Five dominant amino acid substitution signatures shape tumour immunity

Although numerous mutational processes operate in cancer, their functional impacts are unclear. We hypothesized that certain mutation sources preferentially generate amino-acid substitutions that evade immune recognition, producing immune-cold tumours regardless of tissue or mutation load. By analysing 9300 cancer exomes and performing mutagenesis experiments, we mapped links between mutagens, DNA-repair defects, and amino-acid substitution signatures (AAS). Surprisingly, the spectrum collapsed into five recurrent AAS with distinct functional profiles. AAS4—generated by alkylating agents and mismatch-repair (MMR) deficiency and enriched in kidney and liver cancers—is less likely to accumulate hydrophobic residues, yielding poorly immunogenic neopeptides. These tumors display immune-desert microenvironments and respond poorly to immunotherapy. However, certain human leukocyte antigen (HLA) class I variants, such as HLA-B*07:02, correlate with immune-hot tumours in this subgroup. HLA-B*07:02, common in Europeans, presents proline-enriched neopeptides derived from AAS4-mutations. Supporting this, B*07:02-positive cancer cells harbouring AAS4-type mutations stimulated T cell proliferation in vitro. These results show that neoantigen quality, not merely quantity, dictates anti-tumour immunity, explain inconsistent immunotherapy responses in MMR-deficient cancers, and advocate incorporating amino-acid substitution patterns into predictive biomarkers and therapy design.