γδ and αβ T cells are important parts of the vertebrate immune system, expressing a γδ T cell receptor (TCR) or αβ TCR, respectively. We plan to engineer chimeric γδ TCRs to unravel how γδ T cells respond to temporally varying ligand encounter and provide proof-of-principles of new approaches to improve cancer immunotherapy and to combat virus infection. To this end, we plan to use technologies that we have successfully developed for the αβ TCR, and apply them for the first time to the γδ TCR. The planned work is divided into two parts, whose common denominator is the use of novel chimeric γδ TCRs.
Firstly, we will ask how γδ T cells respond to temporally controlled ligand encounter. To this end, we will engineer γδ TCRs to bind to Phytochome B (PhyB) as their ligand by appending a small domain to the TCR that binds to PhyB. The protein PhyB is derived from plants and can be switched between two conformations using red and far-red light, thus either binding or not to the engineered γδ TCR. We will alter the half-life of the γδ TCR-ligand interaction by using different intensities of red light and study T cell activation. These experiments aim to answer an important unsolved question: whether γδ T cells can distinguish between ligands of different affinities using a mechanism known as kinetic proofreading. In fact, we have used this optogenetic system with the αβ TCR, to show that the half-life is the critical parameter in αβ T cell activation (Yousefi et al., eLife, 2019). Under physiological conditions, T cells are activated while exploring the putative target cells, thus having temporal contacts to the antigen-presenting cells separated by pauses of different duration. Our system is unique to find out whether pauses of ligand binding are tolerated by the γδ T cells, and if specific temporal patterns will lead to differential γδ T cell activation.
Secondly, we will equip γδ TCRs with new specificities by fusion of a single chain Fv fragment against tumour antigens to the γδ TCR. Although γδ T cells have the inherent property to recognize tumours, this is often not sufficient to successfully kill the malignant cells. We will evaluate whether this could be used as a novel T cell based therapy against cancer. We have already used this technology with the αβ TCR and shown that the re-programmed αβ T cells are superior in killing tumour cells in vivo than CAR-T cells (Baeuerle et al., Nature Comm, 2019). We will also explore whether chimeric γδ TCRs can be used to redirect γδ T cells to kill virus infected cells.