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. 2020 Mar 11;5:25. doi: 10.1038/s41392-020-0118-x

Fig. 1. CDK4/6 inhibition blocks breast cancer metastasis by inducing ZEB1 protein degradation.

Fig. 1

a Western blotting of ZEB1 expression in MDA-MB-231 and SUM-159 cells after treatment with the indicated concentrations of palbociclib for 48 h. b CHX pulse-chase analysis of ZEB1 protein stability in MDA-MB-231 and SUM-159 cells after treatment with palbociclib at the indicated time points. The results were normalized to the levels of β-actin. c Western blotting of ZEB1 protein expression in MDA-MB-231 and SUM-159 cells after treatment with MG132 in the presence or absence of palbociclib. d Coimmunoprecipitation analysis of ZEB1 protein ubiquitination in MDA-MB-231 and SUM-159 cells after treatment with palbociclib for 48 h. The cells were treated with MG132 for 12 h prior to harvest. e Western blotting of EMT markers in MDA-MB-231 and SUM-159 cells treated with palbociclib for 48 h. f, g Transwell migration (f) and wound-healing (g) assays in ZEB1-expressing MDA-MB-231 and SUM-159 cells treated with palbociclib. Scale bars, 100 μm. **P < 0.01, ***P < 0.001 vs. respective control by unpaired Student’s t-test. h 3D outgrowth invasion assay in ZEB1-expressing MDA-MB-231 cells treated with palbociclib. Scale bars, 20 μm. **P < 0.01 vs. respective control by unpaired Student’s t-test. i, j Representative images and the quantification of lung nodules of BALB/c nude mice that were injected with ZEB1-expressing MDA-MB-231 cells and treated with palbociclib. Scale bars, 100 and 20 μm. *P < 0.05, **P < 0.01, ***P < 0.001 vs. respective controls by unpaired Student’s t-test.