Non-Invasive Detection and Staging of Decubitus and Diabetic Ulcers

项目摘要 我们通过对现有NIA奖（R21 AG 065776）进行竞争性修订来申请资金，以支持 关于COVID-19的研究这些补充资金将使我们能够建立和验证一个 多模式造影剂，其报告存在与血管内皮细胞复杂连接的Mpro-a蛋白酶。 SARS-CoV-2的生命周期现有的检测和监测SARS类病毒的工具是基于 在PCR。虽然是定量的，但这些体外工具在绘制时空分布图的能力方面是有限的。 病毒在活体中的分布缺少的元素是一种造影剂， Mpro成像来绘制和测量SARS-CoV-2感染的这种特定副产物。这项工作 将完成这一点，并报告存在的Mpro与传统的荧光，以及 新型光声成像荧光将允许快速和常规的体外测定 而光声模态将用于通过啮齿动物模型的体内深部组织成像。 Aim 1将基于被Mpro选择性切割的肽序列构建探针， 基于电荷的细胞渗透。探测器将装饰有超声波细胞， 在肽被切割之前，肽处于失活状态。一旦被切割，这些分子产生 荧光和光声信号。目标2将验证这种造影剂的感染性较低 SARS-CovV-2（辛德毕斯病毒）的类似物。辛德毕斯病毒可以在BSL-2设施中轻松处理 并允许立即开始工作。我们将用感染的细胞验证探针， 被感染的动物由于Jokerst博士先前在光学成像方面的工作，这些目标是可行的 以及Siqueira-Neto博士在传染病方面的工作， 用于治疗和病原体的基于图像的筛选工具。这部作品的创新之处是第一位的 造影剂来成像COVID-19感染。重要的是，一旦完成， 社区将有一个强大的化学工具来量化和定位SARS-Cov-2感染， 回答关于这种疾病的关键问题：感染和生物分布的时间过程是什么？ 感染途径如何改变生物分布？是否存在潜在的病源？如何 蛋白酶水平会随着治疗而改变吗？不幸的是，这些问题都不能 因为没有专门针对病毒的体内成像方法，更不用说SARS了- 二型冠状病毒

Project Summary We request funds via a competitive revision to our existing NIA award (R21 AG065776) to support research on COVID-19. These supplemental funds will enable us to build and validate a multimodal contrast agent that reports the presence of Mpro—a protease intricately linked to the life cycle of the SARS-CoV-2. Existing tools to detect and monitor SARS-type viruses are based on PCR. While quantitative, these in vitro tools are limited in their ability to map the spatiotemporal distribution of the virus in living subjects. The missing element is a contrast agent for specific imaging of Mpro to map and measure this specific byproduct of SARS-CoV-2 infection. This work will accomplish this and report the presence of Mpro with conventional fluorescence as well as novel photoacoustic imaging. The fluorescence will allow for rapid and routine in vitro assays while the photoacoustic modality will be used for deep tissue in vivo imaging via rodent models. Aim 1 will build the probe based on a peptide sequence that is selectively cleaved by Mpro with cell penetration based on charge. The probe will be decorated with sonophores that are in a deactivated state until the peptide is cleaved. Once cleaved, these molecules produce both fluorescence and photoacoustic signal. Aim 2 will validate this contrast agent with a less infectious analogue of SARS-CovV-2 (Sindbis virus). Sindbis virus can easily be handled in BSL-2 facilities and will allow work to commence immediately. We will validate the probe with infected cells and infected animals. These aims are feasible because of Dr. Jokerst’s prior work in optical imaging and contrast agent construction and Dr. Siqueira-Neto’s work in infectious disease including image-based screening tools for therapies and pathogens. The innovation of this work is the first contrast agent to image COVID-19 infection. The significance is that, once completed, the community will have a powerful chemical tool to quantify and locate SARS-Cov-2 infection to answer key questions about this disease: What is the time course of infection and biodistribution?; How does biodistribution change by route of infection? Are there latent disease reservoirs?; How do protease levels change in response to therapy? Unfortunately, none of these questions can be answered because there are no in vivo imaging methods specific for viruses much less SARS- CoV-2.