Deciphering the Role of Cancer Stem Cells in Breast Cancer Brain Colonization Using a Novel Patient‐Derived Model

ABSTRACT Brain metastases from breast cancer (BCBM) are fatal and lack effective treatments. Their cellular and molecular drivers remain poorly understood, partly due to limited preclinical models that fail to capture patient tumor heterogeneity. Cancer stem‐like cells (CSCs) are implicated in metastatic dissemination; however, their specific role in brain metastasis remains unclear. In this study, CSCs are isolated from human BCBM specimens and characterized for stem‐like properties, including CD44 and ALDH1 expression, sphere formation, tumorigenicity, and in vitro and in vivo self‐renewal. Intra‐nipple and intra‐cardiac xenograft models demonstrate CSC ability to generate brain and bone metastases that recapitulate patient‐specific dissemination patterns. Transcriptomic and functional analyses reveal cellular heterogeneity and identify a metastasis‐initiating cell (MIC) subpopulation enriched in stemness and adhesion‐related pathways. These MICs exhibit enhanced adhesion to brain endothelium and undergo brain‐specific transcriptomic reprogramming that enables vascular co‐option, resistance to stromal stress, and survival‐promoting interactions with brain‐resident cells. High‐throughput drug screening indicates broad therapeutic resistance within the CSC compartment. Through the comprehensive characterization of BCBM‐derived CSCs, this study establishes a clinically relevant model that identifies CSCs and the MIC subpopulation as key drivers of brain metastatic progression and as promising targets for the development of effective therapeutic strategies.