Fig. 3: Targeting active-EGFR by HX103 in various cancer cells.

a Visualization of HX103 (5 µM, 30 min) uptake in cells by confocal microscopy. Scale bar, 50 µm. The uptake of HX103 in a panel of cells can be blocked by preincubation with gefitinib (50 µM). b Quantification of the probe’s mean fluorescence intensity in cell images from (a). Data represent average values ± SEM. n = 14 for H1975 and Hela (3 biological replicates × 4–5 pictures); n = 12 for HCC827 (3 biological replicates × 4 pictures); n = 15 for A549 and MCF-7 cells (3 biological replicates × 5 pictures); n = 9 for A431 (3 biological replicates × 3–4 pictures). The statistical P values were calculated by the two-tailed Student’s test. c–e Flow cytometric experiments were performed to quantify the labeling of HX103 (c), EGFR antibody (d), and pEGFR antibody (e) in cancer cells. Bar chart revealing normalized MFI in cells. For HX103 (5 µM, 30 min) and pEGFR, the MFI of HCC827 was set at 100% to normalize the MFI of the remaining cells. For EGFR, the MFI of A431 was set at 100% to normalize the remaining cells. Data represent average values ± SEM, n = 3 independent experiments; note that n = 4 independent experiments for PC-9 and A431 in (e). The statistical P values were calculated by the two-tailed Student’s test. f Bar chart showing the percentages of total EGFR that is mutant in cancer cells (Supplementary Fig. 14). Data represent average values ± SEM, n = 3 independent experiments. The statistical P values were calculated by the two-tailed Student’s test. g Bar chart showing percentage values of EGFR(+)pEGFR(+) or EGFR(+)HX103(+) in cells (Supplementary Figs. 9–11). Data represent average values ± SEM, n = 3 independent experiments. h Bar chart showing the percentage of EGFR(+)pEGFR(+) in the presence and absence of EGF or 10% FBS (Supplementary Figs. 16, 17). Data represent average values ± SEM, n = 3 independent experiments. The statistical P values were calculated by the two-tailed Student’s test. Source data are provided as a Source Data file.