P2Y ₂R-mediated inflammasome activation is involved in tumor progression in breast cancer cells and in radiotherapy-resistant breast cancer

摘要

In the tumor microenvironment, extracellular nucleotides are released and accumulate, and can activate the P2Y _(2) receptor (P2Y _(2)R), which regulates various responses in tumor cells, resulting in tumor progression and metastasis. Moreover, the inflammasome has recently been reported to be associated with tumor progression. However, the role of P2Y _(2)R in inflammasome activation in breast cancer cells is not yet well defined. Therefore, in this study, we investigated the role of P2Y _(2)R in inflammasome-mediated tumor progression in breast cancer using breast cancer cells and radiotherapy-resistant (RT-R) breast cancer cells. We established RT-R-breast cancer cells (RT-R-MDA-MB-231, RT-R-MCF-7, and RT-R-T47D cells) by repeated irradiation (2 Gy each, 25 times) in a previous study. In this study, we found that the RT-R breast cancer cells exhibited an increased release of adenosine triphosphate (ATP) and P2Y _(2)R activity. In particular, the RT-R-MDA-MB-231 cells derived from highly metastatic MDA-MB-231 cells, exhibited a markedly increased ATP release, which was potentiated by tumor necrosis factor (TNF)-α. The MDA-MB-231 cells exhibited inflammasome activation, as measured by caspase-1 activity and interleukin (IL)-1β secretion following treatment with TNF-α and ATP; these effects were enhanced in the RT-R-MDA-MB-231 cells. However, the increased caspase-1 activities and IL-1β secretion levels induced in response to treatment with TNF-α or ATP were significantly reduced by P2Y _(2)R knockdown or the presence of apyrase in both the MDA-MB-231 and RT-R-MDA-MB-231 cells, suggesting the involvement of ATP-activated P2Y _(2)R in inflammasome activation. In addition, TNF-α and ATP increased the invasive and colony-forming ability of the MDA-MB-231 and RT-R-MDA-MB-231 cells, and these effects were caspase-1-dependent. Moreover, matrix metalloproteinase (MMP)-9 activity was modulated by caspase-1, in a P2Y _(2)R-dependent manner in the MDA-MB-231 and RT-R-MDA-MB-231 cells. Finally, nude mice injected with the RT-R-MDA-MB-231-EV cells (transfected with the empty vector) exhibited increased tumor growth, and higher levels of MMP-9 in their tumors and IL-1β levels in their serum compared with the mice injected with the RT-R-MDA-MB-231- P2Y _(2)R shRNA cells (transfected with P2Y _(2)R shRNA). On the whole, the findings of this study suggest that extracellular ATP promotes tumor progression in RT-R-breast cancer cells and breast cancer cells by modulating invasion and associated molecules through the P2Y _(2)R-inflammasome activation pathway.