CXCL13 and CCL21 Induce Tertiary Lymphoid Structures and Enhance the Efficacy of Immunotherapy for Melanoma

Tertiary lymphoid structures (TLS) are acquired ectopic lymph follicle-like structures observed inside and around tumors, in which clusters of CD20-positive B lymphocytes are surrounded by CD3-positive T lymphocytes. In many cancers, the existence of TLS is a useful biomarker for better prognosis and better response to immune checkpoint inhibitors (ICI) and plays important roles in activating anti-tumor immunity. In order to induce TLS and enhance the therapeutic effect of ICI, we attempted to induce TLS using multiple chemokines in malignant melanoma, for which there have been no reports of TLS induction previously. Immunohistochemical analysis of tumor samples from 41 melanoma patients treated with ICI revealed TLS in 63.4% of cases. Patients with ≥ 5 TLS exhibited significantly improved disease-specific survival compared to those with fewer or no TLS. Plasma chemokine profiling in 46 samples from 18 melanoma patients showed elevated CC motif chemokine ligand 21 (CCL21) in TLS-positive samples before and after ICI treatment and CXC motif chemokine ligand 13 (CXCL13) significantly increased pre- to post-ICI treatment in paired samples from TLS-positive patients. In a mouse melanoma model, co-administration of CXCL13 and CCL21 alongside anti-programmed death ligand-1 (PD-L1) antibody therapy significantly increased TLS formation and improved tumor growth suppression. Gene expression analysis of human melanoma samples demonstrated that high CXCL13 and CCL21 expression correlated with upregulation of immune response, particularly B cell activation. These findings highlight the potential of chemokine-based therapies. TLS induction using CXCL13 and CCL21 in combination may be useful for enhancing the effects of ICI therapy in melanoma.