Leveraging the sensitivity of antigen-specific T cells to interrogate disease mechanisms and identify therapeutic targets for Long COVID

Project Summary/Abstract: An estimated 400 million people worldwide have experienced persisting and sometimes debilitating symptoms for months and years after SARS-CoV-2 infection, known as Long COVID (LC). No proven therapies exist to treat LC, and the underlying mechanisms driving disease remain poorly understood. Reactivation of herpesviruses such as Epstein-Barr virus (EBV) and varicella zoster virus (VZV) has been linked to LC, but it is unclear how SARS-CoV-2 infection alters immune responses to these common viruses. Recent studies have also provided emerging evidence for persistence of SARS-CoV-2 in some individuals with LC, but key questions remain unanswered, including how viral reservoirs persist, why the immune system fails to clear virus, whether persisting virus drives ongoing immune stimulation, and how therapies to treat LC will affect immune responses to SARS-CoV-2. Efforts to identify effective treatments for LC depend on answering these questions. Our preliminary data demonstrate elevated activation of SARS-CoV-2-, EBV-, or VZV-specific T cells in 40% of LC patients, providing one of the most sensitive measures of immunopathology in LC to date. Thus, there is an urgent need to investigate virus-specific T cell activation in LC to define underlying mechanisms of disease and identify promising therapeutic targets. The proposed research will respond to this need by testing three working hypotheses: first, that SARS-CoV-2-specific CD8 T cells survey tissue reservoirs of SARS-CoV-2 and sense viral antigens, but fail to clear the virus due to impaired functionality in LC; second, that therapies suppressing SARS-CoV-2 replication will reduce T cell stimulation in LC but may not achieve a durable cure; and third, that immune stimulation by common herpesviruses is associated with disease pathology in LC. These hypotheses will be tested by leveraging custom HLA-I/peptide tetramers to identify and sort rare virus-specific CD8 T cells from hundreds of longitudinal blood and tissue samples from people with LC and people who fully recovered after SARS-CoV-2 infection. Cells will be analyzed using spectral flow cytometry and single-cell sequencing approaches to elucidate the mechanisms driving LC pathology, identify promising therapeutic targets for LC, and investigate how immune stimulation by herpesviruses shapes human disease. This research will be integrated with a comprehensive training plan to develop skills in tissue immunology, single-cell sequencing, and bioinformatics, which will be reinforced with targeted coursework and professional development activities in scientific writing, responsible conduct of research, laboratory management, and mentorship of junior personnel. The research and training plan will take full advantage of the outstanding environment at the University of Pennsylvania and the wealth of expertise in Dr. John Wherry's lab. Completing the proposed research and training will substantially advance the human immunology field and form the foundation of my research program as I open my independent laboratory, creating a pathway to my goal of being an impactful independent researcher in human infectious disease immunology.