Abstract: Cancer stem cells (CSCs) have become an important target to overcome the obstacle of tumor therapy. In recent years, many reports have shown that the heterogenicity and plasticity of CSCs, and the microenvironment interact with each other, which become important factors affecting the dynamic conversion of CSCs. Therefore, we should not only focus on the stem characteristics of CSCs, but also consider the factors related to the regulation of the dynamic transformation of CSCs stemness. The signaling pathways, epithelial-mesenchymal transition (EMT) and the niche of CSCs, are proved to play an important role in the regulation of stemness of CSCs. The aspects are important in the studies of tumor therapy by targeting CSCs. This article summarizes the mechanisms for regulation of the stemness of cancer stem cells and the progress in the studies of targeting cancer stem cells with a focus on the CSCs dynamic regulation.