RANKL regulates differentially breast cancer stem cell properties through its RANK and LGR4 receptors

Biochim Biophys Acta Mol Cell Res. 2024 Dec 9;1872(2):119888. doi: 10.1016/j.bbamcr.2024.119888. Online ahead of print.

Abstract

Background: Breast cancer stem cells (BCSC) are a subpopulation responsible for cancer resistance and relapse. The receptor activator of nuclear factor kappa-Β ligand (RANKL) is a cytokine capable of activating RANK and LGR4 receptors. RANKL/RANK signaling maintains the self-renewal of BCSCs, however, the effect of RANKL via LGR4 remains unclear. Evidence from osteoclasts suggests that RANKL/LGR4 axis disrupts RANK signaling, leading to opposing cellular responses. Anti-RANKL inhibitors are potential agents for eradicating CSCs, but their effect on RANKL/LGR4 signal has not been demonstrated.

Objective: This project aimed to elucidate the role of RANKL in regulating stemness depending on the expression of its receptors.

Methods: We use in vitro and in vivo approaches to evaluate the effects of RANKL inhibition in stemness in low or high-LGR4 expressing cells. Furthermore, we analyze the effects of RANKL stimulation on the stemness of LGR4 or RANK overexpressing cells. Additionally, we evaluated the impact of RANKL/LGR4 signaling in the activity of Wnt/β-catenin and NF-κB signaling pathways.

Results: Our findings indicated that elevated RANKL expression is related to a favorable prognosis in patients with high LGR4 levels. Furthermore, RANKL inhibition decreased BCSC properties in LGR4-low cell lines, while it promoted migration in LGR4-high cells. Additionally, the RANKL/RANK axis activated NF-κB signaling and enhanced BCSCs in RANK-overexpressing cells. In contrast, in LGR4-overexpressing cells, RANKL failed to activate NF-κB but instead inhibited the Wnt/β-catenin pathway, leading to a reduction in BCSCs.

Conclusion: Our findings suggest that RANKL exerts different responses according to the expression of its receptors.

Keywords: Cancer stem cells; LGR4; NF-κB.; RANKL; Stemness; WNT.