In vivo PET imaging of novel engineered AAVs informs capsid design

Abstract of: Proposed COVID-19-related supplement to In vivo PET imaging of novel engineered AAVs informs capsid designThere is an urgent need to develop tools to assess viral pathogenesis and the efficacy of potential therapeuticsfor novel viruses such as SARS-CoV-2. While organoid and cell-based assays have been broadly used toassess the candidate receptor of such viruses, these assays cannot answer key questions as to: 1) whethermultiple receptors for the virus exist, 2) the in vivo receptor affinity of the virus and accumulation within theupper respiratory tract, 3) transport of the virus into the vascular system and ultimately to the heart andkidneys, and 4) the resulting transfection of these various sites. Our laboratory has previous developedmethods to label adeno-associated viruses and track their transport following systemic injection. We foundthat these engineered viruses carry cargo attached to the capsid across the blood brain barrier and the cargoaccumulates deep within the brain. Combined optical and PET studies have suggested that binding of thevirus to its receptor results in transcytosis of the intact capsid. We hypothesize that coronaviruses maypossess similar capabilities to be transported across the lung epithelium. We plan to address key issues byassessing receptor binding and transduction using PET and optical imaging. Our resulting specific aims are thefollowing: 1) development and validation of tagging strategies to image pseudotype viral particles at BSL2, 2)development and validation of reporter gene strategies to image transduction of engineered viruses, and 3)application and dissemination of these dual strategies to assess viral transport, transduction and susceptibilityto available therapies including a) antibodies, b) protease inhibitors, and c) fusion inhibitors. We propose to develop and image engineered viruses expressing the spike protein and a reporter gene and track theseviruses within a model of lung fibrosis and a mouse model with a humanized ACE2 receptor. We will leveragethe capabilities to label and track viral capsids and transduction developed within this R01 and key capabilitiesof Stanford University. Pseudotype viruses based on vesicular stomatitis virus (VSV) and lentivirus have beendeveloped with spike proteins corresponding to SARS-CoV or SARS-CoV2. Further, replicons with intact viralproteases have been engineered. We propose to collaborate with those developing and testing engineeredviruses and therapeutics at Stanford, including Jan Carette and Catherine Blish in addition to the key personnelon our parent project. At the conclusion of each phase of this project, we will disseminate strategies for theincorporation of a PET tag, a reporter gene, and dual PET imaging protocols. We hypothesize that these toolscan be disseminated and rapidly modified to assess both SARS-CoV-2 and future viruses. We will make ourtechnology available through commercial and scientific partners.