Endocrine Disruptors Fludioxonil and Fenhexamid Stimulate miR-21 Expression in Breast Cancer Cells

Fenhexamid and fludioxonil are antifungal agents used in agricultural applications, which are present at measurable amounts in fruits and vegetables. Fenhexamid and fludioxonil showed endocrine disruptor activity as antiandrogens in an androgen receptor reporter assay in engineered human breast cancer cells. Little is known about how environmental chemicals regulate microRNA (miRNA) expression. This study examined the effect of fenhexamid and fludioxonil on the expression of the oncomiR miR-21 in MCF-7, T47D, and MDA-MB-231 human breast cancer cells and downstream targets of miR-21 in MCF-7 cells. Fenhexamid and fludioxonil stimulated miR-21 expression in a concentration-dependent manner and reduced the expression of miR-21 target Pdcd4 protein. Antisense to miR-21 blocked the increase in Pdcd4 protein by fenhexamid and fludioxonil. Fenhexamid and fludioxonil reduced miR-125b and miR-181a, demonstrating specificity of miRNA regulation. Induction of miR-21 was inhibited by the estrogen receptor antagonist fulvestrant, by androgen receptor antagonist bicalutamide, by actinomycin D and cycloheximide, and by inhibitors of the mitogen-activated protein kinases and phosphoinositide 3-kinase pathways. Fenhexamid activation was inhibited by the arylhydrocarbon receptor antagonist -napthoflavone. Fenhexamid and fludioxonil did not affect dihydrotestosterone-induced miR-21 expression. Fludioxonil, but not fenhexamid, inhibited MCF-7 cell viability, and both inhibited estradiol-induced cell proliferation and reduced cell motility. Together these data indicate that fenhexamid and fludioxonil use similar and distinct mechanisms to increase miR-21 expression with downstream antiestrogenic activity.