Runx2 is required for activity of CD44⁺/CD24^-/low breast cancer stem cell in breast cancer development

Runx2, a master regulator of osteogenesis, is abnormally expressed in advanced breast cancer. Here we addressed Runx2 contribution to breast cancer cell growth and metastasis. We found that CD44 and Runx2 were both elevated in breast cancer tissues compared with the adjacent normal tissues in breast cancer patients. Runx2 expression was significantly correlated with tumor TNM stage, metastasis and poor prognosis. We then screened several breast cancer cell lines and found that Runx2 expression level was positively related to the malignant level of the cells screened. Knockdown of Runx2 in high metastatic cell line MDA-MB-231 could inhibit breast cancer cell vitality, invasion and clone formation capacity, while overexpression of Runx2 in low metastatic cell line MCF-7 could increase those malignant behaviors. The mechanism might be due to Runx2 positively regulating cancer stem cell properties, as CD44 expression level and CD44⁺/CD24^-/low breast cancer stem cell population were both significantly decreased in Runx2 knockdown cells. Cancer stem cell renewal ability such as soft agar clone formation, mammospheres formation and tumor formation ability in null mice were all decreased after knockdown of Runx2. On the contrary, overexpression of Runx2 could enhance all above stem cell renewal ability. Lastly, we explored how Runx2 changes cancer stem cell population. We found it could affect epithelial mesenchymal transition (EMT). Runx2 could regulate mesenchymal marker and epithelial marker expression and affect activation of Wnt/β-catenin signaling pathway. These results together strongly suggest that Runx2 can promote CD44⁺/CD24^-/low breast cancer stem cell properties and breast cancer tumorigenesis through EMT process.