Session: SAT 292-325-Breast & Prostate Cancer
Poster Board SAT-314
The microRNA, miR-34a, is a component of the p53 tumor suppressor pathway and has been shown to mediate induction of cell cycle arrest, senescence, and apoptosis in cancer cells (3, 4.) Indole-3-carbinol (I3C) derived from cruciferous vegetables and artemisinin isolated from the sweet wormwood plant, Artemisia annua, effect components of the p53 pathway to growth arrest human breast cancer cells, implicating a potential role for miR-34a in their anti-proliferative effects (5, 6.) Both compounds are currently undergoing clinical trials for the treatment of reproductive cancers (7, 8.)
To investigate the possibility of miR-34a regulation by I3C and artemisinin, human breast cancer cells containing wild-type or mutant p53 were treated with either compound under conditions of cell cycle arrest as detected by flow cytometry. Quantitative PCR analysis of mature microRNA levels revealed a time and dose-dependent upregulation of miR-34a that correlated with induction of functional p53 by I3C and decreases in the mRNA and protein levels of CDK4 and CDK6, cyclin-dependant kinases that are known targets of miR-34a inhibition.
Luciferase assays in which cells were transfected with the miR-34a binding site of CDK6 or CDK4 mRNA attached to the firefly luciferase reporter gene confirmed miR-34a reduces CDK6 and CDK4 levels in cells growth arrested by I3C or artemisinin.
miR-34a also appears critical for the anti-proliferative effects of both compounds as cells in which miR-34a levels have been reduced by transfection of non-translatable miR-34a targets did not growth arrest upon treatment with either drug. Levels of CDK6 and CDK4 also remained unchanged in treated miR-34a knockdown cells, suggesting miR-34a is a crucial component of CDK regulation by both phytochemicals.
Interestingly, transfection of dominant negative p53 prevented I3C upregulation of miR-34a in growth arrested cells containing wild-type p53 yet had no effect on artemisinin regulation of miR-34a, indicating a p53-indepedent mechanism of miR-34a regulation. Artemisinin also upregulates miR-34a in breast cancer cell lines containing non-functional p53.
All of these data suggest that miR-34a plays a critical role in the anti-proliferative effects of artemisinin and indole-3-carbinol in human breast cancer cells. Such evidence further elucidates the therapeutic potential of either drug to ectopically express tumor suppressive microRNA while implicating the use of miR-34a expression levels to determine the efficacy of phytochemical treatment.
Nothing to Disclose: KH, LH, GLF
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