Non-Invasive Detection and Staging of Decubitus and Diabetic Ulcers

Project Summary: We request funds via a competitive revision to our existing NIA award (R21 AG065776) to supportresearch on COVID-19. These supplemental funds will enable us to build and validate amultimodal contrast agent that reports the presence of Mpro-a protease intricately linked to thelife cycle of the SARS-CoV-2. Existing tools to detect and monitor SARS-type viruses are basedon PCR. While quantitative, these in vitro tools are limited in their ability to map the spatiotemporaldistribution of the virus in living subjects. The missing element is a contrast agent for specificimaging of Mpro to map and measure this specific byproduct of SARS-CoV-2 infection. This workwill accomplish this and report the presence of Mpro with conventional fluorescence as well asnovel photoacoustic imaging. The fluorescence will allow for rapid and routine in vitro assayswhile 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 withcell penetration based on charge. The probe will be decorated with sonophores that are in adeactivated state until the peptide is cleaved. Once cleaved, these molecules produce bothfluorescence and photoacoustic signal. Aim 2 will validate this contrast agent with a less infectiousanalogue of SARS-CovV-2 (Sindbis virus). Sindbis virus can easily be handled in BSL-2 facilitiesand will allow work to commence immediately. We will validate the probe with infected cells andinfected animals. These aims are feasible because of Dr. Jokerst's prior work in optical imagingand contrast agent construction and Dr. Siqueira-Neto's work in infectious disease includingimage-based screening tools for therapies and pathogens. The innovation of this work is the firstcontrast agent to image COVID-19 infection. The significance is that, once completed, thecommunity will have a powerful chemical tool to quantify and locate SARS-Cov-2 infection toanswer 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?; Howdo protease levels change in response to therapy? Unfortunately, none of these questions canbe answered because there are no in vivo imaging methods specific for viruses much less SARS-CoV-2.