Comparative evaluation of T-cell receptors in experimental glioma-draining lymph nodes.

ABSTRACT

BACKGROUND: Glioblastomas, the most common primary malignant brain tumors, are considered immunologically cold malignancies due to growth in an immune sanctuary site. While peptide vaccines have shown to generate intra-tumoral antigen-specific T cells, the identification of these tumor-specific T cells is challenging and requires detailed analyses of tumor tissue. Several studies have shown that CNS antigens may be transported via lymphatic drainage to cervical lymph nodes, where antigen-specific T-cell responses can be generated. Therefore, we investigated whether glioma-draining lymph nodes (TDLN) may constitute a reservoir of tumor-reactive T cells. METHODS: We addressed our hypothesis by flow cytometric analyses of chicken ovalbumin (OVA)-specific CD8+ T cells as well as T-cell receptor beta (TCRß) next-generation-sequencing (TCRß-NGS) of T cells from tumor tissue, TDLN, spleen, and inguinal lymph nodes harvested from experimental mouse GL261 glioma models. RESULTS: Longitudinal dextramer-based assessment of specific CD8+ T cells from TDLN did not show tumor model antigen reactivity. Unbiased immunogenomic analysis revealed a low overlap of TCRß sequences from glioma-infiltrating CD8+ T cells between mice. Enrichment scores, calculated by the ratio of productive frequencies of the different TCRß-CDR3 amino-acid (aa) rearrangements of CD8+ T cells derived from tumor, TDLN, inguinal lymph nodes, and spleen demonstrated a higher proportion of tumor-associated TCR in the spleen compared to TDLN. CONCLUSIONS: In experimental glioblastoma, our data did not provide evidence that glioma-draining cervical lymph nodes are a robust reservoir for spontaneous glioma-specific T cells highlighting the requirement for detailed analyses of glioma-infiltrating T cells for the discovery of tumor-specific TCR.