Directed evolution of a soluble human DR3 receptor for the inhibition of TL1A induced cytokine secretion

TNF-like 1A (TL1A) is a cytokine belonging to the TNF superfamily that promotes inflammation in autoimmune diseases. Inhibiting the interaction of TL1A with the endogenous deathdomain receptor 3 (DR3) offers a therapeutic approach for treating TL1A-induced autoimmune diseases. Here, we generated improved DR3 variants showing increased TL1A binding affinity and stability using a directed evolution approach. Given the high cysteine content and post-translational modification of DR3, we employed yeast surface display and expression in mammalian cell lines for screening, expression and characterization of improved DR3 variants. A cell-based assay performed with the human TF-1 cell line and CD4⁺ T cells showed that two improved DR3 mutants efficiently inhibited TL1A-induced cell death and secretion of IFN-κ, respectively. These DR3 mutants can be used as drug candidates for the treatment of inflammatory bowel diseases and for other autoimmune diseases, including rheumatic arthritis and asthma.