Next generation tools for imaging bacterial infection and its relationship to the immune system

现代生物医学研究的最大挑战之一是观察生物 动物和人身上的现象。这方面的一个重要例子是我们有限的监控能力 细菌感染的过程。基于图像的细菌感染读数将允许 感染与其他病因的区别，量身定做的抗生素治疗时间，以及 抗生素耐药性的鉴定建议使用适当类别的抗生素。 除了细菌监测的改进对临床和种群的影响 对于感染，基础研究人员没有简单的工具来测量对 细菌感染的地方。同样，通过在体内促进成像，成像适合于解决这个问题 在空间和时间上进行监控。我的工作是开发以成像为基础的化学和 阐明细菌发病机制、对抗生素的反应、 抗生素耐药性的发展，以及细菌与免疫系统的相互作用。我 提出互补的方法来实现这些目标，使用交付的免疫细胞 进入追踪细菌生物标志物的血流--包括细菌表面标志物和 细菌酶--并使用正电子发射断层扫描的直接细菌成像 (宠物)。这些新方法利用了我开发的概念和技术，包括 “细胞-细胞接近报告者”、蛋白质不稳定结构域和基于 抗生素甲氧苄啶(TMP)。使用免疫细胞的优势包括能够产生 多路传感器和报告输出，转录和酶信号放大， 以及免疫细胞贩运的地区性评估。直接细菌成像的一个优点是 成像细菌负荷的能力不依赖于免疫细胞对 感染。这项提议的主要目标是：1)开发新的受体 体内细菌感染的严重程度和种类。2)开发新的笼养小动物班级 利用合成生物学监测免疫细胞-细菌细胞相互作用的分子 原理，以及3)评估我最近开发的一类用于成像的新型PET放射性示踪剂 囊性纤维化(CF)大鼠模型的感染和细菌对放射性示踪剂的摄取 抗菌药物治疗前后的CF患者。 这项工作奠定了监测体内病原菌的基础，涵盖了 从长椅到床边。我希望为科学界提供一整套试剂，包括 编码各种细菌受体的质粒，酶激活的小分子，以及 有用的PET探针，所有这些都是针对活细菌的特定成像。

One of the great challenges of modern biomedical research is observing biologic phenomena in animals and people. An important example of this is our limited ability to monitor the course of bacterial infection. An image-based readout of a bacterial infection would allow differentiation of infection from other etiologies, a tailored duration of antibiotic treatment, and identification of antibiotic resistance suggesting an appropriate class of antibiotic. In addition to the clinical and population implications of improved monitoring of bacterial infections, basic researchers do not have simple tools to measure the immune response to the site of a bacterial infection. Again, imaging is suited to address this problem by facilitating in vivo monitoring over space and time. My work seeks to develop imaging-based chemical and synthetic biology technologies that illuminate bacterial pathogenesis, response to antibiotics, the development of antibiotic resistance, and bacterial interactions with the immune system. I propose complementary approaches to accomplish these goals using immune cells delivered into the blood stream that track bacterial biomarkers –including bacterial surface markers and bacterial enzymes– and using direct bacterial imaging with positron emission tomography (PET). These new approaches leverage concepts and techniques I have developed including “cell-cell proximity reporters”, protein destabilizing domains, and PET imaging based on the antibiotic trimethoprim (TMP). Advantages of using immune cells include the ability to generate multiplexed sensors and reporter outputs, transcriptional and enzymatic signal amplification, and regional assessment of immune cell trafficking. An advantage of direct bacterial imaging is the ability to image the bacterial load that does not depend on immune cell access to the infection. The primary objectives of this proposal are 1) to develop new receptors that can report the severity and species of bacterial infection in vivo. 2) to develop new classes of caged small molecules for monitoring immune cell-bacterial cell interactions using synthetic biology principles, and 3) to evaluate a new class of PET radiotracers I recently developed for imaging infection in a rat model of cystic fibrosis (CF) and measure bacterial radiotracer uptake in patients with CF before and after antibiotics. This work builds a foundation to monitor pathologic bacteria in vivo and spans from bench to bedside. I expect to provide sets of reagents to the scientific community including plasmids encoding receptors for a variety of bacteria, enzyme activated small molecules, and useful PET probes, all geared toward specific imaging of live bacteria.