MicroRNAs (miRNAs) regulate protein expression by targeting messenger RNAs (mRNAs) for cleavage or by inducing translational repression. In breast cancer, estrogen receptor-a (ERa) levels are critical for cancer cell growth, but our knowledge of miRNA-dependent ERa regulation is limited. In order to identify miRNAs impacting ERa expression in breast cancer, we transiently transfected 319 pre-miRs into MCF-7 and BT-474 cell lines and monitored their effects on ERa expression by protein lysate microarrays. There was a significant enrichment of the in silico predicted ERa targeting miRNAs among the miRNAs that experimentally reduced ERa protein expression. The ERa inhibition by 21 pre-miRs was validated with western blotting and qRT-PCR. Proliferation assays demonstrated that the impact of ERa-downregulating miRNAs on the inhibition of estrogen-stimulated growth of MCF-7 cells was as strong as that of an siRNA for ERa. Overexpression of the top pre-miR hits (miR-18a/b, miR-193b, miR-206, and miR-302c) followed by gene microarray profiling revealed downregulation of ERa responsive genes, and ERa 3'UTR-luciferase reporter assays confirmed that these miRNAs directly targeted ERa. Finally, miR-18a and miR-18b showed increased expression in ERa-negative as compared to ERa-positive clinical tumors. Taken together, these results suggest that miRNAs may be useful in the therapeutic inhibition of ERa expression, and that upregulation of certain miRNAs may be involved in the silencing of ERa in the progression towards estrogen-independent breast cancers.
|100th Annual Meeting of the American Association for Cancer Research
|18/04/09 → 22/04/09