Development of Feline Chimeric Antigen Receptor-Expressing Natural Killer Cells (CAR-NK) Against a Feline Coronavirus Spike Protein
Principal Investigator: Cynthia Leifer
DESCRIPTION (provided by applicant):
Feline coronavirus (FCoV) causes a subclinical infection called feline enteric coronavirus [FECV]) that can transition to fatal feline infectious peritonitis (feline infectious peritonitis virus [FIPV]) when mutations occur in the viral surface spike protein. Despite over three decades of research into the virus, there are still no approved treatments for FIP. This is in part due to the fact that Type I FIPV causes most clinical infections but does not grow well in culture and has few reagents available for study while Type II grows well, has more reagents available but does not typically cause clinical disease. Development of novel immunotherapies require significant preclinical study and validation, and due to limitations in studying Type I FIPV, no immunotherapies have been developed for clinical FIP. Here we propose to provide proof-of-principle information on a novel immunotherapy using Type II in vitro, which could lead to therapies for FIP cats caused by Type I FIPV. An immunotherapy that has gained attention in treatment of human cancers is chimeric antigen receptor (CAR)-engineered immune cells. This approach has not yet been developed for acute viral infections like FIPV, nor used in cats to date. CARs are comprised of two main components: a single chain antibody fragment (ScFv) and a signaling domain from an immune costimulatory receptor. We propose to use an ScFv specific for FIPV spike expressed in feline immune cells that could be infused into a cat to seek out and destroy the FIPV-infected cells. Current human CAR therapies require use of autologous T cells since allogeneic T cells kill host cells and result in severe graft -versus-host disease. Natural Killer (NK) cells are gaining attention since NK cells do not induce graft-versushost disease and can thus be used to generate “off-the-shelf” therapies. However, little is known about feline NK cells. We hypothesize that genetically engineered CAR-expressing feline NK cells targeting the spike protein of feline infectious peritonitis virus Type II will recognize and kill FIPV-infected target cells in vitro. To achieve our goals, we will 1) determine NK cell subset cytotoxic activity compared to NKT and T cells to validate feline NK cells as potential CAR carriers, 2) use currently available anti-FIPV Type II spike antibody hybridomas to clone, sequence, and assemble a feline CAR targeting Type II FIPV spike, and 3) validate an in vitro feline NK killing assay and provide proof-of-principle data demonstrating feline CAR-NK kill FIPV Type II infected target cells. Completion of this study will provide proof-of-principle data using Type II FIPV support to development of FIPV CAR-NK therapy for Type I FIPV and thus a novel treatment for this devastating disease.