Knockdown of RET mRNA expression sensitizes endocrine-resistant MCF-7:5C breast cancer cells to 4-hydroxytamoxifen. (a) Rearranged during transfection (RET) protein and mRNA levels in wild-type MCF-7 cells and endocrine-resistant MCF-7:5C cells as determined by western blot analysis and quantitative real-time PCR, respectively. (b) MCF-7:5C cells were transfected with either a RET-targeted siRNA or a nontarget control siRNA for 48 hours and RET protein and mRNA expression levels were determined by western blotting and real-time PCR. (c) Effect of RET suppression on the proliferation of MCF-7:5C cells. Cells were transfected with a RET-targeted or a nontargeting control siRNA, and after 24 hours cells were seeded into 12-well plates and exposed continuously to the indicated concentrations of 4-hydroxytamoxifen (4OHT) or vehicle for an additional 72 hours. Cell proliferation was determined by cell counting using a hemacytometer. Data shown are representative of three independent experiments. UT, untreated.