Abstract
BACKGROUND: Our adoptive cellular therapy (ACT) enhances dendritic cell (DC) and T cell infiltration into tumors, improving survival in murine brain tumor models. However, tumors employ mechanisms to restrict immune surveillance and efficacy of immunotherapy. We hypothesize that tumor-induced DC dysfunction facilitates immune evasion during ACT. This study investigates the mechanisms of DC dysfunction in ACT-escaped gliomas. METHODS: T cell phenotypes were characterized using flowcytometry, and DC function was assessed through T cell activation assays. Gene set enrichment analysis (GSEA) was performed on transcriptomic data to identify enriched pathways. The impact of hypoxia and factors secreted from tumor-T cell interactions on DC tolerance was evaluated using RT-qPCR. In vivo correlations between hypoxia and DC tolerance were analyzed using GeoMx spatial transcriptomics. RESULTS: ACT-escaped tumors retained adoptively transferred cytotoxic but non-exhausted T cells that failed to recognize antigen-shifted tumors. DCs from both untreated primary and ACT-escaped tumors showed impaired T cell activation and reduced expression of antigen-presentation genes. DCs from ACT-escaped tumors exhibited increased expression of tolerance-associated genes, with significant enrichment in hypoxia pathway genes. Hypoxia induced the expression of DC tolerance genes in a HIF1α-dependent manner, impairing T cell activation. Spatial transcriptomics confirmed a strong correlation between HIF1α and ARG1 in tumor-bearing brains. Increased immune infiltration in ACT-treated gliomas exacerbated hypoxia, as shown by HIF1α and CD45 co-expression, further driving DC dysfunction. Inflammatory factors secreted during tumor-T cell interactions activated hypoxia pathways and induced DC tolerance genes. CONCLUSION: Hypoxia-driven DC tolerance is a key driver of immune escape under ACT.