The endosteal niche regulates breast cancer cell dormancy in bone: identification of new molecular determinants.

Abstract

Cellular dormancy compromises the long-term survival of breast cancer patients. Bone represents a frequent site for metastasis, where Spindle-shaped N-cadherin⁺CD45^- osteoblasts (SNOs) hold dormant metastatic cells in the endosteal niche with a Notch2-dependent mechanism. In this work, we excluded the involvement of Notch1 in SNO-induced breast cancer cellular dormancy by immunofluorescence/immunohistochemistry and molecular approaches, using breast cancer tissues and cell lines. RNAdSeq in human bone metastatic MDA-MB231 breast cancer cells sorted for Notch1^HIGH and Notch2^HIGH expression demonstrated that, compared to their low counterpart, only Notch2^HIGH cells expressed enriched pathways relevant for the metastatic process, including pluripotency and Hematopoietic Stem Cell (HSC) gene signatures. They expressed the HSC-associated genes CXCR4, CD34 and TIE2 and MDA-MB231 cells enriched in the encoded proteins showed lesser proliferation ability. Reduced incidence of osteolytic lesions was induced by CXCR4^HIGH cells intratibially injected in immunocompromised mice, while lower lesion extension was induced by CXCR4^HIGH and TIE2^HIGH injected cells compared to CXCR4^LOW and TIE2^LOW cells. Notch2^HIGH cells were enriched in endoplasmic reticulum stress and unfolded protein response genes and overexpressed the CD177 protein, while the CD177 ligands, Plaur, Itgam and Ceacam 1, were highly expressed in SNOs. Kaplan-Meier plots showed positive correlation between high expression of CD177, ITGAM and CEACAM 1 - but not PLAUR - and overall survival of patients. CD177^HIGH cells were also CXCR4^HIGH, CD34^HIGH and Notch2^HIGH and proliferated less than CD177^LOW cells. These results support the relevance of Notch2 in SNO-mediated cellular dormancy and identified new pathways implicated in bone metastatic breast cancer cell quiescence.