A little STING spices up tertiary lymphoid structures: Therapeutic induction of mature TLS by STING and lymphotoxin-beta receptor activation confers immune protection against cancer metastasis

Significant clinical evidence demonstrates an association of tertiary lymphoid structure (TLS) formation with cancer survival. Clinically translatable pharmacologic/biologic agents that induce TLS-rich tumor immune environment could provide a breakthrough strategy to treat therapy-resistant cancers. Here, we show that simultaneous activation of innate immune effectors, STING and lymphotoxin-beta receptor (LTbetaR), by their agonists induces the formation of multiple TLS in TLS-free tumors in a manner dependent on CD4 + and CD8 + T cells. Using this approach, functional TLS were induced in all treated mice, in different tumor types and anatomical sites-TLS developed in adenocarcinomas and sarcomas in the pancreas, mammary gland, skeletal muscle, and subcutaneous tissues. TLS also developed in the brain harboring gliomas. STING activation alone was insufficient for inducing B cell-containing TLS or eliciting long-term therapeutic effects. However, when combined with LTbetaR activation, it improved the fitness of TLS with B cell expansion and maturation to IgG-producing long-lived plasma cells and memory cells, exhibiting lymph node follicle-like germinal center responses within tumors. In addition, the treatment induced high endothelial venule formation, increased CD4 + T cell recruitment, memory CD8 + T cell expansion, and the accumulation of TCF1 + stem-like CD8 + T cells around TLS. These immune responses resulted in strong suppression of tumor growth and metastasis, with excellent survival benefit, demonstrating that the treatment successfully immunized mice against tumor cells. The STING/LTbetaR combination therapy also substantially improved the efficacy of anti-PD-1 immune checkpoint blockade in resistant tumors. The immediate early response to the combination therapy was mediated mainly by CD8 + T cells. The delayed but prolonged response was mediated by humoral immunity of B cells, which exerted a lasting anti-tumor effect together with the cellular immunity of CD8 + T cells and NK cells. Our finding establishes a foundation for the therapeutic induction of TLS in cancer. Unlike other reported strategies, TLS induction by this method does not require viral antigens or artificial genetic manipulations to boost immune response, making it feasible for clinical applications. The high-affinity tumor-specific IgG-producing long-lived plasma cells and memory B and T cells generated in these TLS could provide lifelong protection against tumor recurrence and metastases. Effective tumor immunization by this strategy is expected to improve other cancer immunotherapies and suggests broad therapeutic applications in cancer. Since STING agonists are clinically available and humanized agonistic antibodies to LTbetaR can be developed, this strategy is readily translatable to clinical use for cancer treatment.