Figure 5
Effect of gefitinib on EROD activity. A) EROD assay was performed in intact living cells untreated or treated for 24 h with 0.1 μM gefitinib in the absence or in the presence of 10 μM α-NAP. CYP1A1 activity was measured using 7-ethoxyresorufin as a substrate and expressed as pmol resorufin/mg protein/min. Values given are the means (± SD) of three independent determinations. It is of note that in the gefitinib-resistant cells both the basal and the induced EROD activity were undetectable (not shown) (*P < 0.05; **P < 0.01; ***P < 0.001). B) H322 cells were transfected with siRNA against CYP1A1 or a scrambled negative control with a final concentration of 30 nM for 48 h and then treated for 16 h with 0.1 μM gefitinib before EROD assay was performed. Values given are the means (± SD) of four independent determinations (***P < 0.001). C) EROD assay was performed in intact living cells untreated or treated for 24 h with 0.1 μM gefitinib, 0.1 μM erlotinib, 0.1 μM lapatinib 100 μg/ml cetuximab, 10 μM U0126, 10 μM PD98059, 1 μM Wortmannin, 100 nM RAD001 and 100 nM NVPBEZ235. CYP1A1 activity was measured using 7-ethoxyresorufin as a substrate and expressed as pmol resorufin/mg protein/min. Values given are the means (± SD) of three independent determinations. (**P < 0.01; ***P < 0.001).