Multimodal profiling uncovers tertiary lymphoid structures as a critical determinant of immunotherapy response and prognosis in nasopharyngeal carcinoma

•The immune TME in NPC is unsupervisedly dissected into “Immune Inflamed” and “Immune Deficient” subtypes.•Tertiary lymphoid structures (TLS) serve as better prognostic indicators than individual immune cells.•TLS enhance anti-tumor immunity by increasing immune cell diversity and infiltration.•The prognostic value of TLS applies across various cancer types and immunotherapy responses. Nasopharyngeal carcinoma (NPC), historically termed ‘lymphoepithelioma-like carcinoma’ due to its rich lymphocyte infiltration, benefit from PD-1 blockade treatment. However, a comprehensive understanding of its tumor microenvironment (TME) remains elusive, hindering the identification of effective biomarkers for immunotherapy. We leveraged multimodal profiling data, including gene expression, immunohistochemistry, and multiplex immunohistochemistry, from three independent cohorts of NPC patients with a total of 327 patients to dissect the TME in NPC. Unsupervised hierarchical clustering of TME cell populations in the discovery cohort revealed two novel subtypes with distinct prognosis: ‘Immune Inflamed’ and ‘Immune Deficient’. Intriguingly, the most significant differences between the two subtypes were the abundance of B cells and tertiary lymphoid structures (TLS), with a nearly two-fold increase in TLS presence in the Immune Inflamed subtype. The prognostic significance of TLS was confirmed in three independent NPC cohorts, surpassing the prognostic value of individual immune cell subsets. Mechanistically, TLS enhanced anti-tumor immunity by increasing T and B cell receptor repertoire diversity, promoting infiltration of plasma cells, macrophages, and natural killer cells, and consequently increasing antibody-dependent cell-mediated cytotoxicity and antibody-dependent phagocytosis. Finally, TLS status robustly predicted prognosis in a cohort of NPC patients treated with PD-1 blockade, and its prognostic value was consistent across a pan-cancer immunotherapy cohort of 10 tumors and 1158 patients, although with context-specific effects depending on cancer type and immunotherapy modality. In conclusion, this study provides compelling evidence that TLS is a robust indicator of overall immune response within TME and have great potential to guide individualized immunotherapy.