A relatively rare aldehyde dehydrogenase 1 (ALDH1)-positive "stem cell-like" subpopulation of tumor cells has the unique ability to initiate and perpetuate tumor growth; moreover, it is highly resistant to chemotherapy and significantly associated with poor clinical outcomes. The development of more effective therapies for cancer requires targeting of this cell population. Using cDNA microarray analysis, we identified that the expression of the Caenorhabditis elegans lin-28 homologue (LIN28) was positively correlated with the percentage of ALDH1+ tumor cells; this was further validated in an independent set of tissue arrays (n=197). Both loss-of-function and gain-of-function studies showed that LIN28 plays a critical role in the maintenance of ALDH1+ tumor cells. In addition, we found that there is a double-negative feedback loop between LIN28 and let-7 in tumor cells, and that let-7 negatively regulates ALDH1+ tumor cells. Finally, we report that a LIN28/let-7 loop modulates self-renewal and differentiation of mammary gland epithelial progenitor cells. Our data provide evidence that cancer stem cells may arise through a "reprogramming-like" mechanism. A rebalancing of the LIN28/let-7 regulatory loop could be a novel therapeutic strategy to target ALDH1+ cancer stem cells.