RESUMO
Toll-like receptors (TLRs) are indispensable for the activation, maintenance and halting of immune responses. TLRs can mediate inflammation by recognizing molecular patterns in microbes (pathogen-associated molecular patterns: PAMPs) and endogenous ligands (danger-associated molecular patterns: DAMPs) released by injured or dead cells. For this reason, TLR ligands have attracted much attention in recent years in many cancer vaccines, alone or in combination with immunotherapy, chemotherapy and radiotherapy (RT). TLRs have been shown to play controversial roles in cancer, depending on various factors that can mediate tumor progression or apoptosis. Several TLR agonists have reached clinical trials and are being evaluated in combination with standard of care therapies, including RT. Despite their prolific and central role in mediating immune responses, the role of TLRs in cancer, particularly in response to radiation, remains poorly understood. Radiation is recognized as either a direct stimulant of TLR pathways, or indirectly through the damage it causes to target cells that subsequently activate TLRs. These effects can mediate pro-tumoral and anti-tumoral effects depending on various factors such as radiation dose and fractionation, as well as host genomic features. In this review, we examine how TLR signaling affects tumor response to RT, and we provide a framework for the design of TLR-based therapies with RT.
RESUMO
BACKGROUND: Radioresistance of HNSCCs remains a major challenge for effective tumor control. Combined radiotherapy (RT) and immunotherapy (IT) treatment improved survival for a subset of patients with inflamed tumors or tumors susceptible to RT-induced inflammation. To overcome radioresistance and improve treatment outcomes, an understanding of factors that suppress anti-tumor immunity is necessary. In this regard, regulatory T cells (Tregs) are critical mediators of immune suppression in HNSCCs. In this study, we investigated how radiation modulates Treg infiltration in tumors through the chemokine CCL20. We hypothesized that radiation induces CCL20 secretion resulting in Treg infiltration and suppression of anti-tumor immunity. METHODS: Human and mouse HNSCC cell lines with different immune phenotypes were irradiated at doses of 2 or 10 Gy. Conditioned media, RNA and protein were collected for assessment of CCL20. qPCR was used to determine CCL20 gene expression. In vivo, MOC2 cells were implanted into the buccal cavity of mice and the effect of neutralizing CCL20 antibody was determined alone and in combination with RT. Blood samples were collected before and after RT for analysis of CCL20. Tumor samples were analyzed by flow cytometry to determine immune infiltrates, including CD8 T cells and Tregs. Mass-spectrometry was performed to analyze proteomic changes in the tumor microenvironment after anti-CCL20 treatment. RESULTS: Cal27 and MOC2 HNSCCs had a gene signature associated with Treg infiltration, whereas SCC9 and MOC1 tumors displayed a gene signature associated with an inflamed TME. In vitro, tumor irradiation at 10 Gy significantly induced CCL20 in Cal27 and MOC2 cells relative to control. The increase in CCL20 was associated with increased Treg migration. Neutralization of CCL20 reversed radiation-induced migration of Treg cells in vitro and decreased intratumoral Tregs in vivo. Furthermore, inhibition of CCL20 resulted in a significant decrease in tumor growth compared to control in MOC2 tumors. This effect was further enhanced after combination with RT compared to either treatment alone. CONCLUSION: Our results suggest that radiation promotes CCL20 secretion by tumor cells which is responsible for the attraction of Tregs. Inhibition of the CCR6-CCL20 axis prevents infiltration of Tregs in tumors and suppresses tumor growth resulting in improved response to radiation.
