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. Author manuscript; available in PMC: 2016 Mar 1.
Published in final edited form as: Biochim Biophys Acta. 2015 Jan 21;1849(3):317–327. doi: 10.1016/j.bbagrm.2015.01.003

Figure 5.

Figure 5

Effects of EGFR pathway inhibitors on EGF-induced human BCRP or p-CREB expression in IGROV1 cells. A. Effects of EGFR pathway inhibitors on EGF-induced BCRP promoter activity. Cells were transfected with pGL4 or pGL4/1285 respectively. After 24-hours of culture in complete medium, then overnight starvation in SFM, cells were pretreated with the EGFR selective inhibitor ZD1839 (10 μM), the PI3K inhibitor LY294002 (15 μM), or the MEK inhibitor PD98059 (30 μM) respectively for 1 hour, followed by treatment with EGF (50 ng/mL) for 6 hours. Then, BCRP promoter reporter gene activities were determined by measuring luciferase luminescence as described in Methods. B. Effects of EGFR pathway inhibitors on EGF-induced BCRP mRNA expression. Following treatment with EGF and inhibitors as described for (A) above, total mRNA was isolated and BCRP mRNA expression were determined by real-time qPCR and normalized for the expression of β-actin mRNA (B). Results shown in A and B represent the means ± S.D. from duplicate measurements and three independent experiments for promoter activity and mRNA, respectively. *, significantly (P<0.05) different vs. untreated by student's t-test. C. Effects of EGFR pathway inhibitors on EGF-induced p-CREB expression. Following treatment with inhibitors as described for (A) above, then 30 min treatment with EGF (50 ng/mL), the expression of p-CREB, CREB, and GAPDH (done as loading control) was determined by Western blot. Bands represent results typical for three independent experiments, done on different days.