A2B adenosine receptor agonist induces cell cycle arrest and apoptosis in breast cancer stem cells via ERK1/2 phosphorylation

It has been reported that cancer stem cells (CSCs) may play a crucial role in the development, recurrence and metastasis of breast cancer. Targeting signaling pathways in CSCs is considered to be a promising strategy for the treatment of cancer. Here, we investigated the role of the A2B adenosine receptor (A2BAR) and its associated signaling pathways in governing the proliferation and viability of breast cancer cell line derived CSCs. CSCs were isolated from the breast cancer cell lines MCF-7 and MDA-MB-231 using a mammosphere assay. The effect of the A2BAR agonist BAY606583 on cell proliferation was evaluated using XTT and mammosphere formation assays, respectively. Apoptosis was assessed using Annexin-V staining and cell cycle analyses were performed using flow cytometry. The expression levels of Bax, Bcl-2, cyclin-D1, CDK-4 and (phosphorylated) ERK1/2 were assessed using Western blotting. Our data revealed that the breast cancer cell line derived mammospheres were enriched for CSCs. We also found that A2BAR stimulation with its agonist BAY606583 inhibited mammosphere formation and CSC viability. In addition, we found that the application of BAY606583 led to CSC cell cycle arrest and apoptosis through the cyclin-D1/Cdk-4 and Bax/Bcl-2 pathways, respectively. Notably, we found that BAY606583 significantly down-regulated ERK1/2 phosphorylation in the breast cancer cell line derived CSCs. From our results we conclude that A2BAR induces breast CSC cell cycle arrest and apoptosis through downregulation of the ERK1/2 cascade. As such, A2BAR may be considered as a novel target for the treatment of breast cancer.