To validate the relationship between cancer stemness and ECM stiffness, we investigated expression of the bona fide cancer stem cell marker CD133 (Alamgeer et al, 2013) and again showed positive correlation between increased ECM stiffness (4kPA) and greater expression in H1299control but not in H1299RASSF1A cells, suggesting that RASSF1A prevents cancer stemness in soft ECM (Fig 5C). This evidence concerns the gene PROM1 and cancer.