Spatial self-organization of cancer stem cell niches revealed by live single-cell imaging

• Ultra-wide field microscopy with a stemness reporter enables long-term tracking of breast cancer cell phenotypes, revealing spatial zonation of CSCs and CDCs into niche-like clusters. • Spontaneous reprogramming from differentiated to cancer stem cells occurs even in unperturbed populations, often during the cell cycle, challenging the unidirectional differentiation model. • Phenotypic inheritance is partial, but cell-cell interactions critically influence transitions: neighboring CSCs promote reprogramming while CDCs inhibit it, driving spatial self-organization. • These findings provide a mechanistic framework for understanding tumor heterogeneity and resistance, with implications for developing therapies that target CSC plasticity and niche maintenance.