Discovery and preclinical evaluation of novel peptide inhibitors targeting the TIGIT immune checkpoint

Immune checkpoint (IC) receptors negatively regulate immune responses and play crucial roles in maintaining self-tolerance and preventing autoimmunity. However, tumor cells can exploit these pathways to evade immune recognition and destruction. TIGIT has emerged as a key immune checkpoint receptor, which exerts immunosuppressive effects through direct and indirect mechanisms. While preclinical studies with TIGIT-blocking monoclonal antibodies demonstrated encouraging antitumor activity, clinical trials of anti-TIGIT monotherapy showed disappointing outcomes, shifting attention toward alternative strategies. In this study, we identified potential peptide-based inhibitors against TIGIT. Through computational affinity maturation, starting from the anti-TIGIT antibody MG1131, we identified a library of TIGIT-inhibitory peptides. This process integrated sequence sampling, molecular dynamics, and free energy calculations, guided by Monte Carlo selection. Compared with conventional monoclonal antibodies, these peptides are expected to provide superior tissue penetration, reduced immunogenicity, and an improved safety profile. In this study, two mutant peptides (named T-M2 and T-M3) exhibited stronger inhibition of TIGIT-CD155 interactions than the wild-type peptide in hTIGIT-CHO-K1 cell-based blocking assay. Furthermore, these peptides induced higher levels of IFN-gamma secretion from both human CD4+ and CD8+ T cells. This enhanced immunostimulatory activity was confirmed in a co-culture system of hTIGIT-Jurkat and hCD155-CHO-K1 cells, where the mutant peptides induced greater IL-2 secretion. The in vivo therapeutic efficacy of these peptides, either as monotherapy or in combination with immune checkpoint inhibitors, is currently being evaluated in MC38 tumor-bearing and humanized HCT116 mouse models. Tumor growth dynamics, as well as the phenotype, cytokine profile and activation status of T cells in the tumor microgovernment, tumor-draining lymph nodes, and spleens will be assessed in the treated groups and compared with control groups. Collectively, this study introduces novel TIGIT-blocking peptides as promising candidates for cancer immunotherapy, offering a potential alternative or complementary approach to antibody-based immune checkpoint blockade.