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

现代生物医学研究面临的最大挑战之一是观察生物 动物和人类的现象。这方面的一个重要例子是我们的监测能力有限 细菌感染的过程。基于图像的细菌感染读数将允许 区分感染与其他病因，定制抗生素治疗的持续时间，以及 鉴定抗生素耐药性，提示适当的抗生素类别。 除了改进细菌监测的临床和人口影响外， 感染，基础研究人员没有简单的工具来测量免疫反应， 细菌感染的部位。再次，成像适合于通过促进体内 在空间和时间上进行监控。我的工作旨在开发基于成像的化学和 合成生物学技术阐明了细菌的发病机制，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 提出补充方法来实现这些目标， 进入血液，跟踪细菌生物标志物-包括细菌表面标志物， 细菌酶-并使用正电子发射断层扫描直接细菌成像 （PET）。这些新方法利用了我开发的概念和技术，包括 “细胞-细胞邻近报告物”、蛋白质去稳定化结构域和基于该蛋白质的PET成像。 抗生素甲氧苄啶（TMP）。使用免疫细胞的优点包括产生免疫细胞的能力。 多重传感器和报告输出，转录和酶信号放大， 和免疫细胞运输的区域评估。直接细菌成像的优点是 成像细菌负荷的能力，不依赖于免疫细胞进入 感染该提案的主要目标是：1）开发新的受体， 体内细菌感染的严重程度和种类。2)开发新的笼中小 使用合成生物学监测免疫细胞-细菌细胞相互作用的分子 原则，和3）评估一类新的PET放射性示踪剂，我最近开发的成像 在囊性纤维化（CF）大鼠模型中感染并测量 抗生素治疗前后的CF患者。 这项工作建立了一个基础，以监测病理细菌在体内，并跨越从 长凳到床边我希望向科学界提供成套试剂，包括 编码各种细菌受体的质粒，酶激活的小分子，和 有用的PET探针，都是针对活细菌的特异性成像。

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.