Cell Cycle and Senescence p53/mdm2 Feedback Loop Sustains miR-221 Expression and Dictates the Response to Anticancer Treatments in Hepatocellular Carcinoma

The overexpression of microRNA-221 (miR-221) is reported in several human cancers including hepatocellular carcinoma, and its targeting by tailored treatments has been proposed. The evidence supporting the role ofmiR-221 in cancer is growing and has been mainly focused on the discovery of miR-221 targets as well as on its possible therapeutic exploitations. However, the mechanism sustaining miR-221 aberrant expression remains to be elucidated. In this study, MDM2 (E3 ubiquitin-protein ligase homolog), a known p53 (TP53) modulator, is identified as a direct target of miR-221, and a feed-forward loop is described that sustains miR-221 aberrant expression. Interestingly, miR-221 can activate the p53/mdm2 axis by inhibiting MDM2 and, in turn, p53 activation contributes to miR-221 enhanced expression. Moreover, by modulating the p53 axis, miR-221 impacts cell-cycle progression and apoptotic response to doxorubicin in hepatocellular carcinoma–derived cell lines. Finally, CpG island methylation status was assessed as a causative event associated with miR-221 upregulation in hepatocellular carcinoma cells and primary tumor specimens. In hepatocellular carcinoma–derived cell lines, pharmacologically inducedDNAhypomethylation potentiated a significant increase inmiR-221 expression. These data were confirmed in clinical specimens of hepatocellular carcinoma in which elevated miR-221 expression was associated with the simultaneous presence of wild-type p53 and DNA hypomethylation. Implications:These findings reveal a novel miR-221–sustained regulatory loop that determines a p53-contextspecific response to doxorubicin treatment in hepatocellular carcinoma. Mol Cancer Res; 12(2); 203–16.