Non-Invasive Detection and Staging of Decubitus and Diabetic Ulcers

项目摘要 我们通过对现有NIA奖项(R21 AG065776)进行竞争性修订来申请资金，以支持 关于新冠肺炎的研究。这些补充资金将使我们能够建立和验证 多模式造影剂，报告存在MPRO-一种复杂地与 SARS-CoV-2的生命周期。现有的检测和监测SARS类型病毒的工具基于 在聚合酶链式反应上。虽然这些体外工具是定量的，但它们绘制时空图的能力有限 病毒在活体中的分布情况。缺失的元素是特定的对比剂 对MPRO进行成像，以绘制和测量SARS-CoV-2感染的这种特定副产品。这部作品 将实现这一点并用常规荧光报告MPRO的存在以及 新颖的光声成像技术。这种荧光可以进行快速和常规的体外分析。 而光声模式将用于通过啮齿动物模型进行体内深层组织成像。 目标1将根据MPRO选择性切割的多肽序列构建探针 基于电荷的电池穿透性。探测器将用声学浮标进行装饰，这些声学浮标位于 去激活状态，直到多肽被裂解。一旦被裂解，这些分子就会同时产生 荧光和光声信号。Aim 2将用一种传染性较低的造影剂来验证这种造影剂 SARS-CovV-2(辛德比斯病毒)的类似物。辛德比斯病毒可以在BSL-2设施中轻松处理 并将允许工作立即开始。我们将用受感染的细胞来验证探针 被感染的动物。这些目标是可行的，因为Jokerst博士之前在光学成像方面的工作 造影剂构建和Siqueira-Neto博士在传染病方面的工作包括 基于图像的治疗和病原体筛查工具。这项工作的创新之处是第一 对比剂成像新冠肺炎感染。重要的是，一旦完成， 社区将拥有一种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进程是什么？ 生物分布是如何通过感染途径改变的？是否存在潜在的病源？如何？ 治疗后蛋白水解酶水平会发生变化吗？不幸的是，这些问题都不能回答 因为没有专门针对病毒的体内成像方法，更不用说SARS了- CoV-2。

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.