Development of NK cells expressing a chimeric antigen receptor (CAR-NK) against feline coronavirus spike protein

Project leader: Cynthia Leifer
Sponsor: Cornell Feline Health Center Research Grants Program
Original Name: Development of Feline Chimeric Antigen Receptor-Expressing Natural Killer Cells (CAR-NK) Against a Feline Coronavirus Spike Protein
Project amount: $ 55,200
Project period: July 2021 to June 2022

DESCRIPTION (provided by the applicant):

Feline coronavirus (FCoV) causes a subclinical infection that can develop into FIP's lethal infectious peritonitis. It is caused by the feline infectious peritonitis virus (FIPV), which occurs when a mutation occurs in the spike of the FCoV virus. Despite more than three decades of virus research, there is still no approved treatment for FIP. This is partly due to the fact that FIPV type I causes most clinical infections, but is not very successful in cultures and there are few reagents available for its study, while type II grows well, has more reagents, but generally does not cause clinical infections. disease. The development of new immunotherapies requires significant preclinical study and validation, and due to limitations in the FIPV type I study, no immunotherapies have been developed for clinical FIP. We want to try a new immunotherapy using in vitro FIPV type II research, which could lead to the treatment of FIPV cats caused by FIPV type I. Immunotherapy, which has recently gained attention in the treatment of human cancers, are immune cells treated with a chimeric antigen receptor ( CAR). This approach has not yet been developed for acute viral infections such as FIPV, nor has it been used in cats. CAR consists of two main components: a single chain antibody fragment (ScFv) and a signal domain from the immune costimulatory receptor. We propose to use a FIPV-specific ScFv spike protein expressed in feline immune cells that could be administered to a feline to screen for and destroy FIPV-infected cells. Current human CAR therapies require the use of autologous T cells because allogeneic T cells kill host cells and lead to graft-versus-host disease.
NK cells (natural killers) are of increasing interest because NK cells do not induce graft-versus-host disease and can therefore be used in "conventional" therapies. However, little is known about feline NK cells. We hypothesize that genetically modified feline NK cells expressing CAR targeting the feline infectious peritonitis virus type II spike protein will recognize and kill FIPV-infected target cells in vitro.

To achieve our goals:

  1. determine the cytotoxic activity of a subset of NK cells in comparison with NKT and T cells for the validation of feline NK cells as potential CAR carriers,
  2. for cloning and sequencing, we will use currently available hybridoma spike anti-FIPV type II antibodies to construct a CAR directed against spike type II protein, and
  3. in vitro we validate the NK cell assay and data demonstrating the effect of CAR-NK cells killing FIPV type II infected cells.

The completion of this study will provide data important for the development of CAR-NK therapy for FIPV type I, a completely new treatment for this devastating disease.

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