receptors between the host cell and cytoplasmic inclusion membranes. In Aim Two, we will determine at what time point post-in vitro infection delivery of those nanoparticles is most effective, and whether delivery of antibiotics to infected cells by nanoparticles is effective in clearing C trachomatis infection. In addition, using fluorescence microscopy, we will study the intracellular trafficking of nanoparticles in infected and uninfected cells to understand the mechanism of nanoparticle targeting to inclusions. In Aim Three, using in vivo imaging, molecular genetics, and quantitative analytical methods, we determine if FAR targeted delivery of nanoparticle antibiotics to Chlamydia-infected mice clears synovial infection in vivo. If successful, results of this research will suggest a novel nanotechnology-based therapeutic regimen for effective treatment of an important health care problem. PUBLIC HEALTH RELEVANCE: Chlamydiae are associated with several chronic diseases, including inflammatory arthritis, tubal occlusion leading to ectopic pregnancy, and cervical cancer (C trachomatis); C pneumoniae has been compellingly associated with atherosclerosis, inflammatory arthritis, and temporal arteritis, among several others. The proposed research is expected to positively affect human health, because, it is expected to lead to the development of an effective approach for the treatment of persistent Chlamydial infections.