CDRI-85/287 interferes with MEK/Erk activation to inhibit downstream signaling.

<p>(A) Western blot analysis to see the expression of p-MEK, MEK, p-Erk and Erk. MDA-MB 231 (left panel) and primary breast cancer cells (right panel) were treated with the indicated concentrations of CDRI-85/287 for 24h and 30 µg of whole cell lysate in each lane was probed for the expression of different proteins using specific antibodies. Representative blots are shown in the upper panel and densitometric quantitation of protein expression levels are shown as fold changes in the lower panel. Results are expressed as mean ± SEM, n = 3. p values are a-p<0.001, b<0.01, c-p<0.05 and d-p>0.05 vs. control. (B) Transcriptional activation of the AP-1 promoter complex in transiently transfected MDA-MB 231 cells in response to CDRI-85/287 alone or in the presence of 100 ng/ml EGF. MDA-MB 231 cells were transfected with pAP1-Luc or p-c-fos-Luc or p-c-jun-Luc reporter plasmids and incubated with various concentrations of compound for 24h. pRL-luc plasmid was used as internal control. Results are described as % of normalized relative luciferase unit (RLU). Results are expressed as mean ± SEM, n = 3. p values are a-p<0.001, b-p<0.01, c-p<0.05 and d-p>0.05 vs. control and e-p<0.001, f-p<0.01, g-p<0.05 and h-p>0.05 vs. EGF. (C) Effect of compound on AP-1 dependent proliferation markers. MDA-MB 231 (left panel) and primary breast cancer cells (right panel) were treated with the indicated concentrations of compound for 48h. 30 µg of whole cell lysate in each lane was probed for the expression of different proteins using specific antibodies. Representative blots are shown in the upper panel and densitometric quantitation of protein expression levels are shown as fold changes in the lower panel. Results are expressed as mean ± SEM, n = 3. p values are a-p<0.001, b<0.01, c-p<0.05 and d-p>0.05 vs. control.