Structure‐Guided Engineering of TIGIT to Modulate Its Interaction With Nectin‐4, a Tumour‐Specific Antigen for the Development of Therapeutic Strategy

Nectin and nectin-like proteins are cell adhesion molecules belonging to the immunoglobulin superfamily that also play a crucial role in the process of immune modulation by interacting with immune receptors like TIGIT, DNAM-1, CD96 and PVRIG. Nectin-4 is a tumour-specific antigen that interacts exclusively with the inhibitory immune receptor TIGIT, resulting in inhibition of NK-cell cytotoxicity and facilitating the process of immune evasion by the cancer cells in the tumour microenvironment. Therefore, deciphering the molecular and structural mechanisms underlying this novel interaction can provide insights for the development of targeted immunotherapeutic interventions. In this study, using structure-guided mutagenesis studies, a novel TIGIT mutant is engineered exhibiting enhanced interaction affinity towards nectin-4 and reduced binding to other TIGIT ligands, such as PVR and nectin-2, which, in contrast to nectin-4, are predominantly expressed on healthy cells. Surface plasmon resonance-based biophysical studies were done to characterize and compare the interaction kinetics of the wild-type (WT) and the engineered mutant TIGIT ectodomains with their ligands. We have shown how a single point mutation of an amino acid residue located in the centre of the F strand of TIGIT dictates its interaction affinity with its ligand. These findings can provide a framework for the development of small-sized non-antibody therapeutics specifically targeted towards nectin-4 overexpressing cancer cells with minimal off-target effects on healthy cells.