Tertiary lymphoid structures in Merkel cell carcinoma facilitate naïve and central memory T-cell infiltration linked...

The presence of tertiary lymphoid structures (TLS) in solid tumors, including Merkel cell carcinoma (MCC), is associated with a better prognosis and a better response to immunotherapy with immune checkpoint inhibition (ICI). The detailed mechanisms by which TLS influence antitumor immune responses are only partially understood.

Clinically annotated tumor tissues of 27 patients with MCC were obtained prior to ICI therapy. Tumor samples were subjected to transcriptomic and multiplex immuno-visual profiling, T-cell receptor (TCR) clonotype mapping, as well as-in selected cases-spatial transcriptomics to comprehensively characterize the tumor immune microenvironment.

Weighted gene co-expression network analysis (WGCNA) of transcriptomic data in combination with topological overlap measures indicated a higher abundance of TLS in tumors of patients with MCC responding to ICI therapy. This concept was substantiated through immunomorphological analyses, revealing mature B-cell follicle-like structures characterized by high endothelial venules (HEVs). Further supporting HEVs as critical entry points for naïve T cells, the presence of TLS was correlated with a pronounced infiltration of CD4⁺ and CD8⁺ T cells, exhibiting both naïve and central memory phenotypes. The TCR repertoire of these infiltrates exhibited enhanced richness and diversity with a pronounced reactivity toward Merkel cell polyomavirus-derived T-cell epitopes. Spatially resolved RNA and V(D)J sequencing revealed the expression of genes associated with T-cell recruitment within TLS, alongside the presence of naïve and central memory T-cell markers. Notably, individual clonally expanded TCR transcripts were detected both within TLS and among tumor-infiltrating lymphocytes. The latter were associated with low expression of memory cell markers and high expression of effector cell markers. Additionally, a spatial gradient in the expression of genes linked to immune stress in MCC cells-such as those involved in the interferon-γ response and antigen processing and presentation machinery-originated in proximity to the TLS.

Our findings are consistent with a key role of TLS in shaping immune interactions within the MCC microenvironment, driving the recruitment of diverse tumor-reactive T cells. These insights hold promise for advancing immunotherapeutic strategies.