Gefitinib-dose dependent effects on the phosphorylation of signaling molecules. (A) Experimental results. Phosphorylation of EGFR, Shc, MEK, and ERK in H1299EGFR-WT (green circle) and H1299L858R (red square) cells was measured after 5 minutes of EGF (10 nM) administration, in the presence of gefitinib at the indicated concentrations. Western blots were performed twice, and the mean values and the standard deviations of the signal intensities were calculated. The values were normalized for each protein so that the values without gefitinib were equal to 1. Representative western blot images are shown in Additional file 1, Figure S1C. (B) Simulation results. The gefitinib effect was simulated by multiplying the kinetic parameter of k3 (the rate constant for the forward reaction of EGFR phosphorylation). The multiplying coefficient α is given by α = (k3)/(k3 in EGFR-WT model or L858R models). The green circles (EGFR-WT model), red squares (L858R model A), and blue triangles (L858R model B) show concentrations of phosphorylated EGFR, Shc, MEK, and ERK at t = 5 minutes, plotted with several values of α (1, 0.5, 0.1, 0.05, 0.01, 0.005, 0.001 and 0.0005). α = 1 means without gefitinib.