There is a well-established association between aging and the onset of metastasis. Although the mechanisms through which age impinges upon the malignant phenotype remain uncharacterized, the role of a senescent microenvironment has been emphasized. We reported previously that human epithelial cells that undergo telomere-driven chromosome instability (T-CIN) display global microRNA (miR) deregulation and develop migration and invasion capacities. Here, we show that post-crisis cells are not able to form tumors unless a senescent microenvironment is provided. The characterization of cell lines established from such tumors revealed that these cells have acquired cell autonomous tumorigenicity, giving rise to heterogeneous tumors. Further experiments demonstrate that explanted cells, while displaying differences in cell differentiation markers, are all endowed of enhanced stem cell properties including self-renewal and multilineage differentiation capacity. Treatments of T-CIN+ cells with senescence-conditioned media induce sphere formation exclusively in cells with senescence-associated tumorigenicity, a capacity that depends on miR-145 repression. These results indicate that the senescent microenvironment, while promoting further transdifferentiations in cells with genome instability, is able to propel the progression of premalignant cells towards a malignant, cell stem-like state.

Whereas phenotypic assays such as Boyden chambers and wound healing assays can easily be employed to characterize the migratory potential of cells at the population level, few methods exist that can sort subpopulations of cells based on their migratory behaviour from an initial heterogeneous pool. In this paper, we present an approach to sort migratory cancer and immune cells based on their spontaneous migration in 2D and 3D microenvironments.Using this method, which is easy to implement and readily scalable, millions of live cells can be sorted based on their migratory characteristics and then subjected to downstream genomic, molecular and functional tests. We reveal that enrichment of the most migratory cytotoxic T lymphocytes yields a pool of cells with enhanced cytotoxicity against cancer cells. This new functional sorting method opens new avenues for the precise characterization of the mechanisms underlying hitherto unexplained heterogeneities in migratory phenotypes within a cell population, and for the targeted enrichment of the most potent migratory leukocytes in immunotherapies.